iCoconut Genetic Resources Pons Batugal, V. Ramanatha Rao and Jeffrey Oliver, editors ii COCONUT GENETIC RESOURCES The International Plant Genetic Resources Institute (IPGRI) is an independent international scientific organization that seeks to improve the well-being of present and future generations of people by enhancing conservation and the deployment of agricultural biodiversity on farms and in forests. It is one of 15 Future Harvest Centres supported by the Consultative Group on International Agricultural Research (CGIAR), an association of public and private members who support efforts to mobilize cutting-edge science to reduce hunger and poverty, improve human nutrition and health, and protect the environment. IPGRI has its headquarters in Maccarese, near Rome, Italy, with offices in more than 20 other countries worldwide. The Institute operates through four programmes: Diversity for Livelihoods, Understanding and Managing Biodiversity, Global Partnerships, and Improving Livelihoods in Commodity-based Systems. The international status of IPGRI is conferred under an Establishment Agreement which, by January 2005, had been signed by the Governments of Algeria, Australia, Belgium, Benin, Bolivia, Brazil, Burkina Faso, Cameroon, Chile, China, Congo, Costa Rica, Côte d’Ivoire, Cyprus, Czech Republic, Denmark, Ecuador, Egypt, Greece, Guinea, Hungary, India, Indonesia, Iran, Israel, Italy, Jordan, Kenya, Malaysia, Mauritania, Morocco, Norway, Pakistan, Panama, Peru, Poland, Portugal, Romania, Russia, Senegal, Slovakia, Sudan, Switzerland, Syria, Tunisia, Turkey, Uganda and Ukraine. Financial support for IPGRI’s research is provided by more than 150 donors, including governments, private foundations and international organizations. For details of donors and research activities please see IPGRI’s Annual Reports, which are available in printed form on request from ipgri-publications@cgiar.org or from IPGRI’s Web site (www.ipgri.cgiar.org). The geographical designations employed and the presentation of material in this publication do not imply the expression of any opinion whatsoever on the part of IPGRI or the CGIAR concerning the legal status of any country, territory, city or area or its authorities, or concerning the delimitation of its frontiers or boundaries. Similarly, the views expressed are those of the authors and do not necessarily reflect the views of these organizations. Mention of a proprietary name does not constitute endorsement of the product and is given only for information. Citation: Batugal, P, V Ramanatha Rao and J Oliver, editors. 2005. Coconut Genetic Resources. International Plant Genetic Resources Institute – Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor DE, Malaysia. Cover pictures (clockwise from upper left corner): Dr Pons Batugal, COGENT Coordinator, admiring the PNG Brown Dwarf x Renell Island Tall hybrid produced by PNG’s Stewart Research Station; Vietnamese mother and child proudly showing seedling of local coconut variety they raised for planting; Mr Tiara Mataora, Senior Research Officer, Ministry of Agriculture, Cook Islands, admiring the typhoon- resistant local coconut cultivar; top view of the International Coconut Genebank for Africa and the Indian Ocean hosted by Côte d’Ivoire; Mr Lolo Fili, Researcher of Tonga, showing prized local coconut variety with high husk content. Pictures courtesy of Dr Roland Bourdeix. ISBN 92-9043-629-8 IPGRI-APO PO Box 236, UPM Post Office Serdang 43400 Selangor Darul Ehsan, Malaysia © International Plant Genetic Resources Institute, 2005 iii Contents Contents Foreword Introduction CHAPTER 1. An introduction to the coconut palm M Foale CHAPTER 2. Locating and collecting coconut germplasm Locating coconut genetic diversity V Ramanatha Rao, T Hodgkin and R Bourdeix Mapping of coconut genetic diversity R Bourdeix, L Guarino, PN Mathur and L Baudouin Status, gaps and strategy in coconut germplasm collecting R Bourdeix, L Guarino, V Ramanatha Rao and L Baudouin In vitro collecting of coconut germplasm F Engelmann CHAPTER 3. Germplasm conservation Complementary conservation of coconuts ME Dulloo, V Ramanatha Rao, F Engelmann and J Engels Coconut field genebank V Ramanatha Rao COGENT’s multi-site International Coconut Genebank P Batugal and K Jayashree The International Coconut Genebank for the South Pacific (Papua New Guinea) M Faure x xii 1 13 32 44 65 75 91 106 115 iv COCONUT GENETIC RESOURCES The International Coconut Genebank for South Asia (India) V Rajagopal The International Coconut Genebank for Southeast and East Asia (Indonesia) H Novarianto The International Coconut Genebank for Africa and the Indian Ocean (Côte d’Ivoire) JL Konan Proposal for the establishment of the International Coconut Genebank for Latin America and the Caribbean (Brazil) EA Tupinamba Status of cryopreservation research in coconut F Engelmann, B Malaurie, O N’Nan and M Borges In situ conservation of coconut diversity B Sthapit, V Ramanatha Rao and D Davis Poverty reduction in coconut growing communities: A strategy for coconut in situ/on-farm conservation P Batugal, J Oliver and K Jayashree Global coconut conservation strategy P Batugal and V Ramanatha Rao CHAPTER 4. Characterizing diversity Morphometric methods of determining diversity L Baudouin and G Santos Biochemical and molecular methods for characterizing coconut diversity P Lebrun, A Berger, T Hodgkin and L Baudouin 117 119 121 123 142 149 161 190 209 225 vCHAPTER 5. Germplasm use Conventional coconut breeding P Batugal and R Bourdeix Use of molecular markers for coconut improvement: Status and prospects L Baudouin, P Lebrun, F Rognon and E Ritter Breeding for drought tolerance in coconut: Status and potentials V Rajagopal, KV Kasturi Bai and N Kumar Performance of coconut hybrids in some countries of Asia, Africa and Latin America P Batugal Performance evaluation of coconut varieties and farmers’ varietal preferences P Rethinam, P Batugal and F Rognon Multilocation coconut hybrid trials in three African and three LAC countries P Batugal, JL Konan, A Sanaoussi, AK Kullaya, E Tupinamba, R Castillo and B Been Coconut micropropagation C Oropeza, E Rillo, V Hocher and JL Verdeil CHAPTER 6. Major pests and safe movement of germplasm Coconut lethal yellowing C Oropeza, JA Escamilla, G Mora, D Zizumbo and NA Harrison Status of coconut lethal yellowing in Jamaica B Been and W Myrie Indexing and pathogen characterization S J Eden-Green and AA Mpunami Strategies for safe movement of coconut germplasm M Diekmann Contents 251 268 282 302 309 326 334 349 364 371 390 vi COCONUT GENETIC RESOURCES Pest risk assessment of the International Coconut Genebank for Africa and Indian Ocean, and Latin America and Caribbean H de Franqueville Pest risk analysis and guidelines for the safe movement of germplasm in the International Coconut Genebank of Asia and the Pacific R Ikin CHAPTER 7. Information, public awareness, institutional support and partnerships The International Coconut Genetic Resources Database C Hamelin, R Bourdeix and L Baudouin Public awareness initiatives in coconut J Oliver and P Batugal Standardized catalogues of coconut germplasm: Catalogue of conserved germplasm and farmers’ varieties R Bourdeix and P Batugal Catalogue of high-value coconut products K Jayashree Catalogue of coconut food recipes Zulyana MN CGIAR’S support to coconut research G Persley The International Coconut Genetic Resources Network (COGENT): Its history and achievements P Batugal The Coconut Research for Development Programme (PROCORD) P Batugal and J Oliver 395 411 427 439 456 463 466 473 482 500 vii CHAPTER 8. COGENT’s regional network reports Research on coconut genetic resources in the South Pacific T Osborne Research on coconut genetic resources in South Asia G Kalloo Research on coconut genetic resources in Southeast and East Asia C Carpio, G Santos, E Emmanuel and H Novarianto Research on coconut genetic resources in Africa and the Indian Ocean A Kullaya and JL Konan Research on coconut genetic resources in Latin America and the Caribbean D Zizumbo, B Been, E Tupinamba, R Castillo and C Oropeza CHAPTER 9. Country reports on status of coconut genetic resources research South Asia India V Rajagopal, PM Kumaran, S Arulraj and V Arunachalam Sri Lanka AD Samarajeewa, CK Bandaranayake, CS Ranasinghe, JMDT Everard, LK Weerakoon, WMU Fernando and S Senarathne Bangladesh SA Bhuiyan, A Rashid, Md Nazirul Islam and BC Sarker Pakistan M Hashim Laghari and AH Solangi 513 524 533 546 557 573 581 596 604 Contents viii COCONUT GENETIC RESOURCES Southeast and East Asia Indonesia H Novarianto, RH Akuba, N Mashud, E Tenda and J Kumaunang Thailand P Naka and K Jayashree Vietnam VV Long Malaysia AW Fong, N Kalitu and K Jayashree Philippines C Carpio, E Manohar, E Rillo, C Cueto, O Orense, MB Areza-Ubaldo and AR Alfiler China L Tang and Z Ma Africa and the Indian Ocean Côte d’Ivoire JL Konan Ghana SK Dery, J Owusu Nipah and F Ofori Nigeria CO Okwuagwu Tanzania AK Kullaya Kenya FK Muniu and PK Kibet Mozambique JS Cumbi Seychelles AM Moustache 608 618 625 634 639 648 654 661 667 670 682 688 691 ix Latin America and the Caribbean Brazil EA Tupinambá, JF da Silva, Jr and WM Aragão Mexico R Gonzalez and C Oropeza Guyana O Homenauth Jamaica B Been South Pacific Cook Islands W Wigmore and T Mataora Fiji V Kumar and T Kete Kiribati M Tenang Papua New Guinea M Faure Samoa A Peters and K Jayashree Tonga P Taufatofua and K Jayashree Vanuatu JP Labouisse and J Lahva Index Contents 695 704 709 715 720 725 732 737 744 748 750 762 xCOCONUT GENETIC RESOURCES Foreword Coconut is an important crop for poor people, supporting their livelihoods and the sustainability of their environment. It is a source of materials for food, drink and shelter. As a fundamental element in the food system it provides essential nutrition to people in coconut-growing communities. It stabilizes farming systems, especially in fragile environments such as small island states, atolls, and in coastal zones. And coconut generates much needed income for small growers as well as employment and foreign currency earnings for their countries. Despite the enormous potential of the crop, coconut farmers in the past mostly scraped a living well below the poverty line. They were marginalized. About 96% of the farmers, who collectively grow coconuts on 12 million hectares worldwide, are smallholders tending less than four hectares. Many do not own the land they work, lack the resources to invest in technologies that would improve production, and are considered non-bankable by the formal banking sector. Despite its importance in the economies of many poor countries, the farmers who grow coconut lack a voice to influence government policy or private sector practices. To address these problems, the Consultative Group on International Agricultural Research (CGIAR) decided to include coconut in its research portfolio in 1991. Case studies had indicated that international research on coconut would generate high pay-offs for resource-poor coconut farmers. The International Board for Plant Genetic Resources (IBPGR), now the International Plant Genetic Resources Institute (IPGRI), organized the International Coconut Genetic Resources Network (COGENT) to implement this decision. COGENT started with 15 coconut- growing countries as members and subsequently expanded to 38 member countries. The substantive results of the work of the COGENT and its partner institutions across the developing and the developed world, reported in this book, are testimony to the correctness of the CGIAR’s vision and its implementation by IPGRI. The work has seen a tremendous advance in the scientific basis of coconut conservation and use. It has also generated substantial institutional and funding support from member countries. COGENT’s project on ‘Poverty reduction in coconut growing communities,’ which makes use of coconut diversity to improve livelihoods in 54 poor coconut-growing communities in 15 countries worldwide, is also described in this book. It illustrates the impact that research on coconut genetic resources generates for poor coconut farmers. It also describes the coconut conservation strategy that provides the way forward for this important area of work. xi Foreword At least in some communities, coconut farmers now have the resources they need – genetic, social, legal, political and financial – to make better use of coconut and hence improve life for themselves and their families. The task now is to improve matters for all coconut growers, worldwide. It is hoped that bringing together in this book the results of all COGENT’s work to date will serve as a benchmark for planning new initiatives to address the emerging challenges and opportunities that stem from the continuing erosion of coconut genetic resources, and the constant pressure on the research community to help resource-poor coconut farmers. Emile Frison Director General International Plant Genetic Resources Institute xii COCONUT GENETIC RESOURCES Introduction The coconut industry is at the crossroads. Coconut farmers are suffering due to decreasing farm productivity and income and unstable markets for copra, the traditional dried kernel product. This situation arisen in part by the competition from other vegetable oils and further aggravated by ageing and senile coconut palms, natural calamites such as drought, typhoons, incidence of pests and diseases, lack of resources to invest in innovations to improve yields and incomes and lack of availability of affordable high-yielding and adapted coconut varieties. Important coconut diversity, the basis for sustainable coconut production, is under threat of genetic erosion due to decreasing hectarage caused by urbanization, crop shifts, and natural and human-made calamities. Many governments cannot afford to conserve their diversity. Unless farmers earned more from their coconut, they could not afford to conserve their important varieties. Many have lost interest to plant coconuts and have shifted to other crops but have met the same fate due to lack of production resources. To help address the above problems, the International Plant Genetic Resources Institute (IPGRI) organized the International Coconut Genetic Resources Network (COGENT) in 1992. This network which started with 15 coconut producing countries as members has expanded to 38 members worldwide. In the last 14 years, COGENT has been fully operational with 38 coconut producing member countries in five regions (South Asia; Southeast and East Asia; South Pacific; Africa and the Indian Ocean, and Latin America and the Caribbean). It has developed and promoted the implementation of coconut collecting and conservation strategies. It has successfully developed, in collaboration with the Centre de Coopération Internationale en Recherche Agronomique pour le développement (CIRAD), the International Coconut Genetic Resources Database (CGRD), which has been disseminated to coconut breeders and curators worldwide. The CGRD contains passport and characterization data (morphometric and molecular marker) and some pictures of 1416 accessions which are conserved by national programmes in 28 sites in 23 countries. To further secure and provide access to conserved germplasm, the establishment of a COGENT multi-site International Coconut Genebank has been initiated to conserve 200 important accessions in each region. The identification, characterization and promotion of coconut varieties with multi-purpose uses have been initiated in farmers’ fields in 15 countries. The performances of high-yielding hybrids and farmers’ varietal preferences have been evaluated in nine countries and the results xiii Introduction will be used to help breeders in developing improved varieties. Further testing of 34 high-yielding hybrids were evaluated in multi-location trials involving three African and three Latin America/Caribbean countries to identify suitable varieties and hybrids for resource-poor farmers has been initiated. Diversity-linked income-generating activities are being used as a strategy to promote in situ and on-farm conservation and germplasm utilization in 15 countries. Protocols for in vitro embryo culture, germplasm collecting, cryopreservation, morphometric and molecular marker-based methods for locating and characterizing diversity; pest risk assessment and germplasm health management are being developed, tested and upgraded. Strategies and techniques for farmer participatory research, collecting, characterization and ex situ and in situ conservation are being refined. To strengthen the coconut research capability of COGENT member countries, COGENT/IPGRI has organized 43 country need assessment missions and conducted 45 workshops and meetings involving 1090 coconut researchers to share information and technologies, discuss issues and common problems and opportunities and how to address them; conducted 42 training courses involving 775 participants from 41 countries; supported 288 research and training/capacity building activities in 30 countries worth US$ 2.335 million. To promote the deployment of coconut genetic resource in a wider range of coconut R&D, COGENT led the establishment of the Global Coconut Research for Development Programme (PROCORD) to an appropriate platform. This publication describes the status of coconut genetic resources to date by documenting the work of COGENT and it partner institutions from both the developing and developed world. Chapter 1 provides an introduction of the coconut palm and describing its evolution, taxonomy, diversity and ethnobotany. Chapter 2 describes the strategies of locating diversity, the gaps in collecting and the developments on in vitro technique of collecting germplasm. Chapter 3 describes the strategies and initiatives in ex situ and in situ conservation and the global coconut conservation strategy. Chapter 3 describes the morphometric, biochemical and molecular methods of characterizing diversity. Chapter 5 describes the strategies and research on germplasm use, including the establishment of the International Coconut Genetic Resources Database, the use of conventional and molecular markers in coconut breeding, initiatives on breeding for drought tolerance, and the evaluation of coconut hybrids worldwide and the identification of farmers’ varietal preferences. Chapter 6 describes the status of the lethal yellowing disease and the search for disease resistant varieties, the techniques for indexing and pathogen characterization, the guidelines for safe germplasm movement and the xiv COCONUT GENETIC RESOURCES methods used in conducting pest risk assessment for COGENT’s multi- site International Coconut Genebank. Chapter 7 describes the history of the establishment of COGENT by the CGIAR, its achievements in the last 14 years and its public awareness strategy to maximize its impact. Chapter 8 describes the regional research and capacity building of COGENT’s five regional sub-networks while Chapter 9 contains the reports on the status of research on coconut genetic resources work in 28 COGENT member countries. This publication is the most comprehensive report on the status of coconut genetic resources to date. It is hoped that it will be useful to researchers, policy makers and development organizations as a basis for planning and implementing research and development initiatives to promote the conservation and use of coconut genetic resources. Some of the materials in this book will also be useful to students who choose to work on coconut genetic resources and their use. The editors xv Chapter 1 An introduction to the coconut palm xvi COCONUT GENETIC RESOURCES 1CHAPTER 1: An introduction to the coconut palm An introduction to the coconut palm M Foale Specialist in Coconut and Dryland Soils, Commonwealth Scientific and Industrial Research Organization (CSIRO), Sustainable Ecosystems, Long Pocket Labs, 120 Meiers Road, Indooroopilly, Queensland Australia Evolution of the coconut palm The evolutionary history of a species, which has been pressed since antiquity into the human economy, surviving as wild fragment populations in a minute number of locations, has to be necessarily highly speculative. Nevertheless, the separation of the coconut from a cluster of ancestral palms located on what later became the dispersing land fragments of the super-continent of Gondwana, and the subsequent development of a whole suite of unique and interesting features, excites the scientific imagination to wonder how nature could generate such an outcome (Foale 2004). Through a process of natural selection, over a period of perhaps 80 million years, the coconut developed the means to disperse across vast expanses of ocean and take hold firmly on the perilous boundary between land and sea, adapting to fierce windstorms and periodic inundation, thriving unassisted by any other fauna and flora, and delivering its fruit in turn to the ocean vehicle for further dispersal. The very components of the fruit that enabled this species to successfully conquer the open ocean and take firm hold of the land on arrival became for humans the source of nourishing food and drink, and raw materials for fuel and tools of many kinds ranging from ropes to cups and buttons. The sandy berm on the land edge of the strand (the zone between high and low ocean tides), built of sand by raging storm tides, retains little water or nutrients to support a seedling attempting to become established. The coconut evolved to defeat these environmental shortcomings by developing a huge endosperm, larger than found in any other plant with the exception of the ‘double coconut’ of Seychelles. There is sufficient energy and nutrients in the coconut seed to support growth of the seedling for more than one year, providing the opportunity for the roots to extend through the berm to the underlying soil layer that is bathed by the fluctuating fresh water table, which responds to the twice daily tidal rise and fall of the ocean. The coconut is most ‘at home’ in an environment where the roots are thus intermittently immersed in ground water, which has accumulated the essential plant nutrients released by the decay of plant residues. The endosperm or kernel of the coconut, evolved to enable the palm to colonise new habitats, also proved to be highly supportive of human colonisation of new habitats and a 2COCONUT GENETIC RESOURCES subsequent major support for the prosperity of human communities (Foale 2003; Foale and Ashburner 2003). The cohabitation of human and coconut populations ushered in a new era for the evolution of the coconut. Selection, especially for enhanced food and drink qualities, moved the coconut away from some of the critical traits that had enabled it to successfully disperse over a vast portion of the globe on a geological time scale. Before human arrival, the coconut had undoubtedly reached thousands of islands and mainland shore locations stretching over most of the tropical Pacific Ocean, throughout the islands of Southeast Asia, and probably to some shores of the Indian Ocean. It appears that the lack of an ocean current connection between the Indian and the Atlantic Oceans was responsible for the absence of the coconut on the west coast of Africa and the east coasts of South and Central America prior to its introduction from East Africa and India by the 16th century Portuguese navigators. Taxonomy The Cocos genus is mono-specific, showing recognition by taxonomists that it has no close botanical ‘relatives’. The palm known as the ‘double coconut’ or ‘coco de mer’ (coconut of the sea) produces a fruit that bears some slight resemblance to the coconut, however, it is many times larger and is very different for all other traits (morphology and phenology). The coconut is placed in the Arecaceae family (formerly Palmaceae) and the sub-family Cocoideae which has 27 genera and 600 species (Teulat et al. 2000). The coconut has 32 chromosomes (16 pairs) and may be divided into two distinct groups - Tall and Dwarf, with a further division within the Dwarf group between forms with either fragile or robust trunks. The Tall, in general, is allogamous(out-breeding) because even though the male and female flowers are located close together on each inflorescence, the pollen is shed before the females are ready for it. The fragile Dwarf is mostly autogamous because the male flowers shed pollen freely while the female flowers are receptive. The Dwarf group that possesses a robust trunk and is less precocious that the fragile Dwarf has flower behaviour similar to the Tall. There are no reports of incompatibility between any paired individuals or populations. Tall and fragile Dwarf palms cross readily to produce vigorous and fertile hybrids. Diversity The confinement in nature of the coconut palm close to the strand environment described in the evolution section has placed narrow limits upon its many morphological and some phenological characters. 3CHAPTER 1: An introduction to the coconut palm Successful dispersal depended upon a buoyant fruit that contained adequate kernel in the seed to ensure successful establishment. A variant that bore fruit possessing less husk might not survive lengthy immersion in the ocean without becoming waterlogged. Another variant that produced a smaller seed (nut) might contain insufficient water to last through a sea voyage to retain the vital amount needed to stimulate germination upon arrival. A thin-shelled variant might crack upon falling from the palm and lose its water, and so on. Certainly, it has been reported that some populations of putative wild coconut palms show remarkable physical similarities in fruit characters even though there is molecular evidence of wide evolutionary separation (Ashburner 1995). The aspects of coconut phenology (development) that contribute to the success of the coconut in its unique habitat include late germination (early germination would risk the seedling emerging from the seed before the fruit fell into the ocean) and late onset of flowering. Early flowering risks diversion of growth resources from the development of a sturdy trunk and adequate peripheral roots at the base of the trunk to withstand destructive wind. Besides these easily recognised morphological and phenological traits, there are many others that are not readily observable and that have been discovered only as a result of the movement of populations between diverse habitats. This reference to diversity of habitat is not a contradiction of the earlier assertion that the coconut is confined next to the strand. There is wide scope for variation in the shore environment already described with respect to climate – especially seasonality and quantity of rainfall, and exposure to strong wind; soil type – ranging across sands derived from coral, silica, volcanic ash or rocks and the effluent of streams and various biotic pressures especially from insects and micro-pathogens originating in adjacent vegetation systems. It has become clear during almost one hundred years of coconut research in different parts of the world that there is great diversity for tolerance to water deficit, which is one of the most important adaptations that could contribute to improved productivity. The factors mentioned in the previous paragraph have generated diversity in the coconut through natural selection but the genetic changes have not been accompanied by the appearance of any morphological markers that could assist the observer to know that a particular adaptation is present. In the early days of coconut research, awareness of such genetic diversity came about through the observation of exotic populations in new environments. A few examples will serve to illustrate the existence of ‘invisible’ adaptation in coconut. Recognition of hidden tolerance or partial avoidance of the Brontispa leaf beetle took place in Solomon Islands in 4COCONUT GENETIC RESOURCES the 1920s, when an introduced coconut population from the Federated Malay States was severely damaged in both nursery and early field stages of growth on plantations alongside the almost untouched indigenous population. In Vanuatu in the 1960s, exotic material succumbed fatally to a virus whose presence in the local Tall population was not in any way suspected, due to a lack of any observable symptoms. In Indonesia in the 1980s, many hybrids between West African Tall and Malayan Yellow Dwarf were attacked by a variant of the fungal pathogen Phytophthora palmivora that apparently did not harm local palm populations. Thanks to molecular techniques, it is now possible to associate specific important adaptations with molecular markers that can be accurately identified in the laboratory. This aspect of research technology has the potential to contribute very potently to the success of future breeding endeavours based upon the screening of the assemblages of genetic material that have been gathered under the auspices of International Coconut Genetic Resources Network (COGENT). Some useful definitions to describe different groupings of genetic material, prepared by Bourdeix et al., are presented in Annex 1. There are many other examples of diversity in coconut for which morphological markers are actually present, as it was those very markers that were the traits being sought by human selection. Much of the diversity generated by human intervention is within the distinctive Dwarf sub- group of the coconut. With the exception of the robust Dwarf known as Niu Leka, all Dwarfs are almost completely obligate self-pollinators, leading to the emergence of traits transmitted by recessive genes, particularly a number of orange and yellow colour variants. These colours are particularly useful in eliminating the few hybrid seedlings that arise from uncontrolled pollination of a Dwarf palm in the company of Tall neighbours. Brown and green Dwarf seedlings can be recognised with less accuracy relying on their growth rate as an indicator of dwarfness. In traditional coconut cultures, where the Dwarf is valued for domestic use because of the relative ease of harvesting fresh fruit and the convenience of a small fruit as a source of juice, the Dwarf has been assigned an important role in homes and home-gardens. Another result of human-managed diversity is the so-called niu vai or water coconut associated especially with Polynesian settlements. Whereas, the small-fruited Dwarf is seen as the preferred source of fruit for drinking in the settled environment of the village, the situation is quite different when provisioning is required for a sea voyage. Coastal peoples are invariably mariners, maintaining skills and boats in order to trade along the coast as well as to harvest from the sea, and being prepared for longer voyages in response to invasion or population 5CHAPTER 1: An introduction to the coconut palm pressure. Selection of a coconut fruit suited to such contingencies would have the objective of providing the best possible source of both food and drink on the voyage. An immature fruit, such as the typical ‘juice nut’ has a shelf life of just a few days without refrigeration and is therefore unsuitable for a long voyage. Both food and drink can be provided, with least space required in the sea-going craft (and least weight of ‘cargo’ relative to the amount of vital components), by means of large mature fruit possessing a thin husk. There is evidence of selection for large fruit in many parts of Asia and the nearby island nations, and there are remnants of isolated populations of large-fruited palms on scattered islands across the Pacific Ocean. This association of diversity with specific human needs will be dealt with in more detail in the ethnobotany section below. Ethnobotany This section overlaps somewhat with that on genetic diversity, dealing in more depth with the targeting of specific traits selected to serve for human purposes at the local level, and also in developing or expanding a promising wider market for a coconut product. Whereas, Dwarf populations are distinctive and uniform at the local level with respect to fruit morphology and colour, Tall populations generally look pretty much the same all over the world. Extremes of fruit size are noticeable but otherwise there is usually, within any group of palms, a great diversity of colour, fruit shape and size, whilst palm crown morphology is mostly a matter of age and nutritional status. Ethnobotanical traits (those sought by humans) are elusive in Tall palms because of the lack of markers already noted, and because of the predominant outbreeding behaviour. Unlike inbreeding species such as wheat, rice or sorghum for example, where the selection of a distinctive individual plant offers the opportunity for the traits of that plant to be preserved securely in its progeny, in the Tall coconut a valuable trait identified in a single palm can prove difficult to multiply because of the genetic diversity in which it is embedded. Preservation of traits is much simpler in the Dwarf, suggesting that effort might be made to assist in the transfer of particularly interesting traits such a special aroma, or edibility of the immature husk, from Tall to Dwarf. In general, the coconut is not rich in distinct ethnobotanical traits that are of great commercial value. The relative genetic uniformity of the species compared to most crop plants that have emerged from regions that encompass great biophysical diversity confines coconut to a narrow range of variants. Little variation in the qualities of the fibre, shell, kernel 6COCONUT GENETIC RESOURCES and water has been demonstrated. There is on the other hand great ethnic diversity in the ways that the various coconut resources are produced and used. It is ethno-industry more than ethno-botany that gives rise to the great range of products derived especially from the fibre and shell including exquisitely crafted works or art and many materials of great commercial value. Distribution The natural dispersal of the coconut prior to the colonisation of the entire planet by humans has been dealt with above, but the historic distribution that was limited by the ‘reach’ of viable seeds floating on the ocean has been enormously expanded in recent millennia. Expansion inland from the natural coastal ‘foothold’ has been responsible for a great increase in coconut populations as human populations grew, and there has also been, quite recently, expansion to new lands previously not colonised by the wild coconut. An interesting aspect of coconut distribution arises from the evidence of widespread ‘invasion’ of natural coconut habitats by human settlers. Many coconut populations exhibit a degree of shift from the characteristics of formerly wild coconut populations (attributed to introgression between two populations of different origin – the wild one and the introduced one) that would have taken many generations to achieve. Except for a small number of atolls, where human settlement apparently never took place, or where it was very recent indeed (as for example the Cocos (Keeling) Islands settled in 1827), coconut populations from Asia to the eastern Pacific show signs of introgression between wild and ‘domesticated’ variants. The degree of drift in fruit morphology away from the classical wild traits varies greatly, being most marked in some relatively isolated Polynesian islands such as Rennell in Solomon Islands and Rotuma in Fiji Islands. It is likely that the wild population of coconut palms before the arrival of the Polynesian colonisers (which is known to have been at least 1000 years ago) was quite small, as the rocky coasts of these two islands possess only a few sandy berms where wild palms would have been growing. The genetic traits of the palms planted by the colonisers therefore became dominant within a few generations through a combination of relatively little dilution by the wild genotype and purposeful selection by the people. The final ‘coconut-free zone’ of the earth, being the tropical shores of the Atlantic Ocean, was ‘invaded’ very early in the 16th century by the introduction of seeds from the Indian Ocean shores, brought there by Portuguese navigators returning from voyages in the Indian Ocean. From a base in the Cape Verde Islands (close to the coast of Senegal), seeds 7CHAPTER 1: An introduction to the coconut palm were distributed within a few decades to all the new colonies in the tropical Americas. However, there was already a coconut population on the Pacific coast of Central America with nut characters inconsistent with dispersal from the ocean. One could speculate that the seeds which founded that population had originally arrived by boat prior to the entry of European navigators into the Pacific, from a source in Polynesia. By the end of the 16th century, the coconut could be found throughout the tropics with one notable exception. The tropical coast of Australia was as yet unknown to European navigators, but even when it was charted – mostly in the early 19th century - there was no sign of the coconut palm. A tiny population was later found (in 1848) on an offshore island, during intense local mapping of the northeast coast and the barrier reef. The possible reasons for this strange absence could be attributed to a combination of the consumption of drifted fruit for food by both native people, and also by a particular species of native rat capable of opening a mature fruit with ease (Foale 2003). The coconut can now be found on practically every suitable tropical and sub-tropical coastline worldwide, and has been transported far from the coast in many regions, wherever rainfall is adequate and the altitude does not exceed 1000 m. Although one would expect a species whose native habitat is at the very edge of the ocean to be “ecologically challenged” elsewhere, the coconut has shown a remarkable capacity to thrive on soil textures that range from coarse sand, like that of the berm on the coast, to heavy-textured clay. Certain essential nutrients constrain success of the coconut by their absence, and particular mention should be made of chlorine because its importance to the coconut is less well known compared with the usual major nutrients. Whilst chlorine is generally abundant near the coast, there are many sub-coastal and inland areas where the leaching action of intense rainfall has reduced chlorine availability to the point where it limits coconut growth. In the era of great industrial use of the coconut, commencing in the middle of the 19th century, coconut distribution became much broader as plantations were successfully established on hundreds of thousands of hectares of land previously occupied by rainforest. Provided that the annual rainfall totalled at least 2000 mm, and the season of severe water deficit did not exceed three months duration, the coconut prospered. Many biotic and nutritional challenges were encountered away from the coast but in general, these were overcome and the plantations were productive and profitable for up to 100 years. It has been economic rather than ecological factors that have placed constraints on the distribution of the coconut in recent decades. 8COCONUT GENETIC RESOURCES References Ashburner, GR. 1995. Characterisation, collection and conservation of Cocos nucifera L. in the South Pacific. PhD thesis, The University of Melbourne. Foale, MA. 2003. The coconut odyssey: The bounteous possibilities of the Tree of Life. ACIAR Monograph No. 101, 132 p. Foale, MA. and RA Ashburner. 2003. The coconut palm. Pp. 35-40. In: Plantation crops. In: Series on the impact of biotechnology on agricultural crops. Indian Council for Agricultural Research, New Delhi and Howarth Press, New York. Teulat, B, C Aldam, R Trehin, P Lebrun, JHA Barker, GM Arnold, A Karp, L Baudouin and F Rognon. 2000. Analysis of genetic diversity in coconut (Cocos nucifera L.) populations from across the geographic range using sequence-tagged microsatellites (SSRs) And RFLPs. Theoretical Applied Genetics. 100: 764-771 Foale, M. 2004. The coconut palm: Origin and evolutionary history (in press). In: VL Chopra and KV Peter (eds). Handbook of industrial crops. Haworth Press, Haworth, UK. 9CHAPTER 1: An introduction to the coconut palm Annex 1. Useful definitions of terms and nomenclature by R Bourdeix, G Santos, JP Labouisse and L Baudouin Cultivar/Variety/Ecotype/Population/Variant Definitions: ‘True’ variety: from a strict botanical point of view, there are only three main varieties in coconut: • Tall : high height increment, spaced leaf scars, predominantly allogamous, late bearing. • Dwarf: reduced height, narrow spacing between leaf scars, pre- dominantly autogamous, precocious. • A few intermediate and other types of coconut, such as Niu leka Dwarf (Polynesia) and King Coconut of Sri Lanka. Cultivar: ‘Cultivated variety’ is defined as a group of individuals or plants having similar traits that can reproduce “true-to-type” in the natural (sexual) way from generation to generation. Ecotype: Individual plants or populations which survive as a distinct group through environmental selection and isolation and that is compa- rable with a taxonomic species. Comments: The focus here is on the words: survival, environment, diverse. In most cases, it seems to be difficult to qualify coconut cultivars as true ‘ecotype’. Some possible exceptions are cultivars found in atolls and other environmentally very particular conditions. Vanuatu Tall, for instance, the only cultivar resistant to the foliar decay virus, can be considered as a true ecotype (this is a special case where a pathogen is given the status of a major ecological influence, yet it is a factor that could be eliminated). Population and variant: This can be considered as similar in connota- tion and would refer to a group of individuals obtained from a cultivar. Population refers to any subgroup located in a restricted location, such as one island, atoll or continuous strip of coastline. Variant is narrower than that in the sense that members of the group exhibit a specific trait as stated below. Variant could be preferred for special botanical types which may be found in different cultivar: Makapuno/Kopyor/Coco Gras/Dikiri or Spicata. 10 COCONUT GENETIC RESOURCES Populations could denote minor geographical and/or phenotypic dif- ferentiation within a cultivar. (Population could also refer to palms in a location, whether they be highly heterozygous, as most Tall populations are, or homozygous, as in the self-pollinating Dwarfs). Unfortunately, experience shows that most non-scientific observers and stakeholders do not know or do not appreciate the term “Cultivar”. They frequently use the term ‘variety’ instead of ‘cultivar’. Even in many scientific papers, ‘variety’ remains and used in place of ‘cultivar’. The scientific community has to make an effort to be understood through better communication with the rest of the world. So assuming that the terms of ‘cultivar’ and ‘variety’ are mostly synonyms, the following examples can be proposed: “True” varieties: • Tall : rapid height increment, widely spaced leaf scars, predominantly allogamous, late flowering and bearing. • Fragile Dwarf: reduced height, narrow spacing between leaf scars, trunk diameter about 40% less that Tall, little of no basal bole, pre- dominantly autogamous, precocious (early flowering and bearing). • Robust Dwarf: Very low rate of trunk extension, trunk diameter and flowering behaviour similar to Tall, crown more compact than that of Tall or Fragile Dwarf Cultivar or Variety : West African Tall, Catigan Green Dwarf, and Vanuatu Tall which could also be considered an Ecotype. Population: WAT06 (West African Tall Ouidah from Benin) Variant: Makapuno (Philippines), Dikiri (Sri Lanka), Spicata (different countries), Nawasi (Sri Lanka), Nim (Thailand) Sub-population: Individual plant/palm 11 CHAPTER 1: An introduction to the coconut palm Chapter 2 Locating and collecting germplasm 12 COCONUT GENETIC RESOURCES 13 CHAPTER 2: Locating and collecting germplasm Locating coconut genetic diversity V Ramanatha Rao1, T Hodgkin2 and R Bourdeix3 1Senior Scientist, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia 2Principal Scientist, International Plant Genetic Resources Institute (IPGRI), Rome, Italy 3Coconut Breeder, Centre de cooperation internationale en recherché agronomique pour le developpement (CIRAD), Cedex 5, Montpellier, France Introduction Cocos is a genus in the family Arecaceae (Palmaceae), subfamily Cocoideae, which includes 27 genera and 600 species. Distributed mainly in coastal regions between 20° N and 20° S, from sea level to 1000 m asl, the coconut – Cocos nucifera L. (2n = 2x = 32) – the only species in the genus, is an important perennial tropical plantation crop with no known truly wild forms. The variability of local coconut types is reported to be highest in Southeast Asia (Whitehead 1976). However, it has not been possible to establish either a true centre of diversity or centre of origin for the species. These simple but basic factors are of great importance for understanding the extent and distribution of coconut genetic diversity and for locating useful variation. In 1992, the International Plant Genetic Resources Institute (IPGRI), with the endorsement of the Consultative Group on International Agricultural Research (CGIAR) and its donors, established the International Coconut Genetic Resources Network (COGENT) with the aim of promoting an international collaborative programme on the conservation and use of coconut genetic resources. Collecting, conserving, evaluating and enhancing coconut germplasm of member countries, and locating and characterizing genetic diversity using morphometric and molecular biology techniques, have been some of COGENT’s major concerns (http://www.ipgri.cgiar.org/networks/cogent). Under the auspices of COGENT, the activities related to genetic resources collection and genetic diversity in coconut have been streamlined and significant progress in these areas has been made. Genetic diversity Genetic diversity is usually thought of as the amount of genetic variability among individuals of a variety, or population of a species (Brown 1983). It results from the many genetic differences between individuals and may be manifest in differences in DNA sequence, in biochemical characteristics (e.g. in protein structure or isoenzyme properties), in physiological 14 COCONUT GENETIC RESOURCES properties (e.g. abiotic stress resistance or growth rate) or in morphological characters such as flower colour or plant form (Ramanatha Rao and Hodgkin 2001). Four components of genetic diversity can be usefully distinguished; the number of different forms (alleles) ultimately found in different populations (sometimes referred to as ‘richness’), their distribution (or ‘evenness’), the effect they have on performance (related to population density) and the overall distinctness between different populations. The variation that underpins genetic diversity arises from mutation and recombination. Selection, genetic drift and gene flow act on the alleles present in different populations to cause variation in them. The selection can be natural or it can be artificial, as is the case with much of the variation present in crop species (Suneson 1960; Frankel 1977; Nevo et al. 1984; Brown 1988; Hamrick et al. 1992). It allows species and populations to adapt to changing conditions and provides the basis for the observed differences between different ecotypes, populations or cultivars of coconut. It is generally accepted that the genetic variation in plant populations is structured in space and time (Loveless and Hamrick 1984). The description of the extent and distribution of the different aspects of genetic diversity in a species, and of the way in which it is structured, is an essential prerequisite to determining what to conserve, and where and how to conserve it. To date, most conservation efforts, either in situ or ex situ, have proceeded with little information on the genetic diversity that was being conserved and on what part of the total genetic diversity of a species this constituted. There is an urgent need to remedy this situation by describing the variation observed and identifying the factors likely to affect its distribution. Such factors often include climatic, edaphic and biotic ones as well as those specific to the populations (e.g. population size, selection) or to the species (e.g. ploidy, breeding system, linkage). Where data is available at the genetic level, e.g. from DNA or isozyme studies, direct measures of richness, evenness and distinctness may be obtained (Nei 1973). However, when dealing with morphological data, direct estimates of numbers and distributions of genes or alleles are hardly obtained and analyses of diversity are based on statistical parameters such as means, ranges, standard deviation and variance. Nonetheless, descriptions of morphological characteristics and reactions to pests and diseases of a population, local cultivar or accession remain the most useful information for plant breeders, agronomists and other users. Although these are often obtained in ways that make formal analyses of the extent and distribution of genetic diversity difficult, the information can often provide useful guidance on location of variation of particular characteristics. It can also be combined with other more formal analyses 15 CHAPTER 2: Locating and collecting germplasm to provide an overall view of the ways in which different components of diversity are distributed. Coconut genetic diversity: General considerations Coconut is one of the few major crop species that has no closely related wild relatives. Coconut belongs to the palm family (Palmae or Arecaceae), which has about 2800 species of 190 genera. The Cocoeae tribe with 27 genera and nearly 600 species includes several economically important plants such as Cocos nucifera (coconut), Elaeis guineensis (African oil palm), Attalea cohune (babacu) and Bactris gasipaes (peach palm). Palm species most related to the coconut palm are found in Colombia (Cook 1901). However, there appears to have been no possibility of mating or gene exchange with any related species and all coconut cultivars constitute a single potentially freely intermating genepool. Since coconut is an ancient species and has been under cultivation for several thousand years, it is reasonable to presume that early humans, while developing habitats in coastal areas, must have slowly domesticated any wild form that was present. Current theories mainly suggest that it must have originated in the Indonesian Islands and later spread to become pantropical, although the date of its spread to the Pacific has been under considerable debate (Harries 1990; 1995). It is also reasonable to presume that the spread of coconuts was based on small initial sample size, considering the bulk of the seed material. If the theory about its spread by flotation is true, then the sample size might be limited to one or two nuts and this is especially so in coconut populations found on the mostly uninhabited small islands and atolls. Thus, the bottleneck processes through which the coconut must have undergone through its world-wide spread, either human- assisted or otherwise, may well have resulted in considerable genetic drift in the founding populations. These observations have a significant bearing on the current genetic structure of the coconuts. In order to understand the process of spread, further studies on the historical and pre-historical knowledge of coconut are needed. Current knowledge appears to be weak in many countries and there has been no attempt to carry out a thorough check of the world literature relevant to this subject as suggested by Bourdeix et al. (1999). Although it has been generally agreed that humans must have domesticated the coconut in pre-historical time, it is not clear what was the domestication process involved and how the species evolved under domestication, as there are no ‘wild’ coconuts for comparison. The nature of selection pressure that farmers might have applied is difficult to comprehend taking account of the perennial character of the species. In many cases, there could be more than one human generation in the life 16 COCONUT GENETIC RESOURCES of a coconut palm. This leads to difficulties in trying to determine the farmer practices and their effect on the constitution and characteristics of local populations. This is further complicated by the fact that farmers are unlikely to have replaced individual coconut trees in their prime, let alone a whole orchard or population, with another crop or improved genotype, unless such substitution brought enormous benefits or they were forced to take such measures by external circumstances. This suggests that complex evolution of the different types or genotypes of coconut (with more and less desirable coconut types occurring together) may have been a quite common feature of coconut populations. This would lead to highly heterogeneous populations in which there could have been substantial. Thus the genetic structure of coconut could be fairly complex even if the frequent bottlenecks might result in populations with reduced genetic diversity in individual populations than what occurs in other perennial species that have undergone similar process of evolution, with less stringent bottlenecks. Guarino et al. (1998) suggested that the key features of coconut evolution might be summarized as follows: • Populations initially established by few individuals (founder effect, genetic bottleneck), but often from a variety of sources. • Low but continuing levels of gene migration among wild-type, feral or semi cultivated populations. • High levels of outcrossing within populations. • Selection by local communities, and movement of domesticated germplasm to Africa and the New World. • Continuing introgression of selected local varieties with wild-type populations and hybridization among domesticated varieties. The result, as revealed by genetic diversity studies using a range of morphological, physiological, agronomic, biochemical and DNA characters (see below), is that every region or large island has more or less distinctive populations, (commonly described as ecotypes). Tall ecotypes are highly variable (about 60% of the total diversity is found within Tall ecotypes in the Pacific), while Dwarfs are less variable, probably reflecting the fact that they are autogamous. The distinction between Tall and Dwarf types (which is really a difference in precocity) is sometimes formalised into botanical varieties typica and nana, but the taxonomic validity of this is not universally recognized. Although variation among ecotypes is basically continuous, regional Afroindian, Southeast Asian and Polynesian groupings can be recognized. Sub- groupings are also recognized within the Polynesian germplasm, in particular South Pacific, Northeast Pacific and Marquesas-Hawaii groupings, with the Rennell Island population relatively isolated. 17 CHAPTER 2: Locating and collecting germplasm Morphometric studies of diversity A description of the morphological characteristics and reaction to pests, diseases and stresses of an accession are the most useful information to plant breeders and other users. Such data (in conjunction with locality data) can be used in identifying especially diverse areas and those where specific traits can be found, and in exploring the relationship among accessions. However, using morphological characterization data for locating diversity has a number of limitations. Differential heritabilities, pleiotropic and epistatic effects, polygenic control and genotype x environment (G x E) interactions that are often associated with morphological characters can make estimation of genetic variation difficult. In many cases, long-term crossing and inheritance studies will be needed for precise estimation. There is also the problem that most genetic variation is hidden and is not apparent at the phenotypic level, so that morphologically similar material may in fact be genetically quite different. Despite these drawbacks, morphometric methods have been used to advantage in coconut as well as other crops (N’cho et al. 1993; Akpan 1994; Sugimura et al. 1997; Ashburner et al. 1997). The first publications comparing a large amount of data gathered from coconut accessions came from Africa. The description of most of the accessions from the collection at the Marc Delorme Station (Ivory Coast) has been reported in numerous publications (Nuce de Lamothe and Rognon 1977; Nuce de Lamothe and Wuidart 1979 and 1981; Le saint et al. 1983; Sangare et al. 1984; N’Cho et al. 1988). However, each of these publications produced in a series covered only a limited number of accessions, generally four to six, always compared with West African Tall (for Tall types) or Malayan Yellow Dwarf (for Dwarfs) used as reference controls. Using the same data, 17 Tall coconut ecotypes were assessed taking a biometric approach with the use of 24 major morphological descriptors. A discriminant analysis revealed the relations existing between ecotypes. The resulting dendrogram groups together accessions to the extent of their similarity into nine groups of 1 to 3 (N’cho et al. 1993). Sugimura (1997) carried out a genetic diversity study using botanical and agronomical traits on 39 cultivars of coconut palms which were mainly collected in the Philippines, and statistically analyzed to clarify the variation between and within cultivar groups (typica, nana and javanica). Although there were broad variations in all the traits except for several male flower characters, significant differences among the three cultivar groups were found in a dozen of traits. The variation within a cultivar group was higher in typica and javanica. Nana was noted as an aggregate group, which was distantly far from typica. Javanica was 18 COCONUT GENETIC RESOURCES characterized as the intermediate group having overlapping boundaries with other groups. As noted earlier, although these are not valid taxonomic classes, they seem to be useful for morphological groupings of cultivars. Zizumbo-Villarreal (1998) studied the pattern of morphological variation of coconut in Mexico. They analyzed 41 populations using 17 morphological fruit characters. Principal component and cluster analyses indicated four main groups of coconut populations that showed high similarity with four different genotypes recently imported into Mexico from areas that could be the origin of Mexican coconut populations. These four genotypes were evaluated with regard to lethal yellowing disease in Jamaica and showed a differential susceptibility. Based on the difference in susceptibility of the Mexican genotypes, the analysis of correlation between morphological and geographical distances showed a high positive correlation that supports: 1) historical evidence that indicates early introductions of coconut from different regions of the world, and 2) that on both coasts of Mexico two different patterns of dispersal were involved - continuous and in jumps. It was concluded that collectively these results suggest that the impact of the lethal yellowing disease on coconut populations will vary depending on the specific area and the origin of its coconuts, although it is not very clear how this conclusion could be drawn. This will require some level of follow up. Vargas (2000) evaluated Tall coconut cultivars from the Pacific coast of Costa Rica and the Philippines (San Ramón, Tagnanan and Laguna), for fruit characteristics. Most of the introduced cultivars showed extremely large heterogeneity. A cluster analysis, based on the Ward method (Ward and Neel 1970), classified the palms into four groups with high internal homogeneity. Some of the evaluated coconut palms from the Costa Rican Pacific area had nut characteristics similar to San Ramon (Group 1: large and elongated nuts) and Tagnanan palm (Group 4: heaviest fruits and nuts) groups but not with the Laguna group (Group 3: rounded and small-sized nuts). At the association level used (semipartial R2 = 0.10), another group (Group 2: small size and mildly elongated nuts) that included the remaining palms sampled from the Costa Rican Pacific Coast (Group 2: small-sized mildly elongated nuts) was constituted, thus showing that the Costa Rican types were different from the established cultivars (for detailed treatment, see Baudouin and Santos, Chapter 4). Use of Isozymes This method of genetic variability evaluation is barely developed for coconut. The initial study, undertaken with pollen involved nine enzyme systems (Benoit and Ghesquiere 1984; 1989). After several technical difficulties, only four systems were used to compare eight ecotypes: 19 CHAPTER 2: Locating and collecting germplasm Malayan Yellow Dwarf (MYD); Cameroon Red Dwarf (CRD); Pumilla Green Dwarf (PGD); Niu Leka Dwarf (NLA); West African Tall (WAT); Malayan Tall (MLT); Tahiti Tall (TAT) and Vanuatu Tall (VTT). The eight ecotypes showed a weak enzyme polymorphism, few polymorphic loci per system, and never more than two alleles per locus. The intra-ecotype variability was low for autogamous Dwarfs, higher for the Niu-Leka Dwarf and the Talls, with the exception of the West African Tall, which was monomorphic for the four enzyme systems tested. The low enzyme polymorphism of coconut contrasts with the morphologic diversity within the species and suggests that the marked phenotypic differences could hide homologous genetic structures. The apparent absence of variability in WAT is possibly due to successive bottlenecks in its spread that have led to a high level of consanguinity. Other studies of patterns of isozyme variation were also conducted in Sri Lanka (Fernando 1995) and Indonesia (Asmono et al. 1993) with rather similar results. Villareal et al. (2002) studied the diversity of 22 populations of Mexican coconut and six imported populations using 15 enzymatic systems and the allele frequencies in: peroxides, endopeptidase, glucose 6-phosphate dehydrogenase. They observed very low polymorphism, not more than two alleles per locus. The Wright fixations indices, F(it)= 0.62, F(is) = 0.40 and F(st) =.036, indicated low total heterozygosity and low heterozygosity within populations suggesting inbreeding and genetic drift and a high diversity among populations due to differentiation between Pacific and Gulf of Mexico coastal populations. The phylogenetic tree with values for genetic distance, indicated three groups on the Pacific coast related to Rennell Tall and Polynesian Tall, and two groups on the coast of the Gulf of Mexico, one related to the West African Tall and the other to Mexican Pacific coast populations. This corroborated historical antecedents and morphological and physiological patterns. The Dwarf coconuts were related to the Pacific Tall populations, Rennell Tall and Polynesian Tall. There was no difference between local and imported Dwarf populations. Cardeña et al. (1998) determined electrophoretic patterns of leaf peroxidases, endopeptidases, and Coomassie blue stained proteins were analyzed in four cultivars (West African Tall, Rennell Tall, Malayan Yellow Dwarf, Cameroon Red Dwarf), and in the hybrids PB121 (MYD x WAT) and PB111 (CRD x WAT). Polymorphisms were detected for the expression of two alleles of a dimeric peroxidase, two alleles of monomeric endopeptidase, and a pair of active null alleles of a dimeric peroxidase, two alleles of Coomassie blue stained protein. Four distinctive genotypes were identified, one for each of the Tall cultivars, another for both of the Dwarf cultivars, and the last for both of the hybrids. 20 COCONUT GENETIC RESOURCES Use of polyphenols The analysis of the polymorphism based on the analysis of leaf polyphenol using High Performance Liquid Chromatography (HPLC) provided an original approach to the study of genetic diversity in numerous plant species. The first analysis on coconut has involved the measurement of 16 sufficiently individualized peaks or major items of chromatographic information, each corresponding to a molecule or a few molecules of strong structural affinity (Jay et al. 1989). From 32 ecotypes, 171 palms were sampled in the collection of the Marc Delorme Station in Côte d’Ivoire. The data were subjected to multivariate analysis. The first discriminant analysis showed a clear distinction between Dwarfs and Talls. Only 19 out of 171 individual palms showed atypical behaviour. Certain Tall trees of various ecotypes behaved like Dwarfs: AGT, MLT, RGT, TAGT, RIT, TAT, WAT, PNTO1; while one NLAD tree behaved like a Tall. Most Dwarfs presented common characteristics that clearly distinguished them from Talls as shown in the morphologic and polyphenol analyses. The second analysis consisted of a canonical analysis per ecotype. Data analysis favoured the differences between ecotypes at the expense of intra-ecotype variability. Classification within this analysis was not based on geographic groups; the image obtained, however, permits such an interpretation. Five groups were recognized to classify the collection of the Marc Delorme station: Pacific, Far East, Indian Ocean, Africa and America, the last one being represented by only one ecotype. Among the Tall ecotypes, the representation permitted the determination of three distinct groups corresponding to the Pacific, the Far East and Africa. The ecotypes of the Indian Ocean may be divided between the African and the Far East groups. Certain points precisely strengthen the historical hypothesis. The Ghana Yellow Dwarf (GYD) and Malayan Yellow Dwarf (MYD) are very close, confirming the old hypothesis that the Yellow Dwarf was introduced from Malaysia into Africa during the time of the British colonial rule. Anyway, with the advent of DNA marker technology, the characterization of genetic diversity in coconut germplasm at the DNA level has recently begun to substitute other strategies like isozyme or leaf polyphenol analysis. Molecular studies of diversity The use of molecular techniques in studying genetic diversity in recent years has contributed to better understanding of the genetic diversity of some species (Karp 2002; Hodgkin et al. 2001). The increase in the use of molecular techniques in genetic diversity studies is based on the facts that: 21 CHAPTER 2: Locating and collecting germplasm • Appropriate molecular markers can provide direct estimates of gene and allele frequencies and can detect whether plants are homozygous or heterozygous for given markers; • Molecular techniques make it possible to analyze numerous and independent characters, whereas morphological analysis provides fewer characters, often of dubious homology; • Morphology is prone to considerable convergence while most DNA regions are less so and even if there is some convergence, the genetic basis of convergence in molecules is better understood; and • Molecular markers are relatively independent of the environment (Beckmann and Soller 1986). It has been argued that molecular markers provide a particularly powerful approach to understanding patterns of distribution of genetic diversity that can be used to adjust collecting, evaluating and breeding strategies so as to obtain maximum variation from any given wild population (Morikawa and Leggett 1990). However, it has also been noted that molecular methods should not be used on their own. Thus, Ashburner (1994) emphasized that DNA analysis should not replace currently used characterization methods, but should be used as adjunct when formulating conservation and crossing strategies. Analysis of data can distinguish similarities or differences between coconut populations and thus can be used to prevent duplication in conservation blocks and crossing programmes. However, if two populations appear similar, major adaptive genes may still exist and these may not be picked up by molecular studies. Therefore, collecting priorities should still take account of the need to sample unique environments. Where differences are detected by molecular techniques, there is a greater probability of the presence of different genes resulting from genetic drift, and priority should also be given to their collection. The information from molecular marker studies can also help improve utilization of diversity in coconuts. The data can assist in setting priorities for crossing programmes allowing breeders to maximize genetic distance and take advantage of any heterosis that may occur. Markers can also be used to tag important genes and allow the use of marker-assisted selection. For details on use of molecular markers, see Lebrun et al., Chapter 4. Improving location of diversity A molecular marker kit for COGENT partners Sampling, collecting and maintaining coconuts have always raised substantial logistical problems. The development of in vitro collecting 22 COCONUT GENETIC RESOURCES techniques helps deal with the physical problems of collecting large nuts but the logistical requirements still remain labour intensive and expensive. Currently, fruit component analysis coupled with observing a few other characteristics at the time of collecting are used to get some idea on the population variability at the time of collecting (see Bourdeix et al. in Chapter 2). However, this approach does not really give a measure of the genetic diversity that is being sampled. It was argued that molecular methods based on field collected tissue samples (Adams 1992) provided an efficient way of optimising the diversity collected and minimizing the numbers of new samples that had to be maintained in field gene banks. For this reason, over the last few years, COGENT and a number of other donors have supported the development of a molecular marker kit for coconut. The Bureau for the Development of Research on Tropical Perennial Oil Crops (BUROTROP) and IPGRI/COGENT supported the research by Centre de Cooperation Internationale en Researche Agronomique pour de Developpment (CIRAD) France, with participation from IACR Long Ashton (UK), on developing a microsatellite marker kit and dedicated software for developing countries. As a result, the kit, consisting of 14 microsatellite loci, was developed and tested on 681 coconut palms representing a large range of diversity. A statistical method was devised to identify any small set of individual palms of the same, unknown origin. The method allows the user to compare this sample with a set of reference populations and to rank these populations in order of decreasing probabilities of being the origin of the sample. It is a very efficient tool for diversity studies and identification of germplasm accessions. The transfer of this technology to the countries where the coconut germplasm collections are located will improve efficiency and reduce the cost of conserving, characterizing, managing and utilizing germplasm accessions for breeding improved varieties (see also Chapter 4 by Lebrun et al.). To downstream this technology to developing countries, 18 trainees from Brazil, India, Indonesia, Papua New Guinea, Mexico, Côte d’Ivoire, the Philippines, Portugal and Tanzania participated in a workshop on “Coconut Genetic Resources Management Using a Microsatellite Kit and Dedicated Software” held at CIRAD in Montpellier, France on 15-24 April 2002. Specialists from CIRAD managed the workshop while other specialists from partner institutions, consisting of nine molecular biologists and nine collection managers (representing a team of two participants per country) participated in the activity. The workshop was supported by IPGRI/COGENT, Common Fund for Commodities (CFC), the European Union, BUROTROP and CIRAD. Thus, there is now a tool kit available for estimating the genetic diversity prior to collecting to facilitate the locating of germplasm and 23 CHAPTER 2: Locating and collecting germplasm make appropriate conservation decisions including the identification of on-farm conservation sites. Using GIS tools A Geographic Information System (GIS) may be defined as a database management system which can simultaneously handle spatial data in graphics form - i.e., maps, or the ‘where’ - and related, logically-attached, non-spatial, attribute data - i.e. the labels and descriptions of the different areas within a map, or the ‘what’ (Guarino et al. 2001). It is a tool for managing information of any kind according to where it is located (Treweek 1999). The main elements of a GIS are as follows (Guarino 1995; Guarino et al. 1999): • Data input, verification and editing • Data storage, retrieval and management • Data manipulation and analysis • Output If we have georeferenced information on some level of genetic diversity of coconut based on the characterization and evaluation, including molecular evaluation of the available genetic resources, it would be possible to predict where additional genetic diversity could exist using GIS tools. GIS tools will not be able to measure genetic diversity but will be able to help locate new areas where coconut diversity might exist or the areas for extension of coconut cultivation. This is somewhat a refined way of using pre-existing information. To support this type of analysis, IPGRI and the International Potato Centre (CIP) have collaborated in the development of a software called DIVA-GIS, which calculates diversity indices for all the cells in a user-defined grid given latitude, longitude and characterization data for a set of accessions, and maps the results. They have recently trained a number of plant genetic resources (PGR) workers using this technology. It is expected that new areas of coconut genetic diversity would be located using this technology in the near future. Preliminary studies, using existing data in the Coconut Genetic Resources Database (CGRD) and the specialized GIS tools (FloraMap and DIVA- GIS), have been carried out to map the diversity collected, from different COGENT member countries as well as for diversity analysis for certain important morphological traits and for prediction of similar sites where similar diversity may exist or the sites for coconut cultivation (Prem Mathur 2003, pers. comm). Using these GIS tools, one can also generate climatic database for individual collecting sites and the climatic grids for temperature, precipitation and elevation. Some of the examples are presented in Figures 1 and 2. 24 COCONUT GENETIC RESOURCES Figure 1. Mapping of major coconut cultivation areas, coconut collecting sites and gaps identification in coconut collections in Vietnam 25 CHAPTER 2: Locating and collecting germplasm Figure 2. Mapping coconut diversity for nut weight in the Philippines 26 COCONUT GENETIC RESOURCES Figure 1 presents the mapping of major coconut growing areas and the coconut collecting sites in Vietnam, from which one can easily visualize where the gaps are in their coconut collections and can plan accordingly for more collections from those areas, which have not been surveyed earlier. In Figure 2, using GIS tools to calculate trait location-specific diversity, enabled the identification of sites with high diversity grids where recollecting could be done. Additional historical information on the movement of people, especially ethnic minorities, could provide additional information on genetic diversity as agricultural practices followed (including farmers’ selection) are closely linked to ethnic origins of a community. Quite often, the ethnic composition of the population is a very important factor to be taken in account for locating diversity. For instance, the islands of Rennell, Bellonna and Rotuma are the only ‘Polynesian’ Islands of the ‘Melanesian’ archipelago (Solomon and Fiji) and these islands have provided very important coconut varieties. Bourdeix et al. (1999) recommended that the Farmer Participatory Method (FPM) be used by following a grid based not only on the geographical aspect but also the ethnic aspect. For example, People of Ko Samui Island in Thailand came long time ago from Hainan (China) which is famous for its coconuts and it would be possible that this community in Thailand could be maintaining coconut growing tradition. Additional survey or FPM should be conducted also in Ko Samui Island. Conclusion Locating, maintaining and using genetic diversity of coconut present substantial challenges given the wide dispersal of the species, the limited knowledge of the history of that dispersion and of the current extent and distribution of diversity. The logistical problems conservers and users face when dealing with a perennial species with large recalcitrant seeds added to the complexity of managing coconut germplasm. However, in recent years, substantial progress has been made, at least in part, through the strong support of COGENT partners by establishing an effective framework of knowledge on which to base their activities. Certain general features of the species seem to be important in understanding the picture that is emerging from recent studies. These include the lack of a related wild genepool, small founding populations, human involvement in the selection and spread of the species/cultivars, outbreeding and intercrossing among populations of Talls and low but continuing gene migration among wild type or distant populations. These characteristics provide a general framework for analyzing the data that 27 CHAPTER 2: Locating and collecting germplasm are currently coming from molecular studies. Clear differences are emerging between groups of ecotypes and populations from different areas and expected patterns of migration and transfer are being better described and understood. Further, detailed studies are needed, particularly in high diversity areas. These should focus on ecogeographic aspects of the distribution of diversity and on the location of populations and ecotypes with unique useful traits such as resistance to biotic and abiotic stresses. The use of general, commonly agreed procedures that COGENT has developed and made available will be important to maximize the value of this new information for users and to safeguard the resources needed by poor farmers who still depend for coconut for much of their livelihood. References Adams, RP, N Do and C Ge-lin. 1992. Preservation of DNA in plant specimens from tropical species by desiccation. Pp. 153-181 In: RP Adams and JE Adams (eds). Conservation of plant genes. In: DNA banking and in vitro biotechnology. Academic Press Inc., San Diego, USA. Akpan, EEJ. 1994. Evaluation of tall coconut genotypes within Nigerian coconut germplasm bank. Oleagineux 49:13-20. Antonovics, J. 1968. Evolution in closely adjacent plant populations. VI. Manifold effects of gene flow. Heredity 23:507-524. Ashburner, G.R. and W. Rohde. 1994. Coconut germplasm characterization using DNA marker technology. Pp. 44-46 In: MA Foale and PW Lynch (eds). Coconut improvement in the South Pacific. Proceedings of a workshop held in Taveuni, Fiji Islands. 10-12 November 1993. ACIAR, Canberra. Asmono, D, A Hartana, E Guhardja and S Yahya. 1993. Genetic diversity and similarity of 35 coconut populations based on isoenzyme banding pattern analysis. Buletin Pusat Penelitian Kepala Sawit 1, 1: 39-54. Beckmann, JS and M Soller. 1986. Restriction fragment length polymorphisms and genetic improvement of agricultural species. Euphytica 35: 111-124. Benoit, H. 1979. Isozymic variation within and between coconut populations. Paper presented during the 50th session of the FAO Technical Working Party on coconut production, protection and processing, Manila, 3-8 December 1979. Benoit H. and M Ghesquière. 1984. Electrophorèse, compte rendu cocotier. IV. Déterminisme génétique. Rapport interne IRHO, CIRAD, (FRA). 11 p. Bourdeix, R, L Baudouin, J Ollivier and JP Labouisse. 1999. Consultancy 28 COCONUT GENETIC RESOURCES report on coconut collecting strategies submitted to COGENT/IPGRI. IPGRI, Rome. Brown, AHD. 1988. The genetic diversity of germplasm collections. Pp. 9-11 In: B Fraleigh (ed). Proceedings of a Workshop on the Genetic Evaluation of Plant Genetic Resources, Toronto, Canada. Research Branch, Agriculture Canada, Toronto. Brown, WL. 1983 Genetic diversity and genetic vulnerability: An appraisal. Economic Botany 37(1): 4-12. Cardeña, R, C Oropeza and D Zizumbo. 1998. Leaf proteins as markers useful in the genetic improvement of coconut palms. Euphytica 102: 81-86. Cook, OF. 1901. The origin and distribution of the coca palm. Contributions from the US National Herbarium 7: 257-293. Ferguson, ME, BV Ford-Lloyd, LD Robertson, N Maxted and HJ Newbury. 1998. Mapping the geographical distribution of genetic variation in the genus Lens for the enhanced conservation of plant genetic diversity. Molecular Ecology 7:1743-1755. Fernando, WMU. 1995. Patterns of isozyme variation in Cocos nucifera L. Proceedings of Annual Sessions of Sri Lanka Assoc Advancement Science. 51 : 84-86. Frankel, OH. 1977. Natural variation and its conservation. Pp. 21-44. In: A Muhammed, R Aksel and RC von Borstel (eds). Genetic diversity in plants. Plenum Press, New York. Guarino, L, V Ramanatha Rao and P Batugal. 1998. Collecting coconut genetic diversity: Elements of a strategy for COGENT germplasm collecting activities. Paper presented at COGENT meeting, Kuala Lumpur, Malaysia. Guarino, L. 1995. Geographic information systems and remote sensing for the plant germplasm collector. Pp 315-328. In: L Guarino, V Ramanatha Rao and R Reid (eds). Collecting plant genetic diversity. Technical guidelines. CAB International, Wallingford, U.K. Guarino, L, N Maxted and M Sawkins. 1999. Analysis of geo-referenced data and the conservation and use of plant genetic resources. Pp. 1- 24. In: SL Greene and L Guarino (eds). Linking genetic resources and geography: Emerging strategies for conserving and using crop biodiversity. CSSA Special Publication No. 27. ASA and CSSA, Madison, U.S.A. Guarino, L, A Jarvis, RJ Himans and N Maxted. 2001. Geographic Information Systems (GIS) and conservation and use of plant genetic resources. Pp. 387-404. In: JMM Engels, VRamanatha Rao, AHD Brown and MT Jackson (eds). Managing plant genetic diversity. CABI and IPGRI, Wallingford and Rome. 29 CHAPTER 2: Locating and collecting germplasm Hamrick, JL and MJW Godt. 1990. Allozyme diversity in plant species. Pp. 43-63. In: AHD Brown, MT Clegg, AL Kahler and BS Weir (eds). Plant population genetics, breeding and genetic resources. Sinauer Associates Inc., Sunderland. Hamrick, JL, MJW Godt and SL Sherman-Broyles. 1992. Factors influencing levels of genetic diversity in woody plant species. New Forests 6:95-124. Hamrick, JL and MD Loveless. 1986. The influence of seed dispersal mechanisms on the genetic structure of plant populations. Pp. 211- 223. In A Estrada and TH Fleming (eds). Frugivores and seed dispersal. Dr W Junk, Dordrecht. Harries, HC. 1990. Malesian origin for a domestic Cocos nucifera. Pp. 351- 357. In: P Baas, K Kalkamn and R Geesink (eds). The plant diversity of Malesia. Kluwer Academic Publishers. Amsterdam. Harries, HC. 1995. Coconut. Pp. 389-394. In: J Smartt and NW Simmonds (eds). Evolution of crop plants. Longman, Harlow. Hodgkin, T, R Roviglioni, MC de Vicente and N Dudnik. 2001. Molecular methods in the conservation and use of plant genetic resources. Acta Horticulturae 546: 107-118. Jain, SK and AD Bradshaw. 1966. Evolutionary divergence among adjacent populations. Heredity 21:407-441. Jay, M., R Bourdeix, F Potier and C Sanlaville. 1989. Initial results from the study of the polymorphism of coconut polyphenols. Oléagineux 44: 151-161. Karp, A. 2002. The new genetic era: Will it help us in managing genetic diversity? Pp. 43-56. In: JMM Engels, V Ramanatha Rao, AHD Brown and MT Jackson (eds). Managing plant genetic diversity. CAB International and IPGRI, Wallingford and Rome. Le Saint, JP, M de Nuce de Lamothe and A Sangare. 1983. The dwarf coconut palms at Port Bouet (Ivory Coast) II. Sri Lanka Green Dwarf, and additional information about Malayan Yellow and Red Dwarfs, Equatorial Guinea Green Dwarf and Cameroon Red Dwarf. Oléagineux 38: 595- 606. Loveless, MD and JL Hamrick. 1984. Ecological determinants of genetic structure in plant populations. Annual review of ecology and systematics 15: 65-96. Miller, JC and SD Tanksley. 1990. RFLP analysis of phylogenetic relationships and genetic variation in the genus Lycopersicon. Theoretical and Applied 80(4): 437-448. Moss, H and L Guarino. 1995. Gathering and recording data in the field. Pp. 367-417. In: L Guarino, V Ramanatha Rao and R Reid (eds). Collecting plant genetic diversity. CAB International, Wallingford. 30 COCONUT GENETIC RESOURCES Morikawa, T and JM Leggett. 1990. Isozyme polymorphism in natural populations of Avena canariensis from the Canary Islands. Heredity 64: 403-411. Nabhan, GP. 1991. Wild Phaseolus ecogeography in the Sierra Madre Occidental, Mexico. Systematic and ecogeographic studies of crop genepools No. 5. IBPGR, Rome, Italy. N’cho, YP, JP Le Saint and A Sangare. 1988. The dwarf coconut palms at Port Bouet (Ivory Coast) III. New Guinea Brown Dwarf, Thailand Green Dwarf, Polynesia Red Dwarf. Oléagineux 43: 55-66. N’cho, YP, A Sangare , R Bourdeix, F Bonnot and L Baudouin. 1993. Assessment of a few coconut ecotypes a biometric approach 1. Study of tall populations. Oléagineux 48: 121-132. Nei, M. 1973. Analysis of gene diversity in subdivided populations. Proceedings of the National Academy of Sciences, USA 70:3321-3323. Nei, M. 1977. F-statistics and analysis of genetic diversity in subdivided populations. Annals of Human Genetics 41: 225-233. Nevo, E, A Beiles and R Ben-Shlomo. 1984. The evolutionary significance of genetic diversity: ecological, demographic and life history correlates. Lecture notes on biomathematics 53: 13-213. Nevo E & Beiles A (1989) Genetic diversity of wild emmer wheat in Israel and Turkey. Theoretical and Applied Genetics 77: 421-455. Nuce de Lamothe, MW de and F Rognon. 1977. The dwarf coconut palms at Port Bouet (Ivory Coast) I. Ghana Yellow Dwarf, Malayan Red Dwarf, Equatorial Guinea Green Dwarf and Cameroon Red Dwarf. Oléagineux 32 : 367- 375. Nuce de Lamothe, MW de and W Wuidart. 1979. The tall coconut palms at Port Bouet (Ivory Coast) I. West African Tall, Mozambique Tall, Tahiti Tall, Malayan Tall. Oléagineux 34 : 339-349. Nuce de Lamothe, MW de and W Wuidart. 1981. The tall coconut palms at Port Bouet (Ivory Coast) II. Rennel Tall, Solomon tall, Thailand tall, Vanuatu Tall. Oléagineux 36 : 353-365. Pickersgill, B. 1984. Migrations of chili peppers, Capsicum spp., in the Americas. Pp. 105-123. In D Stone (ed) Pre-Columbian plant migration. Papers of the Peabody Museum of Archaeology and Ethnology. Vol. 76. Harvard University Press, Boston, U.S.A. Ramanatha Rao, V and T Hodgkin. 2001. Genetic diversity and conservation and utilization of plant genetic resources. Plant cell, tissue and organ culture 68: 1-19. Rao, CR. 1952. Advanced statistical methods in biometric research. John Wiley & Sons, New York. Sangare, A, JP Le Saint and MW de Nuce de Lamothe. 1984. The tall coconut palms at Port Bouet (Ivory Coast) III. Cambodia Tall, Tonga tall, Rotuman tall. Oléagineux 39: 205-215. 31 CHAPTER 2: Locating and collecting germplasm Santos, GA, PA Batugal, A Othman, L Badouin and JP Labouisse. 1996. Manual on Standardized Research Techniques in Coconut Breeding. COGENT/IPGRI, Singapore. Sokal, RR and RF Rolf. 1995. Biometry. San Francisco, Freeman. p. 887. Sugimura, Y, M Itano, CD Salud, K Otsuji and H Yamaguchi. 1997. Biometric analysis on diversity of coconut palm: Cultivar classification by botanical and agronomical Traits. Euphytica 98(1-2): 29-35. Suneson, CA. 1960. Genetic diversity: A protection against diseases and insects. Agronomy Journal 52: 319-321. Templeton, AR and DA Levin. 1979. Evolutionary consequences of seed pools. American Naturalist 114:232-249. Treweek, J. 1999. Ecological impact assessment. Blackwell Science, Oxford, U.K. Ward, RH and JV Neel. 1970. Gene frequencies and microdifferentiation among the Makiritare Indians. IV. A comparison of a genetic network with ethnohistory and migration matrices; a new index of genetic isolation. American Journal of Human Genetics. 22: 538-561. Whitehead, RA. 1976. Coconut. Pp. 221-224. In: NW Simmonds (ed). Evolution of crop plants. Longman, London and New York. Wright, S. 1943. Isolation by distance. Genetics 28:114-138. Vargas, A and FA Blanco. 2000. Fruit characterization of Cocos nucifera L.(Arecaceae) cultivars from the Pacific coast of Costa Rica and the Philippines. Genetic Resources and Crop Evolution 47: 483-487. Villarreal, DZ, R Cardenã-Lopez and D Piñero. 2002. Diversity and phylogenetic analysis in Cocos nucifera L. in Mexico. Genetic Resources and Crop Evolution 49: 237-245. 32 COCONUT GENETIC RESOURCES Mapping of coconut genetic diversity R Bourdeix1, L Guarino2, PN Mathur3 and L Baudouin4 1Coconut Breeder and 4Geneticist, Centre de Coopération Internationale en Recherche Agro- nomique pour le Développement (CIRAD), Montpellier, Cedex 5, France 2Plant Genetic Resources Advisor, Secretariat of the Pacific Community (SPC), Suva, Fiji 3Scientist, International Plant Genetic Resources Institute - South Asia Office, New Delhi, India Introduction Mapping of coconut genetic diversity means representing any characteristic of coconut populations on maps, be it their phenotypic or molecular traits; and then studying the links between these traits and any other useful spatial information. According to the needs, the cultivars may be related to their site of origin or to the genebank where they are conserved. This type of analysis can improve the effectiveness of collecting, conservation, management and use of coconut genetic diversity. The mapping studies conducted so far have used data on accessions already collected and conserved in germplasm banks around the world. The latitude and longitude of collection sites have been entered into databases and checked. Then the localities of collection sites are mapped to locate under-represented areas, i.e. areas in which the coconut palms can potentially grow, but where collecting has been inadequate or has not occurred at all. Subsequently, it is possible to identify hotspots of diversity and investigate the geographic distribution of specific traits or combinations of traits using information such as characterization and evaluation, including molecular markers, of the available genetic resources. Georeferencing coconut accessions From 1995 to 2002, coconut researchers of Brazil, China, India, Indonesia, Ivory Coast, Jamaica, Mexico, Papua New Guinea, Philippines, Sri Lanka, Thailand, Vanuatu and Vietnam were trained (Bourdeix 1996; 1997a; 1997b; 1998; Bourdeix et al. 1999; Baudouin 2002) in gathering and inputting data into the Coconut Genetic Resources Database (CGRD) (see Hamelin et al., Chapter 7). Most of the determination and recording of the geographical location of the collecting sites were done in the framework of this work (Bourdeix et al. 1999). This preliminary work was conducted without any sophisticated geographical information system (GIS). It consisted mainly in marking sites of collection by hand on easily available commercial hard-copy maps, preferably with the aid of the researchers who were in charge of the collecting in each country. 33 CHAPTER 2: Locating and collecting germplasm The method of linear approximation was used to determine, as precisely as possible, the longitude and latitude of each collection site on the maps. Later, the Encarta electronic atlas was also used to obtain geographic information more quickly. All the data were systematically entered into the CGRD and then extracted for further geographical analysis. Table 1 presents the status of the geographical localization of coconut accessions according to countries of conservation in the CGRD (version 5.0, December 2002). Table 1. Georeferencing of coconut accessions in CGRD (version 5.0) A total of 1043 accessions in the CGRD are localized by longitude and latitude. Because of duplicates, these 1043 accessions refer to only 710 distinct cultivars or population names. Only 579 collecting sites have a unique combination of latitude and longitude, to the level of minutes. Currently, the CGRD database gives only the geographical localization of the factual female parent, i.e. the location of the palms where the seednuts of the accession have been collected. Many accessions have been moved from one research institute to another; others are rejuvenations of the original population within the same institute. In these two situations, as the female parent is located in a research institute, the geographic coordinates of the institute is given in the CGRD as the ‘collection site’. There is no direct information regarding the collecting site of the original sample of each accession. Let us take a practical example. A researcher wants to know the real Country Number of conservation sites Total number of coconut accessions Number of accessions with geographical coordinates Percentage of accessions with geographical coordinates Benin 1 4 4 100 Bangladesh 1 40 40 100 Brazil 1 16 16 100 China 1 17 17 100 Ivory Coast 1 99 99 100 Mexico 1 20 20 100 Philippines 3 224 224 100 Vietnam 1 31 31 100 Indonesia 4 156 151 97 Vanuatu 1 79 71 90 Papua New Guinea 2 57 51 89 Sri Lanka 1 78 65 83 Fiji 1 11 9 82 Thailand 2 124 97 78 India 1 212 115 54 Solomon Islands 1 21 11 52 Malaysia 2 89 18 20 Jamaica 1 60 4 7 Ghana 1 16 0 0 Pakistan 1 32 0 0 Tonga 1 7 0 0 Western Samoa 1 9 0 0 Total 30 1402 1043 Average 74 34 COCONUT GENETIC RESOURCES origin of the accession ‘NCDP-D9’. This accession is a Tacunan Green Dwarf (originating from the Philippines) but planted in Tanzania in 1989: 1) Looking at the passport data entered in Tanzania, the researcher will see that the accession ‘NCDP-D9’ came from ‘Station Marc Delorme’, a research centre in Côte d’Ivoire. For the accession ‘NCDP-D9’ the latitude and longitude given as collection site in- deed refers to the research station in Côte d’Ivoire, the place where the seednuts were collected. Unfortunately, both fields ‘male par- ent’ accession number and ‘female parent’ accession number have yet to be entered in Tanzania’s collection record. Therefore, the researcher would not know the parental accession of ‘NCDP-D9’. 2) However, looking at all the data from Côte d’Ivoire, a researcher will see that there is only one accession of ‘Tacunan Green Dwarf’ in Côte d’Ivoire: it is the accession ‘SMD NVP3’ planted in 1982. Therefore, she/he will conclude that ‘NCDP-D9’ is the progeny of ‘SMD NVP3’. Looking further at the passport data of Côte d’Ivoire germplasm collection, she/he will see that the accession ‘SMD NVP3’ came from the Davao Research Centre in the Phil- ippines. However, no accession from the Davao Research Centre is yet registered in the CGRD database. 3) Nevertheless, if the researcher is clever and persistent, she/he will check all coconut accessions available in the Philippines. She/he will finally find that the original accession of Tacunan Green Dwarf is also available at the Zamboanga Research Centre, as accession ‘ZRC PD1’ planted in 1977. Based on CGRD, the origi- nal collection site of ‘ZRC PD1’, and therefore ‘NCDP-D9’ is the village of Tacunan, Davao, 007°04’N, 125°36’E. This search process will take a researcher at least 15 minutes, and it will require some luck for complete success. Just one piece of information missing in the whole line and, it becomes difficult to make the links. Another option available in CGRD consists in searching directly all the accessions of ‘Tacunan Green Dwarf’ registered worldwide, but even this does not solve the problem. Both descriptors list and dedicated software have been conceived for managing complete information, and therefore become inefficient if information is incomplete. Data regarding the collection site of the original sample should be recorded in the passport data of each accession. Nevertheless, the status of georeferencing of coconut accessions (Table 1) compares favourably with other crops. In the CGRD, 74% of the accessions have latitude and longitude information. As there are 1402 accessions but only 710 distinct cultivars/population names, it can be said that more than 80% of the coconut cultivars of the world’s germplasm 35 CHAPTER 2: Locating and collecting germplasm banks are geo-referenced. As comparison, only 9% of the accessions of six major genebanks of the United State Department of Agriculture have coordinates, although 50% have a locality description (Greene and Hart 1996). Therefore, in the case of the coconut palm, a large amount of geographical information exists. The challenge is to improve its reliability, to make it more easily available, and to use it to improve coconut genetic resources conservation. Mapping collection sites Mapping of the locations where accessions were collected was done using GIS tools. This may be defined as a database management system which can simultaneously handle spatial data i.e., maps, or the ‘where’– and related, logically-attached, non-spatial, attribute data, and the labels and descriptions of the different areas within a map, or the ‘what’ (Guarino et al. 2002). It is a tool for managing information of any kind according to where it is located (Treweek 1999). The main elements of a GIS are as follows (Guarino 1995; Guarino et al. 1999): • Data input, verification and editing • Data storage, retrieval and management • Data manipulation and analysis • Output The first mapping was done using the Map module of the Corel Quattro- Pro Software (Bourdeix et al. 1999). The data obtained were checked to detect and correct abnormal localizations. As many island countries were involved, a very convenient test was to detect errors such as accessions which appear to have been collected in the open sea (‘sea coconuts’). Sometimes the commercial maps were inaccurate and had to be changed. Lists of errors and corrections were exchanged a number of times with most of the countries in order to reach an acceptable level of precision. This work has not yet been systematically conducted on the entire database. The number of coconut accessions registered in the CGRD database increased from 665 in 1994-1995 to more than 1400 as of 2003. No checking of geographical coordinates was done after 1999 at the database level. There is thus a need to continue this work more efficiently. The International Plant Genetic Resources Institute (IPGRI) and the International Potato Center (CIP) collaborated and developed specialized GIS software called DIVA-GIS that could be downloaded free from Internet at http://diva-gis.org/. DIVA-GIS is dedicated to the analysis of genebank and herbarium databases to elucidate genetic, ecological and geographic patterns in the distribution of crops and wild species. The maps in Figures 1, 2 and 3 have been made using the 579 locations where coconut accessions have been collected. Climatic information from 36 COCONUT GENETIC RESOURCES various sources can be used in conjunction with these georeferenced accessions to determine the zone of cultivation of the coconut palm. A DIVA-GIS module uses the Food and Agricultural Organization (FAO) Ecocrop database of crop climatic and other environmental requirements together with world climatic surfaces to predict the level of crop adaptation, particularly coconut, over geographic areas. By superimposing the theoretical coconut growing area and the location of collecting sites, it is possible to visualize covered geographical regions. Some areas remain clearly under-represented in the national and international coconut Figure 1. Mapping of the locations where coconut accessions were collected in Asia and the Pacific regions. Collection sites of the conserved accessions Theoretical coconut growing area (the darker, the better) Figure 2. Mapping of the locations where coconut accessions were collected in Africa and Indian Ocean regions. Theoretical coconut growing area (the darker, the better) Collection sites of the conserved accessions 37 CHAPTER 2: Locating and collecting germplasm Figure 3. Mapping of the locations where coconut accessions were collected in America. Theoretical coconut growing area (the darker, the better) Collection sites of the conserved accessions germplasm centres. These are areas where coconut palm can grow, but where there are no accession points recorded. For details of this analysis see Chapter 2 on ‘Status, gaps and strategy in coconut germplasm collecting’. DIVA-GIS can also be used to check the coordinates of the collection sites in relation to an administrative boundaries database. In effect, this would automate the data-checking process (including locating ‘sea coconuts’) which used to be done manually. It would therefore be useful in the future to create an interface between CGRD and DIVA-GIS. This would require a module allowing the export of data from the CGRD in a format easily readable by DIVA-GIS. Mapping morphometric characteristics Further analytical functions implemented in DIVA-GIS include mapping the distribution of specific traits and mapping of richness and diversity. Genetic diversity mapping usually begins by dividing the target area (or strata within the target area, e.g. climate zones) into a number of smaller zones, for each of which a measure of diversity can be calculated (Guarino et al. 2002). Different geometric, political or socioeconomic spatial units have been used, but ideally, areas of equal shape and size (to reduce the effect of the area on diversity measures) should be employed, for example square grid cells (Nabhan 1991; Ferguson et al. 1998). One of the important parameters describing the quality of the coconut fruit is the Q factor. The Q factor can be defined as the weight of husk 38 COCONUT GENETIC RESOURCES divided by the weight of the fruit without free water (coconut water inside the nut). The quantity of free water is quite variable according to environmental factors (such as rainfall) and the degree of maturity of the fruit. This is the reason why the Q factor is calculated without taking into account the free water. The larger the Q factor, the higher the proportion of husk in the fruit. Normally, to get a good estimate of fruit composition, a sample of two fruits is analyzed six times a year over four years on each palm, and this must be done on 30 palms to characterize an accession. In CGRD version 5.0, only 32% of the accessions have data for fruit component analysis. But in practice, the data are even less complete. It can be estimated that at least 20% of the fruit component data available in CGRD 5.0 were derived from the analysis of a single fruit sample harvested at one point. This again reinforces the importance of having complete data in order to carry out an analysis that can be really useful in germplasm conservation, management and use. In any case, all the available georeferenced accessions with fruit analysis data were used to carry out some spatial analyses. The first results obtained were not convincing, because they included both Tall and Dwarf cultivars, which have distinct fruit characteristics. Nevertheless, there was a clear geographical pattern, based on the geographical distribution of mean values of Q factor, for Tall cultivars (Figure 4). Accessions originating from India and Africa show a higher Q value than accessions from Southeast Asia and the Pacific region. Figure 4. Geographical distribution of mean values of the Q factor for Tall cultivars 0.35-0.40 0.40-0.45 0.45-0.50 0.50-0.55 0.55-0.60 Means of the Q fruit ratio 39 CHAPTER 2: Locating and collecting germplasm Mapping molecular markers New functionality in version 2 of the DIVA-GIS software includes options of mapping based on molecular markers (DNA) data that are illustrated here by an example. Figure 5 presents the geographical repartition of allele 128 of locus CnCir A3 in the Tall coconut varieties (see Lebrun et al., Chapter 4). Allele 128 is one of the four alleles whose presence at a significant frequency is characteristic of Indo-Atlantic coconuts. Similar (but not identical) pictures could be obtained with any of these markers. Allele 128 is found at a high frequency in an area extending from the Indian sub-continent to the Atlantic coast of America. It is rare or absent in the eastern part of Southeast Asia, in the South Pacific and on the Pacific coasts of America. Figure 5. Geographical distribution of the frequency range of the allele 128 of locus cnCir A3 in the Tall coconut varieties. The average frequency is based on the samples studied, which do not necessarily reflect the relative abundance of the cultivars. Nevertheless, if we consider the picture in more detail, the intermediate frequencies observed in East Africa result most probably from the introduction of coconuts from South Asia by Austronesian navigators, whose language is still spoken in Madagascar. There is also a transition zone in the western part of Southeast Asia, demonstrating some genetic exchange with South Asia. This may have involved floating, but the activity of Arab merchants who have crossed it for several centuries is probably the cause of most of the exchanges across the Indian Ocean. The apparently medium frequency of this allele in Sri Lanka is actually an artefact in the Sri Lanka Tall, by far the most dominant cultivar of 40 COCONUT GENETIC RESOURCES this country. The allele 128 frequency is 0.82, but other cultivars, with a round nut were also sampled, which could be the result of hybridization of local coconuts with planting material imported at different periods from Southeast Asia. Finally, three cultivars with allele 128 frequencies between 0.10 and 0.16 are found in the Pacific Ocean. Considering the three other characteristic (marker) alleles, they are probably not related to the Indo- Atlantic group. The presence of allele 128 at a low frequency in this area is probably a case of homoplasy (i.e., the fact that similar traits – here, fragments with the same length – appear independently by mutation in different genetic groups). Homoplasy is not infrequent in microsatellites. Mapping for collecting genetic diversity The genetic diversity mapping described in this Chapter concerns mainly coconut palms that are already in genebanks and their characteristics, such as fruit composition and molecular marker profiles. Another possible field of investigation could be to study coconut populations in situ and to map their diversity before the collecting of seednuts or embryos. According to Pernes (1984), the best germplasm collecting programmes are carried out in two stages, with a first exploration and preliminary survey used as a basis for studies that will permit better planning of the second, more systematic campaign. Such a two-step programme was done in Mexico (Zizumbo Villarreal et al. 1993) where fruit analyses were made first at 47 localities along the Atlantic and Pacific coasts and in the narrowest part of the country (the States of Oaxaca and Veracruz). Collecting was then carried out in only 19 localities, 90% on the Pacific coast, where the greatest fruit variability was found. However, most budget appropriations seldom permit such organized programme. It is also necessary to underline the great sensitivity of the coconut to environmental variations. The phenotype of a palm at a certain moment is usually not representative of its genetic value. There is much variation linked to heterogeneity of the field, for example in soil fertility or water availability. The growth of the coconut palm is controlled by various rhythms, depending on internal and external factors. Although it may produce continuously, yields are often irregularly distributed over the year. Biannual cycles are superimposed on this variation. These phenomena are rather frequent in young palms or following droughts (Bourdeix et al. 1994). For a proper evaluation, vegetative and yield characteristics of a sufficient number of palms should be measured over a period of several years. Nucé de Lamothe and Wuidart (1982) have emphasized that it is difficult to conduct such a study outside a research station. However, it will be possible to collect leaf samples from surveys, to analyze their DNA and then to sample the field origin where genetic 41 CHAPTER 2: Locating and collecting germplasm diversity is greatest. Up to now, we have no example of such a strategy, but it may happen in the near future (see related articles in Chapter 2). Conclusion Recent developments in GIS technology for mapping genetic diversity is expected to contribute significantly to identify and fill the gaps in coconut collections, enhance the effectiveness of collecting, better manage them in field genebanks, effectively select parents from geographically distinct regions and expand coconut production through site-genotype matching. Figure 6 provides an analogous map of the number of accessions registered in the germplasm banks of the COGENT member countries. An additional effort is needed to make coconut descriptors list and related software better adapted to management of incomplete data. The new version of DIVA-GIS software (Version 4.0) can be downloaded free from the internet (http://www.diva-gis.org), and is easy to learn and use, and is tailor-made for genetic resources applications. Country- level GIS databases can also be downloaded from this site and these databases can be used together with the genetic resources databases that are being mapped and analyzed. These are files with data on administrative boundaries, country boundaries, and first and second level administrative subdivisions for most countries. For all countries, grids are available for altitude, land cover and population density. DIVA-GIS can be used to check existing coordinates and carry out analyses of characterization and evaluation data. Specific software devoted to the management of genetic resources, such as CGRD for coconut, could be improved by allowing easy exporting of data to DIVA-GIS. Figure 6. Geographical distribution of the coconut accessions conserved in COGENT germplasm banks 42 COCONUT GENETIC RESOURCES References Baudouin, L. 2002. Study on genetic resources status in Hainan, China (visit 12-18 December 2000). CIRAD Report (CP SIC no. 1505), France. 50p. Bourdeix, R. 1996. Coconut germplasm in Jamaica, Mexico and Brazil. Mission Report, March 1997. Doc CIRAD-CP no. 548, France. 53p. Bourdeix, R. 1997a. Coconut germplasm in Tanzania, Sri Lanka and In- dia. Mission Report, March 1997. Doc CIRAD-CP no. 739, France. 49p. Bourdeix, R. 1997b. Actions de formation en Afrique et Amérique La- tine/Caraïbes. Cours de formation des formateurs à l’utilisation du manuel Stantech (Techniques Standardisées de Recherches pour l’Amélioration du Cocotier). Mission Report, August 1997. Doc CIRAD-CP no. 857, France. 49p. Bourdeix, R. 1998. Coconut germplasm and breeding in Papua New Guinea and Vanuatu. CIRAD 1997 Annual Review Report, March 1998. Doc CIRAD-CP no. 998, France. 52p. Bourdeix, R., Y.P. N’Cho and A. Sangare. 1994. Rythmes de production chez le cocotier Nain (Cocos nucifera L.): Etude de l’alternance castration-production comme mode de gestion des champs semenciers. Agronomie Africaine VI, 2:77-162. Bourdeix, R, L Baudouin, J Ollivier and JP Labouisse. 1999. Consultancy report on coconut collecting strategies (submitted to COGENT/IPGRI). IPGRI, Rome. Ferguson, ME, BV Ford-Lloyd, LD Robertson, N Maxted and HJ Newbury. 1998. Mapping the geographical distribution of genetic variation in the genus Lens for the enhanced conservation of plant genetic diver- sity. Molecular Ecology 7:1743-1755. Greene, SL and T Hart. 1996. Plant genetic resources collections: An op- portunity for the evolution of global data set. Third International Conference/Workshop of integrating GIS and Environmental Modeling Santa Fe, New Mexico, USA. Guarino, L, A Jarvis, RJ Hijmans and N Maxted. 2002. Geographical Information Systems (GIS) and the conservation and use of plant genetic resources. Pp. 387-404. In: JMM Engels, V Ramanatha Rao, AHD Brown and MT Jackson (eds). Managing plant genetic diver- sity. CAB International, Wallingford. Guarino, L. 1995. Geographic information systems and remote sensing for the plant germplasm collector. Pp 315-328. In: L Guarino, V Ramanatha Rao and R Reid (eds). Collecting plant genetic Diversity. Technical Guidelines. CAB International, Wallingford. Guarino, L, N Maxted and M Sawkins. 1999. Analysis of geo-referenced 43 CHAPTER 2: Locating and collecting germplasm data and the conservation and use of plant genetic resources. Pp. 1- 24. In: SL Greene and L Guarino (eds). Linking genetic resources and geography: Emerging strategies for conserving and using crop biodiversity. CSSA Special Publication No. 27. ASA and CSSA, Madi- son, U.S.A. Nabhan, GP. 1991. Wild Phaseolus Ecogeography in the Sierra Madre Occidental, Mexico. Systematic and ecogeographic studies of crop genepools No. 5. IBPGR, Rome, Italy. Nuce de Lamothe, MW de and F Wuidart.1982. L’observation des carac- téristiques de développement végétatif, de floraison et de production chez le cocotier. Oléagineux 37:290-296. Pernès J. 1984. Gestion des ressources génétiques des plantes. Tome 2 : Manuel. Agence de coopération culturelle et technique, Paris ISBN 92-9028-043-3, éditions Lavoisier, France. Treweek, J. 1999. Ecological impact assessment. Blackwell Science, Ox- ford, U.K. Zizumbo-Villarreal, DF, R Hernandez and HC Harries. 1993. Coconut varieties in Mexico. Economic Botany 47(1): 65-78. 44 COCONUT GENETIC RESOURCES Status, gaps and strategy in coconut germplasm collecting R Bourdeix1, L Guarino2, V Ramanatha Rao3 and L Baudouin4 1Coconut Breeder and 4Geneticist, Centre de Coopération Internationale en Recherche Agro- nomique pour le Développement (CIRAD), Montpellier, Cedex 5, France 2Plant Genetic Resources Advisor, Secretariat of the Pacific Community (SPC), Suva, Fiji 3Senior Scientist, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction The International Coconut Genetic Resources Network (COGENT) Steering Committee decided to promote germplasm collecting in areas at risk of genetic erosion at its first meeting in Singapore in 1992. This was expected to fill the gaps in national collections, developing (and refining) both morphometric and molecular markers techniques for efficiently locating diversity and transferring efficient and practical techniques for collecting. Phase 1 of the COGENT project ‘Coconut Genetic Resources Network in Asia and the Pacific Region’ was completed in July 1997. A regional network consisting of 13 countries was established to foster the conservation and utilization of coconut genetic resources. In December 1998, the Asian Development Bank (ADB) approved Phase 2 of the project. Its objective was to expand the network to 20 countries, to further promote coconut collecting and sustainable conservation, and to strengthen human resources. During 1997-2000, many coconut accessions were collected and planted in field genebanks in all the network member countries. The main objective of this chapter is to review and assess the strategies used in collecting coconut germplasm and make suggestions for future work. Status of coconut germplasm collecting As noted earlier, many coconut accessions have been collected and conserved (see Chapter 5 for more details). Access to information about this coconut germplasm is much better than it was ten years ago. COGENT network members are regularly updating passport information and characterization data of accessions in the Coconut Genetic Resources Database (CGRD). If a new coconut accession is now collected somewhere, there is a reasonably high probability that passport data will be available to the whole network through the CGRD, within one or two years. In the CGRD Version 5.1 (April 2002), the total number of accession was 1416, of which 216 had no registered accession size (number of true- 45 CHAPTER 2: Locating and collecting germplasm to-type living palms in the field). This means either that all palms of these accessions are dead or that data on them is missing. Information on dead accessions is kept in the database because it remains essential to researchers. Some general statistics on coconut collections are given below: • 1186 accessions have a size of one or more palms • There are only 620 distinct names (of cultivars or populations) • 74% of accessions are of the Tall type • 25% are of the Dwarf type, and the remaining 1% are intermediate forms • 140 000 is the total number of ‘living’ palms • The average number of palms per accession is 118 and per culti- var, 225 • About 30% of accessions have already been duplicated in several genebanks or rejuvenated Another very important piece of information is the ‘Date of Last Inventory/Counting’ of each accession. It is the most recent date on which the number of living palms was checked. An examination of this field shows the following disturbing trend for the 1193 accessions, which have an accession size of at least one palm: • For 36%, the Date of Last Inventory (DLI) remains unknown • For 6%, DLI is during the past three years (2000-2002) • For 47%, DLI is between 1996 and 1999 • For 11%, DLI is prior to 1995 During the period 1996-2001, visits were conducted to many countries to train and assist researchers to input data into CGRD (Bourdeix 1996; 1997a; 1997b; 1998; Bourdeix et al. 1999; Baudouin 2001). Although this improved data management, there is a strong need for continued efforts in this regard. In particular, checking and entering DLI should be done at least once a year. In addition, among the 1416 accessions, 120 (of which 16 were from Jamaica, 32 from Pakistan and 29 for Bangladesh, and 43 from various other countries) do not have any registered ‘acquisition date’. The oldest accession registered in CGRD is a Samoan Tall planted in 1912 in the Solomon Islands. Levers Plantations began there around 1905. Coconut research is said to have begun in India in 1916 and a varietal collection was started there in 1921 (Harries 1978). Twenty-three accessions were planted in India between 1934 and 1946 and are registered in CGRD database. An accession from Mapanget, Indonesia is dated 1927. The Coconut Research Scheme was established in Ceylon 46 COCONUT GENETIC RESOURCES – now Sri Lanka - in 1929. The depressed copra market of the 1930s impeded research, and a varietal survey that began in 1939 was terminated after only a few months (W.V.D. Pieris, personal communication, cited by Harries 1978) and the oldest accessions are dated 1954. In Africa, the Marc Delorme Research Centre began its activities in Côte d’Ivoire in the fifties (Nuce de Lamothe and Wuidart 1979). Parham (1960) carried out one of the first scientific surveys intended to collect coconut palms and breadfruit trees in the Pacific. As a result, some coconut varieties with very large fruits, such as the Markham Valley Tall, were introduced to various genebanks throughout the world. Whitehead (1966) conducted a survey in the Pacific searching for varieties tolerant to the Lethal Yellowing disease of Jamaica. An indirect result of this work was to inspire Harries (1978) to develop his theory of evolution and dissemination of the coconut palm. Vanuatu began its germplasm planting in 1963 and the Philippines in 1976. An examination of the CGRD also reveals that from 1912 to date, there have been only 11 years during which 50 or more coconut accessions were collected per year. Five of these 11 years were between 1992 and 1999, (i.e. during the early days of the COGENT). The other years in which at least 50 accessions were collected, were 1981 and from 1983- 87. Around 30% of the registered accessions were planted after the COGENT was established (1992 and later). However, no accession acquired between 2000 and 2003 is registered in the CGRD database at the time of writing this paper. This suggests a significant reduction in collecting activities in the past three years. Gaps in coconut germplasm collecting The foregoing historical survey has established the fact that a substantial number of coconut accessions are being conserved in genebanks around the world. However, there may still be compelling reasons for further collecting. Additional collecting may be justified if: 1. Diversity is still missing or has been lost from existing ex situ col- lections; 2. Diversity is in imminent danger of disappearing from farmers’ fields; and 3. Diversity is needed for immediate use and is not available from existing collections. Related palm species The palm family (Palmae or Arecaceae) counts about 2800 species scattered among 190 genera. The Cocoeae tribe contains 27 genera and nearly 600 species, including several economically important plants such as Cocos nucifera L. (coconut), Elaeis guineensis (African oil palm), Attalea 47 CHAPTER 2: Locating and collecting germplasm cohune (babacu) and Bactris gasipaes (peach palm). Morphologically, the Cocoeae tribe is characterized by having the synapomorphy of presence of three or more pores or ‘eyes’ on the endocarp (Gunn 2002). It comprises of six sub-tribes, among which the Butiinae includes the Cocos genus and seven American genera (plus a recently discovered genus from Madagascar, Voaniola). Since most of the related genera are American in origin, in the past it was speculated that coconut also originated in Americas (Cook 1901). In recent classifications, Cocos nucifera L. is considered as the only species of the genus. It is generally considered that it cannot be crossed with any other species. However, as far as we know, no published report of such an attempt to date. There is thus an opportunity for research in this field, checking for such possibilities, as resistance to lethal yellowing in allied palms closest to coconut may be a revealing exercise. If nothing else, it would establish that the coconut is indeed a botanical and genetic ‘outlier’. Geographical gap-filling Most often, gap-filling collecting focuses on uncovered geographical regions, which may be quite extensive, e.g. a whole country. Figures 1, 2 and 3 in the earlier article ‘Mapping of coconut genetic diversity’ can be used to visualize inadequately covered geographical regions by superimposing the theoretical coconut growing area and the location of collection sites. The zones coloured in grey, which are climatically suitable for coconut, do not seem to have coconut occurring in them, however, this needs to be confirmed by ground truthing (i.e. checking in the field). It must be noted, however, that some areas may be better represented than they might look in these maps. For instance, India is probably better surveyed than the map implies, but Indian researchers have not yet inputted all the geographic coordinates of their national accessions. A collecting mission was conducted in Madagascar in 1999 by Indian researchers, but collecting information remains incomplete. Some other areas are probably of low coconut diversity. For instance, for historical reasons, there is probably a low probability of finding unique diversity in African countries such as Congo, Democratic Republic of Congo, Angola, Ethiopia and Sudan. The same could be true in South America – in the central part of Brazil, and the parts of Peru and Bolivia east of the Andes. Nevertheless, all these zones have never been surveyed for coconut, and exploration would be justified. Some areas remain clearly under-represented in national and international genebanks, which are listed below, in a subjective ranking of priority: 1. The west coast of South and Central America (except Mexico and Panama, which have already been surveyed). Germplasm 48 COCONUT GENETIC RESOURCES collecting is presently being conducted in Guatemala. These studies are essential, considering the problem of the Lethal Yellowing Disease and the history of coconut in this region; 2. A large part of Micronesia, including the Caroline and Mariannas Islands; 3. The eastern part of Polynesia, including the Tuamotu and the Marquesas Islands and Hawaii; 4. Irian Jaya and the Moluccas archipelago; 5. The tropical coasts of Australia and the Cocos/Keelings Islands, where putative wild coconut occurs (Williams 1990; Leach et al. 2003); 6. Madagascar. Seafarers from Southeast Asia reached this island probably around the sixth century AD and settled there. Molecular biology studies show that they probably introduced coconut seednuts with them, and new diversity developed thereafter; and 7. Other more localized areas like Somalia, Myanmar, Laos and Sarawak in Malaysia. Some of the areas that are suggested here (such as Micronesia, eastern Polynesia, and the Cocos/Keelings Islands) represent only a very small part of the coconut world, in terms of cultivated area and economic value. However, these areas could prove to be extremely important for coconut diversity. Pacific Islanders, especially Polynesians, have been involved in coconut cultivation and transportation for a very long time. Coconut diversity is more endangered in these areas, precisely due to its comparatively low economic importance and due to the possible effects of global warming and other human activities. It is interesting to note that the Arab traveller Ibn Batutta reported the presence of coconut in Yemen in 14th century. Climate is considerably drier at present than in antiquity (and probably than at the time of Ibn Batutta), and Yemen is not reported as a producing country. However, contact with local botanists could reveal the presence of a few remnants of this historically interesting population. Targeted surveys and under-represented phenotypes The various existing ex situ collections are still not fully representative of the germplasm available in farmers’ fields, especially with regard to the diversity of climate under which coconut is grown. Occasionally, specific environmental conditions may be targeted. For example, high-altitude or cold-tolerant varieties remain under-represented in coconut collections. Finally, missing genotypes are sometimes targeted, e.g. named varieties of known appearance, which are not found in collections. 49 CHAPTER 2: Locating and collecting germplasm Most of the old surveys, such as those of Parham (1960) or Nuce de Lamothe and Wuidart (1979), intentionally focused on varieties with large, thin-husked fruits. Many farmers indeed prefer big round nuts. However, the use of the coconut husk is making a comeback, and it seems very important for the future to further safeguard and study the thick husked varieties. Coconut from India and Africa has, on average, higher husk content than most of the coconut from Asia and the Pacific. In the Pacific, ‘Niu Kafa’ types are an exception. However, there are references from everywhere, including Southeast Asia, describing a few coconut varieties with a high percentage of husk. In 1978, Harries developed a theory about coconut evolution, dissemination and classification of the coconut. He used the name Niu Kafa to describe a putative wild coconut palm with a large husk. “First came the natural evolution and dissemination by floating of a variety with large, long, angular, thick husked and slow germinating fruits. From this thick-husk type, selection under cultivation produced a spherical fruited variety, not necessarily larger but with increased endosperm, reduced husk thickness, earlier germination and disease resistance” (Harries 1978). However, according to Foale (1987), islanders also selected other palms bearing fruits that contained long fibres to make strong twine and ropes for use in the construction of both buildings and boats. Consequently, the huge fruits presently known as Niu Afa in Samoa, Niu Kafa in Tonga and Magi Magi in Fiji are no longer wild coconuts; they are varieties highly selected by the Polynesians for the utilization of husk. This is particularly clear in Samoa, where the variety seems to occur in its purest form, and where the palms are located near houses and are all of a homogeneous green colour. An important theoretical question that arises is whether there is a link between the Indo-African Coconut group and the Pacific and Asiatic cultivars with high husk content. Molecular techniques may help to resolve such a question. However, so far only a very few samples of the Niu Kafa type have reached laboratories in good order. Only one typical sample could be analyzed, and it appears that it is not closely related to the Indo-Atlantic coconuts. At least 20 to 30 more samples of thick-husked varieties originating from different parts in Asia, the Pacific and Oceania should be collected and DNA-analyzed. These varieties could be of Niu Kafa types, but they may also give smaller fruits of quite different shapes. Some varieties from the Tuvalu archipelago have high husk content but with a shape that, although elongated, is very different from those of Niu Kafa (Labouisse and Bourdeix 2003). It is important to collect different putative ‘wild’ coconut types and analyze them using molecular markers. Such a study may enhance our knowledge about dissemination and help 50 COCONUT GENETIC RESOURCES in refining collecting strategies and even the design of coconut breeding programmes. Another endangered special phenotype is a class of coconut varieties described as ‘Sweet Husk’. The husk of young fruits of this type is soft and sweet and can be chewed like sugarcane. When over-mature, the fruits can be husked easily. Fruits of these varieties are generally eaten by children, flying foxes and rats before nuts mature. It is almost impossible to collect them in a classical survey, as no seed is usually available. Local people are no longer interested in them as in the past as consuming them due to changes in social norms. For example, Tiara Mataora, from the Cook Islands said “I like it but do not want somebody to see me chewing sweet husk, because these people will think I am a poor man”. A special effort to collect and study these types must be made. Such special variants could be useful for making high value products for the tender nut market. Two important collecting programmes were known to focus on particular traits: drought adaptation in Sri Lanka (Liyanage et al. 1988), and selection for Lethal Yellowing Disease (LYD) tolerance in Tanzania (Schuiling et al. 1992). It seems that these two programmes have not really been successful. The accessions from areas in Tanzania with high LYD pressure continue to die from the disease during the next generation. Accessions collected in Sri Lanka from both dry and wet zones were compared under dry conditions, but no significantly different reactions were noted. Other interesting types will probably emerge from the results of the farmer participatory approach (see related section below). Losses from existing ex situ collections The life span of coconut accessions is sometimes shorter in germplasm conservation centres than in farmers’ fields. Some example will illustrate this. Indonesian accessions registered in the CGRD are conserved at four different sites: Mapanget (Manado City), Pakuwon, Bone-Bone and Sikijang, Selakau (West Kalimantan), Makariki (Molluccas) and Marihat (North Sumatra) (Rognon and Batugal 1998). However, Indonesian researchers in Manado informed us that these conservation sites are no longer in use. The remaining accessions in Marihat are said to be original populations and to date, these have not been duplicated anywhere else and thus become important for future rejuvenation and planting in current genebanks. In CGRD Version 5 (2002), 55 Indonesian accessions out of 156 do not have any data for the accession size field (number of living palms) and the date of the last inventory/counting. Some of these accessions, such as the 1995 planting in Manado and those conserved at the Bone- 51 CHAPTER 2: Locating and collecting germplasm Bone Station, appeared to have been destroyed and later was no longer considered as a coconut germplasm centre. According to Indonesian researchers, the 41 accessions (1682 palms planted between 1984 and 1988) are considered lost. At Sikijang, at least 25 accessions, with 100 palms each, were planted in 1998 and 1999. Because of various factors, including fire, in January 2001 (i.e. only 3 years later) 77% of these palms were either dead or in a poor condition. Due to the change of status of Sikijang station, it is assumed that the 30 accessions at that station were mostly lost. However, as some palms remained, they have not been removed from the inventory. Indonesian germplasm now stands at 170 accessions (including some new ones), of which 61 can be considered as lost. Therefore, the real number of living accessions for Indonesia cannot be more than 109, with 4976 palms (on average, only 46 palms per accession). At least 65 accessions from Indonesia are now lost and should be re-collected (after having found a way to safely conserve them for the future). In Papua New Guinea, demonstration plots of various cultivars were planted during the early 1930s at the Bubia Lowland Agricultural Experimental Station. In 1964, it was decided to plant a new trial at Kapogere Agricultural Station in the Central District, Papua. The scope of the trial was broadened to include at least nine foreign introductions: New Hebrides, Solomon Islands, Malaysia, Rennell Island, Singapore, Ceylon-Random, Ceylon-Selected, Maldives and Fiji Talls. The status of these accessions remains unknown. They are not registered in CGRD and they were not transferred to the international collection in Madang. The accessions collected in the past and planted in old, possibly now neglected, field genebanks should be safeguarded. In Thailand, it seems that some old accessions were cut without being rejuvenated in order to plant oil palm experiments. The sustainability of germplasm banks seems better in Côte d’Ivoire, India, the Philippines and Sri Lanka. Targeted exchanges between germplasm conservation centres can help in duplicating accessions in different genebanks for safety and in promoting the sustainability of coconut genetic resources conservation. Exchange of germplasm immediately after a collecting mission is also advantageous as many freshly collected embryos would be available and could be exchanged safely. The exchange of coconut germplasm among coconut-producing countries remains very limited. For example, from 1995 to 1999, only one coconut variety was exchanged between the Philippines and Vietnam. In contrast, more than 80% of the foreign cultivars existing in Brazil, Indonesia, Philippines, Tanzania, Thailand, Sri Lanka and Vietnam came from the Marc Delorme Research Centre in Africa in the past. 52 COCONUT GENETIC RESOURCES India is an exception, with a strong collecting programme abroad. But only a few palms remain from the survey conducted by Indian researchers in Madagascar. Five accessions were collected in 1997 from a single location in Sambava province. Many plantlets died before reaching the field planting stage. These may have to be re-collected to have a representative population of these accessions. More than 3000 coconut embryos were collected from Tuvalu, Cook Islands, Marshall Islands and Kiribati and sent to the Secretariat of the Pacific Community’s (SPC) Regional Germplasm Centre (RGC) in Suva, Fiji. Unfortunately, almost all these embryos died during the in vitro culture and/or the transfer to the International Coconut Genebank (ICG) in Papua New Guinea. The reasons for these losses were the high rate of contamination and low rate of rooting. Some of these accessions need to be collected again. An FAO report by Pieris (1966) indicated that the concern for collecting exotic germplasm was high in the early 60s, as about 30 countries reported seed or pollen exchange. This period contributed indeed to the richness of present genebanks. However, many of the cultivars are no longer reported in the receiving country. For example, the Philippines received planting material from 14 countries primarily for resistance trials against Cadang-Cadang. Apparently, nothing is left from this introduction and some of these cultivars had to be re-sampled about 20 years later. Genetic erosion To understand on a smaller scale the mechanisms that build diversity and the factors that influence the evolution of coconut types, a study was undertaken in Vanuatu, a remote archipelago in the South Pacific (Caillon 2003). There were 60 variants named based on a particular aspect describing distinct character from the rest of the population (Labouisse and Caillon 2001). Of these 60 variants, 45% may not be selected but are still recognized, 20% are chosen for their social importance (e.g. a coconut brought by a local mythical hero), 15% to make copra, 13.3% for their nutritional qualities and 6.7% for non-food uses (e.g. containers, ropes). In a remote village of a northern island (Vanua Lava), where 30 variants are found, only 5% of all the coconuts planted by 25 farmers are named (Caillon, pers. com.). Coconuts selected for their domestic and social interest are the least numerous (7.4% and 8.5% of the planted variants, respectively) whereas 46.9% are planted for food purposes. The most striking example concerns the variant with a large proportion of husk traditionally used to make ropes. These specific coconut types are currently ignored as other types of ropes have become more prominent. At the 53 CHAPTER 2: Locating and collecting germplasm same time, the importance of copra for cash has increased. As a result, truly ‘high husked’ variant can only be found on old plantations dating from the time when farmers still used coconut ropes. This exemplifies genetic erosion due to changes in farmers’ preferences. The number of named variants in a field depends on a farmer’s willingness to select and plant variants with characteristics other than high copra, in order to respond to other uses for food, shelter or social needs. Generally, planting material for new plantation comes from farmer’s own garden or from a nearby plantation. However, the most remarkable variants come from other plantations, sometimes distant, where the farmers might have seen while helping other villagers/farmers making copra and brought a few seednuts back. However, that level of diversity also varies greatly depending on the degree of knowledge of a farmer about his/her own coconuts. Thus, young plantations planted by the current generation owner in which immature fruits are accessible and where copra is frequently made will be the richest ones in terms of genetic diversity. Consequently, the reduction of named variants at a village scale is due to the combination of cultural erosion through the loss of traditional uses and through the younger generations’ loss of ability to identify variants. Such loss caused by social process could further be demonstrated more clearly by molecular techniques to assess real genetic erosion even if variants are not readily identified but are still growing around and are able to exchange genes through allogamy. Such an approach is currently underway. Changes in land use patterns, urban migration, industrialization and replacement with other species (such as oil palm) or with introduced and/or improved varieties (hybrids) are contributing greatly to the loss of coconut diversity. Natural calamities (cyclones, drought, diseases such as cadang-cadang and lethal yellowing) as well as human induced ones (pollution, war, etc.) are also agents of genetic erosion. Strategy in coconut germplasm collecting: Towards a diversity of approaches No single approach is likely to be effective to collect and conserve the full range of variation within a target gene pool and making it available to breeders and other users, and coconut is no exception. Collecting germplasm for ex situ conservation should thus be regarded as simply one of the components in a comprehensive strategy for conservation of the target gene pool. Until recently, coconut surveys were faced with two constraints linked to the biology of the plant. The first is the large size of the fruits; a sample of a hundred fruits often weighs more than 150 kg. The volume of the 54 COCONUT GENETIC RESOURCES fruits considerably restricts the number of samples that can be transported, or leads to a reduction of the effectiveness of the samples. Another constraint is the nature of the seed. The coconut, with recalcitrant seeds (Roberts et al. 1984), looses germination capacity rapidly. Most cultivars have no dormancy period; the seeds start to sprout 1-3 months after reaching maturity. Moreover, the coconut seed is relatively sensitive to cold. Due to these characteristics, numerous samples of coconut varieties have been lost partly or totally for various reasons: survey conditions did not allow for sufficient sampling or ships transporting the fruits passed through zones that were too cold, or duration of transport and customs clearance exceeded the survival time of the seeds. For these reasons, in all research stations some coconut accessions can be found that are represented by numbers that are too low to constitute a good population for conservation, though originally large number of nuts might have been sampled. The application of new technologies makes it possible to get around some of these problems (see Engelmann, Chapter 2). However, much care needs to be exercised to avoid what happened recently in the Pacific. Bourdeix et al. (1999) described case studies that were conducted in 14 countries involved in coconut germplasm surveys during the 1994- 1999. These detailed studies cannot be reproduced in extenso here but some of the most general conclusions and thoughts are discussed in the next section. The Coarse Grid Strategy In 1997, a manual on coconut breeding research techniques (STANTECH) was published and distributed to coconut-producing countries (Santos et al. 1996). This manual describes the bases of the recommended collecting method in its Chapter 3 on ‘Germplasm exploration and collecting’ and Chapter 10 ‘Generalized sampling strategy’. The Coarse Grid Sampling strategy described here has been applied systematically to cover the coconut areas in the Philippines (Santos 1987) and Malaysia (Jamadon 1987). The basic elements of this process is described below by Guarino et al. (1998). As noted earlier, the COGENT member countries have collected significant amount of coconut genetic diversity during 1993-2000, with support from ADB. A research team from the French Agricultural Research Centre for International Development (CIRAD) was mandated to review and assess the effectiveness of the collecting strategies followed in the first phase of this project. This study noted that only one country, the Philippines, made use of grid sampling technique. No country used ‘coconut importance value’ suggested in the collecting strategy. It must 55 CHAPTER 2: Locating and collecting germplasm The Coarse Grid Strategy How can a national, regional or international coconut research programme assess the relative importance of the different reasons for collecting? It will clearly need some basic information on its mandate region: • Where is the crop growing, in relation to agro ecological zones of the region? • How much genetic variation is already present in genebanks? • What are the main agents of genetic erosion and where are they most threatening? • Who are the principal users and what are their needs? The sources of this information will include agricultural censuses and atlases, the databases of genebanks, local extension agents and their records and coconut breeders. Based on this information, it should be possible to identify (and prioritise among) areas of the following types within the mandate region: 1. Under-represented areas. These can be identified by mapping passport data of existing collections, and include areas where collecting has been inadequate or has not occurred at all. 2. Complementary areas. These are areas, which are genetically, or environmentally different from areas from which collecting has already taken place, based on passport and characterization data. 3. Environmentally or genetically diverse areas. In previously uncollected or under-collected areas, it is advantageous to collect over wide range of agroecological conditions because genetic diversity is partially correlated with environmental diversity. Preliminary characterization and evaluation (including genetic diversity studies) of conserved material may have identified areas, which are particularly diverse genetically. 4. Areas with target genetic material. This may be inferred from environmental conditions, known from previous characterization and evaluation work and/or revealed by local knowledge. 5. Threatened areas. These may be identified by local people, repeat visits, etc. Based on the points derived from the brief survey of patterns of genetic diversity in coconut, the following basic elements of a coconut collecting strategy are proposed: Choosing the sites 1. Divide the coconut-growing region in 40x40 km grids. This should be done separately and independently for each sub-regional grouping (stratified sampling). In general, collecting in the SE Asian region should be more intensive, so smaller grid sizes could be used. 2. Superimpose the location of the different types of areas listed above on 56 COCONUT GENETIC RESOURCES the grid. This can be done using a GIS. Calculate a ‘coconut collecting importance value’ (CCIV) for each grid square based on the presence and priority value of each type of area in the grid area. 3. If possible, carry out a preliminary exploratory visit to 2-3 sites per grid square and collect morphological information to complement characterization information from germplasm already conserved. Use this information to further refine the CCIV. 4. Collect germplasm systematically at a minimum of two sites in all grid squares. If the material is of the same ecotype and/or environmental con- ditions are similar, leave a minimum of 15 km between sites. 5. Collect more intensively (up to six sites) in grid squares that have a higher CCIV. however be noted that much of the collecting in Phase I was over in 1997, while the strategy was developed in 1998. Most of the surveys were conducted by following, more or less precisely, administrative divisions such as regions, subregion and districts. Major constraints noted for the implementation of the collecting strategy were the time and capacity to build geographical grids that need well documented information such as climate, soil and population data. CIRAD team then recommended that the International Plant Genetic Resources Institute (IPGRI) should prepare, for national researchers, computerized maps with standardized geographical grids already documented with general information and national researchers to focus on gathering plant-specific information. However, it is not possible for IPGRI to undertake such country specific activity and training national partners to develop their capacity to make the grids, etc., will be more appropriate. This is also appropriate in the light of other developments in the area of climate and other data that are now available on the web (see below). Independent of the report by the CIRAD team, CIP (International Potato Center) and the IPGRI have collaborated since 1999 in developing the software DIVA-GIS. This software is a Geographical Information System tailor-made for genetic resources applications. The DIVA-GIS may be downloaded free from the Internet at http://diva-gis.org/. The question of availability of collecting grids remains open and is currently being discussed with the DIVA-GIS developers. In the future, it will be useful to standardize the use of these grids at global level - not only for the coconut palm, but also for all crops. It is suggested here to use a grid of 20’ of latitude x 20’ of longitude instead of 40 km x 40 km squares. Such a grid is easier to draw using a GIS or even a commercial map, by interpolating available parallels and meridians. At the equator, the side 57 CHAPTER 2: Locating and collecting germplasm of cell is about 1852 km x 20= 37 km. As latitude increases, the N-S sides remain constant, while the E-W sides decrease progressively. However, it is still close to 35 km at latitude of 20°. Thus, at least at subtropical latitudes, it is almost equivalent to using grids measured in km or in minutes. Discussion on this proposal is in progress. Germplasm collecting programmes are best carried out in two stages. The first phase consist of exploration and preliminary survey to collect information on sites and material that occurs in those sites which will permit better planning of the second phase. The second phase is the more systematic collecting mission. Following the geographic grid approach, the first step will be to gather considerable data in situ (such as fruit component analysis, evidence of erosion, etc.) and samples for DNA testing. The data gathered during the exploration phase will then be analyzed, including using GIS tools. The next step will consist of returning to a limited number of specific sites that are expected to have high, unique, new, useful or threatened coconut genetic diversity, based on the information gathered in phase I, in order to harvest seednuts and bring them back to the genebank(s). The information from areas where no collecting takes place will have value for ground-truthing the theoretical distribution of coconut cultivation (see section on geographical gaps in this article), as well as for determining future on-farm conservation sites and monitoring genetic erosion. Up to now, there is no example of such a strategy using both in situ field characterization and DNA analysis as a decision-making process. However, with the microsatellite tool kit ready for use, this is expected to occur in the near future. Although the two-phase collecting as described above would be ideal, for practical reasons including financial and time constraints, it may be impossible to visit the same place twice as suggested. An alternative method would be to collect directly seednuts and/or the embryos, and leaflet samples, at the same time, along with in situ characterization data such as fruit components. Back at the germplasm centre, DNA from the leaflets or from nuts germinated in the nursery should be analyzed to decide on which samples to include in the genebank as ‘accessions’ i.e., all the populations sampled may not be planted in the genebank. The objective is to use the diversity and other observation data to enable planting only the accessions representing particularly high, unique, new, useful or threatened genetic diversity. This is important as the maintenance of large number accessions in field genebanks by national organizations is very difficult and very expensive. Therefore, genebanks with a minimum number of accessions that capture maximum useful genetic diversity are needed. It must be noted, however, that although some samples may not be 58 COCONUT GENETIC RESOURCES included in the genebank, the data (including the collecting data) on all samples would be very useful to maintain for mapping purposes. The farmer participatory approach There is a growing recognition that the effective conservation of biodiversity will depend on the long-term participation and understanding of local communities. Participatory Rural Appraisal (PRA) comprises a set of techniques aimed at shared learning between local people and outsiders (Baker 2000). Collectors require training in specialized participatory methodologies such as PRA, in particular the use of visual methods (sketches, ranking, diagramming, and cognitive mapping). Important considerations include how to choose informants, the best time for consultations, whether individual interviews should be complemented with group discussions, and ethical issues such as informed consent and anonymity (Ramanatha Rao et al. 1998; Eyzaguirre and Batugal 1999). An example from India may reveal a quite surprising aspect of the PRA method, however. This example was found in a research report distributed during the 1998 COGENT Steering Committee meeting held in Kuala Lumpur, Malaysia. The report states that in India, farmer’s participatory survey was conducted in eight sites representing the three major agro-climatic regions of Kerala. At each site, the interaction was based on a semi-structured questionnaire and lasted some 6-8 hours. The popularity of various coconut varieties was evaluated, including: Tall types, Dwarf x Tall hybrids such as COD x WCT (Chowgat Orange Dwarf x West Coast Tall), and the ‘Natural Cross Progeny of the Chowgat Orange Dwarf’ (NCD). According to participants, many farmers produced NCDs by sowing their own Dwarfs nuts and selecting off- types based on their brown petiole colour for their own use as well as for sale within the locality. In all the eight study sites, the participants favoured off-types of COD (NCDs) in place of TxD and DxT hybrids for cultivation. However, these NCDs are nothing more than natural DxT hybrids! The brown colour of NCDs petiole indicates that the Red Dwarf, as mother palm, is naturally crossed with Green or Brown Coconut palms, i.e. the West Coast Tall coconut available all around in farmers’ fields as male parents. So, the two cultivars compared – Hybrids and NCDs – are in fact the same genetic material. This point was not underlined by the researchers in charge of the PRA survey and analysis. Anyway, it demonstrates that the farmers indeed practice a certain amount of crop improvement and are able to generate their own hybrid seednuts. But the only difference between NCDs and Dwarf x Tall hybrids is that research centres release ‘hybrids’, while 59 CHAPTER 2: Locating and collecting germplasm NCDs are selected by farmers in their own gardens. That may explain the farmers’ preference. Application of PRA methods to obtaining crucial information on the origin and extent of the genetic diversity that is being collected would be most useful in areas where people maintain the closest relationship with their coconut palms. Surveys conducted in archipelagos such as Cook Islands and Tuvalu indicate that germplasm diversity and knowledge seem to be higher in the most isolated islands (Labouisse and Bourdeix 2003; Caillon 2003). This type of information helps in the collecting process, in particular: Locating and accessing target areas and material. Locating target germplasm means being in the right place at the right time. Specialist local knowledge is often the best guide not only to where a particular variety may be found, but also to the optimal timing of collecting. Deciding what to collect and how. When material with particular characteristics is being sought, indigenous knowledge can provide crucial clues. Assessing the completeness of collecting. Local men and women know which varieties are grown in their village or district or are being sold in the local markets. A checklist compiled on the basis of such information can act as a guide to collecting in a given area, providing a benchmark for comprehensive sampling of the available diversity. Understanding the origin and distribution of diversity. Landraces are at least partly shaped by what may be referred to as the informal plant breeding and seed production and supply systems. Thus, understanding the diversity within a crop in an area (which is crucial to developing a conservation strategy) means understanding the practices of the people who grow it. Assessing the reasons for, extent and danger of genetic erosion. Oral testimony is often the only source of information on change in the extent of cultivation of a crop, and in the cultural practices being used. Older farmers will sometimes remember the names and attributes of landraces, which they no longer grow, and which may have entirely disappeared from their area. Documenting and using the collection. Local knowledge should form an important part of the documentation of germplasm samples. Farmers are aware of the many characteristics and properties of varieties. 60 COCONUT GENETIC RESOURCES Documenting such local knowledge of the appearance, properties and adaptations of germplasm should be seen as an integral part of the characterization and evaluation process, and as such as an important way of facilitating and accelerating the use of conserved germplasm. Conclusion Though it is now well recognized that a significant amount of coconut diversity has been collected and conserved in several coconut research organizations, especially since the establishment of COGENT, their representation and availability of associated data are still incomplete. There is still substantial uncollected indigenous germplasm, and some of it is under threat of genetic erosion. The most important reason for the continued occurrence of coconut diversity is that farmers have interest in and possess knowledge about their coconut varieties. However, along with the diversity, such knowledge is rapidly eroding in some areas as so-called modernization and globalisation reach into even the most remote parts of the world. Researchers will have to focus on breeding and germplasm utilization to benefit from the investment made in collecting and conserving. Emphasis should be placed on the use of molecular techniques and morphological characterization to rationalize large collections in order to reduce the actual number of cultivars in the germplasm centres from around 350 to 150-200, so that the genebanks are more manageable, both in terms of financial and human resources and scientific backstopping. Then additional collecting, using these new screening techniques, should allow adding 150-200 more priority accessions. The use of Geographical Information Systems tools will facilitate the task of the collectors. Some elements were discussed regarding the effectiveness of targeted collecting, as compared to comprehensive grid sampling and farmer participatory methods. Use of the concept of CCIV could further help in identifying the priority accessions to be included in genebank collections and training to implement collecting strategy and the use of GIS tools is considered important to enhance the efficiency of collecting. Thick-husked varieties from Asia/Pacific and sweet husk varieties are two endangered phenotypes that should be targeted. Surveys that are more systematic should be conducted in areas that have not been covered during previous collecting programmes. Some important accessions that have been lost in collections should also be re-collected. Farmer’s participatory methods should be applied in communities where people know a great deal about every coconut palm in their gardens (such as very isolated islands) to document the knowledge and practices farmers use to maintain coconut diversity in their fields. 61 CHAPTER 2: Locating and collecting germplasm References Ashburner, GR, MG Faure and MA Foale. 1994. Methods for coconut germplasm prospection. Pp. 41-43. In: MA Foale and PW Lynch (eds). Coconut improvement in the South Pacific. Proceedings of a workshop held in Taveuni, Fiji Islands, 10-12 November 1993. ACIAR, Canberra. Baker, JL. 2000. Evaluating the impact of development projects on poverty: A handbook for practitioners. LCSPR/PRMPO, the World Bank, Washington D.C. Baudouin, L. 2001. 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Proceedings of the COGENT Regional Coconut Genebank Planning Workshop, 26-28 February 1996, Pekanbaru, Riau, Indonesia. IPGRI-APO, Serdang, Selangor, Malaysia. Rognon, F and P Batugal. 1998. Evaluation of Indonesia as a regional coconut genebank host country. Pp. 41-47. In: V Ramanatha Rao and P Batugal (eds). Proceedings of the COGENT regional coconut genebank planning workshop held at Pekanbaru, Indonesia, 26-28 February 1996. IPGRI/COGENT, Rome. Pp 41-47. Santos, GA, PA Batugal, A Othman, L Baudouin and JP Labouisse. 1996. Manual on standardized research techniques in coconut breeding. IPGRI-COGENT, Singapore. 46pp. Schuiling, M, DA Kaiza and HC Harries. 1992. Lethal disease of coconut palm in Tanzania. III. Low resistance of imported germplasm. Oléa- gineux 47(12): 693-697. Sokal, RR and RF Rolf. 1981. Biometry. Freeman, San Francisco. Whitehead, RA. 1966. Sample survey and collection of coconut germplasm in the Pacific islands, 30 May- 5 September 1964. Ministry of Overseas Development and HMSO, London. 64 COCONUT GENETIC RESOURCES Williams, DG. 1990. An annotated bibliography of the natural history of the Cocos (Keeling) Islands, Indian Ocean. Atoll Research Bulletin 331:1-17. Zizumbo-Villareal, D, F Hernandez and HC. Harries. 1993. Coconut varieties in Mexico. Economic Botany 47:65-78. 65 CHAPTER 2: Locating and collecting germplasm In vitro collecting of coconut germplasm F Engelmann1, 2 1Institut de Recherche pour le Développement (IRD), 911 Avenue Agropolis, BP 64501, 34394 Montpellier Cedex 5, France 2Honorary Research Fellow, International Plant Genetic Resources Institute (IPGRI), Via dei Tre Denari 472/a, 00057 Maccarese (Fiumicino), Rome, Italy Introduction In vitro collecting (i.e., the utilization of in vitro culture techniques for collecting plant germplasm) offers the plant collector an additional option for solving various problems which can be encountered during collecting expeditions. The application of in vitro collecting is particularly useful for the two main categories of problem crops (i.e., vegetatively propagated species and species with recalcitrant seeds) (Withers 2002). In vitro collecting protocols have now been developed for a number of different species (Pence et al. 2002). In the case of coconut, seeds are bulky and heavy, making them costly to transport. They are also highly recalcitrant (Chin and Roberts 1980). These characteristics limit the amount of material that can be collected and restrict the geographic range of collecting missions. These limitations may have serious consequences for genetic resources conservation, since it is recognized that a large amount of the untapped genetic diversity in coconut is located in remote areas, such as atoll islands. The key to solving these problems, however, lies in recognizing that only the embryo is needed to propagate a coconut palm. Various efficient in vitro culture protocols are available which allow the production of whole plantlets from coconut zygotic embryos inoculated in vitro (Batugal and Engelmann 1998; Engelmann et al. 2002). Status of work Research on the adaptation of in vitro culture techniques to collecting coconut embryos was initiated 15 years ago under the aegis of the IBPGR (International Board for Plant Genetic Resources, the predecessor of International Plant Genetic Resources Institute -IPGRI), with the aim of facilitating not only the collecting but also the international exchange of coconut germplasm. In addition to the advantages offered by this technique for collecting genetic resources, in vitro collecting would also avoid the transmission of important coconut diseases, which do not pass through the embryo. This is particularly important with the expected increase in international exchange of coconut germplasm linked with the establishment of the multi-site International Coconut Genebank 66 COCONUT GENETIC RESOURCES (Ramanatha Rao and Batugal 1998). Various in vitro collecting techniques have been developed by different teams, thereby demonstrating not only the feasibility of collecting isolated embryos, but also the great flexibility that can be exercised within the basic concept (Engelmann 2002). The in vitro culture of coconut embryos has been adapted by several researchers in collecting coconut germplasm from the field. The techniques basically include the following sequence of operations: • Dehusking and cracking open the nut; • Extracting a plug of endosperm containing the embryo by using a cork borer; • Dissecting the embryo from the endosperm; and • Inoculating the embryo into culture. The methods developed differ in the degree to which attempts are made to reproduce laboratory conditions in the field, the amount of in vitro work actually performed in the field, and, therefore, the point at which sterilization is carried out. Their utilization requires varying levels of technical expertise, and the method selected will depend on the circumstances of the collecting mission and on the tissue culture expertise available among the collecting team. The simplest methods, which do not require specific expertise at the collecting site, are one of the two methods developed in Côte d’Ivoire (Assy-Bah et al. 1987) and that established in the Philippines (Rillo and Paloma 1991; Rillo 1995). In the first protocol developed by Assy-Bah et al. (1987), after disinfection, the plugs of endosperm containing the embryos are placed in a solution of KCl (16.2 g l-1), then brought back to the laboratory where they are redisinfected and inoculated in vitro under the laminar flow (see Protocol 1 below). In the protocol developed in the Philippines, plugs of endosperm containing the embryos are extracted in the field, brought to a simple isolation room close to the collecting site, disinfected with alcohol and commercial bleach, placed in sterile plastic bags with sterile, moist cotton and transported in cold storage. Upon arrival in the laboratory, subsequent manipulations are carried out aseptically, under the laminar flow hood. The cylinders of endosperm are resterilized with commercial bleach, and the embryos are extracted and inoculated in vitro for germination and growth. This protocol is used routinely in the Philippines in the framework of programmes for mass production of Makapuno embryos (Rillo 1999). Another protocol, which has been established by Australian researchers, requires some tissue culture expertise because embryos have to be extracted from the albumen immediately after their collection, but allows transport time of up to six weeks (Ashburner et al. 1995, 1996; 67 CHAPTER 2: Locating and collecting germplasm Samosir et al. 1999). Plugs of endosperm are collected in the field and transported to an improvised laboratory close to the collecting site, where the embryos are extracted from the albumen, sterilized with commercial bleach, and inoculated into 2 ml sterile plastic cryotubes containing sterile water. Manipulations after arrival in the laboratory are performed aseptically under the laminar flow hood. The embryos are resterilized and inoculated in vitro for germination and growth In the other protocols (i.e. the second protocol developed in Côte d’Ivoire, see below) (Assy-Bah et al. 1987) and those established by Sossou et al. (1987) and Karun et al. (1993), in vitro inoculation of the embryos is performed directly at the collecting site, thus requiring the relevant expertise to be available within the collecting team. The field equipment requirements are greater than in the protocols described above, but even these methods range in complexity. The technique of Sossou et al. (1987) attempts to simulate laboratory facilities and methods in the field using an inflatable glove box. The protocols established by Assy-Bah et al. (1987) and Karun et al. (1993), however, accept the limitations of working in the field and present a lower-technology approach. Endosperm plugs are extracted from the nuts and disinfected with commercial bleach. The embryos are then dissected and inoculated inside a wooden or plexiglass box (to protect from airborne contaminants) and transferred into sterile culture tubes. With the protocol developed by the research team from India, embryos are either directly inoculated on growth medium or kept for 2-4 months in sterile water (Karun et al. 1996). This protocol has been used successfully by Indian researchers to collect several thousand embryos from remote Indian Ocean islands (Karun et al. 1998; 2002). All these protocols give good results, with contamination percentages below 10% of the inoculated embryos. Detailed description of the in vitro collecting protocols devel- oped by Assy-Bah et al. (1987) Assy-Bah et al. (1987) developed two coconut embryo in vitro collecting protocols - one consisting of storing the disinfected embryos in a KCl solution until they are brought back to the laboratory, where they are redisinfected and inoculated in vitro under sterile conditions, and the other including in vitro inoculation of the embryos in the field. Details of the protocols are as follows: Protocol 1 (inoculation of embryos in the laboratory) Preliminary operations are performed in the open air, on a folding table that has been washed and disinfected with a bleaching solution. 1. Select and dehusk mature nuts. 2. Break nuts open with a clean hammer. 68 COCONUT GENETIC RESOURCES 3. Use a cork borer to remove a cylinder of solid endosperm containing the embryo, and use forceps to transfer the cylinder to a jar containing 500 ml of commercial bleach. Disinfect all instruments with commercial bleach and sterilize in the flame of the gas burner. 4. Immerse batches of 25 cylinders in commercial bleach for 20 minutes. 5. Immediately after disinfecting, transfer endosperm cylinders without rinsing in individual 30 ml containers containing 15 ml KCl solution (16.2 g/l). The following steps are performed in the laboratory, under the laminar airflow cabinet. 1. Remove endosperm cylinders from the KCl solution and immerse in batches of 25 cylinders in commercial bleach for 20 minutes. 2. Place one cylinder in a sterile Petri dish and dissect out the embryo using forceps and a scalpel. Flame dissecting tools before manipulating a new embryo to reduce the risk of cross- contamination. 3. Rinse the embryo once in sterile water (using one flask per embryo to reduce the risk of cross-contamination) and transfer it to solid medium in a culture tube. 4. Seal the tube with cling film and place it on a rack for culture in the growth room. Protocol 2 (inoculation of embryos in the field) Steps 1-5 are the same as in Protocol 1 above. The following steps are performed inside a wooden box, which provides some protection from external contaminants. The inside walls of the box are disinfected with bleach. 1. Place one cylinder in a sterile Petri dish and dissect out the embryo using forceps and a scalpel. Flame dissecting tools before manipulating a new embryo to reduce the risk of cross- contamination. 2. Rinse the embryo once in sterile water (using one flask per embryo to reduce the risk of cross-contamination) and transfer it to solid medium in a culture tube. 3. Seal the tube with cling film and place it on a rack for transport to the laboratory. Using Protocol 2, contamination was around 10%, while it was only around 5% with Protocol 1. No differences were noted in germination 69 CHAPTER 2: Locating and collecting germplasm and development between embryos treated following Protocols 1 and 2. Embryos could be stored for up to 14 days in the KCl solution without any effect on their further development. After direct inoculation in the field (following Protocol 2), embryos could be kept in semi-solid medium under non-controlled environmental conditions for two months before being grown in the culture room of a laboratory (Engelmann and Assy- Bah 1992). These results were confirmed recently by N’Nan (personal communication) following a series of in vitro collecting experiments performed in Côte d’Ivoire in 2001. In vitro collecting has been used routinely to collect and send over 20000 embryos from Côte d’Ivoire to France over the last two years. In vitro culture of embryos After inoculation in vitro, embryos have to be germinated and grown into weanable (acclimatized and hardened) plantlets. Research towards the development of in vitro culture protocols has been performed over the last 30 years by various research teams worldwide. An assessment of the available protocols, carried out during the IPGRI/COGENT-funded International Coconut Embryo Culture and Acclimatization Workshop held in the Philippines in 1997, revealed a large discrepancy in the performance of these in vitro culture protocols, with 14 to 55% of the inoculated embryos giving rise to plantlets growing in vivo (Engelmann 1998). The main bottleneck was the low efficiency of in vitro embryo germination and plantlet development. The protocols developed also differed in the culture conditions, composition and sequence of media employed and the stage of plantlets selected for weaning. Also, these protocols had been tested with a limited number of coconut varieties. In this workshop, which was participated by seven countries, the embryo culture techniques of country participants were compared and good features were adopted to develop an upgraded protocol to be further tested. The results of this workshop were published to guide embryo culture researchers (Batugal and Engelmann 1998). Another IPGRI/ COGENT-coordinated international project, funded by the UK Department for International Development (DFID), was thus implemented to address two main objectives: (1) to improve the maturation and germination of embryos, and their development into plantlets; and (2) to determine and select the most efficient in vitro culture protocol and to test it with a large number of varieties (Batugal and Engelmann 1998). At the end of this project, the success of coconut embryo in vitro culture was significantly improved, with 31 to 81% of inoculated embryos developing into plantlets in vivo (Engelmann and Batugal 2002). A large diversity of coconut germplasm was employed since the tissue 70 COCONUT GENETIC RESOURCES culture protocols have been tested with over 20 varieties. These experiments also revealed a very strong genotypic effect in response to in vitro culture. No optimal protocol was identified due to the high variability of the responses obtained in the different laboratories involved in the project. However, the ‘hybrid protocol’ proposed by one laboratory, which combines the most efficient steps of the four protocols tested, seems to hold good promises for further improving the performances of coconut embryo in vitro culture. Zygotic embryos have also been employed as starting material for large-scale propagation of coconut genotypes through somatic embryogenesis (Verdeil et al. 1999). However, the reactivity of coconut tissues to in vitro manipulation is very low, and only few plantlets have been obtained from a limited number of coconut accessions. Additional research is therefore required before large-scale propagation of coconut through somatic embryogenesis can be undertaken. Conclusion and prospects The various examples of in vitro collecting protocols developed for coconut embryos range from extreme simplicity to a relatively high level of sophistication and illustrate the flexibility and adaptability of the basic concepts of the procedure. It is with coconut that the largest amount of research has been directed towards the establishment of in vitro collecting protocols because of the particular difficulties encountered with germplasm collecting and exchange for this species. In vitro collecting is currently used on a routine basis for coconut more than with any other species. The utilization of this technique is expected to increase with the establishment of COGENT’s multi-site International Coconut Genebank, thus making coconut one of the best models for the application of in vitro collecting. References Ashburner, GR, MG Faure, DR Tomlinson and WK Thompson. 1995. A guide to the zygotic embryo culture of coconut palms (Cocos nucifera L.). ACIAR Technical Report No 36, Canberra, Australia. Ashburner, GR, MG Faure, DR Tomlinson and WK Thompson. 1996. Collection of coconut (Cocos nucifera L.) embryos from remote locations. Seed Science Technology 24:159-169. Assy-Bah, B, T Durand-Gasselin and C Pannetier. 1987. Use of zygotic embryo culture to collect germplasm of coconut (Cocos nucifera L.). Plant Genetic Resources Newsletter 71:4-10. Batugal, PA and F Engelmann (eds). 1998. Coconut embryo in vitro culture. Proceedings of the first workshop on embryo culture, Banao, 71 CHAPTER 2: Locating and collecting germplasm Guinobatan, Albay, Philippines, 27-31 October 1997. IPGRI-APO, Serdang, Malaysia. Chin, HF and EH Roberts (eds). 1980. Recalcitrant crop seeds. Tropical Press Sdn. Bhd., Kuala Lumpur, Malaysia. Engelmann, F. 1998. Current state of the art and problems in in vitro culture of coconut embryos. Pp. 6-11. In: PA Batugal and F Engelmann (eds). Coconut embryo in vitro culture. Proceedings of the first workshop on embryo culture, Banao, Guinobatan, Albay, Philippines, 27-31 October 1997. IPGRI-APO, Serdang, Malaysia. Engelmann, F. 2002. Coconut. Pp. 68-71. In: VC Pence, JA Sandoval, VM Villalobos and F Engelmann (eds). In vitro collecting techniques for germplasm conservation. IPGRI Technical Bulletin N°7. IPGRI, Rome, Italy. Engelmann, F and B Assy-Bah. 1992. Maintenance of coconut genetic resources: In vitro techniques for medium and long-term conservation. Pp. 63-69. In: Papers of the IBPGR Workshop on coconut genetic resources, Cipanas, Indonesia, 8-11 Oct. 1991. International Crop Network Series No. 8. IBPGR, Rome. Engelmann, F and P Batugal. 2002. Background on the development and implementation of the coconut embryo in vitro culture project. Pp. 1- 4. In: F Engelmann, P Batugal and JT Oliver (eds) Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Malaysia. Engelmann, F, P Batugal and JT Oliver (eds). 2002. Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Malaysia. Engelmann, F and B Assy-Bah. 1992. Maintenance of coconut genetic resources: In vitro techniques for medium and long-term conservation. Pp. 63-69. In: Papers of the IBPGR Workshop on coconut genetic resources, Cipanas, Indonesia, 8-11 Oct. 1991. International Crop Network Series No. 8. IBPGR, Rome, Italy. Karun, A, KK Sajini and RD Iyer. 1996. In vitro active conservation of coconut zygotic embryos. Journal of Plantation Crops 24 (Suppl.): 586-593. Karun, A, KK Sajini and VA Parthasarathy. 2002. Increasing the efficiency of embryo culture to promote germplasm collecting in India. Pp. 7-26. In: F Engelmann, P Batugal and JT Oliver (eds). Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Malaysia. Karun, A, A Upadhyay and VA Parthasarathy. 1998. Status of research on coconut embryo culture and acclimatization techniques in India. Pp. 29-36. In: PA Batugal and F Engelmann (eds). Coconut embryo in vitro culture. Proceedings of the first workshop on embryo culture, Banao, Guinobatan, Albay, Philippines, 27-31 October 1997. IPGRI- APO, Serdang, Malaysia. 72 COCONUT GENETIC RESOURCES Karun, A, S Shivashankar, KK Sajini and KV Saji. 1993. Field collection and in vitro germination of coconut embryos. Journal of Plantation Crops 21 (Suppl.): 291-294. Pence, VC, JA Sandoval, VM Villalobos and F Engelmann (eds). 2002. In vitro collecting techniques for germplasm conservation. IPGRI Technical Bulletin N°7. IPGRI, Rome, Italy. Ramanatha Rao, V and P Batugal (eds). 1998. Proceedings of the COGENT regional coconut genebank planning workshop, Pekanbaru, Riau, Indonesia, 26-29 February 1996. IPGRI-APO, Serdang, Selangor, Malaysia. Rillo, EP and MB Paloma. 1991. Storage and transport of zygotic embryos of Cocos nucifera L. for in vitro culture. Plant Genetic Resources Newsletter 86: 1-4. Rillo, EP. 1995. Embryo culture of coconut: A laboratory manual. Philippine-German Coconut Tissue Culture Project, PCA Albay Research Center, Banao, Guinobatan, Albay, Philippines. Rillo, EP. 1999. Coconut embryo culture. Pp. 279-288. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardeña and JM Santamaria (eds). Current advances in coconut biotechnology Kluwer Academic Publishers, Dordrecht, The Netherlands. Samosir, YMS, ID Godwin and SW Adkins. 1999. A new technique for coconut (Cocos nucifera L.) Germplasm collection from remote sites: Culturability of embryos following low-temperature incubation. Australian Journal of Botany 47:69-75. Sossou, J, S Karunaratne and A Kovoor. 1987. Collecting palm: In vitro explanting in the field. Plant Genetic Resources Newsletter 69:7-18. Verdeil, JL, R Hornung, HJ Jacobsen, E Rillo, C Oropeza, R Bourdeix, YP N’Cho, V Hocher, S Hamon and A Sangaré. 1999. Recent progress on coconut micropropagation through a joined effort involving different countries. Pp. 391-405. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardeña and JM Santamaria (eds). Current advances in coconut biotechnology. Kluwer Academic Publishers, Dordrecht, The Netherlands. Withers, LA. 2002. In vitro collecting: Concept and background. Pp. 16- 25. In: VC Pence, JA Sandoval, VM Villalobos A and F Engelmann (eds). In vitro collecting techniques for germplasm conservation. IPGRI Technical Bulletin N°7. IPGRI, Rome, Italy. 73 CHAPTER 3: Germplasm conservation Chapter 3 Germplasm conservation 74 COCONUT GENETIC RESOURCES 75 CHAPTER 3: Germplasm conservation Complementary conservation of coconuts ME Dulloo1, V Ramanatha Rao2, F Engelmann3 and JMM Engels5 1Scientist, 3Honorary Research Fellow and 5Genetic Resources Management Advisor, Inter- national Plant Genetic Resources Institute (IPGRI), via dei Tre Denari, Maccarese, Rome, Italy 2Senior Scientist, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia 4Institut de recherche pour le Developpement (IRD), BP 64501, Cedex 5, Montpellier, France Introduction The main objective in any plant genetic resources (PGR) conservation programme is to maintain the highest possible level of genetic variability present across the genepool of a given species or crop both in its natural range and in a germplasm collection. The importance of conserving genetic variability or diversity is well recognized and such defense mechanisms need to be introduced into modern cultivars to make them sustainable (Martin et al. 1991; Chang 1994; Kannenberg and Falk 1995). Countries that are signatories to the Convention of Biological Diversity endowed with significant amount of genetic and species diversity have a responsibility to the world at large to conserve them and make them available for use (Ramanatha Rao 1999). It is generally recognized that the two approaches of conservation, ex situ and in situ, are both important in the conservation and use of genetic diversity and should be regarded as complementary to each other (Maxted et al. 1997; Dulloo et al. 1998; Ramanatha Rao 1998; Engels and Wood 1999). The ultimate purpose of germplasm conservation is use and, consequently, any conservation strategy should include mechanisms that will ensure access to the germplasm by relevant stakeholders. Other important issues that must be addressed in a conservation strategy include issues related to policy and legal frameworks, documentation, socioeconomic aspects, infrastructure and networks. Since needs of users and technologies may change over time influencing the ways in which genetic resources are conserved and used in future and hence, should be taken into consideration when designing a conservation strategy. At an in house meeting organized by IPGRI in 2002, a complementary conservation strategy was defined as “the combination of different conservation actions, which together lead to an optimum sustainable use of genetic diversity existing in a target genepool, in the present and future.” A conservation strategy for coconut has been discussed in the past (Ramanatha Rao and Engelmann 2000; Ramanatha Rao et al. 1998) and the current status of the various conservation methods available for coconuts are described in this chapter. This paper discusses the constraints 76 COCONUT GENETIC RESOURCES and advantages of these methods, the elements for a complementary conservation strategy and attempts to provide a framework from which a working strategy for conservation and use of coconut germplasm could be taken forward. Methods for conserving coconut germplasm As noted earlier, PGR are commonly conserved using ex situ or in situ approaches. Ex situ refers to their conservation outside their natural habitat in facilities such as in seed banks, field genebanks, in vitro collections, botanic gardens, with germplasm conserved in the form of plants, seeds, pollen, tissues, cells or DNA. In contrast, in situ conservation is conserving germplasm in the natural habitat where the target species is found, and in habitats such as farms and home gardens, where the species have developed their distinctive properties as a result of long- term selection by humans. The latter applies particularly to cultivated plants and their cultivars, landraces and weedy forms. Generally, there are three categories of in situ reserves: namely, those which maintain optimum conditions such as national parks and nature reserves, those which allow a range of economic activities by indigenous people as in extractive reserves national forests and Biosphere reserves, and a third category where local people act as custodians for the traditional varieties and selections contained in home gardens and farms (Damania 1996). Furthermore, IUCN (1994) classifies protected areas into six categories according to broad management objectives. The previous chapters have described in detail the current status of conservation techniques coconut germplasm, an analysis of which could help in developing a complementary conservation strategy for coconuts. Table 1 examines the feasibility of different techniques, while Table 2 summarizes the constraints and advantages with regard to each of these methods. These two tables provide a comparative framework on which a complementary conservation strategy for coconuts could be based. It is important to emphasize that the information in Table 1 is based on our current knowledge, which could rapidly change in the near future, due to progress made in the development of the conservation methodologies. The biological characteristics of coconut and their compatibility with different options available are briefly discussed below. It is important to underline a fact at this point that when several options are combined to bring about a complementary strategy, we also bring along their advantages and disadvantages with the expectation that a synergistic effect is achieved. The options for conserving coconuts are dependent on the biological characteristics of the whole plant and its component organs and tissues, 77 CHAPTER 3: Germplasm conservation as well as on the state of the technology as applied to coconuts. Coconut, a perennial palm, with exception of most of the Dwarfs, is an outbreeding species. It bears large size seeds that exhibit recalcitrant storage behaviour, rendering seed conservation not possible. Being a perennial crop, coconuts can be conserved ex situ as live plants in field genebanks or botanic gardens or in situ either on farm or in home gardens or on remote islands and atolls. Botanic gardens have limited capacity to conserve a broad range of genetic diversity due to the low number of plants that they can maintain. Field genebanks (attached to a coconut improvement organization) have been the preferred mode of coconut conservation to date, as they can be integrated into institutions and do not require highly technically skilled workers (Ramanatha Rao et al. 1998). However, field collections have some major disadvantages (Table 2). Coconuts are generally outbreeding, especially the Tall types, and requires wither spatial isolation or assisted-pollination. There are still some important research questions to be addressed in regard to collection management such as minimum number of trees needed to maintain representative genetic diversity, field plot techniques for characterization and evaluation and economics of collection maintenance. For details on field genebanks and the ICG, see related articles in this chapter. On-farm conservation, where traditional crop cultivars or landraces and/or farming systems by farmers within traditional agricultural systems are maintained (Hodgkin et al. 1993; Jarvis 1999), has been gaining importance over the last decade. For coconuts, this method is particularly advantageous since most of the stands in South and Southeast Asia are in more or less intensively managed areas. For effective on-farm conservation, knowledge on the effects of farmers’ practices on the extent and distribution of genetic diversity information on history of coconut cultivation and indigenous knowledge and actual genetic diversity measurements may be required. It is now possible to monitor and estimate genetic diversity using molecular markers for coconuts (Foale 1992; Ashburner and Rhode 1994; Lebrun et al. 1998; Mpunami et al. 1998; Perera et al. 1999). People’s participation and cooperation among local people, researchers and conservationists and non-governmental organizations (NGOs), are essential ingredients of success for the sustainability of on-farm conservation efforts. Furthermore, any in situ conservation programme must benefit the local communities. Establishment of areas of intensive management or high yielding plantations would assist long-term sustainability of in situ conservation programmes. This is not to replace, but to bring a balance between high- yielding types for purely commercial purpose and landraces to satisfy all the personal and social needs of farmers. Such a balance is essential to 78 COCONUT GENETIC RESOURCES promote conservation of landraces in the absence of any specific additional benefits to growers. This can attract commercial and private agencies to be partners in on-farm conservation efforts and can lead to much wanted linkages between public, community and private sectors in PGR conservation. For naturally occurring coconuts palms other forms of in situ conservation such as island reserves, biosphere reserves may have very important complementary value in conserving unique diversity as for example populations isolated on small uninhabited islands and atolls (Ramanatha Rao et al. 2000). For more detailed description of on-farm conservation, see related articles in this chapter. Progress achieved in recent years in in vitro culture and cryopreservation as potential methods for conserving coconut germplasm augurs well for the future. Research on the development of such techniques has been performed with zygotic embryos, somatic embryos, pollen, apices and DNA material (Assy-Bah and Engelmann 1993). In vitro culture of zygotic embryos has been significantly improved and is now operational in an increasing number of laboratories (Engelmann et al. 2002). An efficient cryopreservation protocol has been developed for zygotic embryos (Assy-Bah and Engelmann 1992), which needs to be refined and tested on a range of ecotypes before becoming fully operational. Somatic embryos cannot be used for germplasm conservation since in vitro propagation of coconut using somatic embryogenesis is not yet functional. Cryopreservation of apices (‘plumules’) sampled from zygotic embryos is also possible (Hornung et al. 2001; Malaurie et al. 2002) but regeneration of whole plants from such explants is difficult. At the moment, plumules are of no use for germplasm conservation except possibly in case it would be proven that diseases (MLOs) can be transmitted through the embryo. DNA material can be cryopreserved easily and can be of great value in genetic diversity studies. However, regeneration into whole plants is problematic, if at all possible. For more details, see related articles in this chapter. Conservation of coconut pollen is an additional option. Pollen can be dried and stored under vacuum for a short period of time (2-6 months) in a domestic deep freezer (Rognon and de Nucé de Lamothe 1978). Freeze-drying experiments showed no viability loss after 3 and 6 months (Whitehead 1966; Benard 1973) of storage at room temperature. Coconut pollen is highly tolerant to desiccation and preliminary experiments have demonstrated that coconut pollen could be successfully cryopreserved (Dr. Assy-Bah, unpublished results). Long-term storage of coconut pollen under cryopreservation would represent an important additional technique for genetic resources conservation, by allowing conservation of genes. However, additional research is still needed to further develop and refine a cryopreservation protocol. 79 CHAPTER 3: Germplasm conservation Considerations for complementary conservation strategy The knowledge of the biological characteristics of coconuts and how they can be conserved, as discussed above, is just one of the many elements for developing a sustainable complementary conservation strategy. This section discusses some other important elements, which need to be taken in to consideration. Conservation objective The most central element for developing a strategy is to define precisely what the objectives are. In this case, the general objective is to conserve and utilize maximum coconut genetic diversity. However, there would be other minor objectives for the establishment of a coconut genebank such as for immediate utilization, conservation for the long term, focusing on characterization and evaluation, etc. Strategy applied will also depend on what one would want to conserve, i.e. genes or genotypes. The strategy will be very different if the objective is to completely stop the evolutionary processes (e.g. cryopreservation) or in case the evolutionary processes need to be maintained (as in in situ conservation). Thus, if the promotion of conservation of landraces becomes the main objective, conservation on farm becomes the choice strategy for coconut, which also provides an opportunity for coconut to evolve under natural and farmer-imposed conditions. However, there is a need to accumulate more evidence on the role of farmer selection in a perennial crop like coconut. At the same time, with many farmers interested in increasing the productivity of coconut and income generation, breeding for higher yields and multiple uses becomes priority and hence ex situ conservation in field genebanks, which enhance the access to diversity by the coconut improvement scientists, becomes the choice for conserving and using maximum genetic diversity. Genetic diversity The major objective of any conservation effort, especially the one for long- term, is the conservation of maximum genetic diversity in a crop gene pool and this is true for coconut as well. Hence, the factors that contribute to the maximization of genetic diversity in a coconut collection (only infraspecific diversity in the case of coconut) have significant bearing on the balance of options chosen for inclusion in a conservation strategy. Coconut belongs to a monotypic genus and hence all its genetic diversity is in one species, i.e. Cocos nucifera. The diversity in coconut is mainly in the different ecotypes/landraces, i.e. conservation of genotypes and, consequently, using the field genebank allows conservation of most genetic diversity in the gene pool. Since very little information is available on the extent and distribution of coconut genetic diversity within and between 80 COCONUT GENETIC RESOURCES populations and the genetics of useful traits, probability theory and random sampling (at times modified to include some level of bias for elite material, which is generally the norm for horticultural and perennial species) and larger populations are used to locate and conserve the desired level of genetic diversity. Under the ADB-funded project of COGENT, 28 countries have collected and conserved coconut germplasm in national field genebanks and a multi-site International Coconut Genebank (ICG) has been established, which makes the access to genetic diversity easy (see Batugal and Kanniah in this chapter). At the same time, COGENT also recognizes the limitations of the field genebanks. By using on-farm conservation, it is possible to conserve more diversity, especially that diversity which is directly useful to farmers. To do proper on-farm conservation, essential information on the extent and distribution of genetic diversity on farms is being generated. The limited observations to date have shown that very few farmers seem to pay any special attention to phenotypic and other differences in coconut types that they grow. Most often coconuts are just planted and little attention is paid later on. Hence, the so called indigenous knowledge on coconuts seems to be limited. Nevertheless, there are some who recognize this well, and hence should be targets for on-farm conservation efforts. Field genebanks require a substantial number of individual genotypes to be an effective conservation measure. Thus, extensive network of farm sites will be able to complement conservation of genetic diversity in coconut. Stakeholders Conservation of any genepool is not just a responsibility of an organization or individual. Several interested organizations and individuals are involved, including those who were responsible for the generation of the variability in the first place. Thus, in the case of coconut, the interests of small coconut farmers, organizations interested in their welfare and coconut research organizations/scientists and at the end the consumers, etc., need to be considered. For example, coconut farmers for whom coconut growing is a way of life and in some instances, growing the specific landraces or ecotypes, on-farm conservation takes precedence over the other approaches. This needs to be strengthened and complemented by other stakeholders who can play an important role in conserving that part of coconut diversity that might not be conserved on-farm due to reasons such as genetic erosion and utilization, using other complementary approaches such as conservation in field genebanks. Infrastructure The infrastructure needed and their availability determine the option to be chosen. Hence, the infrastructure needs for each of the option and 81 CHAPTER 3: Germplasm conservation their availability and resources required needs to be documented and analyzed. For example, the establishment of a field genebank for coconut genetic resources requires land, labour, good agronomy, facilities for exchange of germplasm, well-trained staff, etc. In vitro culture and cryopreservation would also require specific infrastructure and highly trained skilled staff. For on-farm conservation, identification of sites with high levels of genetic diversity, committed community-based organizations, staff skilled in working along with partners and farmers, access to conservation sites, monitoring mechanisms etc, have to be in place. Once such baseline information is available, then it should be possible to determine which approach will be used to particular part of coconut genetic diversity. Socioeconomic aspects The social considerations probably are more important in implementing on-farm conservation and less so while establishing ex situ conservation facilities. However, the economic aspect would be a key determinant in what methods are utilized. While planning for the former, several issues related to socioeconomics of coconut farming, indigenous knowledge, community participation, etc. have to be considered that make the on- farm conservation sustainable. Such considerations also make germplasm conserved on farm accessible for use by the farmers and communities as well as national agricultural research systems. Generally speaking, in the countries that are interested in conservation and use of coconut, the cultivation of coconut is not greatly threatened and will continue in the end. This consideration is important as establishing either ex situ or in situ conservation programmes are expensive and must be compatible with national objectives. Therefore, it is important to allow the increase of genetic diversity that is actually being planted by farmers to the extent possible. In this respect, a close cooperation between in situ and ex situ efforts is critically important. Network Any complementary conservation efforts for coconut at the national level have to be multidisciplinary and multi-stakeholder driven in order to conserve maximum diversity. Thus, an in-country network consisting of interested individuals, organizations (both public and non-governmental) and farmers is required. Similarly, developing a complementary conservation strategy at an international level requires coordination and collaboration among interested countries, as demonstrated by the International Coconut Genetic Resources Network (COGENT), as the genetic diversity that needs to be conserved is spread across borders. 82 COCONUT GENETIC RESOURCES COGENT has been able to complement the establishment of ICGs with efforts at community level that lead to on-farm conservation of coconut genetic resources. For example, the efforts to promote the cultivation of identified elite germplasm (landraces) from the genebank at sites where poverty reduction work are underway in Bangladesh, India, the Philippines, etc. This will ensure sustained conservation of landraces and at the same time benefit the poor coconut farmers. Costs and risks The options in any complementary conservation strategy need to be weighed against each other keeping in mind the relative costs, benefits and risks. With currently available methods, it has been generally agreed that the establishment of ex situ genebanks is relatively cost-effective and less risky (Pardey et al. 1999). However, using this one method, it would not be possible to conserve all the coconut genetic diversity that might be required in the future, especially when the number of accessions that could be maintained and managed in a field genebank is finite. Hence, a complementation by on-farm conservation of the material that would be difficult to bring to genebank becomes economical. In situ conservation option needs to be incorporated into the strategy. Such efforts also promote conservation through use. In addition, the analysis of the genetic diversity of coconut has shown that significant genetic diversity might exist in remote areas and atolls, collecting of which could be very expensive. One could argue that the germplasm located at these sites are relatively safe except for unfavourable climatic changes (e.g. sea level rise) which may be a risk and has to be considered. If resources are available, efforts should be made to collect and secure them in ex situ collections. Policy/Legal issues Without any doubt, for any conservation approach to be in place, much depends on the type of legal arrangements that can be put in place for transfer and access to genetic material and for sharing of benefits arising out of their use. In many countries, there may not be specific laws that prevent or promote the conservation of coconut genetic resources, but policies in a country could influence the importance accorded to such an effort. Thus, before venturing to establish a conservation strategy for coconut, it is important to check on the priority accorded to coconut at national level. For example, in most countries in the Asia Pacific, high priority is accorded to this crop and hence the efforts on its conservation and use are generally in line with the national policies. As noted earlier, conservation is mainly to make and keep the genetic resources accessible for use by users (researchers, farmers etc.). Hence the policies that promote 83 CHAPTER 3: Germplasm conservation the accessibility and transfer of material and ‘information’ are important for successful implementation of the different types of conservation. For example, if the national laws are very strict about collecting and using the material from farmers (as in the case of Philippines), conservation on farm may be the better option, especially for the new diversity. To establish a regional or international genebank, it is important that the partner countries policies do not hinder the exchange of coconut genetic resources, as exemplified by the agreement of participating countries in the establishment of the ICG (Ramanatha Rao and Batugal 1998). Framework for complementary conservation strategy of coco- nut germplasm It is evident from the above discussions that the options for conserving coconuts germplasm are rather limited (Table 1). The current practice, as already noted, is the use of field genebanks. On-farm conservation appears to have a great potential for such a perennial species as coconut. The perenniality, however, is also a constraint, as the information required for scientifically sound on-farm conservation would be limited. This is mainly because the information on farmers’ practices in terms of selection and genetic diversity is limited since the crop’s life might span over a couple of lifetimes of its growers. At present, in vitro collecting and in vitro culture of zygotic embryos that also facilitate movement of germplasm (phytosanitary aspects and cost) are fully operational. Cryopreservation, which ensures safe and cost-effective long-term storage, is expected to be operational soon after minor improvements to the existing protocol. The establishment of cryopreserved collections could be envisaged on a regional basis (e.g. one cryopreserved collection linked with each ICG site) or even on a global basis (one or two cryopreserved collections at sites agreed by COGENT partners) as a measure of long- term backup. The balance between the different methods employed for coconuts would depend on many factors such as the intended use of the conserved germplasm, the method of maximizing the diversity of coconuts, the available infrastructure and human resources, space availability, accessibility and so on. Based on these elements and on the state of knowledge and the options available to date, a framework for complementary conservation strategy can now be developed. It is not envisaged here to develop a full strategy for coconut, but rather to propose a framework and the elements as how such a strategy could be developed at different levels: national, regional or international. The framework can be seen as a series of steps (Figure 1). At each step information is gathered, specific actions taken and/or decision made. 84 COCONUT GENETIC RESOURCES The first step would be to organize the stakeholders into a network, as has happened in COGENT. This should be facilitated by a lead agency to enable its creation and be established with a steering committee composed of representatives of the various stakeholders. This would then be the decision making body to develop the strategy and take the decision on its content and implementation. The stakeholders would then be responsible to define objectives and sub-objectives according to its mandate. This would for example in the case of coconuts be to conserve and utilize the maximum genetic diversity in Cocos nucifera. A number of sub-objectives could also be elaborated such as the long-term conservation of coconut germplasm, conservation of specific ecotypes or characterization of germplasm, as mentioned earlier. For each specific objective, the conservation options available should then be analyzed in terms of their feasibility and requirement in infrastructure, human resources, land, costs, accessibility and the risks involved. In relation to coconuts, we have seen that field genebanks and on farm conservation represent the best conservation methods but have certain limitations in the long term (Table 2). Other options like in vitro techniques and cryopreservation of zygotic embryos, for example, should be pursued in the future. The advantages and disadvantages of each of the possible options (Table 2) must be weighed against each other. This kind of analysis would provide the basis for taking decisions on which conservation options to be followed for given specific objectives. The next important step in the process would be setting up the enabling environment to allow the conservation options to be implemented. This would involve, as discussed earlier, the policy issues in terms of legislation, germplasm exchange, benefit sharing and also most importantly the sources of funding. Once these are agreed upon and put into place, a strategic action plan can be developed and implemented (steps 6 and 7). For each step, the steering committee would examine the issues and take the relevant decisions and assign responsibilities to the various relevant players. In conclusion, a complementary conservation strategy for coconuts requires a lot of efforts and commitment from many different stakeholders, who must work together with a common objective. A proper enabling environment, including inter alia policy, finances, incentives and good collaborative spirit, is crucial for its success. 85 CHAPTER 3: Germplasm conservation Table 1. Comparison of conservation options for coconuts In situ on farm /Home Gardens/natural habitats Botanic Gardens (Living plants in gardens/ greenhouses) ‘Conventional’ Genebanks (seed banks, field genebanks) Slow growth conditions (short-term) Cryopreservation - liquid N (long-term) Mature plants ; Coconuts conserved on farm widely and in home gardens and natural stands exist on small isolated islands and atolls ; Occurs in botanic gardens but limited scope for conserving genetic diversity ; Field genebank most widely used conservation method so far. National and international coconut field genebanks exist ⌧Not applicable ⌧Not applicable Seeds and zygotic embryos ⌧ Not feasible- seeds are recalcitrant, no natural soil seed banks ⌧Not feasible ⌧ Seeds are recalcitrant and too large; seed conservation not feasible ;Field collecting protocol established for zygotic embryos; In vitro culture functional ; Cryo- preservation protocol has been established for zygotic embryos; suitable for long term conservation Somatic embryos ⌧ Not applicable ⌧ Not applicable ⌧ Not applicable ⌧ Mass propagation problematic; Not applicable ⌧ not applicable Pollen ⌧ Not applicable ⌧ Not applicable ; Possible, for short term conservation (2-6 months) ⌧Not applicable ; Coconut pollen can be cryopreserved and could be suitable for long term conservation Apices ⌧ Not applicable ⌧ Not applicable ⌧ Not applicable ⌧ Not applicable ; cryopreservation protocol established; relatively low survival and regeneration of plants very difficult DNA ⌧ Not applicable ⌧ Not applicable ; Storage as DNA libraries exists – value not known ⌧Not applicable ; Long-term storage possible (LN or –80°C freezer). Use of stored DNA questionable. 86 COCONUT GENETIC RESOURCES Table 2. Relative advantages and disadvantages of conservation methods for coconut Method Advantages Disadvantages Research needed Field genebank • Easy access for characterization, evaluation and use • Simple infrastructure needs • Does not require highly skilled manpower • Space limitation compounded by need to maintain safe isolation distance between trees, especially for the Tall types that out cross frequently • Labour intensive; High risk in mislabelling • Vulnerability to biotic and abiotic factors • Exchange of germplasm • Participation with end users difficult • Legal issues as related to land ownership • Minimum number of palms needed to maintain representative genetic diversity • Filed plot techniques for proper characterization and evaluation • Economics of coconut field genebank maintenance In vitro collecting and culture of zygotic embryos • Well established protocols • Facilitates germplasm exchange • Only short-term storage • Relatively high infrastructure needs • High maintenance cost • Less accessible to users • Testing of optimized in vitro culture protocol Cryopreservation • Feasible for long term secure storage • Easy to maintain, low costs • Protocol for coconut embryos has been developed • Not labour intensive • Requires skilled labour • High initial investment cost for Infrastructure • More work required to refine cryopreservation protocol Pollen conservation • Large number of samples can be maintained in small space • Easy to handle • Useful for crosses • Can be cryopreserved allowing long term storage • Not yet feasible for long term • Only conserve part of diversity • Cannot be used to conserve specific genotypes • Refinement on cryopreservation protocol • Desiccation tolerance On-farm • Dynamic conservation in relation to environmental changes • Participation of local communities and stakeholders made easier • Conserve a much larger genetic diversity overall • Highly suitable for coconuts • Difficult to exchange germplasm • Vulnerable to natural and man-directed disasters, e.g. fire, cyclones, vandalism, change in land use, deforestation etc. • Materials not easily available for utilisation • Appropriate management regimes poorly understood • Require active supervision and monitoring • Genetic diversity scattered • Little information on status of genetic diversity across coconut stands. • Systematic documentation of farmers knowledge is needed • Several issues related to socioeconomics of coconut farming, indigenous knowledge, community participation in relation to on-farm conservation • On farm conservation methodologies need further work • Ways and means to enhance benefits for promoting conservation on farm • Piloting in situ methods for locating, measuring and monitoring genetic diversity 87 CHAPTER 3: Germplasm conservation References Ashburner, GR and W Rhode. 1994. Coconut germplasm characteriza- tion using DNA marker technology. Pp. 44-46. In: MA Foale and PW Lynch (eds). Coconut improvement in the South Pacific. Proceedings of a workshop held in Taveuni, Fiji, 10-12 November 1994. Canberra, ACIAR Proceedings No. 53. Assy-Bah, B, T Durand-Gasselin and C Pannetier. 1987. Use of zygotic embryo culture to collect germplasm of coconut. FAO/IBPGR Plant Genetic Resources Newsletter 71:4-10. Assy-Bah, B and F Engelmann. 1992. Cryopreservation of mature em- bryos of coconut (Cocos nucifera L.) and subsequent regeneration of Figure 1. Framework for developing a complementary conservation strategy Step 1 Networking of stakeholders at national, regional or international, as the case may be Step 2 Definition of objectives and sub-objectives Step 3 Analysis of the feasibility of each option for each sub-objective in terms of infrastructure needs, costs, risks involved, etc. Step 4 Decision on conservation options for each objective/sub-objective Step 5 Setting up enabling environment – policy/legal issues, funding, etc Step 6 Elaboration of Strategic Action Plan by stakeholder/s Step 7 Implementation process Ð Ð Ð Ð Ð Ð 88 COCONUT GENETIC RESOURCES plantlets. Cryo-Letters 13: 117-126. Assy-Bah, B and F Engelmann. 1993. Medium-term conservation of ma- ture embryos of coconut. Plant Cell, Tissue and Organ Culture 33:19- 24. Benard, G. 1973. Quelques aspects de la lypholysation du pollen de co- cotier. Oléagineux 28:447-551. Damania, AB. 1996. Biodiversity conservation: a review of options com- plementary to standard ex situ methods. Plant Genetic Resources Newsletter 107: 1-18. Dulloo, ME, L Guarino, F Engelmann, N Maxted, HJ Newbury, F Attere and BV Ford Lloyd. 1998. Complementary conservation strategies for the genus Coffea: A case study of Mascarene Coffea species. Ge- netic Resources and Crop Evolution 45:565-579. Engelmann, F. 2002. Coconut. In vitro germplasm collecting techniques. IPGRI Technical Bulletin N°6. IPGRI/FAO, Rome. (In press). Engelmann, F, P Batugal and J Oliver (eds). 2002. Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Selangor, Malaysia. Hodgkin, TH, V Ramanatha Rao and KW Riley. 1993. Current issues in conserving crop landraces. Presented at the FAO-IBPGR On-Farm Conservation Workshop, 6-8 December 1993, Bogor, Indonesia. Engels, JMM and D Wood. 1999. Conservation of agrobiodiversity. Pp. 355-385. In: D Wood and JM Lenné (eds). Agrobiodiversity charac- terisation, utilisation and management. CAB International, Wallingford, UK. Foale, MA. 1992. Coconut genetic diversity- Present knowledge and fu- ture research needs. Coconut Resources. Pp. 46-58. In: Papers of an IBPGR workshop, Cipanas, Indonesia, 8-11 October 1991. Interna- tional Crop Network Series No. 8. Rome, IBPGR. Hornung, R, R Domas and PT Lynch. 2001. Cryopreservation of plumular explants of coconut (Cocos nucifera L.) to support programmes for mass clonal propagation through somatic embryogenesis. CryoLetters 22:211-220. IUCN. 1994. Guidelines for Protected areas management categories. Gland Switzerland, 261p. Jarvis, DI. 1999. Strengthening the scientific basis of in situ conservation of agricultural biodiversity on-farm. Botanica Lithuanica Suppl. 2: 79-90. Kannenberg, LW and DE Falk. 1995. Models for activation of plant ge- netic resources for crop breeding programs. Canadian Journal of Plant Science 75(1): 45-53. Lebrun, P, YP N’cho, M Seguin, L Grivet and L Baudouin. 1998. Genetic diversity in coconut (Cocos nucifera L.) revealed by restriction frag- 89 CHAPTER 3: Germplasm conservation ment length polymorphism (RFLP) markers. Euphytica 101: 103-108. Malaurie B, M Borges and O N’Nan. 2002. Research of an optimal cryopreservation process using encapsulation-osmoprotection-dehy- dration and encapsulation-osmoprotection-vitrification techniques on caulinary meristems of coconut (Cocos nucifera L.). In: Abstracts IV Jornada Científica IIA”Jorge Dimitrov”, Bayamo, Cuba, 19-21 Sept. 2002. Martin, JM, TK Blake and EA Hockett. 1991. Diversity among North American spring Barley cultivars based on coefficients of parentage. Crop Science 31: 1131-1137. Maxted, N, BV Ford-Lloyd and JG Hawkes. 1997. Complementary con- servation strategies. Pp. 15-39. In: N Maxted, BV Ford-Lloyd and JG Hawkes (eds). Plant genetic conservation: The in situ approach. Chapman and Hall, London. Mpunami, A, S Sinje, S Chalamila, P Tembo, J Mugini, P Jones, A Tymon and M Dickinson. 1998. Application of molecular methods for diag- nosis of Lethal Disease of coconut palms in Tanzania. Pp. 518-526. In: CP Topper, PDS Caligari, AK Kullaya, SH Shomari, LJ Kasuga, PAL Masawe and AA Mpunami (eds). Trees for life: The key to de- velopment. Proceedings of the International Cashew and Coconut Conference, 17-21 February 1997, Dar es Salaam, Tanzania. BioHybrids International Ltd., Reading, UK. Pardey, PG, B Skovmand, S Taba, M Eric Van Dusen and BD Wright. 1999. Costing the ex situ conservation of genetic resources: Maize and wheat at CIMMYT. Londres, Mexico, International Food Policy Research Institute (IFPRI) and Centro Internacional de Mejoramiento de Miz y Trigo (CIMMYT). p43. Perera, L, JR Russell, J Provan and W Powell. 1999. Identification and characterization of microsatellite loci in coconut (Coos nucifera L.) and the analysis of coconut populations in Sri Lanka. Molecular Ecology 8:335-346. Ramanatha Rao, V. 1998. Strategies for collecting of tropical fruit spe- cies germplasm. Pp. 73-78. In: RK Arora and V Ramanatha Rao (eds). Tropical fruits in Asia: Diversity, maintenance, conservation and use. Proceedings of the IPGRI/ICAR/UTFANET Regional Train- ing Course on the Conservation and Use of Germplasm of Tropical Fruit in Asia, 19-31 May 1997, IIHR, Hesaragatta, Bangalore, India. IPGRI South Asia Office, New Delhi, India. Ramanatha Rao, V. 1999. Complementary conservation strategy. Pp. 139- 150. In: B Mal, PN Mathur and V Ramanatha Rao (eds). Proceedings of the Fourth Meeting of SANPGR, 1-3 September 1998, Kathmandu, Nepal. IPGRI South Asia Office, New Delhi, India. 90 COCONUT GENETIC RESOURCES Ramanatha Rao, V and P Batugal (eds). 1998. Proceedings of the CO- GENT Regional Coconut Genebank Planning Workshop, 26-28 Feb- ruary 1996, Pekanbaru, Riau, Indonesia. IPGRI-APO, Serdang, Selangor, Malaysia. Ramanatha Rao, V, D Jarvis and B Sthapit. 2000. Towards in situ conser- vation of coconut genetic resources. Paper prepared for 9th COGENT Steering Committee Meeting, 20-12 July 2000, Chennai, India Ramanatha Rao, V, KW Riley, JMM Engels, F Engelmann and M Diekmann. 1998. Towards a coconut conservation strategy. In: V Ramanatha Rao and P Batugal (eds). Proceedings of the COGENT Regional Coconut Genebank Planning Workshop, 26-28 February 1996, Pekanbaru, Riau, Indonesia. IPGRI-APO, Serdang, Selangor, Malaysia. Rognon, F and MW de Nucé de Lamothe. 1978. Récolte et conditionne- ment du pollen pour la pollinisation des champs semenciers de coco- tier. Oléagineux 33: 17-23. Whitehead, RA. 1966. Progress in the freeze drying of coconut pollen. Oléagineux 21: 281-284. 91 CHAPTER 3: Germplasm conservation Coconut field genebank V Ramanatha Rao Senior Scientist, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction The two basic approaches to conservation of plant genetic resources (PGR) are termed ex situ and in situ. Ex situ approach involves conserving the genetic resources outside their original habitat in the form of seed, embryos, tissues or plants. Methods of ex situ conservation can include cold storage, in vitro storage or field genebanks, depending on the propagules used. In contrast, in situ conservation involves the maintenance of genetic diversity of a species or genepool in the habitat in which the diversity evolved. In the definition of the Convention on Biological Diversity (CBD), it includes the maintenance of diversity in farmers’ fields and orchards, thus it includes the so-called on-farm conservation. It is now well recognised that for any given genepool, a number of different approaches and methods will be necessary for efficient and cost-effective conservation. Such a strategy is termed as complementary conservation strategy (Ehsan et al. 2003). However, with the current level of conservation options for coconut, field genebanks still play the major role in their conservation and use. It is presently the most feasible ex situ conservation method that can be used for coconut. This chapter attempts to look at the general nature of conservation of PGR in field genebank and looks specifically on how coconut fits into this context. Field genebanks Many important varieties of field and horticultural crops including coconuts are either difficult or impossible to conserve as seeds (i.e. no seeds are formed or if formed, the seeds are recalcitrant) or the species are vegetatively propagated. Conservation in field genebanks (FGBs) is necessary because some species have short-lived seeds (recalcitrant) such as cocoa, coconut, oil palm, rubber and many tropical fruits like mango, mangosteen, jackfruit, durian and rambutan. Seeds of some recalcitrant species can only be stored without desiccation for a few days, weeks or months (Roberts et al. 1984). Even if technology for conserving recalcitrant seeds is developed, there is still the problem with long regeneration cycle and which constraint utilization (Hawkes 1982). Hence, they are conserved in FGBs. FGBs may run a greater risk of being damaged by natural calamities, infection, neglect or abuse. Ex situ conservation of 92 COCONUT GENETIC RESOURCES tree species using FGBs requires a substantial number of individual genotypes to be an effective conservation measure. Thus, FGBs require more space, especially for large plants such as coconut and they may be relatively expensive to maintain, depending upon the location and the complexity of alternative techniques available. However, FGBs provide easy and ready access to conserved material for research as well as for use. The advantages and drawbacks of FGBs are well debated (Engelmann 1999; Epperson et al. 1997; Saad and Ramanatha Rao 2001; Dulloo et al. in this chapter). However, it must be noted that, for a number of plant species, the alternative methods have not been developed to the stage where they can be effectively used (Ramanatha Rao et al. 1998; Engelmann and Engels 2002). Thus, the establishment of FGBs will play a major role in any conservation strategy for PGR. Considerations for conservation of coconut germplasm in field genebanks There are many field collections of coconuts in various countries, usually connected with coconut research institutes. According to the International Coconut Genetic Resources Network’s (COGENT) Coconut Genetic Resources Database (CGRD), over 1416 accessions are collected and conserved in different genebanks around the world and all of these are maintained and managed in FGBs. At this stage, it must be noted that many of the plantings in these genebanks are often quite old and may originally have developed in a somewhat haphazard manner. Although they represent a commendable effort, there is a need to update the collections in a scientific manner, with due regard to thorough documentation of populations, and correct and foolproof labelling. It is clear from the above discussion that the establishment of FGBs is the immediate option for the conservation and utilization of coconut genetic resources. In most annual crops like cereals and legumes, seed storage, multiplication and regeneration methods have been agreed upon (FAO/IPGRI 1994), but for crops like coconut, there are no such agreed methods. Hence, there is a need to look closely at scientific and practical criteria to be considered in establishing and managing a coconut field genebank. In the following sections, some of the issues involved in developing a successful field genebanking technique for coconut germplasm are discussed, much of which are based on Ramanatha Rao et al. (1998). Genetic considerations To effectively collect and conserve PGR, there is a need to have a sufficient understanding of some of the conservation genetic principles, especially those related to the structure and distribution of the genetic diversity of 93 CHAPTER 3: Germplasm conservation species to be conserved as well as the genetic diversity of the materials that are being conserved (for general principles, see Mohd Shukor Nordin and Mohd Said Saad 2001). Ensuring genetic integrity and maintenance of genetic diversity Breeding system. Generally speaking, populations of inbreeding species have a simple genetic structure, consisting of a number of inbred lines, genetically homozygous, several individuals representing each line in the population, some variations between lines, and very little heterozygosity. This makes conservation of mostly genetic resources self-pollinated species (in the case of coconut, the Dwarfs) much simpler, compared to outbreeding species (as Talls) in which, the population in general has higher within population variability. Maintaining this variation within a cross-pollinating species will be complex in terms of both germplasm collecting and regeneration and sampling for utilization. Isolation. To be able to maintain the genetic integrity in an outbreeding species like coconut, especially the Talls, growing of accessions or populations in isolation should be considered. However, for practical reasons different accessions of coconut have to be planted together. Available information indicates that coconut pollen under natural conditions could travel over 300 metres (Mantriratne 1965; de Nucé de Lamothe and Rognon 1975) or assisted pollination (without bagging) results in pollen contamination, unless vary large plot sizes are used (de Nucé de Lamothe and Rognon 1975). Such large plot sizes may be impractical for a genebank. Sampling and selecting entries. To establish a coconut collection, accessions displaying a range of diversity need to be planted. For this reason and from the point of population genetics, the principle of random sampling of genotypes from a given source population should be followed. However, in practice, both elite materials and genetic stocks are planted. Therefore, it is essential to sample a range of diversity to be represented in the FGB, including that of elite material. Thus, a coconut FGB can include populations/genotypes representing a range of diversity, elite materials, genetic stocks and some unique materials. Sample and plot size. The size of the plot depends mainly on the breeding system and diversity in the sample and on the number of palms planted. It will be most appropriate if the material planted in a field genebank can be representative of the source population. For raising seedlings and 94 COCONUT GENETIC RESOURCES transplanting, the best methods should be used to ensure maximum survival and vigour, any loss at seedling stage represents genetic drift (see below). Large plot sizes with high number of plants are appropriate for outcrossing species like coconuts, but for practical reasons, smaller plots are acceptable. However, if characterization and evaluation activities are combined with conservation then larger plot sizes, as dictated by statistical principles, should be used to avoid biases due to competition and xenia. It is well established that square plots will be better than row planting for reducing pollen contamination (Breese 1989). The minimum number of palms per accession is determined by the number needed to represent the genetic diversity of the population while the resources available determine the maximum. Assuming that the material will be conserved for the next 200 years (with a frequency of re-planting in a FGB being 30 years for Dwarfs and 40 years for Talls), this gives about 4-6 regenerations (re-plantings). If the decision is to have a 90% probability of maintaining the alleles with a frequency of >5%, a population of about 40-60 will be needed per accession, and for an accurate characterization, about 90 palms are preferred. This is also based on the assumption that the seedlings for exchange or re-planting will be produced by (hand) pollinating the female parents with the mixed pollen of the accession/ population in question or by using methods such as chain crossing or pair crossing (For more information see Breese 1989; Crossa and Vencovsky 1994; Gale and Lawrence 1984; Gregorius 1991). Production of seednuts through randomly chosen parents ensures a balanced representation of male parents thus increasing the effective population size (lowering the effect of drift) and provides pedigree information for subsequent breeding. This number needs to be supplemented with a few additional palms to compensate for any losses that may occur. From a conservation point of view, it may be better to plant and maintain equal number of plants for each accession, keeping in mind the need for representation of genetic variation. However, from the practical point of view, the number of palms of certain highly productive accessions may have to be increased to make the FGB commercially viable. Additional numbers of these accessions may best be planted in a separate area in the FGB. Drift. Another consideration is avoidance of genetic drift or loss of rare but important genes or alleles from the population or accessions. For coconut, a minimum of 40 to 60 individuals are required to maintain alleles with 0.05 frequency with reasonable degree of confidence, to reduce the effects of genetic drift (again depending on the original sample 95 CHAPTER 3: Germplasm conservation size) and maintain genetic integrity. Assuming pollination is not controlled, there may be a need for some buffering or isolation to minimize random mating among the individuals of an accession. However, isolation between plots is hardly practised, hence, only a small number of plants in the middle of the plot may represent the gene frequencies of original populations. This would imply, especially if the original population size is large, that the total number planted in one block should be high enough to give sufficient number of plants to get an effective population size of at least 30, so as to sample/maintain alleles with frequency greater than 0.05. This would translate into a minimum of 100 palms in two blocks or replications of 50 each per accession, when no isolation or hand pollination is practised for the production of offspring generation (see below). It will be possible to have 60 palms in two replications of 30 each per accession, if isolation and/or hand pollination is practised. Layout and plot management. Although as a general principle, it is best not to mix conservation and evaluation, planting equal numbers of palms per plot assists in evaluation and characterization of the material as well. Evaluation for important traits which are affected by environmental variation can only be carried out using replicated plots and also it must be kept in mind that the FGB needs to be protected from biotic and abiotic stresses, as far as possible, and this might result in poor quality of characterization and evaluation data. If evaluation is to be carried out in the FGB, it will be a good idea to plant each accession in at least two and ideally four replications depending on the total number of plants per accession. The accessions within each replication must be randomized. It is important to realize that, for purposes of conservation, planting sufficient number of plants (40-60) in a square block with a few metres border will be appropriate to reduce the chances of pollen contamination. Additionally, if these are planted in at least two replications, then evaluation of differences between populations, as well as selection of superior palms within populations may be combined with conservation. Since characterization and evaluation is involved, one has to consider soil heterogeneity as well. In controlling the soil heterogeneity, normally, allocating genotypes in a block is very effective if the number of genotypes involved is small. The effectiveness of controlling soil heterogeneity will be very much reduced if the number of genotypes is more than 20, especially for perennial crops that required a big plot size such as coconut. Under this situation, incomplete block designs could be used (Yap and Saad 2001). 96 COCONUT GENETIC RESOURCES Supply material from the FGB From germplasm conservation and use point of view, it is important to supply a sample that represents the population conserved. If the progeny from an accession in the FGB is required, then it will be desirable to carry out hand pollination among the identified parent materials of an accession to produce the required offspring. Otherwise, from the standing population one will only get offspring resulting from open pollination, which may be contaminated from another accession. As described above, pollen of coconut can travel to significant distances resulting in considerable pollen adulteration. Coconuts are also known to be pollinated by insects such as bees, wasps and ants. However, it was observed that the insects tend to return to the same plant or to neighbouring plant, effecting mostly selfing or sibling, if the neighbouring plants belonged to the same population (Child 1964). Theoretically, if the plot size is very large, and the plots are separated by about 500 m, it may be possible to obtain less contaminated (by foreign pollen) nuts from the middle of the plot. However, such large plot sizes and growing in isolation are not practical. Thus, the most practical is hand pollination and bagging. From the point of genetic principles underlying regeneration, promoting random mating (i.e. using a mixture of pollen from different plants in the population, in this case, the plot) is recommended to increase the effective population size which can reduce the effects of genetic drift, while using pollen from single parent may be useful from the breeding point of view (Dr L Baudouin 1996, personal communication). It must also be noted that, in making controlled pollinations and comparing the offspring with its parents, one is not only able to preserve the genetic information of each individual but one may be able to keep the genetic information of the population as a whole. One of the first to consider the genetic factors in maintaining living plant collections was Esser (1976). He concluded that true genetic conservation is not possible but one should know the boundaries and be able to channel plant conservation based on the knowledge and application of genetic parameters. Agronomic considerations Security from natural disasters and safety duplication Despite many problems discussed, FGB is the current method for medium to long-term conservation and use. So the security of the site must be assured. Whenever possible, it may be appropriate to replicate the collections in more than one location. It is important to establish adequate safety duplicate collection(s) of the material maintained in FGB. Along with abiotic stresses like hurricanes, cyclones, drought, fire, etc., the biotic stresses such as pests and diseases can be continuous and serious threats 97 CHAPTER 3: Germplasm conservation to the germplasm being conserved. Therefore, the establishment of safety duplicate collections should be regarded as routine and budgeted for accordingly. The site should not be located in an area known for natural calamities or other disasters. This will help not only for effective monitoring and management of the FGB but also for the long-term safety of the material conserved. Ecological adaptation Close relationship between some characters in a population and its habitat in which the characters or traits have evolved and expressed has been reported many times in the literatures. Collections made from separate geographical areas can differ substantially. For instance, some characters are common in accessions collected from certain regions but not in the other region. Adaptability of the species or the accession to the location may be an important point to consider when long-term conservation is involved, especially in the case of regional genebanks. However, this may not be always possible as FGB may contain introduced or unadapted material. For efficient maintenance as well as from the point of view of use of the material conserved, it is important that the plants in the FGB be able to produce flower, fruit and set viable seed. If the site for the genebank is located in an area with (a potential for) commercial orchard plantation, then the value of the genebank would be even greater, in terms of use of the conserved material. The genebank can act as the nucleus and provide planting material for commercial plantations. Minimal pest occurrence It is essential to establish FGBs in areas that are relatively free from pests and diseases, especially those that are transmitted through propagules. This aspect will be discussed further under the section on germplasm health issues. Also, the site should be protected from animal pests such as wild pigs, porcupines, elephants etc. Access to the FGB To facilitate protection and management of the population stands, continued access to them must be assured. Therefore, the site chosen for FGB should be accessible and be near to the research station so that the material available can be effectively used. Also the material in the FGB can be monitored frequently. It should be possible for the genebank staff and other researchers to reach the site easily. From a practical point of view, this probably has an overriding importance. 98 COCONUT GENETIC RESOURCES Choice of material Entries in regional genebanks The major factors that control the choice of material into any genebank is determined firstly, by the needs of the users of that particular genebank and secondly, to have as much representation of genetic diversity and ecotypes that are available in the region. This requires giving emphasis to regionally recognized accessions. Therefore, there may be a tendency to acquire/assemble mostly the well-known elite accessions. However, it is important to have a balance between elite lines and accessions that represent a broader range of genetic variation from within the country as well as from the region. Giving due regard to the points mentioned earlier, it is important at the time of FGB establishment to plan for the materials to be included and lay out sufficient space. Care must be taken to insure that each accession included is unique and is not a duplicate. Need for a national collection Decisions will have to be taken on which accessions are to be maintained at the regional and national level. Some nationally important accessions or accessions representing national or narrow diversity may not be accommodated in the regional genebank for lack of space and resources. Such accessions must continue to be maintained in the national collection. In addition, it is essential to make sure that each accession is duplicated in another FGB for safety reasons, while other methods of conservation are being considered. The situation in any genebank is dynamic as new materials are collected for conservation. Therefore, it is essential to maintain space in the FGB for new accessions. Policy and management issues National, regional and international collections Conservation of genetic resources is a long-term responsibility and requires long-term commitment of institutions and governments. It is for this reason that any conservation effort should be conducted within the framework of a national programme, that clear institutional responsibilities are assigned as part of a national mandate and that a reliable budget-line is established for continuous funding. At the regional and international level, the situation is different since it is not easy to assign clear mandates and responsibilities for the conservation of worldwide genetic resources of a specific genepool. The international germplasm collections held by the International Agricultural Research Centres are an exception as they have been placed under the auspices of the Food and Agriculture 99 CHAPTER 3: Germplasm conservation Organization (FAO) as part of the International Network of Ex Situ Collections. Individual centres have accepted responsibility of conserving global genetic diversity for one or more specified genepools as part of their broader mandate to deal with the improvement of the so-called mandate crops. Sustained commitment It is important to critically examine the existing arrangements with regard to mandate and responsibilities at both the national and regional level for coconut genetic resource conservation. For example, the multi-site International Coconut Genebank (ICG) has been established in respective countries after an assessment of the level of governmental and institutional commitment to the maintenance of the collections. Only when an effective governmental commitment exists should the establishment or extension of a collection be considered. This is especially important for crops like coconut which needs a relatively large area in order to plant sufficient number of plants/trees necessary to represent the genetic diversity. Initial establishment costs, which in some cases can be very high, and recurring costs for maintenance of the collection should be considered at the planning stage and should be provided for. In many cases, the latter is ignored and the collections can run into problems within a few years of their establishment. Given this background, the establishment and maintenance of FGBs, appears to be more easily organized at national level, as part of national PGR programmes, rather than at regional or international level. In the case of regional or international efforts, it is essential to obtain the full support and commitment of the government of the host country in which the FGB is to be set up and to obtain commitments from the individual member states of the network to financially support the effort. For any emergency situation, provisions have to be made as to how and where the collection can be duplicated, if so decided. The role of the cooperating international institutions needs to be defined as well. Legal issues Since the CBD has come into force, countries now have the sovereign rights over the biological diversity present within their borders. In view of this, a clear consensus must be reached by all the member countries of a given crop genetic resources network with regard to sharing the benefits derived from the germplasm conserved, as well as, on access conditions to the conserved germplasm and information related to it as was done in the case of ICGs. The necessary agreements and mechanisms on access to and provision of accessions should be in place before the establishment 100 COCONUT GENETIC RESOURCES of a regional or international genebank. Within these mechanisms, there might be a need to develop some form of material transfer agreement to accompany germplasm accessions being sent to researchers/breeders within and outside the network. Considering the current legal situation with regard to genetic resources, one of the options for regional or global PGR networks is to consider the possibility of placing their germplasm collections under the auspices of FAO, thus becoming part of the FAO International Network of Ex Situ Collections. In doing so, the host country which acts as trustee of the germplasm on behalf of all the member countries, agrees not to claim ownership over the germplasm and not to claim any form of intellectual property protection to the material or on any information related to it. The host country will also ensure that any further recipients of the germplasm are bound to the same conditions as mentioned above (FAO 1995). The ICGs are managed under the auspices of FAO. Germplasm health issues There are two reasons for establishing FGBs in areas free from important pests and diseases. One is the risk of the entire collection, or part thereof, being destroyed by pests or diseases. The other is the risk of spreading pests and pathogens to new areas, which may easily happen with germplasm (Hewitt and Chiarappa 1977). An effective quarantine system should act as a filter, and should not be a barrier to germplasm exchange. However, as some countries have stronger controls than others, breeders and the germplasm community have a certain responsibility to give due attention to pathogens. For example, FGB managers should apply restrictions to the international (or even national in the case of large countries and where there are clear regional differences in the occurrence of a particular pest) movement of seednuts and choose instead the movement of embryo cultures even if local quarantine authorities do not impose such restrictions. Obviously, before establishing a field genebank, a critical evaluation of the disease situation in the location concerned will be required. Often parts of countries are free from a reported pathogen (e.g. Cadang-cadang) is not reported in Mindanao and the northern part of Luzon in the Philippines (Hanold and Randles 1991a) or Kerala wilt is only reported to occur in parts of Kerala and Tamil Nadu (Frison et al. 1993). A list of coconut diseases of uncertain etiology is given by Frison et al. (1993) and special care needs to be taken with regard to these diseases. An inverse case exists with the reports of viroid-like sequences in coconuts, which could not yet be linked with clear disease symptoms (Hanold and Randles 1991b; Fassil and Diekmann 1995). In the case of lethal yellowing and 101 CHAPTER 3: Germplasm conservation related diseases, it appears that a group of at least three closely related strains of phytoplasmas can be considered the causal agents (Jones et al. 1995). The general recommendation is to move embryo cultures or pollen, and not seednuts. Based on this, establishing embryo culture facilities in connection with FGB and providing the necessary training becomes very important. Germplasm health aspects need to be considered not only at the point of exchange, but also at any stage of germplasm management. During collecting, care must be taken that germplasm is collected only from healthy palms. In the regeneration and multiplication process, plant protection measures including pesticide application may be required. If an evaluation of traits like resistance to pathogens is done under conditions of high disease pressure (e.g. with artificial inoculation), a careful evaluation of the material with regard to its use in regeneration or exchange is essential. Summary and conclusion In the discussion presented here, it has been assumed that the collecting has been effectively carried out; keeping the sampling of genetic diversity in mind, and that all quarantine requirements have been completed. To establish, maintain and manage a FGB for coconut (national or regional collection), the following critical steps (checklist) are suggested (Ramanatha Rao et al. 1998): A. Agreement on precise functions of the collection: 1. Selection of site, based on the criteria established; 2. Agreement on obligations and responsibilities of organizations and/or countries involved; 3. Establishment of infrastructure and facilities; and 4. Legal aspects and exchange protocols (ownership, conditions of release, IPR issues, benefit sharing, use of MTA and other mechanisms) as agreed by all partners; B. Establishment of a coconut FGB: 5. Assure comprehensiveness of collection by including as much genetic diversity as appropriate; 6. Consider carefully the sampling techniques (random vs. non- random, and the need for deviation); 7. Assure that there is no duplication of accessions as this directly increases cost of FGB; 8. Determine the need for having replications, the number etc., based on the objectives of FGB; 102 COCONUT GENETIC RESOURCES 9. Determine through discussions and by actual visitations the accessions and the number of accessions to be included in FGB, and number of plants per plot; 10. Establish nursery of vigorous and healthy seedlings raised from nuts produced through hand pollination, determine planting conditions, etc. depending on the location of FGB; 11. Lay out square blocks of equal size; 12. Plan space for present and future accessions (as much as possible) to be randomized in the FGB; 13. Follow all protocols for safe movement of germplasm; 14. Ensure that embryo culture/tissue culture facilities can be put in place for exchange of material; and 15. Accept more material into FGB as they become available by going through all the steps discussed; C. Maintenance of a coconut FGB: 16. Take all the necessary agronomic and plant protection measures to maintain a healthy stand of coconut palms; 17. Take all the measures feasible to protect FGB from adverse environmental conditions, physical stresses, etc; 18. Make sure that a safety duplication is established and all the needs of health care are fulfilled; 19. Document all accessions as well as activities carried out in FGB by establishing and running an appropriate information management system; and 20. Provide linkages to other methods of conservation, if any, such as in vitro conservation of zygotic embryos, pollen preservation, etc; D. Access to material in FGB: 21. Ensure physical availability of the material; 22. Keep the plants in healthy condition; 23. Facilitate nut/seedling production through hand pollination for distributing germplasm as agreed at the time of the establishment of the genebank; 24. Characterize/evaluate the material in FGB according to agreed principles; 25. Provide for production of nuts through hand pollination, rather than harvesting from the centre of plot to be certain of purity of the material; 26. Make available the information on the material conserved in the FGB to all users; and 27. Exchange material using embryo culture rather than seednuts. 103 CHAPTER 3: Germplasm conservation This is by no means an exhaustive list of steps to be taken but only the important considerations that determine the effectiveness and sustainability of the FGB. It was also assumed that the issues considered in the earlier discussion were taken into account, decisions have been made and the consequences noted. Some of the principles of agronomy, nursery management, etc. can be found in Santos et al. (1996). References Breese, EL. 1989. Regeneration and multiplication of germplasm resources in seed genebanks: The scientific background. IBPGR, Rome, Italy. Child, R. 1964. Coconuts. Longman, London. Crossa, J and R Vencovsky. 1994. Implications of the variance effective population size on the genetic conservation of monoecious species. Theoretical and Applied Genetics 89:936-942. de Nucé de Lamothe, M and F Rognon. 1975. Assisted pollination and contamination by undesirable pollens. Oléagineux 80(8-9):359-364. Ehsan, D, V Ramanatha Rao, F Engelmann and J Engels. 2004. Complementary conservation strategy for coconut. Pp. 63-75. In: P Batugal, JT Oliver and V Ramanatha Rao (eds). Coconut genetic resources. IPGRI-COGENT, Serdang, Selangor, Malaysia. Engelmann, F (ed). 1999. Management of field and in vitro germplasm collections. Proceedings of a consultation meeting, 15-20 January 1996, CIAT, Cali, Colombia. Rome, Italy, IPGRI. 165p. Engelmann, F and JMM Engels. 2002. Technologies and strategies for ex situ conservation. Pp. 89-104. In: JMM Engels, V Ramanatha Rao, AHD Brown and MT Jackson (eds). Managing plant genetic diversity. CABI, Wallingford, UK and IPGRI, Rome, Italy. Epperson, JE, DH Pachio and CL Guevara. 1997. A cost analysis of maintaining cassava plant genetic resources. Crop Science 37:1641- 1649. Esser, K. 1976. Genetic factors to be considered in maintaining living plant collections. Pp. 185-198. In: JB Simmons, RI Beyer, BE Brondham, GL Lucas and VTH Parry (eds). Conservation of threatened plants. Plenum Press, New York. FAO and IPGRI. 1994. Genebank Standards. FAO-IPGRI, Rome, Italy. FAO. 1995. The international network of ex situ collections and the CGIAR centres. Joint report by FAO and the International Plant Genetic Resources Institute (on behalf of the CGIAR Centres) on the implementation of the agreement signed between FAO and the CGIAR Centres on 26 October 1994. CPGR-6/95/12 ADD. 1. Commission 104 COCONUT GENETIC RESOURCES of Plant Genetic Resources, Sixth Session, Rome, 1-30 June 1995. Fassil, H and M Diekmann. 1995. Safe movement of coconut germplasm and coconut cadang-cadang viroid-related sequences. Plant Genetic Resources Newsletter 104:29-30 Frison, EA and CAJ Putter (eds). 1993. FAO/IBPGR technical guidelines for the safe movement of coconut germplasm. FAO/IBPGR, Rome. Gale, JS and MJ Lawrence. 1984. The decay of variability. Pp. 77-101. In: JHW Holden and JT Williams (eds). Crop genetic resources: conservation and evaluation. George Allen, London. Gregorius, HR. 1991. Gene conservation and the preservation of adaptability. Pp. 31-47. In: A Seitz and VJ Loeschcke (eds). Species conservation: A population-biological approach. Birkhäuser Verlaag, Basel, Switzerland. Hanold, D and JW Randles. 1991a. Coconut cadang-cadang disease and its viroid agent. Plant Disease 75:330-335. Hanold, D and JW Randles. 1991b. Detection of coconut cadang-cadang viroid-like sequences in oil and coconut palm and other monocotyledons in the Southwest Pacific. Annals of Applied Biology 118:139-151. Hawkes, JG. 1982. Genetic conservation of recalcitrant species: An overview. Pp. 83-92. In: LA Withers and JT Williams (eds). Crop genetic resources: The conservation of difficult materials. IUBS Series B42, Paris. Hewitt, WB and L Chiarappa (eds). 1977. Plant health and quarantine in international transfer of genetic resources. CRC Press, Cleveland, USA. Jones, P, A Mpunami and A Tymon. 1995. Mycoplasmalike organisms as pathogens of coconut palms. Pp. 35-42. In: C Oropeza, FW Howard and GR Ashburner (eds.) Lethal yellowing: Research and practical aspects. Kluwer Academic Publishers, Dordrecht, The Netherlands. Mantriratne, MAPP. 1965. Coconut pollen. Ceylon Coconut Quarterly 16(3/4):102-110. Mohd Shukor Nordin and Mohd Said Saad. 2001. Genetic considerations in field genebank conservation. Pp. 66-72. In: MS Saad and V Ramanatha Rao (eds). Establishment and management of field genebank: A training manual. IPGRI-APO, Serdang, Selangor, Malaysia. Ramanatha Rao, V, KW Riley, JMM Engels, F Engelmann and M Diekmann. 1998. Towards a coconut conservation strategy. Pp. 4- 20. In: V Ramanatha Rao and PA Batugal (eds). Proceedings of the regional coconut genebank planning workshop, 26-28 February 1996. Pekanbaru, Indonesia. IPGRI APO, Serdang, Selangor, Malaysia. 105 CHAPTER 3: Germplasm conservation Roberts, EH, MW King and RH Ellis. 1984. Recalcitrant seeds: Their recognition and storage. Pp. 38-52. In: JHW Holden and JT Williams (eds). Crop genetic resources conservation and evaluation. George Allen and Unwin, London. Santos, G, P Batugal, A Othman, L Baudouin and JP Labouisse. 1996. Manual on standardized research techniques in coconut breeding. IPGRI-COGENT, Singapore. 46p. Yap, TC and Mohd Said Saad. 2001. Factors in field genebank layout. Pp. 73-76. In: MS Saad and V Ramanatha Rao (eds). Establishment and management of field genebank: A training manual. IPGRI-APO, Serdang, Selangor, Malaysia. 106 COCONUT GENETIC RESOURCES COGENT’s multi-site International Coconut Genebank P Batugal1 and K Jayashree2 1Coordinator and 2Scientific Assistant, International Coconut Genetic Resources Network (COGENT), International Plant Genetic Resources Institute – Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Background of COGENT In 1992, the Consultative Group on International Agricultural Research (CGIAR) decided to include coconut in its research portfolio after studies indicated that international support and global coordination of research in coconut is essential to make coconut more productive and beneficial to small-scale coconut farmers. The CGIAR and its Technical Advisory Committee (TAC) recognized that international support to coconut research was needed as many coconut-producing countries lacked both the human and material resources to conduct expensive and time- consuming research. Thus, it tasked the International Plant Genetic Resources Institute (IPGRI) to undertake research on coconut genetic resources, which the CGIAR identified as one of the five priority research areas that deserved international support. Accordingly, IPGRI included coconut genetic resources in its plant genetic resources research programme and organized the International Coconut Genetic Resources Network (COGENT) to implement this mandate. Starting with 15 countries, COGENT has rapidly developed into an active global network currently involving 38 coconut-producing countries (Table 1). Table 1. COGENT member countries Southeast and East Asia South Asia South Pacific Africa/Indian Ocean Latin America/ Caribbean 1. China 2. Indonesia 3. Malaysia 4. Myanmar 5. Philippines 6. Thailand 7. Vietnam 1. Bangladesh 2. India 3. Pakistan 4. Sri Lanka 1. Cooke Islands 2. Fiji 3. Kiribati 4. Papua New Guinea 5.Solomon Islands 6. Tonga 7. Vanuatu 8. Samoa 1. Benin 2.Cote d’Ivoire 3. Ghana 4. Kenya 5. Madagascar 6. Mozambique 7. Nigeria 8. Seychelles 9. Tanzania 1. Brazil 2. Colombia 3. Costa Rica 4. Cuba 5. Guyana 6. Haiti 7. Honduras 8. Jamaica 9. Mexico 10.Trinidad- Tobago 107 CHAPTER 3: Germplasm conservation COGENT’s goal is to improve coconut production on a sustainable basis and increase income of coconut farmers and growers in developing countries through improved cultivation of the crop and efficient utilization of its products. The objectives of COGENT are: 1) To establish and maintain an international database on existing and future collections; 2) To encourage the protection and use of existing germplasm col- lections; 3) To identify and secure additional threatened diversity by developing and adopting suitable technologies and conservations strategies; 4) To promote greater collaboration among research groups in producer countries and advance technology sources in the exchange of germplasm and the development of new techniques; 5) To conduct appropriate training and information dissemination; and 6) To secure necessary funding for network activities. In the last 12 years, COGENT has generated modest but significant achievements. The network has been successfully established with a Steering Committee serving as its supervisory and policy making body and is fully operational with 38 coconut producing countries as members. The International Coconut Genetic Resources Database (CGRD) has been established that currently contains passport and characterization data of 1416 accessions conserved in 25 sites in 23 countries. In addition, 278 ecotypes from the Asia Pacific region have been collected and conserved. To further secure conserved germplasm, a multi-site International Coconut Genebank (ICG) is being established to conserve 200 important accessions in the regions in which COGENT operates, which is hosted by India, Indonesia, Papua New Guinea, and Côte d’Ivoire and Brazil (under negotiation). An additional 212 farmers’ varieties have been identified and are currently being characterized. Multipurpose uses of coconut varieties are also being documented and promoted. The performance of 34 high-yielding hybrids are being evaluated in multilocation hybrid trials in four African and three Latin America/ Caribbean countries to select varieties and hybrids that are suited to particular agroecological conditions and to determine germplasm x environment interaction. Farmers’ varietal preferences in 15 countries are being evaluated. Diversity-linked income-generating activities have been initiated in 15 countries as part of a strategy to promote in situ and on-farm conservation, and germplasm utilization. Protocols for embryo culture, cryopreservation, morphometric and molecular marker-based methods for locating and characterizing diversity, assessing pest risks 108 COCONUT GENETIC RESOURCES and managing germplasm health are being developed, tested and upgraded. Strategies and techniques for farmer participatory research, collecting, characterization, and ex situ and in situ conservation are being refined. To strengthen coconut research capability of COGENT member countries, IPGRI and COGENT have, as of 2003: organized 39 country missions involving 28 experts to help COGENT member countries conduct research needs assessment and to identify priority research and training activities; conducted 41 workshops and meetings involving 994 coconut researchers to share information and technologies, discuss issues and common problems and opportunities and how to address them; conducted 40 training courses involving 765 participants from 41 countries; supported 180 research projects in 30 member countries; and led in establishing the Global Coconut Research for Development Programme (PROCORD), a global coconut research alliance with the Bureau for the Development of Research on Perennial Tropical Oil Crops (BUROTROP) and the Asian and Pacific Coconut Community (APCC). COGENT’s current priority involves the further promotion of more effective conservation and use of coconut genetic resources, both regionally and globally. This includes the establishment and operation of COGENT’s multi-site International Coconut Genebank (ICG). Integrated approach to coconut conservation COGENT’s conservation strategy is anchored on promoting the sustainable protection of diversity as well as maximizing germplasm use. In developing its conservation strategy, COGENT recognized that no one method or approach of conservation can meet all conservation needs and that there is a need to employ a combination of methods to ensure the sustainable conservation of as much genetic diversity as possible. It actively encourages the participation of its member country governments, partner organizations in both developing and developed countries, non- government organizations (NGOs) and coconut farmers themselves in conserving germplasm. The components of COGENT’s conservation strategy consist of: 1. Conservation in national collections; 2. Conservation in the multi-site ICG; 3. In vitro embryo culture and cryopreservation; 4. In situ and on-farm conservation; and 5. Promoting conservation through use by developing and implementing a globally-coordinated coconut breeding pro- gramme, establishing farmer community-managed coconut seedling nurseries in at least 25 countries, linking germplasm con- servation and use with the broader areas of research and 109 CHAPTER 3: Germplasm conservation development assigned to BUROTROP (agro-physiology and crop protection) and APCC (processing and marketing), developing and disseminating catalogues of conserved germplasm and farmers’ varieties, and upgrading and widely disseminating the CGRD. Conservation in the multi-site International Coconut Genebank Rationale World coconut production is declining due to ageing palms, natural calamities, inadequate replanting programme, lack of suitable planting materials, poor crop management, population pressures causing crop shifts, and lack of capital for farmers to invest in coconut production. The development and use of improved coconut cultivars can markedly help solve these problems and promote increased coconut production. However, the landraces of coconut (ecotypes), which contain important genetic characters for yield, disease and pest resistance and adaptation, are under treat to genetic erosion and need to be collected, conserved, evaluated and shared more widely to develop improved varieties. Conservation and use of a wide range of coconut diversity is faced with several constraints. First, while national coconut field genebanks are important sources of germplasm for exchange among COGENT member countries, many countries still lack the necessary economic and technical capacities to maintain their conserved germplasm. Second, many countries do not have the capacity to evaluate the performance of their germplasm while the data obtained are often not comparable. Third, multi-country negotiations for obtaining germplasm are often difficult for national breeding programmes needing to import germplasm that belong to several countries. Fourth, many researchers, who may want to share their germplasm, do not have the needed policy cover and their countries generally lack the needed facilities for ensuring the safe movement of coconut accessions. Fifth, COGENT does not have a concrete mechanism that would facilitate access and safe movement of germplasm to its member countries. To address these constraints, the COGENT Steering Committee decided to establish a multi-site ICG in 1995. Subsequently, site assessment surveys were conducted to evaluate the suitability of proposed regional genebank sites in the five host countries of Indonesia, India, Papua New Guinea, Côte d’Ivoire and Brazil. During the International Coconut Genebank workshop held from 26 to 28 February 1996 at Pekanbaru, Riau, Indonesia, representatives of IPGRI, the Centre de Cooperation Internationale en Researche Agronomique pour le Development (CIRAD) and the World Bank worked with representatives of COGENT member 110 COCONUT GENETIC RESOURCES countries in developing a series of legal agreements, initial work plans and proposed budgets, using national funds for each of the initial four genebanks to be hosted by Indonesia for Southeast and East Asia, Papua New Guinea for the South Pacific, India for South Asia and Côte d’Ivoire for Africa and the Indian Ocean. Objectives and initial activities The objectives of the ICG are: 1) to conserve nationally- and regionally- identified diversity; 2) to conserve internationally identified diversity; 3) to further assess the diversity, evaluate the performance of the conserved germplasm and disseminate related information to coconut-producing countries; 4) to make germplasm materials available to interested coconut- producing countries in accordance with existing protocols; and 5) to conduct research and training in relation to the above. Memoranda of Agreements (MOAs) for hosting of the ICGs for Southeast and East Asia (Indonesia), South Asia (India), South Pacific (PNG) and Africa and the Indian Ocean (Côte d’Ivoire) were developed and signed by the host countries and IPGRI on behalf of COGENT, with the Food and Agriculture Organization (FAO) of the United Nations serving as trustee. All MOAs were worded similarly (see MOA for ICG- SP in Annex 1.1). Negotiations are underway for Brazil to host the ICG for Latin America and the Caribbean. The host countries agreed to commit resources for their establishment maintenance and data gathering. The existing national field collections of Côte d’Ivoire and Papua New Guinea were donated to the ICG. However, COGENT is also exerting efforts to source additional funds for the maintenance of collections. COGENT has developed a sustainability strategy for the ICGs consisting of the following: 1) MOA committing host countries to maintain the field genebanks; 2) Negotiations for income from the ICG to be used for maintenance; 3) Superimpose research and training onto the ICG to share the cost of administration and maintenance; 4) Charge requesting countries for the cost of preparation, shipment and maintenance of germplasm, the latter on a pro-rata basis; 5) Undertake income generating activities in ICG plantations such as the production and marketing of high-value products from all parts of the coconut and integrate with intercropping and livestock raising as appropriate; 6) Generate external donor support; and 7) Generate national and provincial/state funding and institutional support. The sites for ICG were chosen based on surveys conducted by coconut experts who considered and evaluated several important selection criteria. 111 CHAPTER 3: Germplasm conservation Thus, the basic needs of field genebanks such as safety, security, accessibility, environment, etc. have been established. Among several items that were considered, two principles were highlighted. First, the choice of material in the ICG was determined by the needs of the users and by the need for as much representation of genetic diversity and ecotypes as possible. The importance of having a balance between elite lines and accessions that represent a broad range of genetic diversity from within the country as well as from the region was recognized from the beginning. Care has been taken to ensure that each ICG accession is unique and is not a duplicate. Thus, current accessions are being further validated using molecular marker studies to eliminate duplicates. Second, decisions were made as to which accessions would be maintained regionally and nationally. It was agreed that the nationally important accessions that cannot be accommodated in the regional genebanks would be maintained in the collections of strong national programmes. Thus, from the beginning it was apparent that national collections and the ICG would complement each other to accommodate as much coconut genetic diversity as possible. It is envisioned that the ICG at each regional site will conserve in field genebanks about 200 accessions which are important to the region. The ICG field genebanks are part of the ex situ collection under the International Undertaking on Plant Genetic Resources. The designated germplasm are conserved in the field genebanks and shared with coconut growing countries based on material transfer agreements. The field genebanks are established and managed by national programmes under the oversight of COGENT and IPGRI. Laboratories and facilities will also be developed to further locate diversity, identify and eliminate duplicates, conduct disease indexing, process pollen and embryos for export, conduct cryopreservation and train coconut researchers from member countries in evaluating, conserving and using germplasm. Thus, each site of the ICG will be developed as Centres of Excellence in concurrence to IPGRI’s initiatives of building and upgrading the capacity of partner institutions. Germplasm conservation and sharing In the next seven years (2004-2010), the ICG host countries aim to conserve in respective regional field genebanks a maximum of 200 accessions each, which will be contributed by coconut-producing countries in each region. Accessions will be imported in the form of excised embryos, grown in vitro in the embryo culture laboratory, transferred into pots in the greenhouse and eventually transplanted in the field. These accessions, which will be planted in the field genebank of about 200 hectares, will be characterized and evaluated using agronomic and molecular data to 112 COCONUT GENETIC RESOURCES determine their diversity, performance and potential for improvement work. Four types of coconut accessions will be conserved in the ICG: 1) major varieties (parents of existing hybrids and advanced generations of selected cultivars); 2) varieties/cultivars threatened with genetic erosion or total loss; 3) varieties/cultivars with special traits/genetic markers; and 4) genotypes being used for current genetic diversity studies using molecular markers. Member countries of each region can access germplasm belonging to different countries by negotiating with each ICG host country. The requested accessions will be sent in the form of embryos or pollen to interested countries after disease indexing to ensure safe movement. Requesting countries will be charged the cost of producing the seednuts and for preparing the embryos as well as the pro-rata cost of maintenance, disease indexing and shipping. These germplasm transfers will be covered by Material Transfer Agreements (MTA). Initial achievements Under COGENT, ICG sites in four host countries have been strengthened to some extent [i.e., ICG-South Asia (India), ICG-Southeast and East Asia (Indonesia), ICG-South Pacific (Papua New Guinea), and ICG- Africa and the Indian Ocean (Côte d’Ivoire)]. IPGRI has supported the ICGs in capacity building for embryo culture technology, in terms of materials, skills and laboratory upgrading to prepare them for importing and maintaining germplasm from network member countries in their respective regions. They have also been trained on germplasm collecting, morphometric and molecular marker (microsatellite kits) methods of germplasm characterization, genebank management and on cryopreservation. Since COGENT is currently an open network, it was proposed to further strengthen germplasm conservation by executing a formal Memoranda of Agreement with COGENT member countries, at the highest government level, to formalize their membership in COGENT and to formally commit access to their coconut germplasm. Despite meager resources, the ICGs have made some significant achievements. Table 2 shows the date of signing of the hosting agreements and the status of conserved germplasm in each of the host countries. Plan of action for the International Coconut Genebank As part of the above-described conservation strategy for coconut, IPGRI and COGENT would like to undertake the following plan of action for the upgrading of the ICG in the next seven years: 1. Regeneration of old palms of 50 accessions in the ICG for Africa and the Indian Ocean; 2. Additional morphometric and molecular marker characterization 113 CHAPTER 3: Germplasm conservation of the 1416 accessions conserved in the national collections of 23 countries to select other entries for the ICG and to upgrade the CGRD; and of the 224 accessions in the ICG to identify duplicates; 3. Integration of the CGRD with the System-wide Information Network for Genetic Resources (SINGER), the CGIAR-supported system-wide genetic resources programme; 4. Importation and establishment of additional accessions into the ICG sites to complete the 200 accessions per site; 5. Upgrading of pollen processing and embryo culture laboratories, net houses and coconut seedling nurseries in each ICG site; 6. Establishment of the needed facilities in the ICG host countries (i.e., molecular marker laboratories, except for ICG-Africa and Indian Ocean; disease indexing laboratories; training and dormitory facilities); 7. Research and training support for the following strategic activities: somatic embryogenesis, embryo culture, molecular marker/ microsatellite research, pest risk assessment and germplasm health management, germplasm x environment interaction and genetic distance analysis of conserved germplasm, and globally- coordinated coconut breeding; and 8. International Coconut Genebank evaluation and meeting of stakeholders. Conclusion Two of the major priorities of IPGRI and COGENT are: (1) saving threatened diversity and (2) promoting the use of conserved materials for developing improved varieties for national programmes and small- scale farmers. Thus, accelerated effort is being placed on the movement of germplasm from COGENT’s member countries to their regional ICG and the provision of breeding materials from the older ICG (i.e., Côte * Includes additional accessions entered into the ICG after the signing of the MOA Name of Genebank Date of MOA signed Initial number in list of designated germplasm Designated germplasm currently conserved International Coconut Genebank for the South Pacific (Papua New Guinea) 30 September 1998 55 50 International Coconut Genebank for South Asia (India) 30 October 1998 49 46 International Coconut Genebank for Southeast and East Asia (Indonesia) 26 May 1999 52 29 International Coconut Genebank for Africa and The Indian Ocean (Côte d'Ivoire) 14 October 1999 49 99* Table 2. Germplasm conserved in the multi-site ICG 114 COCONUT GENETIC RESOURCES d’Ivoire and Papua New Guinea) to member countries and soon, from the other ICG host countries where some of the new conserved materials are now starting to bear fruits. While IPGRI/COGENT desires to implement a progressive germplasm movement initiative, at the same time, it would like to ensure that this is done in a safe manner to protect the coconut industry of receiving countries. Thus, IPGRI approached the Australian Centre for International Agricultural Research (ACIAR) to fund the development and publication of a manual on Germplasm Health Management for COGENT’s multi-site International Coconut Genebank. ACIAR has agreed to support this very important and strategic initiative. This manual will be useful as a guide to genebank managers and plant quarantine officers worldwide in making informed decisions on the safe movement of coconut genetic resources. 115 CHAPTER 3: Germplasm conservation The International Coconut Genebank for the South Pacific (Papua New Guinea) M Faure Coconut Breeder, Cocoa and Coconut Institute (CCI), Madang, Papua New Guinea In August 1995, the COGENT Genebank Task Force visited PNG to evaluate the suitability of the proposed site and the commitment of the PNG government. The proposed genebank site is the Murunas plantation currently named Stewart Research Station of the PNG Cocoa and Coconut Institute (CCRI) in Madang, which has a total area of 450 ha, 200 ha of which was made available for the ICG. The CCRI staff and laboratories in Rabaul will provide support to the field genebank in Madang. The larger vegetation was cleared in 1993 and the secondary growth will be cleared as needed. Drainage canals will also be constructed as needed. The annual rainfall is 3500 mm, evenly distributed, and the soil is mostly silty clay loam. Following the successful site suitability evaluation and COGENT’s acceptance, the PNG Stewart Station’s coconut genebank has been transformed into the International Coconut Genebank for the South Pacific (ICG-SP), which to date conserves a total of 50 designated germplasm. The Memorandum of Agreement to establish the ICG-SP was signed in November 1998 between the Government of PNG, IPGRI on behalf of COGENT and the Food and Agriculture Organization (FAO) of the United Nations serving as trustee. The list of initial designated germplasm as stipulated in the signed MOA is shown in Annex 1.2. Since 1994 to 2004, 10 specialists visited PNG on eight technical assistance missions including assessing the country’s coconut R&D capability and assist the national programme in identifying common problems and opportunities for network collaboration, identifying a suitable site for ICG-SP, evaluating embryo culture laboratories and training their staff, evaluating COGENT’s germplasm collecting and conservation strategies, assessing the pest risk for the ICG-SP, assisting in the installation of machineries and training in the production of coconut virgin oil, fiber-based products and coconut candies. To date, IPGRI/COGENT has helped the ICG-SP established an embryo culture laboratory which is currently fully operational with additional stocks of glassware and chemicals purchased and a seedling nursery for ex vitro seedling production established in June 2001. It has also supported the training of ICG staff on embryo culture, genebank management, germplasm characterization using morphometric methods 116 COCONUT GENETIC RESOURCES and molecular marker (microsatellite kits) methods. In addition, six local coconut researchers have undergone staff development training sponsored by COGENT on topics such as the standardized research techniques in coconut breeding (STANTECH); coconut collecting and conservation; coconut data analysis; computer use, documentation, data analysis, dedicated statistical software; and coconut cryopreservation. The ICG-SP contains important germplasm from the fragile ecosystem of the South Pacific. These include typhoon-resistant accessions with big trunks and fruits, which are suitable for the Pacific islands. In the last two years, through COGENT-CIRAD collaboration, precious coconut populations from Cooke Islands, Fiji, Kiribati, Marshall Islands and Tuvalu have been collected, which were not previously available. A total of 13 accessions from four atoll countries (Tuvalu, Kiribati, Cook Islands and Marshall Islands) were collected by the CIRAD/COGENT team, embryo cultured and initially grown in vitro at the laboratories of the Secretariat of the Pacific Community (SPC) in Suva, Fiji and subsequently sent to the ICG–SP in 2000/2001. These accessions, which were collected to prevent losing them from the threat of global warming and possible water rise, were grown in the embryo culture laboratory and nursery and planted in the field. Other germplasm from other Pacific countries will also be imported when funds become available. A total of 62 embryos from one accession (Fiji Tall) were provided by Fiji in March 2002. The imported germplasm are currently being maintained in the laboratory using the upgraded coconut embryo culture protocol. Most of the evaluation work on the performance of the germplasm is being carried out in the field. Recently, a report was received from ICG-SP Papua New Guinea stating that frequent power outages, have been posing a serious threat to the operations of the embryo culture laboratory and causing damage to the embryo-derived plantlets. The PNG Cocoa and Coconut Institute (CCI) and the Secretariat of the South Pacific Commission requested the assistance of COGENT and IPGRI to enable the institute to purchase a standby generator. In response to this request, IPGRI/COGENT co- financed with CCI the purchase and installation of a standby generator. The generator is currently being used to support the air conditioners and other equipment of the embryo culture laboratories in case of power interruptions. 117 CHAPTER 3: Germplasm conservation The International Coconut Genebank for South Asia (India) V Rajagopal Director, Central Plantation Crops Research Institute (CPCRI), Kasaragod, Kerala, India The Central Plantation Crops Research Institute (CPCRI) hosts the International Coconut Genebank for South Asia (ICG-SA). The field genebank in Kidu Farm, Karnataka, which is the ICG-SA field genebank, is supported technically by the laboratory facilities at CPCRI, Kasaragod. CPCRI maintains the world’s largest assemblage of germplasm by undertaking the planting and maintenance of the field genebank and activities on embryo culture, assessment of diversity using molecular markers and disease indexing. The National Bureau of Plant Genetic Resources (NBPGR), New Delhi, collaborates with CPCRI on cryopreservation activities. In July 1995, the COGENT Task Force evaluated the forested area adjacent to the CPCRI Seed Farm in Kidu, the genebank site proposed by the government of India, and found it suitable. The Kidu field genebank is situated in Dakshina Kannada District of Karnataka about 90 km east of Mangalore and about 100 km east of Kasaragod. The farm lies between 12.30°N and 75.20°E at an elevation of 291 msl. The summer temperature range between 33 and 40°C and the winter temperature is between 22 and 18°C. The soil is mostly red lateritic, changing to alluvial laterite towards the riverbank. The average annual rainfall is 2900 mm with a river on the southern farm boundary as perennial source of irrigation water. Irrigation is essential as the site has a distinct dry period. Since the proposed site is within a forest without any coconut plantation nearby, the risk of disease spread from neighboring plantations is minimal. The nearest root wilt affected area is 650 km from the site and the disease is said to have moved only 100 km during the last 120 years. The Memorandum of Agreement for the establishment of the ICG- South Asia was signed by the Government of India, IPGRI on behalf of COGENT and FAO as trustee in October 1998. The list of initial designated germplasm during the signing of the MOA is shown in Annex 1.3. To date, India has conserved a total of 46 of designated germplasm in the ICG-SA. Nearly 30 ha of forestland have been cleared and surrounded with electric fencing for planting. Furthermore, drip irrigation has been provided to 2700 seedlings that have been planted there. IPGRI/COGENT has helped the ICG-SA in collecting germplasm from the Indian Ocean Islands of Maldives, Comoros, Madagascar, Reunion and Seychelles. A total of 746 embryos were collected from Sri Lanka as 118 COCONUT GENETIC RESOURCES of February 2001. Out of these, a total of 396 embryos were damaged. A total of 401 embryos were collected from Bangladesh during November to December 2001, of which 157 embryos survived. The embryos were collected from the following varieties: Chinasukanya, Chinasukanya Dwarf Orange, Pubail Tall, Kayemkola Tall, Bagharpara Tall, Rupdia Tall, Khairtala Tall and BARI Narikel-I, BARI Narikel-II, Uzirpur Tall and Agailjhara. In 2003, 34 collections were added to the existing collection. These included five collections from Goa, six from Maharastra, eight from Assam, four from Sri Lanka and 11 from Bangladesh. The collections from Sri Lanka and Bangladesh were in the form of zygotic embryos. These embryos cultured in vitro were rooted and later planted in pots. Conservation of coconut germplasm in the form of in vitro culture is being attempted at CPCRI, Kasaragod. Furthermore, a total of 4962 inter- crossed nuts from 31 accessions were also sown to generate planting materials at the ICG-SA. To produce additional seednuts, a total of 3004 female flowers were pollinated from eight accessions for regeneration. From 1995 to the present, seven coconut specialists visited India to help identify a suitable site for the ICG-SA; evaluate COGENT’s collecting strategies; and conduct a pest-risk assessment for the genebank. In 2000, a regional training course on In Vitro Conservation and Cryopreservation on PGR was conducted by NBPGR with seven participants from five COGENT member countries. Another eight local staff from various collaborating institutions and NARS were trained on various topics including the use of the manual on standardized research techniques in coconut breeding (STANTECH), in vitro embryo culture and cryopreservation techniques as well as the use of the microsatellite kit and dedicated statistical software. 119 CHAPTER 3: Germplasm conservation The International Coconut Genebank for Southeast and East Asia (Indonesia) H Novarianto Director, Indonesian Coconut and Palm Research Institute (ICOPRI), Manado, Indonesia The International Coconut Genebank for Southeast and East Asia (ICG- SEEA) is hosted by the Indonesian Agency for Agricultural Research and Development (AARD) using the field genebank in Pekanbaru, Riau Province; experimental gardens in Manado, North Sulawesi; and AARD laboratory facilities in Bogor, West Java and in Manado, North Sulawesi. In July 1995, the COGENT Task Force evaluated the proposed site at Sikijang Mati, Pekanbaru, Riau Province in Central Sumatra and found it to be generally suitable and made some suggestions for improvement. The site is located 20 km from the city of Pekanbaru, the capital of Riau. Pekanbaru has regular flights from Jakarta (1.5 hours) and Singapore (30 minutes), as well as other cities in Sumatra. The annual rainfall is about 2000 mm, well distributed over the year. The topography of the area is undulating and most of the land is covered by secondary forest, with small rivers. The soils are yellow to yellow-red podzolic, low in organic matter and with pH of around 5.0. The soils are generally very poor and unsaturated but they make a good substratum for the crop to grow and respond well to the application of fertilizers, which are readily absorbed by the crop. Since the area was not very uniform, the Task Force recommended that a detailed survey be undertaken to select only those areas where soils are generally good and more than one meter deep to avoid the hard pan. About 1000 ha of secondary forest has been offered by the Government of Indonesia which could be used for the ICG (200 ha) and the rest for production area to generate income for the maintenance cost of the ICG. The Memorandum of Agreement for the establishment of the ICG- Southeast and East Asia was signed by the Government of Indonesia, IPGRI on behalf of COGENT and FAO as trustee in May 1999. The function of the coconut collection at Sikijang was not only for germplasm conservation, but also for genetic evaluation and utilization. To date, Indonesia has conserved a total of 29 of the designated germplasm in the International Coconut Genebank for Southeast and East Asia at Sikijang. The list of initial designated germplasm during the signing of the MOA is shown in Annex 1.4. Due to the financial crisis in 1997 and the resulting lack of government budget, there was slow development of the Sikijang area, resulting in the squatting of the remaining areas by surrounding inhabitants and 120 COCONUT GENETIC RESOURCES migrants. Two extension ICG areas have therefore been identified: the Paniki Experimental Garden (100 ha) located beside the Indonesian Coconut and Other Palmae Research Institute (ICOPRI) in Manado, and the Pandu Experimental Garden (80 ha) which is about 18 km from the ICOPRI office and belonging to the Balai Pengkajian Teknologi Pertanian (BPTP). The soil and climate there are very suitable for coconut growing. Therefore, it was recommended that the main part of the ICG-SEEA be moved from Sikjiang to North Sulawesi. However, the 29 accessions which have been collected will remain in Sikijang and maintained by the Indonesian Government. To date, a total of four accessions have been received from Malaysia, six from China and 10 each from the Philippines, Thailand and Vietnam, respectively. A total of at least 100 accessions have been conserved from Indonesia from 1996 to 2001. Twenty-nine of these 100 accessions have been planted at Sikijang, Pekanbaru, Riau, which was the initial identified site for the ICG-SEEA. In addition, a total of 460 embryos of Malayan Tall and 469 embryos of Malayan Green Dwarfs were received from Malaysia and successfully cultured in vitro. IPGRI/COGENT has supported the ICG-SEEA in collecting germplasm from the Moluccas Island, East Timor, West Nusa Tenggara, Sangir Talaud Islands, Salibabu Island, Buol District, Central Sulawesi, Sangir Talaud district and North Sulawesi. From 1995 to 2000, 11 specialists have visited Indonesia to help the country in its coconut PGR activities. These include identifying a suitable site for the ICG-SEEA, collecting leaf samples for electron microscopy detection of mycoplasma, identifying marketable alternative products for coconut as well as suitable varieties for these products, evaluating COGENT’s collecting and conservation strategies, assessing pest risk and evaluating the progress of the ICG, and assisting in the installation of equipment for feasibility studies. Four training courses were held in the country, whereby 52 researchers from nine countries attended. The training courses, which were funded by the Asian Development Bank (ADB), were hosted by ICOPRI (formerly the Research Institute for Coconut and Palmae or RICP). IPGRI/COGENT has also sponsored 20 local researchers and specialists for staff development training in coconut data analysis, coconut collecting and conservation, embryo culture, technical writing/seminar presentation and proposal writing, the use of the microsatellite kit and others which are related to the COGENT’s poverty reduction project. 121 CHAPTER 3: Germplasm conservation The International Coconut Genebank for Africa and Indian Ocean (Côte d’Ivoire) JL Konan Head, Coconut Research Programme, Centre National de Recherche Agronomique (CNRA), Marc Delorme Station, Côte d’Ivoire The ICG-AIO is hosted by the Marc Delorme Coconut Research Station in Côte d’Ivoire which has a total area of 1200 ha. The soil of the station is composed of alluvial deposits of tertiary sands with 8-10% clay, poor in organic matter and minerals. The climate is characterized by two dry seasons of different lengths, one from December to April and the other in August to September which alternate with two rainy seasons. The mean annual rainfall is 1800 mm. The MOA for the establishment of the ICG-AIO was signed in October 1999 by the Government of Côte d’Ivoire, IPGRI on behalf of COGENT and FAO as trustee. At the time of the signing of the MOA, the coconut genebank of the Marc Delorme Coconut Research Station was converted into the ICG-AIO. The list of initial designated germplasm during the signing of the MOA is shown in Annex 1.5. To date, ICG-AIO has a total of 99 accessions. Furthermore, five Tall varieties from Sri Lanka, Tonga, Vanuatu, Tagnanan and Rotuma were received and planted on eight hectares for their renewal. A total of 3400 embryos were provided to CIRAD/IRD Montpellier for in vitro culture technique development. Seednuts were also provided to participating countries in the CFC-funded multilocation hybrid trials of IPGRI/ COGENT. A researcher from the Centre National Agronomique (CNRA) has visited the five other participating countries (Benin, Tanzania, Brazil, Mexico and Jamaica) to help in the project trial implementation. Two researchers from Marc DELORME visited the western region of Ghana as part of its collaborative research activity on lethal yellowing disease. Twelve kilograms of VTT (Vanuatu Tall) pollen were also provided to the Ghana coconut programme per year to produce lethal yellowing-tolerant hybrids. For 2004, nine Dwarf varieties from the ICG- AIO were selected to be tested also against the disease. Marc Delorme Station also received two research teams, from Senegal and Mayotte Island to help them in coconut development. A total of 70 800 seednuts of improved varieties were produced for smallholder farmers and the industrial sectors in the country. For Nicaragua (Coconut Research Institute or CRI), 1200 grams of Panama Tall Monagre pollen have been provided per year to allow appropriate hybrids production. About 9500 122 COCONUT GENETIC RESOURCES seedlings of improved Mawa (PB121) hybrids were provided to Guinea in 2000 and 2001 for commercial planting. In 1999, one COGENT-commissioned expert visited Côte d’Ivoire to conduct a pest risk analysis of the ICG-AIO. Two training courses were hosted by the Centre National de Recherche Agronomique (CNRA) in the Côte d’Ivoire up to 2002. Researchers representing 11 countries participated in the training courses. Furthermore, another two local staff underwent IPGRI/COGENT-sponsored staff development training at CIRAD in Montpellier, France on the use of molecular markers (microsatellite kit and associated statistical software), and on cryopreservation. Currently, the application of microsatellite analysis is ongoing at the central biotechnology laboratory of the Centre National de Recherche Agronomique. Leaf samples of important varieties from Ghana will also be collected and analyzed. These activities are funded by IPGRI/COGENT. Leaf samples of Cameroon Red Dwarf x Rennell Island Tall were provided to CIRAD in France and Max Planck Institute in Germany for coconut map construction, in collaboration with the Mikocheni Agricultural Research Institute in Tanzania (MARI), Philippine Coconut Authority (PCA) and NEIKER in Spain. For this hybrid, agronomic evaluation is being undertaken in Marc Delorme. About 7200 seednuts of Talls (seven varieties), Dwarf s(nine varieties) and hybrids (seven crossings) were produced by assisted and controlled pollination for Mozambique for coconut seedgarden establishment. For germplasm exchanging, the Coconut Research Institute of Sri Lanka is sending a research team to Marc Delorme Station in August 2004 to bring embryos of three varieties (Nawasi Tall, King coconut and Ran Thambili) to the ICG-AIO. In return, embryos of seven varieties will be collected from ICG-AIO and brought to Sri Lanka for conservation. 123 CHAPTER 3: Germplasm conservation Proposal for the establishment of the International Coconut Genebank for Latin America and the Caribbean (Brazil) EA Tupinamba Coconut Researcher, Centro de Pesquisa Agropecuaria dos Tabuleiros Costreiros - Empresa Brasileira de Pesquisa Agropecuaria, Aracaju, Sergipe, Brazil During the COGENT Steering Committee meeting in November 1998, the representative of Empresa Brasileira de Pesquisa Agropecuaria (EMPRAPA) presented Brazil’s proposal to host the International Coconut Genebank for Latin America and the Caribbean (ICG-LAC). Subsequently, a site suitability and pest risk assessment survey was undertaken in April 1999 to evaluate the suitability and pest risk of the ICG if situated in Itaporanga, west of Aracaju; the Neopolis plateau, northeast of Aracaju; and Betume, located between Neopolis and Ilha das Flores. Due to ownership problems, the Neopolis Plateau was dropped as a prospective site. Likewise, due to distance problem, the Betume Station was also not found suitable. Thus, the Itaporanga station was subsequently identified as the proposed site for the ICG- LAC. Itaporanga is 20 km from the city of Aracaju, located 10o 55’ South Latitude and 37o 03’ West longitude, with an elevation of only one meter above sea level. Its predominant soil is ferric with good drainage. The climate is generally warm with the temperatureof the coldest month higher than 15oC. The average annual rainfall is 1643 mm. The area is flat and about 100 ha is available for establishing Tall accessions. Additional areas to plant additional accessions should be identified. In 1999, COGENT commissioned one expert to go to Brazil to conduct a pest risk analysis of the proposed site of the ICG-LAC. Two local staff were sponsored by COGENT to attend staff development training course on the use of the standardized techniques in coconut breeding (STANTECH), microsatellite kit (molecular marker), dedicated statistical software, technical writing/ seminar presentation and proposal writing. Several meetings and communications were conducted between EMBRAPA and COGENT to discuss issues related to the hosting of the ICG-LAC which includes the issues of derivatives, compliance to Brazil’s legislation on intellectual property rights and funding. Embrapa has finally agreed to host the ICG-LAC and the Memorandum of Agreement will be signed soon. 124 COCONUT GENETIC RESOURCES Annex 1.1. MOA for the establishment of the International Coconut Genebank for the South Pacific AGREEMENT BETWEEN THE GOVERNMENT OF PAPUA NEW GUINEA, THE INTERNATIONAL PLANT GENETIC RESOURCES INSTITUTE (IPGRI) AND THE FOOD AND AGRICULTURE ORGANIZATION OF THE UNITED NATIONS (FAO) PLACING COCONUT GERMPLASM COLLECTIONS UNDER THE AUSPICES OF FAO PREAMBLE The Government of Papua New Guinea (hereinafter referred to as ‘Host Country’), hosting the International Coconut Genebank for the South Pacific, the International Plant Genetic Resource Institute (hereinafter referred to as ‘IPGRI’, one of the Cen- tres of the Consultative Group on International Agricultural Research), acting on behalf of the International Coconut Genetic Resources Network (COGENT), as described in the attachment ‘Background to the Agreements’) and the Food and Agriculture Organization of the United Nations (hereinafter referred to as ‘FAO’); Considering the importance to humanity of protecting and conserving coconut germplasm for future generations; Considering the International Undertaking on Genetic Resources adopted by the FAO Conference at its 22nd Session in 1983 (Resolution 8/83) and in particular Article 7 thereof; and the Annexes of the Undertaking adopted by the FAO Conference in 1989 and 1991; Considering that the FAO Commission on Genetic Resources for Food and Agricul- ture (hereinafter referred to as the “Commission”), as the relevant intergovernmental body in this field, has the responsibility for monitoring the implementation of Arti- cle 7 of the International Undertaking on Plant Genetic Resources; Considering the Memorandum of Understanding Between the Food and Agricul- ture Organization of the United Nations and the International Board for Plant Genetic Resources (IBPGR) legally succeeded by IPGRI, dated September 21, 1990, on the respective roles of the two organizations in establishing, maintaining and managing germplasm collections and setting standards for these collections; Considering the importance of the International Coconut Genebank held by the Government of Papua New Guinea within COGENT and supported by IPGRI, as part of a global strategy for germplasm conservation; Considering that the Coconut germplasm accessions have been donated to the In- ternational Coconut Genebank for the South Pacific on the understanding that these accessions will remain freely available; Considering that any country that so desires may participate in COGENT; 125 CHAPTER 3: Germplasm conservation Considering that the Government of Papua New Guinea has expressed the wish that the designated coconut germplasm accessions, kept in the International Coconut Genebank for the South Pacific, be recognized as part of the International Network of Ex Situ Collections (as per the International Undertaking on Plant Genetic Resources) under the Auspices of FAO; - Taking note of the provisions of the Convention on Biological Diversity, particularly those pertaining to affirmation of sovereign rights of nations over their biological resources and access and benefit sharing mechanisms. - Also taking note of the ongoing process of harmonisation of the International Undertaking on Plant Genetic Resources with the CBD, and the request of the Conference of the Parties to the Convention on Biological Diveristy to the governments to speed up this process. Have agreed as follows: Article 1 APPLICATION OF THIS AGREEMENT This Agreement shall be construed and applied in a manner consistent with the provisions of the Convention on Biological Diversity and the International Undertaking on Plant Genetic Resources. Article 2 BASIC UNDERTAKING The Government of Papua New Guinea hereby places under the auspices of FAO, as part of the International Network of Ex Situ Collections provided for in Article 7 of the International Undertaking on Plant Genetic Resources, the accessions of coconut genetic resources listed in the Appendix hereto (hereinafter referred to as the “designated germplasm”), in accordance with the terms and conditions set forth in this Agreement. The List of designated germplasm will be updated every two years as new accessions are added to the collection. Article 3 STATUS OF DESIGNATED GERMPLASM a) The Government of Papua New Guinea shall hold the designated germplasm in trust for the benefit of all countries in accordance with the International Undertaking on Plant Genetic Resources and the terms and conditions set out in this Agreement. b) The Government of Papua New Guinea shall not claim legal ownership over the designated germplasm, nor shall it seek any intellectual property rights over that germplasm or related information. 126 COCONUT GENETIC RESOURCES Article 4 PREMISES a) The premises, i.e., land and/or laboratories, in which the designated germplasm is conserved, shall remain in the charge of the Government of Papua New Guinea. b) FAO shall have a right of access to the premises at any time and the right to inspect all activities performed therein directly related to the conservation and exchange of the designated germplasm. Article 5 MANAGEMENT AND ADMINISTRATION a) The Government of Papua New Guinea undertakes to manage and administer the designated germplasm in accordance with Internationally Accepted Standards, including standards as agreed upon by COGENT, and the Interna- tional Genebank Standards, endorsed by the Commission, where these are applicable to coconut, and ensuring that all the designated germplasm is duplicated in order to ensure its safety. b) FAO may recommend action, if it considers such action to be desirable, to ensure the proper conservation of the designated germplasm. c) If the orderly maintenance of the designated germplasm is impeded or threatened by an event, including force majeure, and the Government of Papua New Guinea does not have the capacity to take appropriate preventive or curative action, FAO and IPGRI shall seek the necessary resources from the international community for action to ensure the safety of the designated germplasm, including if necessary by its evacuation and transfer. Article 6 POLICIES The Government of Papua New Guinea and IPGRI recognize the intergovernmental authority of FAO and its Commission in setting policies for the International Network of Ex Situ Collections referred to in Article 7 of the International Undertaking and undertake to consult with FAO and its Commission on proposed policy changes related to the conservation of, or accessibility to, the designated germplasm, subject, always to the provisions of Article 9 hereinafter. The Government of Papua New Guinea and IPGRI shall give full consideration to any policy changes proposed by the Commission. Article 7 STAFF a) Staff responsible to manage and administer the designated germplasm shall be employed and remunerated by the Government of Papua New Guinea. b) As and when deemed appropriate, FAO and IPGRI shall furnish technical backstopping on request by the Government of Papua New Guinea and COGENT. 127 CHAPTER 3: Germplasm conservation Article 8 FINANCES The Government of Papua New Guinea shall remain responsible for financing the maintenance of the designated germplasm. Article 9 AVAILABILITY OF DESIGNATED GERMPLASM AND RELATED INFOR- MATION Subject to the provisions of Article 10 below, the Government of Papua New Guinea undertakes to make samples of the designated germplasm and related information available directly to all countries participating in COGENT, for the purpose of scientific research, plant breeding or genetic resource conservation, without restric- tion. Article 10 TRANSFER OF DESIGNATED GERMPLASM AND RELATED INFORMA- TION Where samples of the designated germplasm and/ or related information are transferred to any other person or institution the Government of Papua New Guinea shall ensure that such other person or institution, and any further entity receiving samples of the designated germplasm from such person or institution, is bound by the conditions set out in Article 3 (b) and, in the case of samples duplicated for safety purposes, to the provisions of Article 5 (a). This provision shall not apply to the repatriation of germplasm to the country that provided such germplasm. Article 11 DURATION a) This Agreement is concluded for a period of 4 years and shall be automatically renewed for further periods of 4 years unless notice of non-renewal is given in writing by either party not less than 2 years before the end of any 4-year period. b) This Agreement shall be revised, if necessary, in accordance with the provisions of the revised International Undertaking. Article 12 TERMINATION a) Either FAO or the Government of Papua New Guinea may terminate this Agree- ment at any time by giving notice to the other, two years in advance of the termination date. b) FAO, the Government of Papua New Guinea and IPGRI, shall, in such case, take all necessary measures to wind up joint activities in an appropriate manner and, within the limits of their respective competencies, to ensure the continued con- servation of and access to the designated germplasm. 128 COCONUT GENETIC RESOURCES Article 13 SETTLEMENT OF DISPUTES a) Any dispute concerning the implementation of this Agreement shall be settled by mutual consent. b) Failing mutual consent, such dispute may be submitted, at the request of either FAO, or the Government of Papua New Guinea or IPGRI, to an arbitral tribunal composed of four members. Each party shall appoint one arbitrator. The three arbitrators thus appointed shall designate by mutual consent the fourth arbitrator, who will act as the presiding arbitrator of the tribunal. In case of equal division of votes the presiding arbitrator will have a second vote. c) If within two months after the receipt of a party’s notification of the appointment of an arbitrator one or both of the other parties has/have not notified the first party of the arbitrators they have appointed, the first party may request the Secretary-General of the United Nations to appoint arbitrators to represent par- ties that have not appointed an arbitrator. d) If within two months after the appointment of the three arbitrators they have not agreed on the choice of the presiding arbitrator, such presiding arbitrator shall be designated by the Secretary-General of the United Nations at the request of either party. e) Unless the parties to the dispute decide otherwise, the tribunal shall determine its own procedure. f) A majority vote of the arbitrators shall be sufficient to reach a decision which shall be final and binding for the parties to the dispute. Article 14 AMENDMENT a) FAO, the Government of Papua New Guinea or IPGRI may propose that the Agreement be amended by so informing the other parties b) If there is mutual agreement in respect of a proposed amendment, the amendment shall enter into force on whatever date is set, and be reported to the next session of the Commission. Article 15 DEPOSITARY The Director-General of FAO shall be the Depositary of this Agreement. The Depository shall: a) Send certified copies of this Agreement to the Member Nations of FAO and to any other Government which so requests; b) Arrange for the registration of this Agreement, upon its entry into force, with the Secretariat of the United Nations in accordance with Article 102 of the Charter of the United Nations; c) Inform FAO Member Nations of: i) The signature of this Agreement in accordance with Article 16; and ii) The adoption of amendments to this Agreement in accordance with Article 14. 129 CHAPTER 3: Germplasm conservation Article 16 COMING INTO FORCE This Agreement shall come into force upon signature by the authorized representative of FAO, the Government of Papua New Guinea and IPGRI. The Food and Agtriculture Organization of the United Nations By: DAVID A. HARCHARIK (Signature) Date: The International Plant Genetic Resources Institute (IPGRI) By: GEOFFREY HAWTIN (Signature) Date: The Government of Papua New Guinea Hon. Tukape Masane, M.P. Minister of Agriculture and Livestock By: Date: 130 COCONUT GENETIC RESOURCES Appendix 1 List of germplasm accessions covered by this Agreement Accessions Source Accessions Source East New Britain 31. Saiho Oro 1. Pellavarua - Gazelle Peninsula 32. Ajoa 2. Raulawat 33. Kikibator 3. Natava 34. Siagara Milne Bay 4. New Massava 35. Bubuleta 5. Natava Many 36. Baibara Central 6. Fruited 37. Hisihu 7. Gaungo West New Britain 38. Poligolo 8. Naviro 39. Miha Kavava Gulf -Vailala 9. Talasea Red 40. Keakea New Ireland 41. Iokea - Iokea 10. Karu village - Namatanai 11. Kenapit 42. Severimabu Western (Kiwai Tall) 12. Sohu 43. Boze 13. Etalat - Mussau Is. Exotic Talls 14. Lawes Manus 44. Rennell - Rennell Tall 15. Lako 16. Baluan 45. PNG Yellow Local Dwarfs 17. Wutung Sandaun 46. PNG Red 1 18. Hawain East Sepik 47. PNG Red 2 19. Yangoru 48. Rabaul Red 20. Vokio 49. PNG Brown 21. Marineberg 50. Iokea Red 22. Guanaga Madang (Karkar Tall) 51. Malayan Yellow Exotic Dwarfs 23. Kinim 52. Malayan Red 24. Ulatava 53. Nias Green Morobe 54. Nias Yellow 25. Markham Farm - Markham Tall 55. Nias Red 26. Liara village East New Britain 27. Raulawat Yellow - Gazelle Peninsula 28. Raulawat Red 29. Natava Yellow 30. Natava Red 131 CHAPTER 3: Germplasm conservation Additional list of international germplasm to be established Locations where materials are held: • 1 – 55 Stewart Research Station; Papua New Guinea Cocoa & Coconut Re- search Institute Madang, Madang Province • 56 – 82 To be established at Stewart Research Station, Madang, PNG Ecotype Source 56. Rotuma Tall Fiji 57. Tonga Tall Tonga 58. Kiribati Tall Kiribati 59. Rangiroa Tall Tahiti 60. Vanuatu Tall Ivory Coast, PNG 61. Western Samoan Tall Western Samoa 62. Samoan Yellow Dwarf Western Samoa 63. Nui Leka Green Dwarf Fiji 64. Fiji Tall Fiji 65. Niu Vai Western Samoa 66. Niu Afa Western Samoa 67. Christmas Is. Tall Kiribati 68. Kiribati Green Dwarf Kiribati 69. New Caledonia Tall New Caledonia 70. Vanikoro Tall Solomon Island 71. Solomon Tall Solomon Island 72. Niu-bubu, or Pine or Mami Kokonas PNG & Solomon Islands Other Ecotypes 73. Cameroon Red Dwarf Ivory Coast 74. Salak Green Dwarf Indonesia 75. Pilipog Green Dwarf Philippines 76. Tacunan Green Dwarf Philippines 77. Aromatic Green Dwarf Thailand 78. Catigan Green Dwarf Philippines 79. Brizilian Green Dwarf Ivory Coast 80. West African Tall Ivory Coast 81. Sri Lankan Tall Sri Lanka 82. Panama Tall Jamaica 132 COCONUT GENETIC RESOURCES Appendix 2 Background to the agreements The Coconut Genetic Resources Network (COGENT) was established in 1992 to improve coconut production on a substantial basis and to increase incomes in developing countries through improved cultivation of the coconut and efficient utilization of its products. COGENT is actively undertaking an international collaborative programme with member countries to improve the conservation and use of coconut genetic resources in the following areas: 1) Establishing and maintaining an International Database on existing and future collections; 2) Encouraging the protection and utilization of existing germplasm collections; 3) Identifying and securing additional threatened diversity through the development and adoption of suitable technologies and con- servation strategies; 4) Promotion of greater collaboration among research groups in producer countries and advanced technology sources in the exchange of germplasm and the development of new techniques; and 5) Appropriate training, information dissemination and securing the necessary funding. COGENT operates through a steering committee comprised of two members from each of the five sub-networks namely Southeast Asia, South Asia, Pacific, Africa and Latin America/Caribbean, and a full time coordinator based in the Asia, Pacific and Oceania Regional Office of the International Plant Genetic Resources Institute (IPGRI-APO) in Serdang, Malaysia. COGENT’s membership has now grown to 38 coconut-producing countries, with each country having to agree to provide access to its coconut germplasm and data as one of the conditions for membership. The member countries are shown in the table below. Southeast and East Asia South Asia South Pacific Africa/Indian Ocean Latin America/ Caribbean 1. China 2. Indonesia 3. Malaysia 4. Myanmar 5. Philippines 6. Thailand 7. Vietnam 1. Bangladesh 2. India 3. Pakistan 4. Sri Lanka 1. Cooke Islands 2. Fiji 3. Kiribati 4. Papua New Guinea 5.Solomon Islands 6. Tonga 7. Vanuatu 8. Samoa 1. Benin 2.Cote d’Ivoire 3. Ghana 4. Kenya 5. Madagascar 6. Mozambique 7. Nigeria 8. Seychelles 9. Tanzania 1. Brazil 2. Colombia 3. Costa Rica 4. Cuba 5. Guyana 6. Haiti 7. Honduras 8. Jamaica 9. Mexico 10.Trinidad- Tobago 133 CHAPTER 3: Germplasm conservation Under the mandate of the CGIAR, the IPGRI established COGENT with the endorsement of the Technical Advisory Committee. IPGRI functions as the executing institution for COGENT and provides administration and technical support and advice. An essential component for sustainable production and improvement in coconut is the availability of a wide diversity of germplasm from around the world for use as introductions or in coconut breeding programmes to develop improved coconut varieties and hybrids for coconut producing countries. To further ensure the security of germplasm in national collections which are important to each region and to provide member countries with germplasm for developing better varieties and hybrids, COGENT will establish an international multi-site genebank consisting of a regional genebank in each of the five COGENT regions. The host country will benefit from the use of the entire germplasm collection, and duplicates supplied from the other regional genebanks, in its breeding programme to develop high-yielding and adapted coconut varieties. The host countries have agreed to a 10-point criterion which includes, among others, access of member countries to the held germplasm and commitment to gather and submit data and to maintain the collection. The Convention on Biological Diversity (CBD) is a legally binding international agreement that sets out the sovereign rights of countries over their genetic resources as well as the responsibilities of states to conserve and to share these resources and benefits arising from their use. The Food and Agriculture Organization (FAO) is in the process of establishing Global Network of Ex Situ Collections. In December 1994, close to half a million germplasm accessions of food crops held by 12 International Agricultural Research Centres under the CGIAR were placed under FAO trusteeship through a series of agreements signed by FAO and the chairman of the CGIAR acting on behalf of each of the 12 Centres. These agreements were developed in accordance with the CBD. During a COGENT workshop held on 26-28 February 1996 at Pekanbaru, Riau, Indonesia, representatives of IPGRI, CIRAD and World Bank participated with COGENT members in developing a series of legal agreements, seven-year workplans and proposed budgets for each of the initial four genebanks to be hosted by India for South Asia, Indonesia for Southeast Asia, Papua New Guinea for the Pacific and Côte d’Ivoire for Africa. The following three agreements, which are considered consistent with the CBD and necessary to facilitate access to coconut genetic resources of which individual countries agree to designate to the international genebanks, are enclosed. These agreements follow closely those agreed 134 COCONUT GENETIC RESOURCES to by FAO and the CGIAR centres, with two important changes. First, each host country holding the designated accessions is to be a party in signing the tripartite agreement, and IPGRI is the second party, acting on behalf of COGENT. (a) The tripartite agreement [Agreement between {Name of Host Country}, the International Plant Genetic Resources Institute (IPGRI) and the Food and Agriculture Organization of the Uni- ted Nations (FAO) Placing Coconut Germplasm Collections under the Auspices of FAO], provides a list of designated accessions for each genebank, and spells out the rights and obligations of the parties to the agreement. (b) The Germplasm Acquisition Agreement sets out the terms and conditions of movement of coconut germplasm accessions from the providing country to each of the international genebanks. (c) A standard Material Transfer Agreement (MTA) specifies that the recipient agrees not to claim legal ownership over the designated germplasm or take out any intellectual property rights over that germplasm or related information. Furthermore, the recipient also undertakes to pass the same obligations to all fu- ture recipients of designated germplasm. The MTA will be used for designated germplasm. 135 CHAPTER 3: Germplasm conservation Annex 1.2 List of designated germplasm for the International Coconut Genebank for the South Pacific Accessions Source Accessions Source East New Britain 31. Saiho Oro 1. Pellavarua - Gazelle Peninsula 32. Ajoa 2. Raulawat 33. Kikibator 3. Natava 34. Siagara Milne Bay 4. New Massava 35. Bubuleta 5. Natava Many 36. Baibara Central 6. Fruited 37. Hisihu 7. Gaungo West New Britain 38. Poligolo 8. Naviro 39. Miha Kavava Gulf -Vailala 9. Talasea Red 40. Keakea New Ireland 41. Iokea - Iokea 10. Karu village - Namatanai 11. Kenapit 42. Severimabu Western (Kiwai Tall) 12. Sohu 43. Boze 13. Etalat - Mussau Is. Exotic Talls 14. Lawes Manus 44. Rennell - Rennell Tall 15. Lako 16. Baluan 45. PNG Yellow Local Dwarfs 17. Wutung Sandaun 46. PNG Red 1 18. Hawain East Sepik 47. PNG Red 2 19. Yangoru 48. Rabaul Red 20. Vokio 49. PNG Brown 21. Marineberg 50. Iokea Red 22. Guanaga Madang (Karkar Tall) 51. Malayan Yellow Exotic Dwarfs 23. Kinim 52. Malayan Red 24. Ulatava 53. Nias Green Morobe 54. Nias Yellow 25. Markham Farm - Markham Tall 55. Nias Red 26. Liara village East New Britain 27. Raulawat Yellow - Gazelle Peninsula 28. Raulawat Red 29. Natava Yellow 30. Natava Red 136 COCONUT GENETIC RESOURCES Additional list of international germplasm to be established Note: Locations where materials are held: • 1 – 55 Stewart Research Station; Papua New Guinea Cocoa & Coconut Research Institute Madang, Madang Province • 56 – 82 To be established at Stewart Research Station, Madang, PNG Ecotype Source Other Ecotypes Source 56. Rotuma Tall Fiji 73. Cameroon Red Dwarf Ivory Coast 57. Tonga Tall Tonga 74. Salak Green Dwarf Indonesia 58. Kiribati Tall Kiribati 75. Pilipog Green Dwarf Philippines 59. Rangiroa Tall Tahiti 76. Tacunan Green Dwarf Philippines 60. Vanuatu Tall Ivory Coast, PNG 77. Aromatic Green Dwarf Thailand 61. Western Samoan Tall Western Samoa 78. Catigan Green Dwarf Philippines 62. Samoan Yellow Dwarf Western Samoa 79. Brizilian Green Dwarf Ivory Coast 63. Nui Leka Green Dwarf Fiji 80. West African Tall Ivory Coast 64. Fiji Tall Fiji 81. Sri Lankan Tall Sri Lanka 65. Niu Vai Western Samoa 82. Panama Tall Jamaica 66. Niu Afa Western Samoa 67. Christmas Is. Tall Kiribati 68. Kiribati Green Dwarf Kiribati 69. New Caledonia Tall New Caledonia 70. Vanikoro Tall Solomon Island 71. Solomon Tall Solomon Island 72. Niu-bubu, or Pine or Mami Kokonas PNG & Solomon Islands 137 CHAPTER 3: Germplasm conservation Annex 1.3 List of designated germplasm for the International Coconut Genebank for South Asia KASARAGOD 1. Borneo 2. Standard Kudat 3. Java 4. Malayan Orange Dwarf 5. Malayan Green Dwarf 6. F.M.S. 7. S.S. Green 8. S.S . Apricot 9. Philippines Lono 10. San Ramon 11. Cochin China 12. Lifou Tall 13. British Solomon Islands 14. Jamaica Sanblas 15. St. Vincent 16. Blanchissuse 17. Kenya Tall 18. Camaroon Dwarf 19. West African Tall 20. Mawa Hybrid (PB 121 21. Zanzibar Tall 22. Ceylon Tall 23. King Coconut 24. Kappadam 25. Spicata 26. Ayiramkachi 27. Kulasekharam Green Dwarf 28. Kulasekharam Yellow Dwarf 29. Kulasekharam Orange Dwarf 30. Calangute 31. Nadora Tall 32. Andaman Giant 33. Andaman Ranguchan 33. Car Nicobar 34. Auck Chung 35. Tamaloo 36. Kimos 37. Kimmai 38. Katchal 39. Campbell Bay 40. Lakshdweep Micro KIDU 1. West Coast Tall 2. Andaman Ordinary 3. Benaulim 4. Tiptur Tall 5. East Coast Tall 6. Chowghat Green Dwarf 7. Malayan Yellow Dwarf 8. Philippines Ordinary 138 COCONUT GENETIC RESOURCES Annex 1.4 List of designated germplasm for the International Coconut Genebank for Southeast and East Asia Cultivars Code Source 1 Malayan Tall MLT Malaysia 2 Malayan Yellow Dwarf MYD Malaysia 3 Malayan Red Dwarf MRD Malaysia 4 Malayan Green Dwarf MGD Malaysia 5 Eo Brown Dwarf EOD Vietnam 6 Xiem Green Dwarf XGD Vietnam 7 Tam Quan Yellow Dwarf TYD Vietnam 8 Ta Tall TAAT Vietnam 9 Dau Tall DAUT Vietnam 10 Bung Tall (Bi Tall) BIT Vietnam 11 Giay Tall GIT Vietnam 12 Pluak Wan (Edible husk) PKWT Thailand 13 Pak Chok Tall PCKT Thailand 14 Maphrao So Tall SOXT Thailand 15 Kalok Thailand Tall KLKT Thailand 16 Thalai Roi Thailand Tall TLRT Thailand 17 Nalike Dwarf NKED Thailand 18 Maphrao Fai FAID Thailand 19 Bali Tall BAT Indonesia 20 Tenga Tall TAT Indonesia 21 Palu Tall PUT Indonesia 22 Sawarna Tall SAT Indonesia 23 Riau Tall RUT Indonesia 24 Mapanget Tall MTT Indonesia 25 Takome Tall TET Indonesia 26 Nias Yellow Dwarf NYD Indonesia 27 Bali Yellow Dwarf BYD Indonesia 28 Bali Green Dwarf BYD Indonesia 29 Jombang Green Dwarf JGD Indonesia 30 Sagerat Orange Dwarf SOD Indonesia 31 Salak Green Dwarf SGD Indonesia 32 Raja Brown Dwarf RBD Indonesia 33 Tagnanan Tall TAGT Philippines 34 Macapuno Tall MACT Philippines 35 Laguna Tall LAGT Philippines 36 Baybay Tall BAYT Philippines 37 Bago-Oshiro Tall BAOT Philippines 38 San Ramon Tall SNRT Philippines 39 Catigan Green Dwarf CATD Philippines 40 Pilipog Green Dwarf PILD Philippines 41 Aromatic Dwarf AROD Thailand 42 Hainan Tall HAT China 139 CHAPTER 3: Germplasm conservation 43 Cambodia tall KAT Côte d’Ivoire 44 West African Tall W AT Indonesia 45 Rennel Island Tall RIT Indonesia 46 Cameroon Red Dwarf CRD Indonesia 47 Tahiti Tall TAT Indonesia 48 Panama TaII PNT Côte d’Ivoire 49 Niu Leka Dwarf NLAD Côte d’Ivoire 50 Vanuatu Tall VTT Côte d’Ivoire 51 Indian West Coast Tall WCT India 52 Sri Lanka Tall SLT Sri Lanka 140 COCONUT GENETIC RESOURCES Annex 1.5 List of designated germplasm for the International Coconut Genebank for Africa and Indian Ocean Cultivars Code Source 1. Andaman Giant Tall AGT India 2. Andaman Ordinary Tall ADOT India 3. Baybay Tall BAYT Philippines 4. Cambodia Battambang Tall KAT09 Cambodia 5. Cambodia Koh Rong Tall KAT10 Cambodia 6. Cambodia Ream Tall KAT07 Cambodia 7. Cambodia Sre Cham Tall KAT08 Cambodia 8. Cambodia Tuk Sap Tall KAT02 Cameroon 9. Cameroon Kribi Tall CKT Cameroon 10. Cameroon Red Dwarf CRD Cameroon 11. Catigan Green Dwarf CATD Philippines 12. Comoro Moheli Tall CMT Comoro 13. Equatorial Guinea Green Dwarf EGD Equatorial Guinea 14. Gazelle Peninsula Tall GPT Papua New Guinea 15. Kappadam Tall KPDT India 16. Karkar Tall KKT Papua New Guinea 17. Kinabalan Green Dwarf KIND Philippines 18. Laccadive Micro Tall LMT India 19. Laccadive Ordinary Tall LCT India 20. Madang Brown Dwarf MBD Papua New Guinea 21. Malayan Green Dwarf MGD Malaysia 22. Malayan Red Dwarf MRD Malaysia 23. Malayan Tall MLT Malaysia 24. Malayan Yellow Dwarf MYD Malaysia 25. Markham Valley Tall MVT Papua New Guinea 26. Mozambique Tall MZT Mozambique 27. Niu Leka Dwarf NLAD Fiji 28. Palu Tall PUT Indonesia 29. Pilipog Green Dwarf PILD Philippines 30. Rangiroa Tall RGT French Polynesia 31. Rennell Island Tall RIT Solomon Islands 32. Rotuman Tall RTMT Fiji 33. Solomon Island Tall SIT Solomon 34. Sri Lanka Green Dwarf PGD Sri Lanka 35. Sri Lanka Tall Ambakelle SLT02 Sri Lanka 36. Tacunan Green Dwarf TACD Philippines 37. Tagnanan Tall TAGT Philippines 38. Tahitian Red Dwarf TRD French Polynesia 39. Tahitian Tall TAT French Polynesia 40. Takome Tall TKT Indonesia 41. Tenga Tall TGT Indonesia 42. Ternate Brown Dwarf TBD Indonesia 141 CHAPTER 3: Germplasm conservation 43. Thailand Green Dwarf THD Thailand 44. Thailand Tall Ko Samui THT04 Thailand 45. Thailand Tall Sawi THT01 Thailand 46. Tonga Tall TONT Tonga 47. West African Tall Akabo WAT03 Côte d’Ivoire 48. West African Tall Mensah WAT04 Côte d’Ivoire 49. West African Tall Quidah WAT06 Benin 142 COCONUT GENETIC RESOURCES Status of cryopreservation research in coconut F Engelmann1, 2, B Malaurie3, O N’Nan4 and M Borges5 1Institut de Recherche pour le Développement (IRD), 911 Avenue Agropolis, BP 64501, Cedex 5, Montpellier, France 2Honorary Research Fellow, International Plant Genetic Resources Institute (IPGRI), Via dei Tre Denari 472/a, Maccarese, Rome, Italy 3Senior Scientist, Institut de Recherche pour le Développement (IRD), 911 Avenue Agropolis, BP 64501, Cedex 5, Montpellier, France 4Researcher, Centre National de Recherche Agronomique (CNRA), Station de Recherche Marc Delorme, Port Bouët - 07 BP 13 Abidjan 07, Côte d’Ivoire 5Senior Research Officer, Instituto de Investigaciones Agropecuarias “Jorge Dimitrov” (IIA-JD), Bayamo, Granma, Cuba Introduction Seeds cannot be used for coconut germplasm conservation owing to their large size and their highly recalcitrant storage behaviour (Chin and Roberts 1980), which renders their storage under conventional dry and low-temperature conditions impossible. Genetic resources of coconut are thus traditionally maintained in field genebanks. There are many field collections of coconuts in various countries, usually connected with coconut research institutes, which conserve a total of 1416 accessions. This number is projected to increase over the next few years, with the establishment of the multi-site International Coconut Genebank or ICG (see preceding article) under the coordination of the International Coconut Genetic Resources Network (COGENT). In some ways, field genebanks offer a satisfactory approach to conservation. The genetic resources under conservation can be readily accessed and observed, permitting detailed evaluation. However, there are certain drawbacks that limit their efficiency and threaten their security (Withers and Engels 1990). The genetic resources are exposed to pests, diseases and other natural hazards such as drought, weather damage, human error and vandalism. Nor are they in a condition that is readily conducive to germplasm exchange. Field genebanks are costly to maintain and, as a consequence, are prone to economic decisions that may limit the level of replication of accessions, the quality of maintenance and even their very survival in times of economic stringency. Even under the best circumstances, field genebanks require considerable inputs in the form of land, labour, management and materials (see article on ‘Coconut field genebank’ in this chapter). It is now well recognized that the efficient and cost-effective conservation of any given genepool can be achieved only through the 143 CHAPTER 3: Germplasm conservation implementation of a complementary conservation strategy integrating in situ and ex situ approaches and utilizing relevant storage methods (Maxted et al. 1997). In this context, in vitro culture techniques have great potential for the collecting, exchange and conservation of plant germplasm, especially for problem plant species, i.e. those with recalcitrant seeds and those that are propagated vegetatively (Engelmann 1997; 2000). Tissue culture systems allow propagation of plants with high multiplication rates in an aseptic environment. Virus-free plants can be obtained through meristem culture and thermotherapy, thus ensuring the production of disease-free stocks and simplifying quarantine procedures for the international exchange of germplasm. The miniaturization of explants allows genebank managers to reduce space requirements and consequently, the labour costs for the maintenance of germplasm collections. For long-term conservation, cryopreservation, i.e. conservation at ultra-low temperature, usually immersion in liquid nitrogen (-196oC), is the only method currently available for problem species. Cryopreservation protocols have been developed for a wide range of plant species, an increasing number of which are from the tropics (Engelmann and Takagi 2000). In the case of coconut, in vitro culture of zygotic embryos is now routinely applied in numerous laboratories in the framework of germplasm collecting and exchange activities (Batugal and Engelmann 1998; Engelmann et al. 2002). This paper summarizes the present status of coconut cryopreservation research. Status of cryopreservation research In the case of coconut, only a limited amount of research has been conducted towards the development of cryopreservation protocols, involving research teams in Malaysia, Côte d’Ivoire, France and the UK. Cryopreservation experiments have been performed with zygotic embryos, plumules and pollen. Chin et al. (1989) first reported the survival of one single coconut embryo 15 months after freezing using a classical protocol (cryoprotection with DMSO + slow freezing). Assy-Bah et al. (1992a,b) reported high recovery of frozen embryos using the pregrowth/ desiccation technique. In the UK, Hornung et al. (2001) obtained callus growth from plumules of one coconut variety after cryopreservation using an encapsulation/dehydration protocol, including preculture of encapsulated plumules for 72-96 h in medium with 0.75 M sucrose followed by desiccation with silica gel to around 30% moisture content and rapid freezing. Research on cryopreservation of plumules using the encapsulation-dehydration and vitrification techniques has also been initiated in France (Malaurie et al. 2002b). Finally, preliminary 144 COCONUT GENETIC RESOURCES unpublished experiments performed in Côte d’Ivoire by the late Béatrice Assy-Bah showed that coconut pollen is amenable to cryopreservation after partial desiccation. Immature embryos Experiments were performed with immature embryos sampled from seednuts of the hybrid PB121 (Malaysian Yellow Dwarf x West African Tall) 7 to 8 months after pollination (Assy-Bah and Engelmann 1992a). It was decided to start working with immature embryos, on the assumption that they would be more likely to withstand cryopreservation than mature ones, owing to their smaller size and lower degree of differentiation (Engelmann 1992). For cryopreservation, embryos were placed for 4 h on pretreatment medium containing 600 g L-1 glucose and glycerol, sorbitol or polyethyleneglycol (PEG) 6000 at various concentrations and then immersed directly in liquid nitrogen. After rapid thawing in a water- bath at 40°C, embryos were cultured on standard medium (Assy-Bah et al. 1989) for recovery. After one month, the survival of non-cryopreserved embryos was high for all preculture conditions tested, ranging from 73 to 100%. In the case of cryopreserved embryos, lower survival, ranging between 10 and 43%, was obtained when glycerol at 5 or 10% or sorbitol at 10% was used in the preculture medium. However, numerous abnormalities were observed in the further development of non-cryopserved and cryopreserved embryos, and only few fully developed, normal plantlets could be obtained. This was because conditions for their in vitro culture were not mastered (Engelmann and Assy-Bah 1992). Mature embryos Experiments were performed with mature embryos sampled from the seednuts of the hybrid PB121; Cameroon Red Dwarf (CRD); Rennell Tall (RT); and Indian Tall (IT), 10 to 12 months after pollination (Assy-Bah and Engelmann 1992b). For cryopreservation, embryos were placed in open Petri dishes without culture medium and dehydrated for 4 h in the air current of the laminar flow cabinet at room temperature. They were then transferred to open Petri dishes with the medium employed for pretreatment of immature embryos containing 600 g L-1 glucose and 15% glycerol (Assy- Bah and Engelmann 1992a), and dehydrated for an additional period of 11 to 20 h. Hence, the total duration of the pretreatment ranged from 15 to 24 h. Embryos were then placed in 2 ml cryotubes and immersed directly in liquid nitrogen. After rapid thawing in a 40°C water-bath, embryos 145 CHAPTER 3: Germplasm conservation were cultured on standard medium (Assy-Bah et al. 1989) for recovery. The initial moisture content of embryos, which was very similar in all four varieties, averaged 78.4%. It decreased rapidly during the first 15 h of pretreatment to an average of 11.4% and then more slowly, reaching 6.4 % after 24 h. Larger embryos (RT and IT) dehydrated more slowly than smaller ones (PB 121 and CRD). Survival of non-frozen embryos remained very high (>70 %) after pretreatment. By contrast, no survival was noted after cryopreservation without pretreatment. For varieties with relatively larger embryos (RT, IT and PB 121), survival after cryopreservation increased in line with increasing pretreatment durations, whereas it reached an optimum after 17 h for the variety CRD, which has the smallest embryos. Under optimal pretreatment conditions, survival ranged between 76-100 % with non- frozen embryos and between 73-93 % with cryopreserved ones. Most embryos considered alive after one month germinated and the same proportion of non-cryopreserved and cryopreserved embryos developed into whole plantlets. The main differences between control and cryopreserved embryos were the non-development of the haustorium and a delay of 1 to 2 months in the development of cryopreserved ones. These results were validated by N’Nan (1997) with embryos of two ecotypes, West African Tall and Malayan Yellow Dwarf, and recently confirmed (N’Nan et al. 2003) on a total of 10 ecotypes, including 5 Talls and 5 Dwarfs, originating from Africa, Latin America-Caribbean, South Asia, Southeast Asia and the South Pacific, with 44-100% of cryopreserved embryos giving rise to whole in vitro plantlets. Plumules Plumules represent a potentially interesting material for cryopreservation because they are of small size (< 1mm3), they are mostly composed of meristematic cells and it is possible to regenerate whole plantlets from in vitro cultured plumules (Malaurie et al. 2002b). Cryopreservation experiments were performed using the encapsulation-dehydration and encapsulation-vitrification techniques. With the encapsulation-dehydration technique, excised plumules were encapsulated in alginate beads, pregrown for 2-3 days in medium containing 0.5 to 1.0 M sucrose, desiccated to 0.5-0.2 % moisture content and cryopreserved. Depending on the experiments, survival after cryopreservation could reach up to 67%, but only a limited number of frozen plumules could give rise to whole in vitro plantlets (Malaurie and Borges 2001; N’Nan et al. 2002). Preliminary experiments performed with the encapsulation-vitrification technique (Sakai et al. 2000) showed that up to 20% of cryopreserved plumules could survive after freezing 146 COCONUT GENETIC RESOURCES (Malaurie et al. 2002a). Conclusion and prospects These preliminary results demonstrate the great potential of cryopreservation for the long-term conservation of coconut genetic resources. Additional research has to be performed to further refine and standardize the protocols developed for embryos and plumules, to test the improved protocols with additional genotypes before their large-scale application in the genebank context can be envisaged. Long-term storage of coconut pollen under cryopreservation would represent an important additional technique for allowing conservation of genes. Research is needed to further develop and refine an appropriate technique. In view of the very positive results described above, it is clear that, in a not too distant future, cryopreservation will play a greater role in the overall approach in the conservation of coconut genetic resources. References Assy-Bah, B and F Engelmann. 1992a. Cryopreservation of immature embryos of coconut (Cocos nucifera L.). CryoLetters 13:67-74. Assy-Bah, B and F Engelmann. 1992b. Cryopreservation of mature embryos of coconut (Cocos nucifera L.) and subsequent regeneration of plantlets. CryoLetters 13:117-126. Assy-Bah, B, T Durand-Gasselin, F Engelmann and C Pannetier. 1989. Culture in vitro d’embryons zygotiques de cocotier (Cocos nucifera L.). Methode, révisée et simplifiée, d’obtention de plants de cocotiers transférables au champ. Oléagineux 44:515-523 Batugal, PA and F Engelmann (eds.). 1998. Coconut embryo in vitro cul- ture. Proceedings of the First Workshop on Embryo Culture, Banao, Guinobatan, Albay, Philippines, 27-31 October 1997. IPGRI-APO, Serdang, Selangor, Malaysia. Chin, HF and EH Roberts (eds.). 1980. Recalcitrant crop seeds. Tropical Press Sdn. Bhd., Kuala Lumpur, Malaysia. Chin, HF, B Krishnapillay and YL Hor. 1989. A note on the cryopreservation of embryos of coconut (Cocos nucifera L. var. Mawa). Pertanika 12:183-186. Engelmann, F. 1992. Cryopreservation of embryos. Pp. 281-290. In: Y Dattée, C Dumas and A Gallais (eds). Reproductive biology and plant breeding. Springer Verlag, Berlin. Engelmann, F. 1997. In vitro conservation methods. Pp. 119-162. In: BV Ford-Lloyd, JH Newburry and JA Callow (eds). Biotechnology and plant genetic Resources: Conservation and use. CAB International, Wallingford, UK. 147 CHAPTER 3: Germplasm conservation Engelmann, F. 1999. Cryopreservation of coconut germplasm. Pp. 289- 236. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardeña and JM Santamaria (eds). Current Advances in Coconut Biotechnology. Kluwer Academic Publishers, Dordrecht. Engelmann, F. 2000. Importance of cryopreservation for the conserva- tion of plant genetic resources. Pp. 8-20. In: F Engelmann and H Takagi (eds). Cryopreservation of tropical plant Germplasm: Current research progress and applications. JIRCAS, Tsukuba and IPGRI, Rome. Engelmann, F and B Assy-Bah. 1992. Maintenance of coconut genetic resources: In vitro techniques for medium and long-term conserva- tion. Pp. 63-69. In: Coconut genetic resources. Papers of an IBPGR Workshop, Cipanas, Indonesia, 8-11 October, 1991. International Crop Network Series No.8. IBPGR, Rome. Engelmann, F and H Takagi (eds). 2000. Cryopreservation of tropical plant germplasm: Current research progress and applications. JIRCAS, Tsukuba and IPGRI, Rome. Engelmann, F, P Batugal and JT Oliver (eds). 2002. Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Selangor, Malaysia. Hornung, R, R Domas and PT Lynch. 2001. Cryopreservation of plumular explants of coconut (Cocos nucifera L.) to support programmes for mass clonal propagation through somatic embryogenesis. CryoLetters 22:211-220. Malaurie, B and M Borges. 2001. Cryopreservation of coconut (Cocos nucifera L.) plumules by encapsulation/dehydration. Pp. 59. In: Abstracts “Bioveg 2001”. International Workshop on Plant Biotechnology – Plant Breeding and Biotechnology. Centro de Bioplantas, Ciego de Avila, Cuba, April 16-20, 2001. Malaurie, B, M. Borges and O. N’Nan. 2002a. Research of an optimal cryopreservation process using encapsulation-osmoprotection- dehydration and encapsulation-osmoprotection-vitrification techni- ques on caulinary meristems of coconut (Cocos nucifera L.). Abstracts IV Jornada Científica IIA”Jorge Dimitrov”, Bayamo, Cuba, 19-21 Sept. 2002. Malaurie, B, O N’Nan, V Hocher, P Ilbert, F Grosdemange, JL Konan, N Zakra and JL Verdeil. 2002b. State of research on culture and cryopreservation of zygotic coconut embryos at IRD/CIRAD (France). Pp. 146-156. In: F Engelmann, PA Batugal and JT Oliver (eds). Coconut embryo in vitro culture: Part II. IPGRI-APO, Serdang, Se- langor, Malaysia. Maxted, N, BV Ford-Lloyd and JG Hawkes. 1997. Complementary con- servation strategies. Pp. 15-39. In: N Maxted, BV Ford-Lloyd and JG 148 COCONUT GENETIC RESOURCES Hawkes (eds). Plant genetic resources conservation. Chapman and Hall, London. N’Nan, O. 1997. Recherche d’une méthode de déshydratation simple, favorable à la survie et à la régénération des embryons zygotiques matures cryoconservés de cocotier (Cocos nucifera L.). Diplôme d’Etu- des Approfondies, Université Abidjan-Cocody. N’nan, O, JL Verdeil, V Hocher, JL Konan, N Zakra and B Malaurie. 2002. Mise au point d’une méthode de cryoconservation d’apex caulinaires de cocotier (Cocos nucifera L.). Pp. 152-153. In: I El Hadrami (ed). Biotechnologies Végétales: Valorisations pour une Agriculture Durable, VIIIèmes Journées Scientifiques du réseau “Biotechnologies, Amélioration des Plantes et Sécurité Alimentaire” de l’Agence Uni- versitaire de la Francophonie, 7-9 Octobre 2002. Marrakech, AUF, Paris. N’Nan, O, M Borges, JL Verdeil and B Malaurie. 2003. Is cryopreservation of mature zygotic embryos of coconut (Cocos nucifera L.) the easiest and safest way to preserve coconut germplasm? Paper presented during the 4th International Workshop on Plant Biotechnology and Sustainable Development “BioVeg 2003”, 14-19 April 2003, Centro de Bioplantas, Universidad de Ciego de Avila, Ciego de Avila, Cuba. Ramanatha Rao, V and P Batugal (eds). 1998. Proceedings of the COGENT Regional Coconut Genebank Planning Workshop, Pekanbaru, Riau, Indonesia, 26-29 February 1996. IPGRI-APO, Serdang, Selangor, Malaysia. Sakai, A, T Matsumoto, D Hirai and T Niino. 2000. Newly developed encapsulation-dehydration protocol for plant cryopreservation. CryoLetters 21:53-62. Withers, LA and JMM Engels. 1990. The test tube genebank: A safe alternative to field conservation. IBPGR Newsletter for Asia and the Pacific 3:1-2. 149 CHAPTER 3: Germplasm conservation In situ conservation of coconut diversity B Sthapit1, V Ramanatha Rao1 and D Jarvis2 1Senior Scientists, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia 2Senior Scientist, International Plant Genetic Resources Institute (IPGRI), Maccarese, Rome, Italy Introduction Coconut genetic diversity present in farming systems has been maintained through the combined action of natural and human selection and management. Food culture of specific communities also affects selection of preferred culinary traits. In the process of planting, managing, selecting seednuts, harvesting and marketing- farmers, in turn, make decisions on their crops that affect the genetic diversity of the crop populations. Over time, a farmer may alter the genetic structure of a crop population by selecting for plants with preferred agro-morphological or quality characteristics (Jarvis and Hodgkin 2000). Thus, coconut landraces may be a product of farmer selection as well as farmer breeding (Riley 1996). Coconut varieties are grown by resource poor farmers around the world for a diversity of uses. Home consumption, local markets, for industrial processing, and medicinal use are only a few. Farmers search for locally adapted coconut cultivars for diverse environmental niches. Many varieties are adapted to particular micro-niches including climatic and edaphic stressed environments. Farmer preferences for specific size, aroma, nut water quality, colour, taste and type also demand diverse coconut varieties. The continued use of landraces contributes to stable food production and income especially in marginal environments where impacts of modern varieties are limited or less effective. The Convention on Biological Diversity (CBD) has recognized the continued maintenance of traditional varieties in situ as an essential component of sustainable agricultural development. Diversity of local coconut varieties is the foundation upon which coconut breeding depends for the creation of new varieties and is therefore, a critical aspect of food security for coconut- based economies. To develop successful conservation approach, knowledge of plant biology is essential. Coconut (Cocos nucifera L.) is essentially a tree crop of the humid tropics. It is able to adapt to a wide range of soil and climatic conditions. The natural habitat of coconut is the coastal belt of the tropics where it flourishes in sea-washed littoral sand with constant motion of underground current of water in a saline atmosphere (Khan et al. 1994). 150 COCONUT GENETIC RESOURCES It is monoecious with numerous male and female flowers in each spadix. Tall and Dwarf are distinct natural population of the coconut gene pool; Tall coconut trees are usually cross-pollinated and consequently, are usually heterogeneous (Nair and Ratnambal 1994; Iyer and Dhamodran 1994; Batugal and Ramanatha Rao 1998). The inflorescence, 1-2m long, consists of a central rachis, with up to 40 lateral branches, which bear 200-300 male flowers above, opening from the tip of branches downwards, and one or more female flowers at the base which are receptive after the pollen has been shed. Flowering starts at 6-12 years of age for Tall types (Purseglove 1975) and 3-4 years for Dwarf types (Nair and Ratnambal 1994). The male flowers are the first to open, beginning at the top of each branch and proceeding towards the base. After the pollen has been shed in the bud, the female flowers open and remain receptive for 24 hours. The flowers are nectiferous and sweet-scented and visited by a range of pollinators, particularly bees, flies and ants, so that a fair amount of insect pollination is possible. The pollen of coconut is dry and therefore, some wind pollination may also occur. Coconut must be propagated by seednuts and cannot be vegetatively propagated. The first European explorers in Asia and the Pacific found coconuts well established in almost all tropical coastal areas and it is believed that ocean currents carry coconuts, and they become established on open coasts without the aid of human beings. Edmonson (1941) reported that coconuts are capable of germinating after floating in the sea for periods of up to 110 days, during which time they could have travelled 3000 miles in favourable currents. Furthermore, coconut can tolerate saline conditions because of their root structure and is also adapted to high humidity and constant supply of water. What is in situ conservation? In situ conservation is defined by the CBD (Article 2) as “…the conservation of ecosystems and natural habitats and the maintenance and recovery of viable populations of species in their natural surroundings and, in the case of domesticates or cultivated species, in the surroundings where they have developed their distinctive properties” (UNCED 1992). The definition of in situ conservation used in the CBD encompasses two processes: the conservation of wild species in natural ecosystems or reserves, and the maintenance of crops genetic diversity on-farm. On- farm conservation is generally used to describe the processes by which farmers maintain their traditional crop varieties that they have developed and which they continue to manage and improve. These processes have led to the evolution and adaptation of crops to changing environments and socio economic conditions. For coconut genetic diversity, the 151 CHAPTER 3: Germplasm conservation maintenance of local coconut cultivars within the traditional farming systems is still little studied and understood. Why in situ conservation? For the last decades, agricultural scientists have responded to the threat of genetic erosion by developing a worldwide network of genebanks, field genebanks and botanical gardens for conserving the available useful genetic resources ex situ (Bommer 1991). Since the establishment of the International Coconut Genetic Resources Network (COGENT) by IPGRI in 1992, a total of 14 coconut field genebanks have been established (IPGRI 2001). COGENT (2000) coordinated the establishment of a multi-site International Coconut Genebank (ICG), an international Coconut Genetic Resources Database (CGRD) and the studies of diversity in coconut traits including drought tolerance, suitability for high-value products and compatibility for intercropping. COGENT has 38 member countries and these countries have agreed to share and exchange germplasm by putting selected national accessions in the multi-site ICGs for Southeast Asia (located in Sikijang, Indonesia), South Asia (Karnataka, India), the South Pacific (Madang, PNG) and Africa (Abidjan, Côte d’lvoire). In India, coconut-breeding programme has been utilizing farmer’s varieties to develop hybrids from Dwarf x Tall genotype (Ratnambal and Nair 1998). Understanding of coconut genetic resources and its value in traditional farming systems of Sri Lanka, Indonesia, Vietnam, Philippines, Thailand, Fiji, Vanuatu, Côte d’Ivoire, Benin, Ghana, Tanzania, Mexico, Nigeria and Jamaica has improved significantly over time and mother palm selection of preferred coconut genetic resources is a common feature (Batugal and Ramanatha Rao 1998). While this form of conservation remains no doubt an important method, it does not conserve the evolutionary process of local adaptation of crops to their environments. In situ conservation has the potential to: (1) conserve the evolutionary processes of local adaptation of crops to their environments, (2) conserve diversity at all levels – the ecosystem, the species, and the genetic diversity within species, (3) conserve ecosystem services critical to the functioning of the earth’s life-support system, (4) improve the livelihoods for resource-poor farmers through economic and social development, (5) maintain or increase farmers’ control over and access to crop genetic resources, (6) ensure farmers’ efforts are an integral part of national genetic resources systems and involve farmers directly in developing options for adding benefits of local crop diversity, and (7) link farming community to field genebank for conservation and utilization (Jarvis et al. 2000). 152 COCONUT GENETIC RESOURCES Basic information needed to implement in situ conservation programme To implement a coconut in situ conservation programme, it is necessary to understand where, when and how in situ conservation of the coconut crop will be effective, who will maintain the material, and how the maintained material will benefit the stakeholders. Four types of information are needed to design an effective programme: • The extent and distribution of the coconut genetic diversity maintained by farmers over space and over time; • The processes used to maintain the coconut genetic diversity on farm; • The persons who maintains coconut genetic diversity (custodians of genetic diversity) within the farming communities; and • The factors (market, non-market, social, environmental) that in- fluence farmer decisions on maintaining traditional coconut varieties. Information on these topics is needed to develop methods for mainstreaming the use of local crop genetic resources into the agricultural development arena. Building and implementing on-farm conservation programme On-farm conservation involves partnerships among individuals and institutions. A project dominated by conservationist may fail to emphasize farmers’ livelihoods, while a project dominated by development workers may fail to emphasize conservation. A project without ecologists may neglect the importance of ecosystem services that the crop might be providing (Jarvis et al. 2000). Once understanding among institutions, collaborators and farming communities has been reached, existing data, such as descriptor lists, databases of ex situ germplasm collections, herbarium collections, published literature in the natural and social sciences should be reviewed, together with unpublished information, including the personal knowledge of local extension and NGOs. Site selection criteria and farmer selection criteria should then be defined followed by the training of local research teams in participatory methods for information collection from the local communities. After site selection, communities need to be sensitized to the aims and objectives of on-farm conservation programme; and sampling scheme for data collection should be formulated. Broadly speaking, the criteria for site selection should be based on the extent of genetic diversity, accessibility and interest of the farmers to continue to grow coconut varieties. Jarvis and colleagues (2000) spelt this out below in more detail. 153 CHAPTER 3: Germplasm conservation Ecosystems. It will be important to select sites in diverse agroecosystems preferably with different ecotypes. Traditional coastal home gardens are important ecosystem for on-farm management of farmer-preferred coconut diversity in situ. This will increase the chances of conserving genetic diversity, as this may be associated with agroecosystem diversity. Intra-specific diversity within target species. It is important that the areas selected are grown to different coconut landraces. Specific adaptations. Efforts should be made while selecting different agroecosystems to select sites with extreme environmental conditions (high soil salinity, cold temperatures, etc) and variation in pests. This will help to include types with specific adaptations. Genetic erosion. It is better to select sites with less threat of genetic erosion to increase the life of conservation efforts. Diverse use values. It is possible to ensure conservation of hidden genetic diversity by selecting sites with diverse use values of crops for food and other uses. It is important to note that for many farming communities, a crop is not just a matter of food production but also of investing and maintaining social relations and religious rituals. Farmers and communities. Farmers’ interest and willingness to participate are keys in site selection. This may require preliminary work in community sensitisation on the benefits to farmers of conserving crop varieties. Site selection should also include areas with: socio-cultural and economic diversity; diversity of livelihoods, and importance of target crops for various ways of life; farmers’ knowledge and skills in seed selection and exchange; and market opportunities. Partners. Partners with interest in community empowerment, capacity building and development agenda, and experience in conservation interventions will be beneficial to the programme. Partners with distinct community participation expertise will have comparative advantage in dealing with community. The concept of commodity chain, which allows farmers to use multiple parts of coconut trees, increases the chance of conserving in situ. This concept of value addition requires the involvement of a full range of partners from different disciplines, who are not usually involved in agricultural biodiversity research. Logistics. These would include mainly the accessibility of the site throughout the year and availability of resources. The former is very 154 COCONUT GENETIC RESOURCES important for a successful in situ conservation programme and is essential in monitoring and sharing of information back to the community. The existing data should be combined with an exploratory survey using Rapid Rural Appraisal (RRA) and Participatory Rural Appraisal (PRA) approaches. The community needs to be sensitized to issues on hand and for this purpose, the use of participatory approach is recommended (Friis-Hansen and Sthapit 2000). The following broad steps are essential for the effective implementation of on-farm conservation programme (Sthapit et al. 2000) and may need refinement according to local context: • Locating coconut diversity, ecosystem and community (e.g. ecogeographic survey); • Creating (or using existing) institutional framework and participatory planning process; • Site selection (low cost in situ conservation site having both high public as well as farmers’ utility value of crop genetic resources); • Community sensitization and strengthening local capacity; • Locating coconut diversity and custodians of unique and rare types (e.g. diversity fair); • Measuring and assessing local coconut diversity in terms of richness, evenness and uniqueness (e.g. consistency on farmer’s unit of diversity, molecular markers, genetic indices); • Understanding the perceived value of coconut diversity (e.g. four square method); • Monitoring diversity (e.g. community biodiversity register) and sharing information; • Developing strategy for options of on-farm conservation actions; • Diversity utilization and monitoring of intervention impacts; and • Mainstreaming information for development and policy reforms. A number of participatory tools are developed to implement on-farm conservation activities at local level by the farming community themselves, namely: • Local knowledge base: Understanding of local crop diversity and social networks of germplasm and knowledge flow and storage methods; identify technical gaps and strengthen local seed system; • Diversity fair: Local community can organize this fair for locating diversity and custodians, sensitizing community and policy makers and promoting access of information and materials; and • Community biodiversity register (CBR): Recording inventory of lo- cal crop diversity and associated local knowledge, and monito- ring the increase and decrease of number of landraces and mo- 155 CHAPTER 3: Germplasm conservation dern varieties and their distribution pattern within households (by area) or between households within community. The above activities will raise awareness on local crop diversity and help to understand the value of local crop diversity. Diversity fair and CBR are a few participatory methods, which can strengthen the local capacity to document taxonomic data and traditional knowledge on crop genetic resources (CGR) with the following specific objectives of: • Creating awareness and developing sense of community ownership on biodiversity; • Locating unique, rare and culturally significant cultivars and their custodians; • Enhancing access of genetic materials and information on local crop diversity; • Developing options of adding benefits and support biodiversity- based livelihoods; • Building local capacity for monitoring diversity in situ and promoting on-farm management of local crop diversity; • Making aware on and protecting economically important biowealth against biopiracy. The successful implementation of CBR will depend upon how the approach could provide direct benefits to farming community. One of the direct benefits is that it may help to establish a network of key households, which maintain rare, unique and rich local crop diversity resulting into a network of planting material to form a decentralized community seed bank. It is important to focus not only in scientific understanding of the project but also to develop institutional capacity to run internally driven on-farm conservation programmes. The value of such decentralized CBR will be clearer when activities such as diversity kits, Participatory Variety Selection (PVS) and Participatory Plant Breeding (PPB) (Sthapit et al. 1996; Sthapit et al. 2000; Witcombe et al. 1996) are integrated into community-based informal seed management and exchange programme. The PPB and the deployment of diversity kits will strengthen the capacity of the farmers to search, select, maintain and exchange genetic resources for obtaining both genetic and socioeconomic benefits for themselves and for the society. In situ conservation and its benefits to the community The effective management and conservation of genetic resources on farm takes place where the resources are valued and used to meet the needs of 156 COCONUT GENETIC RESOURCES local communities (Jarvis et al. 2000). In order for local coconut farming systems to be maintained by farmers, the genetic resources must have some value and/or be competitive to other options a farmer might have. Understanding the contribution of coconut cultivars to livelihoods, nutrition and food culture is needed to formulate plans that will: (i) support local germplasm supply systems, (ii) improve PVS and PPB, (iii) develop new markets for coconut cultivars and plant parts, (iv) promote appropriate conservation value and education, (v) create methodologies for integrating locally adapted coconut cultivars and farmer preferences into development and extension projects, and (vi) advise on appropriate policies that support the management and use of crop diversity in agroecosystems (IPGRI 2001). Two options were used in adding benefits: the first, on adding benefits through participatory variety selection and plant breeding, seednuts networks and grassroots strengthening; and the second, on adding benefits through public awareness, better processing, marketing, policy incentives, and education in the formal sector (Jarvis et al. 1998). The first option is to seek improved quality, disease resistance, high yield, better taste, ease in harvesting and other preferred traits through breeding; seed networks and modified farming systems. In modern agricultural production systems, Smale and her colleagues (2001) argued that crop genetic resistance to disease can be enhanced by policies that encourage: (1) cultivation of a mosaic of varieties with different genetic mechanisms for combating a pest, (2) cultivation of specific varieties that contain multiple genetic mechanisms for resistance, or (3) continual replacement of varieties in farmers’ fields by more recent releases or exchange of farmers’ cultivars that carry new genetic sources of resistance. The second option includes adding value to coconut resources so that the demand for the material or some derived product may be increased. These diverse options will emerge when the community, researchers and developmental institutions are directly involved in monitoring local crop diversity using CBR and link with crop improvement, seed and market networks for adding benefits on local resources. If diversity can be more highly valued in the marketplace through the creation of consumer demand for certain products, and farmers can access those markets, their incentives to maintain diversity may be increased. The concept of commodity chain applies with coconut as the approach is not restricted to increase in productivity alone but rather that crop is considered ‘as a whole’ in all aspects of a chain (or a system), from its production through its consumption. This concept of value addition adds new dimensions to the traditional agricultural research agenda and it implies the involvement of a full range of new partners, who are not 157 CHAPTER 3: Germplasm conservation usually involved in agricultural biodiversity research. Measures to conserve coconut diversity in situ include: • Creation of economic incentives mechanisms (such as identifica- tion of new products and markets, increase competitiveness of local cultivars, introduction of supportive policies) and other measures to promote cultivation of diverse local coconut culti- vars. IPGRI (2000) has documented at least 12 marketable high- value products and market locations for coconut. They include: tender nuts, palm sugar, desiccated coconut, milk/cream, milk powder, fresh coconut, makapuno coconut dessert, coconut water, nata de coco, coir fibre, fibre dust, shell charcoal and activated carbon. Hence, the goal of in situ conservation is to encourage farmers to select and maintain local crop diversity to benefit themselves as well as the community at large. Understanding of local food culture reveals the need for a range of coconut culti- vars in home gardens. • Increase profile of non-monetary benefits, which include increased access to information and technologies arising from the use of exchanged information, enhanced research and development capacity of local institutions, low food cost and materials, public recognition, an improved quality of life through access of natural chemical free foods, public awareness, environmental benefits such as the protection of habitats and ecosystems (Raymond and Flower 2001). • Strengthening local capacity to document, manage and share in- formation of local diversity for the benefits of the community and individual so that the community has the capacity to develop options for on-farm conservation actions. All this requires greater collaboration between formal and informal sectors with more benefit-oriented activities. Promising results are emerging from all countries and many methods and approaches have been developed which are compiled now to publish guidelines for on-farm conservation of agrobiodiversity (Jarvis et al. 2000). These outputs must be evaluated and monitored in terms of effectiveness and sustainability of coconut genetic resources conservation and utilization. References Altieri, MA and LC Merrick. 1988. In situ conservation of crop genetic resources through maintenance of traditional farming systems. Economic Botany, 41:86-96. Batugal, PA and V Ramanatha Rao (eds). 1998. Coconut breeding. Papers 158 COCONUT GENETIC RESOURCES presented at a workshop on Standardization of Coconut Breeding Research Techniques, 20-25 June 1994, IPGRI-COGENT, Malaysia. Bommer, DFR. 1991. The historical development of international colla- boration in plant genetic resources. In: Th JL van Hintum, L Frese and PM Perret Wageningen (eds). Searching for new concepts for collaborative genetic resources management: Papers of the EUCARPIA/IBPGR Symposium. The Netherlands, 3-6 Dec 1990, International Crop Networks Series No. 4, and IPGRI, Rome. Brush, SB. 1995. In situ conservation of landraces in Centres of crop diversity. Crop Science. 35: 346-354. Brush, SB (ed). 2000. Genes in the field. On-farm conservation of crop diversity. IPGRI/IDRC/Lewis Publishers. Edmonson, CH. 1941. Variability of coconut seed after floating in the sea. Occas. Papers Bernice P. Bishop Museum, Hawaii 16: 293-304. Friis-Hansen, E and B Sthapit (eds). 2000. Participatory approaches to the conservation and use of plant genetic resources. IPGRI/CDR, Rome, Italy. Harlan, JR. 1992. Crops and man. American Society of Agronomy and Crop Science Society of America, Madison, WI. IPGRI. 2001. Regional report APO 1999-2000. IPGRI-APO, Serdang, Se- langor, Malaysia IPGRI. 2001. On-farm management of crop genetic diversity and the Convention on Biological Diversity’s Programme of work on agricultural biodiversity. A synthesis paper prepared by the IPGRI: Overview of crop genetic resources in agrobiodiversity. CBD operational objectives, and principals and best practices, 8-10 November 2001, Montreal, Canada. Iyer, RD and S Dhamodran. 1994. Improvement of coconut. Pp. 217- 242. In: KL Chadha and P Rethinam (eds). Advances in Horticul- ture, Vol 9: Plantation and Spice Crops Part 1. Malhotra Publishing House, India. Jarvis, D and T Hodgkin, 2000. Farmer decision-making and genetic diversity: Linking multi-disciplinary research to implementation on farm. In: S Bush (ed). Genes in the field: Issues in conserving crop diversity on farm. IDRC/IPGRI/Lewis Publishers, Washington DC. Pp 261-278. Jarvis, D, B Sthapit and L Sears (eds). 2000. Conserving agricultural biodiversity in situ: A scientific basis for sustainable Agriculture. Proceedings of a workshop, Pokhara, Nepal. IPGRI Jarvis, DI, L Myer, H Klemick, L Guarino, M Smale, AHD Brown, M Sadiki, B Sthapit and T Hodgkin. 2000. A training guide for in situ conservation on-farm. IPGRI, Rome, Italy. 161pp. 159 CHAPTER 3: Germplasm conservation Jarvis, D, T Hodgkin, P Eyzaguirre, G Ayad, B Sthapit and L Guarino. 1998. Farmer selection, natural selection and crop genetic diversity: The need for a basic data set. Pp. 5-19. In: D Jarvis and T Hodgkin (eds). Strengthening the scientific basis of in situ conservation of agricultural biodiversity on farm: Options for data collecting and analysis. Proceedings of a workshop to develop tools and procedures for in situ conservation on farm, 25-29 August 1997, Rome Italy. IPGRI, Rome, Italy. Khan, HH, CC Biddappa and SR Cecil. 1994. Nutrition of coconut. Pp. 375-394. In: KL Chadha and P Rethinam (eds). Advances in horti- culture. Vol 9. Plantation and spice crops Part 1. Malhotra Publishing House, India. Purseglove, JW. 1975. Coconut. Pp. 440-478. In: Tropical crops monocotyledons, Longman, UK. Maxted, N, BV Ford-Lloyd and JG Hawkes. 1997a. Complementary con- servation strategies. In: N Maxted, BV Ford-Lloyd and JG Hawkes (eds). Plant genetic conservation: The in situ approach. Chapman and Hall, London. Maxted, N, JG Hawkes, BV Ford-Lloyd and JT Williams. 1997b. A practical model for in situ genetic conservation. Pp. 339-367. In: N Maxted, BV Ford-Lloyd and JG Hawkes (eds). Plant genetic conser- vation: The in situ approach. Chapman and Hall, London. Pp. 339- 367. Nair, MK and MJ Ratnambal. 1994. Genetic resources of coconut. In: KL Chadha and P Rethinam (eds). Advances in horticulture. Vol 9. Plan- tation and spice crops Part 1. Malhotra Publishing House, India. Pp. 50-63. Ramanatha Rao, V and BR Sthapit. 2001. In situ conservation: A component of complementary conservation strategy. Pp 123-135. In: V Ramanatha Rao and D Campilan (eds). Proceedings of the Asian Network for Sweet Potato Genetic Resources (ANSWER) Workshop on exploring the potential for in situ (on-farm) conservation of sweet potato genetic resources. Bali, Indonesia, 2-4 October 2001. IPGRI- APO, Serdang, Malaysia. Ratnambal, MJ and MK Nair. 1998. National coconut breeding pro- gramme in India. Pp. 1-14. In: PA Batugal and V Ramanatha Rao (eds). Coconut breeding. Papers presented at a workshop on Standardization of Coconut Breeding Research Techniques, 20-25 June 1994. IPGRI-COGENT, Serdang, Selangor, Malaysia. Raymond, R and C Flower. 2001. Sharing the non-monetary benefits of agricultural biodiversity. Issues in genetic resources No. 5, Sept 2001. IPGRI, Rome, Italy. 160 COCONUT GENETIC RESOURCES Riley, KW. 1996. Decentralised breeding and selection: tool to link diversity and development. Pp. 140-157. In: L Sperling and M Loevinsohn (eds). Using diversity: Enhancing and maintaining genetic resources on-farm. International Development Research Centre IDRC, New Delhi, India. Smale, M, M Bellon, D Jarvis and B Sthapit. 2001. Economic concepts for designing policies to conserve crop genetic resources on farms. IPGRI ISSUES Paper, 17 May 2001, IPGRI, Rome, Italy. Sthapit, BR and D Jarvis. 1999. Participatory plant breeding for on-farm conservation. LEISA 15:40-41. Sthapit, BR, P Sajise and D Jarvis. 1999. Strengthening scientific basis of in situ conservation on-farm: Learning experiences from Nepal and Vietnam. Pp. 338-361. In: Xu Jianchu (ed). Links between cultures and biodiversity. Proceedings of the Cultures and Biodiversity Congress 2000, 20-30 July 2000, Yunnan, China. Sthapit, BR, A Subedi, D Rijal, R Rana and D Jarvis. 2002. Strengthening community-based on-farm conservation of agricultural biodiversity. In: Conservation and sustainable use of agricultural biodiversity: A sourcebook. UPWARD, Los Baños, Laguna, Philippines. Witcombe, JR, A Joshi, KD Joshi and BR Sthapit. 1996. Farmer participatory crops improvement I. Varietal selection and breeding methods and their impact on biodiversity. Experimental Agriculture 32:445-460. 161 CHAPTER 3: Germplasm conservation Poverty reduction in coconut growing communities: A strategy for coconut in situ / on-farm conservation P Batugal1, J Oliver2 and K Jayashree3 1Coordinator, 2Communications Assistant and 3Scientific Assistant, International Coconut Genetic Resources Network (COGENT), International Plant Genetic Re- sources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction About 96% of coconuts are grown by smallholders tending four hectares or less of land which many of them do not own. About 85% of the 12 million hectares of coconuts are grown in the Asia Pacific region. Coconut farmers are marginalized: they grow coconut and associated crops in rainfed and often environmentally fragile areas; most live below the poverty line; are resource-poor; considered non-bankable by the credit sector; and they do not have political clout to influence public and private sector policy. Even in many of the large coconut producing countries, research support to this sector is inadequate if not nil. IPGRI believes that if resource-poor farmers are empowered, they could improve their lives and lift their socioeconomic status over the poverty line. To address the urgent need of empowering poor coconut farmers and helping the long neglected coconut sector, IPGRI requested - and the Asian Development Bank (ADB) awarded - a Regional Technical Assistance (RETA) grant to IPGRI (RETA 6005 for 2000-2004) entitled, ‘Developing sustainable coconut-based income-generating technologies in poor rural communities’. IPGRI coordinated the project involving eight national coconut research agencies, three non-governmental organizations (NGOs) and 25 community-based organizations (CBOs) in eight Asia Pacific countries (Bangladesh, India, Sri Lanka, Indonesia, the Philippines, Vietnam, Fiji and Papua New Guinea) as shown in Annex 1. Objective The project objective is to develop efficient village-level, income-generating technologies and strategies that are technically feasible, financially viable, socially acceptable and environmentally safe, using COGENT’s three- pronged strategy: 1) production and marketing of high-value coconut products from all parts of the coconut – the kernel, husk, shell, wood, water and leaves); 2) intercropping cash and food security crops with 162 COCONUT GENETIC RESOURCES coconut and integrating livestock/fodder production; and 3) establishing community-managed nurseries to propagate and sell quality planting materials of farmer-selected local and introduced high-value varieties and conserve them on farm. The project also identified, enhanced and provided access to the five essential capitals (physical, natural, financial, social and human) needed to convert these income generating interventions into sustainable livelihoods. For details about the project framework, see COGENT publication entitled, ‘Poverty Reduction in Coconut Growing Communities, Volume I: The Framework and Project Plan’. Activities Prior to the release of project funds to IPGRI in May 2002, the UK’s Department for International Development (DFID) supported the identification and socioeconomic profiling of 89 coconut growing communities from which 24 communities were chosen as project sites. IPGRI and its partner organizations also organized the project team and technical support groups in each country using IPGRI’s and national counterpart funds. Upon release of the ADB funds, the following activities were conducted: 1. Establishment and strengthening of 25 CBOs to manage the project at the community level. Special emphasis was made on the design of the CBOs to ensure broad access and participation of several categories of stakeholders, including women; 2. Establishment of a microcredit system and provision of initial revolving fund for each of the 25 CBOs; 3. Market surveys to identify marketable products and development of market channels to make such markets sustainable; 4. Development and implementation of farmers’ and women’s action plans for income-generating activities; 5. Development of training manuals on income-generating technologies and the development of instruments for analysis and promotion of viable technologies; 6. Development of community-managed income-generating coconut seedling nurseries and the documentation, enhancement and conservation of selected and promising local and introduced coconut varieties; 7. Training of coconut farmers, women and village-level entrepreneurs on income-generating technologies; 8. Evaluation of inexpensive village-level oil mills and equipment for producing high-value coconut products; 9. Development and viability testing of the production and marketing 163 CHAPTER 3: Germplasm conservation of identified marketable high-value coconut products from the kernel, husk, shell, water, wood and leaves; and promotion of varieties suitable for such high-value coconut products; 10. Pilot production and marketing of high-value products from the coconut’s kernel, husk, shell, wood, water and leaves; 11. Development and viability testing of: (a) coconut-based intercropping technologies for enhancing incomes and food security; and (b) livestock and fodder production to boost total farm productivity and nutrition; and 12. Promoting the use of research results through field days and the replication and adoption of resulting viable development interventions by national governments, development organizations and NGOs. The project inception and stakeholders’ meeting was held on 25 February – 1 March 2002 in Ho Chi Minh City, Vietnam, hosted by the Oil Plants Institute (OPI); the second project meeting on 20 – 24 August 2003 in Davao City, Philippines, hosted by the Philippine Coconut Authority; and the final project meeting on 27–30 September 2004 in Ho Chi Minh City, Vietnam, and hosted by OPI once more. For details of the project activities and target outputs, please refer to Annex 2 and the COGENT publication entitled, ‘Poverty Reduction in Coconut Growing Communities, Volume II: Mobilizing for Action’. Project outputs The results of the project proved that poor coconut farmers’ and socioeconomically disadvantaged women’s lives could be improved if they were properly trained, empowered and given access to opportunities and resources, in this case the coconut-based village-level income generating activities, technologies and related support systems. In a period of only three years, 25 farmer CBOs in eight countries with a total of 5715 members were established and strengthened; 17 392 farmers and women trained on various income-generating activities; 43 community- managed coconut and 14 multi-purpose seedling nurseries established; 65 501 coconut seednuts of local varieties sown in community-managed nurseries; 64 521 coconut seedlings of farmers’ and introduced high-value varieties planted and conserved on-farm; 1593 farmers and women involved in coconut-based livestock production trials, 4039 in intercropping trials and 2005 in production of high-value coconut products; and about 140 public awareness materials developed and disseminated. For details of these achievements, please see Annex 3. In the production of high-value coconut products, more than 2000 164 COCONUT GENETIC RESOURCES CBO members, 74% of whom are women, participated. There were 210 participants in Bangladesh, 17 in Fiji, 615 in India, 100 in Indonesia, 89 in Papua New Guinea, 378 in the Philippines, 168 in Sri Lanka and 428 in Vietnam. CBO members, trained and working as individuals or in groups, produced cooking oil, virgin coconut oil for body and hair lotion, kernel-based detergent and bath soaps, fibre-based ropes, doormats and geotextile, shell- and wood-based cooking utensils and exportable handicrafts, coconut water- and sap-based vinegar and sugar, and coconut leaf-based decorative baskets, hats and other handicrafts. Depending on how many capable members of the participating families were involved, they increased their income by 3-5 times compared to their previous income from copra, securing for them a steady source of additional income and helping them rise above the poverty line. Equally important, this project intervention has provided employment opportunities to formerly unemployed and underemployed rural women resulting in enhanced self-esteem, and economic and social empowerment. For intercropping, 4039 farmers and their households participated in intercropping trials consisting of 115 in Bangladesh, 454 in Fiji, 759 in India, 748 in Indonesia, 418 in Papua New Guinea, 473 in the Philippines, 328 in Sri Lanka, and 744 in Vietnam. Net incomes from planting cash crops in small plots of land between coconuts have significantly increased. Not only were income and total farm productivity enhanced, but also food security and nutrition since families planted, grew and ate their own produce. For animal production, a total of 1593 CBO members, 58% of whom are women, raised a variety of livestock like quails, poultry, ducks, rabbits, goats, swine and cattle. There were 185 participants in Bangladesh, 32 in Fiji, 370 in India, 82 in Indonesia, 126 in Papua New Guinea, 334 in the Philippines, 197 in Sri Lanka and 267 in Vietnam. The integration of livestock production in coconut farming is still in its early stages, but many CBO members have already adopted the animal production technologies introduced by the project as components of their sustainable livelihood activities. The initial results showed tremendous potential not only in generating income but more so in improving nutrition. To support the conservation and promotion of coconut diversity, 43 community-managed coconut and 14 multi-purpose seedling nurseries were established; 65 501 coconut seednuts of local varieties sown in these nurseries; and 64 521 coconut seedlings of farmers’ and introduced high- value varieties planted and conserved on farm. The 24 communities (excluding the Maitum site in the Philippines) also identified and characterized 89 important local varieties through farmers’ diversity fairs. 165 CHAPTER 3: Germplasm conservation Through this participatory intervention, the farmers themselves characterized and identified suitable, high-yielding and high-value local varieties. The source palms of the selected varieties were paint-marked and the seednuts harvested from these palms were propagated in the nurseries. These community-managed nurseries are envisioned to provide a steady supply of high-quality planting materials for the communities. Project benefits To determine the benefits and the initial impact of the project, a two- stage assessment was carried out: (1) rapid assessment survey involving project leaders and heads of implementing research agencies, NGOs and CBOs; and (2) more detailed survey involving farmers and other members of the participating communities in the project. Based on these surveys, the following project benefits were identified: 1. The project provided an effective IARC-NARS-CBO mechanism for promoting income generating activities in previously resource- poor coconut growing communities in their countries; and in providing the five needed capitals for sustainable livelihoods (i.e., physical, natural, financial, social and human capitals) to make these income generating activities sustainable. Most of these technologies and resources were not available to the 25 project participating communities in eight countries before the project; 2. The project provided farmers access to efficient but affordable village-level coconut processing equipment, machinery and technologies for producing high-value coconut products which were sourced from several COGENT member countries. In some communities, the local government provided the needed infrastructure and other facilities such as roads, training centres and electrical power connections to support the project; 3. The project enriched the communities’ natural capital in the form of important local coconut varieties which farmers identified and characterized with the help of researchers and breeders and propagated them in community-managed nurseries. The project also facilitated the introduction of high-value coconut varieties in the community thereby enhancing the diversity of their coconut germplasm; 4. The project enhanced the communities’ social capital by organizing the farmers into CBOs and strengthening and enabling these organizations to effectively plan, manage and implement income generating activities for its constituents; 5. The project provided the needed financial capital in the form of 166 COCONUT GENETIC RESOURCES collateral-free revolving funds for the 25 CBOs to establish their own microcredit systems, enabling poor farmers and women without land or assets to engage in income generating activities; and 6. The project enhanced the human capital of the communities, empowering them through training to effectively and efficiently carry out coconut-based sustainable livelihood activities. The project trained over 17 000 community members on establishing and managing CBOs and microcredit system, producing high- value coconut products, intercropping cash and food security intercrops, raising livestock in a coconut-based farming system and producing feed/ fodder, and establishing and managing seedling nurseries. Project impact The same surveys indicated how the benefits generated by the project affected the lives of coconut farmers, particularly their socioeconomic status. The identified initial impacts of the project include: Impact on farm households At the farm household level, income generating skills have been enhanced, providing capable members of the family the opportunity to earn in each of the four stages of the commodity chain - production, processing, marketing and consumption– thereby increasing farm incomes by three to five-fold compared to pre-project earnings. And because of additional incomes and savings, more families were able to send their children to school. Intercropping cooking banana, cassava, sweet potato, taro, yams, maize, etc has enhanced food security, while raising vegetables and livestock (native chicken for meat and eggs, goat and cattle for meat and milk, ducks, etc) has improved nutrition. Engaging in collective work has also promoted family cohesiveness. Impact on communities At the community level, the impact on women has been particularly tremendous. The project enabled previously destitute and unemployed or underemployed women to earn money, shed inhibitions and empowered them to make informed decisions affecting their lives and, in the process, raising their self-esteem. Unemployment rate in the communities was significantly reduced as formerly idle labour was put into productive use in various coconut-based income generating activities. The project also encouraged community members to work in groups as agricultural entrepreneurs, developing their business and group problem solving skills. 167 CHAPTER 3: Germplasm conservation The project also increased farmers’ capabilities in optimally and profitably managing their coconut farms, with many adopting the modern, integrated coconut farming systems technique introduced by the project and discarding their traditional methods which mainly revolved around coconut monoculture. Due to the actual socioeconomic benefits experienced by project participants in a period of three years, the project has enhanced the attractiveness of coconut as a commercial crop and has convinced farmers to plant and invest more in coconuts. The project also increased the awareness of community members on the importance and the need to conserve and promote diversity which is imperative for sustained farm productivity. Because of its unique combination of poverty reduction interventions, the project attracted the attention and, therefore, the support of government, donors and other stakeholders which benefited the participating communities. Its strategic public awareness strategy made popular the participating communities, so much so that others are looking up to them as models of how farmers, working in unison towards a common goal, would be able to make a significant difference in their own lives. Because of the participatory approaches adopted by the project, the farmers’ sense of ownership and community belonging were enhanced. The project not only improved the quality of life of resource-poor farmers in coconut growing communities but also prepared them socially, psychologically and emotionally for longer-term socioeconomic development. Impact on NARS The survey respondents agreed that the project was able to enhance the service capacity of the research implementing agencies in each of the eight countries. The project improved the ‘bridging’ or facilitating role of the implementing research institute by providing their scientists, researchers and extension workers with the opportunity to test and disseminate to resource-poor coconut growing communities recommended technologies produced by their research programmes. This has given the implementing agencies’ staff the needed exposure to actual grassroots work and boost their confidence about their research. Some project leaders also said that because of the various training and other capacity-building activities conducted under the project, many of their staff acquired new knowledge and skills, thereby “making them better scientists and researchers”. The establishment of community-managed nurseries enabled the research institutes to conserve important local and introduced high-value coconut varieties and promote their in situ and on-farm conservation efforts. Also, the establishment and effective management of village-level 168 COCONUT GENETIC RESOURCES seedling nurseries, as demonstrated by the communities, could lessen the burden on the part of the NARS and the government in establishing and maintaining a formal seed and seedling distribution system. If replicated and scaled-up, this activity could help provide the framework for establishing a community-managed informal seed distribution system which is self-sustaining – a system in which the communities themselves raise and propagate high-value and suitable varieties, providing the necessary inputs, manpower and land while gaining income from them as well. This could free up the NARS and the government from providing the needed staff and other resources for this purpose and instead realign them into other research areas that would benefit the poor coconut farmers. Strong linkages between government research institutions and poor coconut growing communities have been established, effectively mobilizing the former to help the latter. This has motivated government researchers to deploy more research results, enhance and expand their coconut-based farming systems research, and link with other relevant research and development organizations. As a result of the three years of research at the village level, CBO members are now more cooperative with and trusting of government organizations to help them. For the government coconut research institutions, this project has given them an important experience and impetus for developing a research agenda to refine and scale up similarly designed poverty reduction research in coconut growing communities in the future. Impact on the coconut industry As the project was implemented in only three to four communities per country, the impact on the industry would not really be obvious at this stage. However, potential impacts have been recognized. The project demonstrated that diverse high-value products can be made from all parts of the coconut (kernel, husk, shell, wood, water, leaves) and that these could be marketed, thereby expanding the value of coconut as a commodity crop and enhancing the competitiveness of its value-added products in the global market. This could expand existing local and international markets and create demand for new products. This diversity of products that can be profitably produced by farmers at the village level serves as an attractive incentive for them to plant more coconuts, thereby making coconut production and marketing more equitable and sustainable. In conclusion, this farmer participatory poverty reduction project has shown that poor coconut farmers could effectively manage their coconut 169 CHAPTER 3: Germplasm conservation and associated production systems to improve their lives. The project has also shown in eight participating countries and other member countries of COGENT that the coconut could be conserved and at the same time optimally utilized to help improve the socioeconomic and environmental conditions of poor rural coconut-growing communities and countries. Based on the results of this project, IPGRI and COGENT will convince and help these countries institutionalize and scale-up this poverty reduction research intervention to maximize its benefits and expand its impact. Sustainability Sustaining impact when a project phases out is a major concern especially of those dealing with action research for development, in this case, reducing poverty through the introduction and testing of various coconut- based income-generating technologies. Under the project, this concern was consciously addressed by adopting and integrating into its design the sustainable livelihoods framework, which essentially calls for identifying and implementing interventions to enhance the five capitals (i.e., human, natural, social, physical and financial) as well as formulating sustainability indicators for each intervention. Human capital In the project, human capital was enhanced through the conduct of various training and capacity-building activities aimed at two essential objectives: (1) to organize farmers into formal CBOs and strengthen these organizations so that they would be able to effectively manage their own affairs without the help of ‘outsiders’ even after the project terminates; and (2) to build the capacity and skills of individual farmers to undertake various income-generating activities to enable them to fully and profitably engage in coconut-based enterprises in the future. Of course, building- up the capacity of CBOs and its members through training is just one aspect of enhancing human capital to ensure the sustainability of project impact. Another equally important aspect is giving farmers a sense of self-confidence, self-realization and fulfillment, which goes hand-in-hand with training to tap into their potentials and develop their skills, which the project helped achieve. Once these values are ingrained in them, they would have the motivation and the spirit to continue to further develop themselves and thereby contribute to the development of the community in general. 170 COCONUT GENETIC RESOURCES Natural capital It is an accepted fact that in order for the benefits and impact of this project to endure long after it has terminated, the component central to its strategy (i.e., coconut diversity) will have to be continuously enhanced. Thus, the project has established a mechanism through community- managed seedling nurseries that would assure the continuous on-farm propagation of high-value and suitable coconut varieties. Catering to both coconuts and various other intercrops, these integrated nurseries would form the platform on which a self-sustaining, non-government dependent, informal and village-level seed and seedling distribution system could be built on. Such a system, which would benefit both the farmers – as they would also earn from them – and the government – they would free scarce resources needed to maintain such nurseries – would ensure that coconut diversity, as well as the diversity of various cash, food security and feed crops, in the communities would continue to improve and flourish. Social capital The whole gamut of people, tradition and culture comprise what we call a community’s ‘social fabric’. In the implementation of the project, the enhancement of the social capital was taken into account by introducing activities and technologies that are socially acceptable – those that do not run counter to the existing norms of the people and the community. This ensures that no friction is created between the project interventions and the people who were supposed to adopt them. In Huntu, for example, a predominantly Muslim project site in Indonesia, swine was not introduced as livestock despite market surveys showing that pork has a high demand in the neighbouring villages as this is in conflict with the dominant religion in the area. Efforts like this would ensure that interventions introduced would continue to be supported by the community. Under the project, individual farmers were encouraged to form common-interest groups in undertaking similar income-generating activities, which promoted social cohesiveness and group unity. In the coconut fibre processing community of Tam Quan Nam in Vietnam, for example, CBO members established a common area for dehusking, decorticating and collecting fibre. Each member of the group takes turns in operating and maintaining the machinery and the work area itself, and as ‘payment’ they get processed coconut fibre which they then bring home to use in producing ropes, mats, geotextile and other fibre-based products. These are then collected and sold by the CBO on behalf of its members, with part of the proceeds going to the further upkeep of the 171 CHAPTER 3: Germplasm conservation machinery and equipment. Such a practice ensures that that machinery and the processing area are well-kept, which would have been very difficult if only the CBO did the job itself. This has proven to be so successful that they are now constructing another building (shed) to house one more set of fibre processing machinery, even after the project has phased out in December 2004. Additionally, there was conscious effort that technologies introduced by the project would be simple enough that all the members of the family would be able to take part in them. In raising poultry under coconut for example, young children could feed the chickens before going to school and after classes. In making single-ply fibre ropes, the technology is so uncomplicated that even the grandparents could learn how to spin coconut fibre, and the machine is very simple and inexpensive that a family could own two or more units. In all, the project interventions and activities promoted unified family work and bound the family closer together with the common objective of contributing to the upliftment of their plight. Social cohesiveness is vital if continuous community development is to be achieved. Physical capital It goes without saying that infrastructure, machinery, equipment and other physical facilities are important in a technology-based poverty reduction project such as this. Under the project, the physical capital of the communities was enhanced by introducing simple, inexpensive, village-level machinery and equipment to process coconuts into high- value, marketable products. This was a strategy adopted by the project to ensure that successful technologies would easily be replicated, adapted and expanded by the communities themselves at minimal cost but with maximum benefit to its constituents, even when the project ended. The project also promoted the interchange of technologies between and among countries. The Vietnam model of the simple single-ply rope making machine could now be seen in other countries which participated in the project and who did not have this machine before. Another example is the simple coconut virgin oil extracting machine of Sri Lanka, which has been disseminated to other countries as well. Some of these technologies and machines, shared in good faith by countries, have been modified and adapted to suit the local conditions of the communities, usually through the initiative of the CBOs themselves and with some support from the national implementing agencies. These simple but highly efficient and effective technologies, machinery and equipment would provide the motivation for the communities and the farmers to continue to adopt them since they provide additional income at minimum cost – 172 COCONUT GENETIC RESOURCES an almost no lose situation for them. To protect the interest of the inventors or developers of these village-level machineries and equipment, a Material Transfer Agreement which binds recipients not to patent these resources has been developed by IPGRI-COGENT. Also, because of the project’s achievements, the local government units have taken notice of the project sites that some of them provided the needed infrastructure to support their development. Some have given diesel-powered generators to run the dehusking and decorticating machinery provided by the project, while others have improved the farm to market roads going in and out of the project sites to help farmers transport their products. Others have also promised to install transformers to provide three-phase electrical line to a project site so that the CBO members could cheaply operate coconut processing machinery which presently runs on a diesel generator. These physical and infrastructure improvements would go a long way in making sure that the project’s interventions and impact would continue. Financial capital One of the main reasons why small-scale coconut farmers remain poor is because they have very limited access to financial resources to diversify and invest in higher-return, high-value income generating activities or enterprises. As mentioned, most coconut farmers are marginalized – with no land and collateral to obtain loans and are considered non- bankable by formal lending institutions. This is also the reason why most ‘relief’ or dole-out projects fail – farmers have a very simple concept of money - when you have it, you spend it, without consideration of paying it back since it was ‘given’. Although the project provided funds to the communities, these were mostly in the form of revolving funds for microcredit that the CBO used to loan-out to members which were repaid at nominal interest. At the onset of the project, the CBOs were told that “nothing is free in this project”, that everything would have to be paid back, except training and capacity building. With this principle in mind, the CBOs were encouraged to develop their own microcredit lending system, which would make their revolving fund grow. The establishment of these microcredit systems provided the ‘non-bankable’ farmers access to capital which they used to finance their various coconut-based income- generating activities. This initiative proved successful that some CBOs have doubled or almost tripled the original seed money given to them by the project. Moreover, the microcredit system inculcated in the farmers a sense of ‘pay back’ – meaning they have to work to repay what they owe and not merely spend their money at whim. Such a system would 173 CHAPTER 3: Germplasm conservation ensure that farmers have continuous access to needed financial resources to further expand their activities and at the same time catalyze the fiscal growth of the CBO to serve more poor members. Lessons learned The lessons learned from the Poverty Reduction project were consolidated from the various communications, correspondences and meetings with the Country Project Leaders, Community Coordinators and project participants; and from the reports submitted, and discussions with other project staff and stakeholders, including partners and collaborators in the project sites. Some of the lessons are not entirely new, and many cut across various topics. There are 33 lesson points, which are listed below and classified under eight major headings: On implementing the project 1. A special arrangement to expedite the transmittal and release of funds from the Implementing Agencies (IA) to the CBOs was necessary for more efficient project implementation; 2. Staff assigned to the project should have been freed of some of their regular institutional workload so that they could have focused more on implementing and monitoring the activities of the project; 3. Formalizing the assignment of staff to the project (i.e., Programme Leader, the Project Leader, Community Coordinators and members of the Technical Support Team), indicating terms of reference, duties, responsibilities and obligations, would have helped mitigate the delays and difficulties caused by unexpected staff movement; 4. Having a senior staff of the IA as head of the Project Management Team (i.e., Programme Leader) was advantageous in terms of project implementation and team coordination; and 5. A comprehensive and exhaustive orientation and levelling off to get a clear grasp of the nature, concept, rationale, strategy, objectives and overall goal of the project could have prevented the confusion as to the true nature of the project that was evident among some project implementers and participants. On transferring coconut-based income-generating technologies through on farm and off farm trials 6. Different people had different but equally valid reasons and motivations for choosing to be involved or not to be involved in certain project-related activities; therefore, not all people tried all the technologies being tested or introduced or, conversely, not all technologies introduced suited all project participants. The reasons 174 COCONUT GENETIC RESOURCES and motivations of farmers for testing and adopting or not adopting project interventions need to be further identified and studied; 7. Testing and introduction of improved livestock breeds, specifically of poultry, did not always translate to immediate adoption of the technology by farmers as the latter have a different set of criteria than researchers or scientists in selecting the poultry type they would adopt; 8. Exchange visits were effective mechanisms in promoting and disseminating various coconut-based income generating technologies and in catalyzing in-country and international partnerships and collaboration; 9. Making people fully understand the purpose and advantages of forming focused groups and letting them do so at their own choice and pace was better than ‘forcing’ them to establish or join a group just for the sake of meeting the project’s requirements but not really understanding why; and 10. A ‘blue print’ project design was not always suitable or applicable to different communities in different countries with highly-diverse peoples and distinct cultures. On managing community-based organizations 11. The success of the CBO, as with any other organization, hinged much on the leadership qualities and dedication of its officers; 12. Since the project both strengthened CBOs in areas where one already existed and established new CBOs in communities where there was none before, it was recognized that the former was more advantageous in terms of producing more results given the activities and time frame of the project; and 13. The CBO, to continue to effectively function as a social organization, also needed to earn income to sustain itself. On monitoring and evaluating the project 14. Designing a project Monitoring and Evaluation (M&E) system and its tools without the inputs or participation of those who were supposed to regularly implement them, especially Project Leaders and Community Coordinators, was ill-advised as monitoring and reporting became unfocused; 15. Limited, and oftentimes difficult, accessibility to some of the project sites posed some challenges to project implementation, monitoring and evaluation; 16. The country project leader and project support/ technical team should be physically located as near as possible to each other and the project sites; 175 CHAPTER 3: Germplasm conservation 17. The project Community Coordinators should be located in the same community where the project is being implemented; 18. Project IA staff (other than the Community Coordinators) should be extensively immersed in the field to get the “feel” of the community and to earn the trust and respect of its members; and 19. Documenting project success stories and lessons are effective approaches in highlighting critical changes in the participants’ lives, in rallying support for the communities and in learning from past mistakes and successes to make the future, and ourselves, better. On building the capacity of CBO members 20. Purposeful hands-on training is the most effective method to maximize farmer learning; 21. Men and women have different reasons and considerations in joining project activities. Therefore, training requires gender-specific approaches which take into account such differences; 22. Participation in training activities is not always possible for those who live far from the project community proper; 23. Transfer of technologies within the CBO – members training other members – was a long and often arduous process which largely depended on the kind of the technologies to be transferred; 24. Building the organizational management capacity of both IA staff and CBO officers to implement and manage project interventions should have been given top priority; and 25. Linking the project with providers of training and capacity building support services could make capacity building activities sustainable. On marketing 26. Encouraging and helping farmers diversify into more than one income- generating activity or enterprise was better than persuading them to pursue just one. This not only provides them with more earning options but more importantly, it spreads out the risks in case one activity fails or if the market becomes saturated with one of the products; 27. Poor transport infrastructure (farm-to-market roads) and facilities negate the benefits realized from increased farm productivity; and 28. Linking with specialized associations or organizations dealing with product and market development was a good approach in developing and promoting the products of the CBOs. On establishing and managing a village-level microcredit system 29. The community microcredit systems that were generally successful usually shared common characteristics, which are: (a) provision of 176 COCONUT GENETIC RESOURCES small, non-collateralized loans in cash or in-kind; (b) flexible repayment in cash or in-kind; and (c) with technical backstopping and training support; 30. Employing people who are well-respected and highly-regarded in the community increased the recovery rate of loans; and 31. An elaborate yet simple screening process for borrowers and applying social pressure to collect payments helped in recovering loans. On promoting coconut diversity (planting of coconut seedlings) 32. Involving the landowners in communities where most farmers are tenants proved to be beneficial in promoting the diversity of coconuts and other crops; and 33. Land tenure, farm size and perceived benefits were major determining factors in encouraging or discouraging farmers to plant coconut seedlings. Constraints and recommendations Constraints 1. The delayed release of donor funds to IPGRI and consequently to the implementing agencies which delayed the overall implementation of the project. This was alleviated by initiating the project using the counterpart funds of IPGRI and the eight participating national research organizations; 2. The replacement of the project leader of the Philippines in the middle of the project and of Fiji close to the end of the project. This was remedied by the accelerated support to the project through the effort of consultants and the Project Coordinator; 3. Reservation of some countries in sharing their coconut processing equipment, machineries and technologies. This was remedied through the development of a Material Transfer Agreement (MTA) binding recipient countries not to acquire patents to transferred equipment and machineries. Interest to share technologies was increased by convincing more countries to also share their processing technologies on a reciprocal basis; and 4. Lack of technical staff from national coconut research organizations to provide adequate technical support to the project. This will be remedied in the future projects by negotiating with the implementing agency to provide this much needed technical support. 177 CHAPTER 3: Germplasm conservation Recommendations 1. IPGRI and its partner implementing agencies should regularly monitor the status of project assets which were transferred to the 25 CBOs including their use of the project machineries and revolving funds for income generating activities. 2. The activities in the 25 communities as indicated in item 1 above should be complemented by additional project activities under the newly approved IFAD-funded Technical Assistance Grant to IPGRI entitled ‘Overcoming poverty reduction in coconut growing communities’ which will involve 15 countries including four of the previous ADB-funded RETA. 3. IPGRI should institutionalize and enhance sustainability elements by linking the communities and research organizations with support groups in the public and private sector. 4. IPGRI should scale up the project by helping countries develop their research agenda on poverty reduction in coconut growing communities and loan-based project proposals in order to develop a critical mass of research to help the neglected coconut sector. Acknowledgment IPGRI and COGENT would like to thank the eight NARs organizations, the three NGOs and the 25 CBOs which collaborated with IPGRI to implement the project; IFAD in funding the first initiative which identified initial high-value coconut products and production systems for increasing farmers’ incomes; DFID in supporting the identification and socioeconomic profiling of the 25 project sites; and ADB in supporting the large-scale testing of technologies and strategies, empowerment and capacity building and overall management of the project. Last but not least, IPGRI-COGENT would like to thank the over 17 000 poor coconut farmers and socioeconomically disadvantaged women who organized themselves and collaborated with each other to improve their lives. 178 COCONUT GENETIC RESOURCES Annex 1. List of participating national coconut research agen- cies, non-governmental organizations and community-based organizations in the Poverty Reduction in Coconut Growing Communities Project National Research Institutes: 1. Bangladesh Agricultural Research Institute (BARI), Bangladesh 2. Central Plantation Crops Research Institute (CPCRI), India 3. Coconut Research Institute (CRI), Sri Lanka 4. Indonesian Center for Estate Crops Research & Development (ICECRD), Indonesia 5. Philippine Coconut Authority (PCA), Philippines 6. Oil Plant Institute of Vietnam (OPI), Vietnam 7. Ministry of Agriculture, Sugar and Land Resettlement (MASLR), Fiji 8. Cocoa and Coconut Institute (CCI), Papua New Guinea Non-governmental Organizations: 1. Banchte Shekha Foundation, Sri Lanka 2. Peekay Tree Crops Development Foundation, Vayalar, India 3. Siyath Foundation, Sri Lanka Community-based Organizations: Bangladesh 1. Bandabila Coconut Community, Bandabila, Jessore District 2. Chandrapara Coconut Community, Chandrapara, Barisal District 3. Banchte Shekha (BS) Coconut Community, Jamira, Khulna District India 4. Ariyankuppam Community Coconut Farmers’ Association, Ariyankuppam 5. Pallikkara Community Coconut Development Centre, Pallikkara 6. Vayalar Community Development Centre, Vayalar, Kerala Sri Lanka 7. Thuthipiritigama Entrepreneurship Development Society, Thuthipiritigama, Hettipola, Kurunugala, Northwestern Province 8. Womens Savings Effort, Wilpotha, Puttalam, Western Province 9. Dodanduwa Womens Collective, Dodanduwa, Galle, Southern Prov- ince 179 CHAPTER 3: Germplasm conservation Indonesia 10. Kelompok Tani Kelapa Harapan Wori, Wori, Wori District, Minahasa Regent, North Sulawesi 11. Kelompok Tani Kelapa Momosad Nonapan I, Nonapan, Poigor Dis- trict, Bolaang Mongondow Regent, North Sulawesi 12. Kelompok Tani Kelapa Huyula Huntu, Huntu-Batudaa, Bongomeme District, Donggala Regent, Gorontalo Province Philippines 13. Malapad Integrated Livelihood Cooperative, Malapad, Real, Quezon 14. Bahay Patol Agrarian Reform Beneficiaries Multi-Purpose Coopera- tive, Caliling, Cauayan, Negros Occidental 15. Linabu Coconut Planters’ Association, Linabu, Misamis Oriental 16. Fleischer Estate Integrated Marketing Cooperative, Old Poblacion, Maitum, Saranggani (associated CBO) Vietnam 17. Hung Phong/Phong Nam Coconut Communes, Hung Phong and Phong Nam, Giong Trom District, Ben Tre Province 18. Xuan Dong Coconut Community, Xuan Dong, Tien Giang Province 19. Tam Quan Nam Coconut Community, Tam Quan Nam, Binh Dinh Province Fiji 20. Tukavesi Development Association, Tukavesi, Savusavu 21. Belego Multiracial Farmers Association, Belego, Wailevo, Savusavu 22. Cicia Women’s Group, Cicia Island Papua New Guinea 23. Murukanam Community Association, Murukanam, Madang Prov- ince 24. Transgogol Community Association, Transgogol, Madang Province 25. Last Karkar Community Association, Last Karkar, Madang Province 180 COCONUT GENETIC RESOURCES A nn ex 2 . Su m m ar y of a ct iv iti es a nd ta rg et o ut pu ts o f t he e ig ht c ou nt rie s pa rt ic ip at in g in C O G EN T’ s Po ve rt y R ed uc tio n Pr oj ec t ( Ja nu ar y 20 02 to D ec em be r 2 00 4) Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a I. C ap ac ity B ui ld in g A. E st ab lis hm en t & st re ng th en in g of C om m un ity -B as ed O rg an iz at io ns (C B O s) (C B O in co rp or at ed a nd re gi st er ed w ith ap pr op ria te g ov er nm en t ag en cy ) 3 C B O s 2 C B O s 1 C BO 3 C B O s 3 C B O s 3 C B O s 3 C B O s 3 C B O s B . T ra in in g of of fic er s/ le ad er s an d m em be rs o n : (1 ) C B O m an ag em en t an d re sp on si bi lit ie s; an d (2 ) M ic ro cr ed it sy st em an d m an ag em en t P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s/ le ad er s ƒ a t l ea st 1 00 m em be rs P er p ro je ct si te : ƒ 1 0 of fic er s / le ad er s ƒ a t l ea st 1 0 0 m em be rs C . I de nt ifi ca tio n of m ar ke ta bl e pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu e co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts P er p ro je ct si te : A t l ea st : ƒ 3 h ig h va lu co co nu t pr od uc ts ƒ 3 c ro p pr od uc ts ƒ 3 li ve st oc k pr od uc ts D . E va lu at io n of o il ex tr ac tio n m ac hi ne s P er c ou nt ry : 2 m ac hi ne s P er c ou nt ry : 2 m ac hi ne s - P er c ou nt ry : 2 m ac hi ne s P er c ou nt ry : 2 m ac hi ne s P er c ou nt ry : 2 m ac hi ne s P er c ou nt ry : 2 m ac hi ne s P er c ou nt ry : 2 m ac hi ne s E. E va lu at io n of o th er pr oc es si ng m ac hi ne s (E ac h co un try s ho ul d ev al ua te a t l ea st 3 o f 5 m ac hi ne s fo r m ea t, fib re , sh el l, le af & w oo d pr od uc ts ) P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo r m ea t, fib re , sh el l, le af o r w oo d pr od uc t P er p ro je ct si te : A t l ea st 1 m ac hi ne ry fo m ea t, fib re , sh el l, le af o r w oo d pr od uc 181 CHAPTER 3: Germplasm conservation Target Outputs India Project Activities Sri Lanka c/o CPCRI (2 sites) c/o Peekay Tree (1 site) Bangladesh Indonesia Philippines Fiji Papua NeGuinea F. D ev el op m en t o f a ct io n pl an s fo r i nc om e- ge ne ra tin g ac tiv iti es 1. F ar m er s’ a ct io n pl an 2. W om en ’s a ct io n pl an 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct s ite 1 pe r p ro je ct 1 pe r p ro je ct G . F ab ric at io n of s el ec te d in ex pe ns iv e pr oc es si ng m ac hi ne s fro m (E ) a bo ve P er c ou nt ry : 1- 3 m ac hi ne s Pe r p ro je ct s ite : 1- 3 m ac hi ne s Pe r p ro je ct s ite : 1- 3 m ac hi ne s P er c ou nt ry : 1- 3 m ac hi ne s P er c ou nt ry : 1- 3 m ac hi ne s P er c ou nt ry : 1- 3 m ac hi ne s P er c ou nt ry : 1- 3 m ac hi ne s P er c ou nt ry : 1- 3 m ac hi ne II. T ra in in g of F ar m er s an d W om en A . H ig h- va lu e co co nu t pr od uc ts Pe r p ro je ct s ite : ƒ 1 00 pa rti cip an ts w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs Pe r p ro je ct s i ƒ 1 00 pa rti ci pa nt s w ith a t l ea s 20 % w om e ƒ a t l ea st 2 re se ar ch er s an d 4 ex te ns io n w or ke rs B . In te rc ro pp in g Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s i ƒ 1 00 pa rti ci pa nt s w ith a t l ea s 20 % w om e ƒ a t l ea st 2 re se ar ch er s 4 ex te ns io n w or ke rs C . Li ve st oc k pr od uc tio n Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s ite : ƒ 1 00 pa rti ci pa nt s w ith a t l ea st 20 % w om en ƒ a t l ea st 2 re se ar ch er s & 4 ex te ns io n w or ke rs Pe r p ro je ct s i ƒ 1 00 pa rti ci pa nt s w ith a t l ea s 20 % w om e ƒ a t l ea st 2 re se ar ch er s 4 ex te ns io n w or ke rs 182 COCONUT GENETIC RESOURCES Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a D. F ee d fo rm ul at io n Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s ite : A t l ea st 5 fa rm er s tra in ed in it em (C ) ab ov e Pe r p ro je ct s i A t l ea st 5 fa rm er s tra in in it em (C ) ab ov e III . Pr od uc tio n an d M ar ke tin g A. I de nt ifi ca tio n, pr od uc tio n an d m ar ke tin g of h ig h- va lu e co co nu t p ro du ct s Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s ite : A t l ea st 3 di ffe re nt h ig h- va lu e co co nu t pr od uc ts (o ne fro m m ea t, fib re , sh el l, le af o r w oo d) Pe r p ro je ct s i A t l ea st 3 di ffe re nt h ig h va lu e co co nu pr od uc ts (o n fro m m ea t, fi sh el l, le af o r w oo d) IV . I nt er cr op pi ng T ria ls A. C as h cr op s Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s i ƒ 3 ty pe s of an nu al s ƒ a t l ea st 1 pe re nn ia l cr op ƒ a t l ea st 50 fa rm er s in vo lv ed B . Fo od s ec ur ity c ro ps Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s ite : ƒ 3 ty pe s of cr op s ƒ a t l ea st 5 0 fa rm er s in vo lv ed Pe r p ro je ct s i ƒ 3 ty pe s of cr op s ƒ a t l ea st 50 fa rm er s in vo lv ed C . So ur ci ng a nd pr op ag at io n of q ua lit y pl an t m at er ia ls fo r in te rc ro ps id en tif ie d in (A ) a nd (B ) a bo ve ƒ C R IS L (fo r co co nu t se ed nu ts ) ƒ A gr ar ia n se rv ic es ce nt er s ƒ C P C R I ƒ K er al a A gr i. U ni ve rs ity (K A U ) ƒ T am il N ad u A gr i. U ni ve rs ity (T N A U ) ƒ S ta te G ov ’t C en te rs ƒ C oc on ut D ev el op m en t B oa rd ƒ C PC R I ƒ A gr ic ul tu ra l U ni ve rs ity D ep ar tm en t ƒ B A R I ƒ B A D C ƒ D AE ƒ H or t C en tre ƒ A gr ic ul tu ra l U ni ve rs ity an d S ta te G ov er nm en t ƒ P C A ƒ W ai ni ga ta R es ea rc h S ta tio n ƒ lo ca l ex te ns io n se rv ic es co m po un d ƒ P N G C C R I ƒ N AR I ƒ D AL 183 CHAPTER 3: Germplasm conservation Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a V. L iv es to ck A. L iv es to ck p ro du ct io n Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s i 3 ty pe s of liv es to ck in vo lv in g at le as t 5 0 fa rm er s B . Pa st ur e, fo dd er a nd le gu m es Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s ite : A t l ea st 1 ty pe ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s Pe r p ro je ct s i A t l ea st 1 ty p ea ch o f pa st ur e, le gu m e & fo dd er , in vo lv in g at le as t 5 0 fa rm er s C . Lo ca l f ee d fo rm ul at io n by e ac h C B O Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s ite : A t l ea st 1 fe ed fo rm ul at io n pe r ty pe o f liv es to ck Pe r p ro je ct s i A t l ea st 1 fe e fo rm ul at io n p ty pe o f liv es to ck D . S ou rc in g an d m ul tip lic at io n of q ua lit y liv es to ck fo r l oa n or di st rib ut io n Su gg es tio n: C B O s sh ou ld ex pl or e th e po ss ib ilit y of lin ki ng w ith a pp ro pr ia te N G O s fo r t hi s ac tiv ity ƒ C R IS L ƒ V R IS L ƒ L oc al liv es to ck di st rib ut io n pr og ra m m es ƒ K A U ƒ T N A U ƒ S ta te G ov er nm en t C en te rs ƒ K A U ƒ A ni m al H us ba nd ry D ep ar tm en t ƒ N at io na l ba nk s’ lo an pr og ra m m es ƒ N ab ar d ƒ L iv es to ck R es ea rc h In st itu te ƒ L iv es to ck S ta te G ov er nm en t C en te r ƒ D A R c at tle di sp er sa l pr og ra m m e ƒ A ni m al H ea lth & P ro du ct io n D iv is io n ƒ D ep t. of A g & L iv es to ck re se ar ch 184 COCONUT GENETIC RESOURCES Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a VI . C on se rv at io n an d En ha nc em en t o f C oc on ut D iv er si ty A. C ha ra ct er iz at io n of ex is tin g c oc on ut va rie tie s in e ac h pr oj ec t s ite u si ng : 1. F ar m er s’ p ro to co l ( to be p ro vi de d by C O G E N T) 2. S TA N TE C H p ro to co l 3. M ol ec ul ar m ar ke rs - C O G E N T to p ro vi de ad di tio na l b ud ge t (e xc ep t t ho se w ith as te ris k* ) C R IS L sc ie nt is ts to ch ar ac te riz e co co nu t va rie tie s in ea ch s ite C PC R I sc ie nt is ts to ch ar ac te riz e co co nu t va rie tie s in e ac h si te S ur ve y to b e co nd uc te d B A R I s ci en tis ts to c ha ra ct er iz e co co nu t va rie tie s in ea ch s ite u si ng th e S TA N TE C H M an ua l R IC P a nd C R IE C sc ie nt is ts to ch ar ac te riz e co co nu t va rie tie s in e ac h si te P C A -Z R C to co nd uc t va rie ta l su rv ey s in ea ch s ite Ta ve un i C oc on ut C en te r & W ai ni ga ta R es ea rc h S ta tio n sc ie nt is ts to ch ar ac te riz e co co nu t va rie tie s in e ac h si te * S ur ve y to b e co nd uc te d B . I de nt ifi ca tio n of h ig h- yi el di ng (H Y) a nd hi gh -v al ue (H V) co co nu t v ar ie tie s Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s ite : ƒ 3 H Y ƒ 2 H V Pe r p ro je ct s i ƒ 3 H Y ƒ 2 H V C . E st ab lis hm en t o f co m m un ity -m an ag ed nu rs er ie s fo r pr op ag at io n & sa le o f: 1 C oc on ut s 2. In te rc ro ps 3. P as tu re /F od de r/ L eg um es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd e r/ le gu m es Pe r p ro je ct s ite : ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at ed nu rs er y fo r in te rc ro ps an d pa st ur e/ fo dd er / le gu m es Pe r p ro je ct s i ƒ 2 c oc on ut nu rs er ie s ƒ 1 in te gr at e d nu rs er y fo r in te rc ro ps an d pa st ur e fo dd er / le gu m es D . P la nt in g of 5 c oc on ut se ed lin gs p er y ea r P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er /w om en pa rti ci pa nt s P er p ro je ct s ite pe r y ea r: 5 se ed lin gs p er 10 0 fa rm er / w om en pa rti ci pa nt s P er p ro je ct s pe r y ea r: 5 se ed lin gs p 10 0 fa rm er / w om en pa rti ci pa nt s VI I. D is se m in at io n an d Pr om ot io n of R es ea rc h R es ul ts A. D ev el op m en t o f Te ch no g ui de s (T G s) / Ex te ns io n B ul le tin s (E B s) P er p ro je ct s ite pe r y ea r: ƒ 3 T G s (in E ng lis h & na tio na l la ng ua ge ) ƒ 1 b ro ch ur e on H VP P er p ro je ct s ite pe r y ea r: ƒ 6 T G s/ EB s re le va nt la ng ua ge s P er p ro je ct s ite pe r y ea r: ƒ 3 E Bs (l oc al la ng ua ge ) P er p ro je ct s ite pe r y ea r: ƒ 2 T G s (lo ca l di al ec ts ) ƒ 2 E Bs P er p ro je ct s ite pe r y ea r: ƒ 2 T G s/ E Bs (re le va nt la ng ua ge s/ di al ec ts ) P er p ro je ct s ite pe r y ea r: ƒ 1 5 TG s (lo ca l di al ec ts ) P er p ro je ct s ite pe r y ea r: ƒ 2 T G s/ EB s (re le va nt la ng ua ge s/ di al ec ts ) P er p ro je ct s pe r y ea r: ƒ a t l ea st 3 TG s/ EB s 185 CHAPTER 3: Germplasm conservation Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a B. S em in ar /p re se nt at io n ab ou t pr oj ec t A t l ea st o nc e a ye ar A t l ea st o nc e a ye ar 2 se m in ar s pe r ye ar 4 se m in ar s pe r ye ar A t l ea st o nc e a ye ar A t l ea st o nc e a ye ar A t l ea st o nc e a ye ar A t l ea st o nc e ye ar C . Pu bl ic at io n of s ci en tif ic pa pe r A t l ea st 1 sc ie nt ifi c pa pe r pe r y ea r A t l ea st 1 sc ie nt ifi c pa pe r pe r y ea r A t l ea st 1 sc ie nt ifi c pa pe r pe r y ea r A t l ea st 1 sc ie nt ifi c pa pe r pe r y ea r 3 sc ie nt ifi c pa pe rs p er ye ar 2 sc ie nt ifi c pa pe rs p er ye ar 3 sc ie nt ifi c pa pe rs p er ye ar 3 sc ie nt ifi c pa pe rs p er ye ar D . Pu bl ic at io n of p ub lic aw ar en es s m at er ia ls in th e na tio na l d ai lie s A t l ea st 2 p er ye ar 6 in th e lo ca l & na tio na l d ai lie s pe r y ea r 3 in th e lo ca l & na tio na l d ai lie s pe r y ea r A t l ea st 2 p er ye ar 6 in th e lo ca l & na tio na l d ai lie s pe r y ea r 6 in th e lo ca l & na tio na l d ai lie s pe r y ea r 3 in th e na tio na l d ai lie s pe r y ea r 3 in th e lo ca l na tio na l d ai li pe r y ea r E. F ie ld d ay s A t l ea st tw ic e pe r y ea r p er si te A t l ea st tw ic e pe r y ea r p er si te At le as t t w ice pe r y ea r p er s ite At le as t t w ice pe r y ea r p er s ite At le as t t wi ce pe r y ea r p er s ite A t l ea st tw ic e pe r y ea r p er si te A t l ea st tw ic e pe r y ea r p er si te A t l ea st tw ic e pe r y ea r p er si te VI II. P ro je ct M ee tin gs a nd S ite V is its A. M ee tin g of C B O of fic er s/ le ad er s Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e a m on th w ith re po rt su bm itt ed Pe r p ro je ct s i O nc e a m on t w ith re po rt su bm itt ed B . M ee tin g of C B O m em be rs Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s ite : O nc e pe r y ea r w ith re po rt su bm itt ed Pe r p ro je ct s i O nc e pe r y e w ith re po rt su bm itt ed C . Si te v is its b y pr oj ec t le ad er Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s ite : A t l ea st o nc e ev er y 2 m on th s Pe r p ro je ct s i A t l ea st o nc e ev er y 2 m on D . G ro up m ee tin gs o f co un tr y pr oj ec t l ea de r an d th re e co m m un ity co or di na to rs Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye a r (s em ia nn ua l E. M ee tin gs o f p ro je ct le ad er w ith p ro je ct te ch ni ca l s up po rt te am Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye ar (s em ia nn ua l) Tw ic e a ye a r (s em ia nn ua l 186 COCONUT GENETIC RESOURCES 1 T he c om pu te r an d co lo ur p ri nt er a re fo r th e ex cl us iv e us e of t he p ro je ct le ad er s fo r of fic ia l c om m un ic at io ns /e -m ai l, re po rt p re pa ra ti on s, d es ig n an d pr in ti ng o f T ec hn og ui de s/ E xt en si on B ul le ti ns ( w it h co lo ur ed c ov er p ag es ), pr od uc ti on o f t ra in in g m at er ia ls a nd o th er p ub lic a w ar en es s m at er ia ls a nd /o r fo r ot he r pu rp os es r el at ed t o th e im pl em en ta ti on of t he p ro je ct . If t he p ro je ct l ea de r is c ha ng ed o r re pl ac ed , th e sa id e qu ip m en t m us t be t ur ne d ov er t o hi s/ he r re pl ac em en t. 2 T he m ot or cy cl es a re in te nd ed fo r th e ex cl us iv e us e of t he c om m un it y co or di na to rs fo r or ga ni zi ng , m on it or in g an d im pl em en ti ng p ro je ct a ct iv it ie s in t he ir r es pe ct iv e pr oj ec t si te s. T he c ou nt ry c oo rd in at or s ar e ex pe ct ed t o m ai nt ai n an d us e th e m ot or cy cl e fo r pu rp os es r el at ed t o th e pr oj ec t on ly . I n ca se t he c om m un it y co or di na to r is c ha ng ed o r re pl ac ed , th e m ot or cy cl e sh ou ld b e tu rn ed o ve r to h is /h er r ep la ce m en t. IX . R ep or tin g A. R ep or t o f c om m un ity co or di na to r t o pr oj ec t le ad er 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th 30 th o f e ve ry m on th B . Te ch ni ca l a nd fin an ci al re po rt o f pr oj ec t l ea de r t o IP G R I E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d 15 D ec E ve ry y ea r: 15 J un e an d D ec C . In te gr at ed d on or re po rt of IP G R I t o AD B 30 th J un e an d 30 th D ec em be r o f e ve ry p ro je ct y ea r X. P ro vi si on o f e qu ip m en t A. C om pu te r 1 1 un it - 1 un it 1 un it 1 un it 1 un it 1 un it 1 un it B . M ot or cy cl es 2 3 un its 2 un its - 3 un its 3 un its 3 un its 3 un its 3 un its Ta rg et O ut pu ts In di a Pr oj ec t A ct iv iti es Sr i L an ka c/ o C PC R I (2 s ite s) c/ o Pe ek ay Tr ee (1 s ite ) B an gl ad es h In do ne si a Ph ili pp in es Fi ji Pa pu a N e G ui ne a 187 CHAPTER 3: Germplasm conservation A nn ex 3 . Su m m ar y of a ch ie ve m en ts o f C O G EN T’ s Po ve rt y R ed uc tio n pr oj ec t ( Ja nu ar y 20 02 - D e- ce m be r 2 00 4) C ou nt ry a nd n am e of c om m un ity Na m e of C B O No . o f m em be rs N o. o f tra in in g pa rt ic ip an ts * Nu rs er ie s es ta bl is he d (c oc on ut / in te gr at ed ) Se ed nu ts pr op ag at ed Se ed lin gs pl an te d on fa rm Fa rm er s in vo lv ed in liv es to ck tr ia ls Fa rm er s in vo lv ed in in te rc ro pp in g tr ia ls Fa rm er s in vo lv ed in h ig h- va lu e pr od uc ts P aw a ac tiv m at pu b di ss e B an gl ad es h 32 5 49 80 9 51 00 34 73 18 5 11 5 21 0 • Ba nd ab ila , Ba gh ar pa ra , Je ss or e Ba nd ab ila C oc on ut C om m un ity 10 0 16 60 3 17 00 11 52 42 43 60 • C ha nd ra pa ra , Ba bu go nj , Ba ris al C ha nd ra pa ra C oc on ut C om m un ity 10 0 16 60 3 17 00 57 1 61 39 50 • Ja m ira , Ph ul ta la m , Kh ul na Ba nc ht e Sh ek ha (B S) C oc on ut C om m un ity 12 5 16 60 3 17 00 17 50 82 33 10 0 Fi ji 45 3 19 19 2 10 00 12 57 32 45 4 17 • Tu ka ve si , C ak au dr ov e Tu ka ve si D ev el op m en t As so ci at io n 12 8 55 3 1 50 0 46 1 5 12 8 - • Be le go W ai le vu Be le go M ul tir ac ia l Fa rm er s As so ci at io n 10 5 45 6 1 50 0 59 6 27 10 6 12 • C ic ia Is la nd C ic ia W om en ’s G ro up 22 0 91 0 0 0 20 0 - 22 0 5 In di a 16 92 32 69 15 56 00 88 00 37 0 75 9 61 5 • Va ya la r, Ke ra la Va ya la r C om m un ity D ev el op m en t Pr oj ec t 90 0 16 15 9 20 00 70 00 23 0 22 7 31 5 • Ar iy an ku pp am , Po nd ic he rry Ar iy an ku pp am C om m un e C oc on ut Fa rm er s As so ci at io n 32 0 92 5 3 18 00 18 00 12 0 60 30 0 • Pa llik ar a, Ka sa ra go d Pa llik er e C om m un ity C oc on ut D ev . C en tre 47 2 72 9 3 18 00 - 20 47 2 In do ne si a 74 8 10 09 9 67 27 69 75 82 74 8 10 0 • W or i, W or i D is tri ct , M in ah as a R eg en t Ke lo m po k Ta ni Ke la pa H ar ap an W or i 25 7 32 8 3 22 27 14 96 20 25 7 36 • N on ap an , P oi go r D is tri ct , B ol aa ng M on go nd ow R eg en t Ke lo m po k Ta ni Ke la pa M om os ad N on ap an I 35 5 35 7 3 25 00 42 89 50 35 5 23 188 COCONUT GENETIC RESOURCES • H un tu , Bo ng om em e D is tri ct , D on gg al a R eg en t Ke lo m po k Ta ni Ke la pa H uy ul a H un tu 13 6 32 4 3 20 00 11 90 12 13 6 41 Pa pu a N ew G ui ne a 52 4 35 8 3 30 00 99 37 12 6 41 8 89 • M ur uk an am , M ad an g Ba re m C om m un ity As so ci at io n 22 6 17 1 1 10 00 77 57 13 12 0 38 • Tr an sg og ol , M ad an g Tr an sg og ol C om m un ity As so ci at io n 19 8 10 9 1 10 00 11 80 11 3 19 8 30 • La st K ar ka r, M ad an g La st K ar ka r C om m un ity As so ci at io n 10 0 78 1 10 00 10 00 - 10 0 21 Ph ili pp in es 74 0 16 09 5/ 2 27 6 74 22 7 28 33 4 47 3 37 8 • M al ap ad , R ea l, Q ue zo n M al ap ad In te gr at ed Li ve lih oo d C oo pe ra tiv e 14 5 58 3 2/ 0 65 89 16 10 10 1 29 0 12 3 • C al ilin g, C au ay an , N eg ro s O cc id en ta l Ba ha y Pa to l A R B (A gr ar ia n R ef or m Be ne fic ia rie s) M ul tip ur po se C oo pe ra tiv e 12 9 38 0 1/ 1 15 35 24 46 12 5 71 49 • Li na bu , Ba lin ga sa g, M is am is O rie nt al Li na bu C oc on ut Pl an te rs A ss oc ia tio n 10 9 38 6 1/ 1 16 0 00 18 6 72 45 53 12 5 • O ld P ob la ci on , M ai tu m , Sa ra ng ga ni Fl ei sc he r E st at e In te gr at ed M ar ke tin g C oo pe ra tiv e 35 7 26 0 1/ 0 35 50 - 63 59 81 Sr i L an ka 78 0 12 87 0/ 6 99 00 80 41 19 7 32 8 16 8 • D od an du w a, G al le D is tri ct D od an du w a W om en ’s C ol le ct iv e 42 0 41 0 0/ 1 32 90 91 0 62 21 4 62 • Th ut tir ip iti ga m a, H et tip ol a Th ut tir ip iti ga m a En tre pr eu rs hi p D ev el op m en t So ci et y 16 0 43 0 0/ 2 33 05 34 81 77 63 53 C ou nt ry a nd n am e of c om m un ity N am e of C B O N o. o f m em be rs N o. o f tra in in g pa rt ic ip an ts * Nu rs er ie s es ta bl is he d (c oc on ut / in te gr at ed ) Se ed nu ts pr op ag at ed Se ed lin gs pl an te d on fa rm Fa rm er s in vo lv ed in liv es to ck tr ia ls Fa rm er s in vo lv ed in in te rc ro pp in g tr ia ls Fa rm er s in vo lv ed in h ig h- va lu e pr od uc ts P aw a ac tiv m at pu b di ss e 189 CHAPTER 3: Germplasm conservation *S om e fa rm er s at te nd ed m ul ti pl e tr ai ni ng a ct iv it ie s ** A dd it io na l p ro je ct fa ct s he et s fo r ea ch c om m un it y ha ve b ee n di ss em in at ed d ur in g th e fie ld d ay s in F eb ru ar y- M ar ch 2 00 3 (I nd ia - 3 , S ri L an ka - 3 , V ie tn am – 3 , I nd on es ia - 3) ; al so i nc lu de s m ed ia ( T V a nd r ad io ) co ve ra ge C ou nt ry a nd n am e of c om m un ity N am e of C B O N o. o f m em be rs N o. o f tra in in g pa rt ic ip an ts * Nu rs er ie s es ta bl is he d (c oc on ut / in te gr at ed ) Se ed nu ts pr op ag at ed Se ed lin gs pl an te d on fa rm Fa rm er s in vo lv ed in liv es to ck tr ia ls Fa rm er s in vo lv ed in in te rc ro pp in g tr ia ls Fa rm er s in vo lv ed in h ig h- va lu e pr od uc ts P aw a ac tiv m at pu b di ss e • W ilp ot ha , Pu tta la m D is tri ct W om en ’s S av in gs Ef fo rt 20 0 44 7 0/ 3 33 05 36 50 58 51 53 Vi et na m 45 3 29 61 0/ 6 65 00 33 10 26 7 74 4 42 8 • H un g Ph on g & Ph on g N am , G io ng T ro m , Be n Tr e H un g Ph on g/ Ph on g N am C oc on ut C om m un ity 1 03 10 0 73 0 51 3 0/ 3 0/ 2 50 00 50 0 20 00 40 0 10 2 56 31 9 23 8 88 17 5 • Xu an D on g, T ie n G ia ng Xu an D on g C oc on ut C om m un ity 10 0 32 1 0 41 0 72 24 • Ta m Q ua n N am , Bi nh D in h Ta m Q ua n N am C oc on ut C om m un ity 15 0 13 97 0/ 1 10 00 50 0 37 16 3 16 5 TO TA L fo r e ig ht co un tr ie s 25 C B O s/ fa rm er s’ as so ci at io ns 57 15 17 3 92 43 /1 4 65 5 01 64 5 21 15 93 40 39 20 05 190 COCONUT GENETIC RESOURCES Global coconut conservation strategy P Batugal1,2 and V Ramanatha Rao2 1COGENT Coordinator and 2Senior Scientists, International Plant Genetic Resources Institute – Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction In 1992, at the request of the Consultative Group on International Agricultural Research, the International Coconut Genetic Resources Network (COGENT) was launched by the International Plant Genetic Resources Institute (IPGRI). Starting with 15 countries, COGENT rapidly developed into an active global Network currently involving 38 coconut producing countries (Table 1). The establishment of COGENT paved the way for the global and regional coordination of coconut conservation efforts.. However, with the increasing threat of genetic erosion and the increasing poverty in coconut growing communities, there is a need to further support these initial initiatives of coconut growing countries to upgrade the collections and to enhance and accelerate the documentation, evaluation, conservation and use of coconut genetic resources. Table 1. COGENT member countries Southeast and East Asia South Asia South Pacific Africa/Indian Ocean Latin America/ Caribbean 1. China 2. Indonesia 3. Malaysia 4. Myanmar 5. Philippines 6. Thailand 7. Vietnam 1. Bangladesh 2. India 3. Pakistan 4. Sri Lanka 1. Cooke Is. 2. Fiji 3. Kiribati 4. Papua New Guinea 5. Solomon Is. 6. Tonga 7. Vanuatu 8. Samoa 1. Benin 2. Cote d’Ivoire 3. Ghana 4. Kenya 5. Madagascar 6. Mozambique 7. Nigeria 8. Seychelles 9. Tanzania 1. Brazil 2. Colombia 3. Costa Rica 4. Cuba 5. Guyana 6. Haiti 7. Honduras 8. Jamaica 9. Mexico 10. Trinidad- Tobago Over the past three years, COGENT has conducted several consultations on the conservation and use of coconut diversity to assist coconut growing member countries to develop a progressive conservation strategy. Such a strategy aims to optimize the conservation of as much representative diversity as possible in the most cost-effective manner for the short, medium and long term. These consultations led to a draft Global Coconut Conservation Strategy. In November 2004, the Global Crop Diversity Trust supported a meeting with of the major coconut producing countries to review and update the strategy and identify priority conservation activities. 191 CHAPTER 3: Germplasm conservation The updated strategy was referred to the COGENT Steering Committee, to coconut growing countries and COGENT partner research organizations and, based on the feedbacks received, this revised draft was produced. The strategy will be finalized at the next COGENT Steering Committee meeting in India in November 2006. The resulting Global Coconut Conservation Strategy defines the framework for promoting the effective conservation and use of coconut genetic resources over the next 10 years to guide coconut producing countries in developing their own conservation strategies. This strategy and the identified priority activities are described below. Integrated approach to coconut conservation The coconut conservation strategy is anchored in promoting the sustainable protection of diversity and maximizing its use. In developing the conservation strategy, coconut growing countries recognized that no one method of conservation can meet all conservation needs and that there is a need to employ a combination of methods to ensure the sustainable conservation of as much diversity as possible. The strategy encourages the participation of governments, partner organizations in both developing and developed countries, NGOs and coconut farmers themselves. The components of the conservation strategy are: 1) conservation in national field collections; 2) conservation in the multi-site International Coconut Genebank; 3) in vitro embryo culture and cryopreservation; 4) In situ and on-farm conservation; 5) Promoting conservation through use by : a) developing and implementing a globally coordinated coconut breeding programme, b) establishing farmer community-managed coconut seedling nurseries in at least 25 countries, c) linking germplasm conservation and use with the broader areas of research and development assigned to CIRAD (agro-physiology and crop protection) and APCC (processing and marketing) by PROCORD; d) developing and disseminating catalogues of conserved germplasm and farmers’ varieties; and e) upgrading and wider use of the International Coconut Genetic Resources Database (CGRD). Scope and status of coconut conservation Conservation in national field collections National governments, through their coconut research institutes or their equivalents hold important coconut germplasm collections in their research stations. To date, 22 countries have conserved, characterized and registered their data on a total of 1416 accessions in the International 192 COCONUT GENETIC RESOURCES Coconut Genetic Resources Database (CGRD, Table 2). Gaps in collections in the South Pacific Island countries, especially in the atolls, have been identified in a CIRAD survey, and in the Indian Ocean countries through a CPCRI-India survey. There is a need to enhance collecting and conservation from these climate change-vulnerable island countries to address the threat of genetic erosion due to global warming. There is also a need to identify, collect and conserve germplasm with resistance/ tolerance to the dreaded lethal yellowing disease in Africa and in the LAC region; to the foliar decay disease in Vanuatu; and the cadang- cadang disease in the Philippines. Due to climate change, there is a need to identify, collect and conserve drought-tolerant germplasm, noting that almost all coconuts are grown in rainfed conditions. Finally, there is a need to collect and conserve materials with specific characteristics for coconut product value addition that could produce high-value products for poor coconut farmers. Conservation in COGENT’s multi-site International Coconut Genebank While national coconut field genebanks are important sources of germplasm for exchange, germplasm exchange among countries has been limited due to political and technical constraints. To address these constraints, and to foster an efficient and effective system of germplasm exchange and conservation, the COGENT Steering Committee decided to establish a multi-site International Coconut Genebank (ICG) in 1995. The ICG today comprises 4 regional genebanks hosted by Indonesia for Southeast and East Asia, Papua New Guinea for the South Pacific, India for South Asia and Côte d’Ivoire for Africa and the Indian Ocean. Negotiations are underway to establish a 5th regional genebank in Brazil. The ICG field genebank collections are held in trust under the auspices of FAO. The designated germplasm is shared under the terms of material transfer agreements agreed with FAO. The ICG field genebanks are established, maintained and managed by national programmes with guidance from COGENT and IPGRI. The conservation strategy envisions that the ICG laboratories and facilities will be further developed and upgraded the enable them to further locate representative diversity, identify and eliminate duplicates, conduct disease indexing, process pollen and embryos for export, conduct cryopreservation and train coconut researchers from member countries in evaluating, conserving and using germplasm. The mandates of the International Coconut Genebank are: 1) to conserve nationally and regionally identified priority diversity; 2) to 193 CHAPTER 3: Germplasm conservation Site Number of accessions E=0 0 5 weeks Palms show the death symptoms, drying of the spindle leaf. Disastrous 285 CHAPTER 5: Germplasm use lowest in the hybrids WCT x PHOT and WCT x GBGD and the highest in WCT x MOD (Ramadasan et al. 1991). Repellin et al. (1994) also placed PB - 121 under drought tolerant group while WAT was classified as moderately tolerant based on a study of the effect of edaphic drought on the leaf water status, gas exchange and membrane lipids at the nursery stage of five coconut varieties. Kasturi Bai et al. (2001) reported the extent of variation in physiological characters among the parents and hybrids at the nursery stage. In Sri Lanka, the drought tolerant Tall x Tall palms were selected based on the mean yield and genotypic adaptation to changes in climate over a 15-year period. Characterization of drought in different coconut growing areas in six states of India falling under different agroclimatic zones revealed that the dry spell length and intensity of stress adversely affect the coconut and consequently nut yield (Naresh Kumar et al. 2003). In this study, high adverse effect of length of dry- spell on nut yield was noticed up to four years, with more impact during the fourth year. Apart from this, adverse impact on the current year nut yield was also observed. Physiological responses to drought stress Coconut palm responds to drought stress by stomatal regulation and deposition of leaf surface wax (ECW) to maintain leaf water potential. Osmotic adjustment, by accumulation of organic solutes, also plays an important role in the drought tolerance mechanism in coconut (Kasturi Bai and Rajagopal 2000). Stomatal regulation. Seasonal variations in climatic conditions viz., solar radiation, air temperature and relative humidity influence xylem tension by stomatal regulation, a key factor controlling the water balance in coconut (Milburn and Zimmermann 1977; Rajagopal et al. 1986). Development of stress was monitored in coconut during wet and dry seasons by estimating stomatal regulation (Kasturi Bai et al. 1988; Juma et al. 1997), which varied among cultivars (Voleti et al. 1993a). The leaf to air vapour pressure deficit (LAVPD) and leaf to air temperature difference (rT) influenced the stomatal conductance (gs) and water relations during day time and thus predominantly determined the variations in photosynthetic efficiency of coconut in irrigated and rainfed conditions (Rajagopal et al. 2000b). Jayasekara et al. (1993) identified the drought tolerant genotypes based on the stability in physiological parameters viz., transpiration rate, diffusive resistance and leaf water potential during the stress period. Leaf water potential. Leaf water potential (Øleaf), an indicator of plant water status, showed a vertical gradation from middle leaf upwards, the 286 COCONUT GENETIC RESOURCES magnitude being higher under rainfed condition (Voleti et al. 1993b). Seasonal variation among the cultivars for Øleaf was also noted (Voleti et al. 1993a; Shivashankar et al. 1991). A rapid screening method was developed based on Øleaf in excised leaflets (Rajagopal et al. 1988) for easy handling of a large number of genotypes. The Øleaf declined with time to different degrees among the genotypes, indicating the degree of tolerance/susceptibility to stress. These were in conformity with the field- testing conducted for drought tolerance. Passos and da Silva (1991) established the relation between the hydric state of the tree and the diameter of its trunk by dendrometry. From the studies in a group of Dwarf and Tall palms on the root system, stomatal conductance and water potential, it was concluded that the behaviour of palms in drought conditions depends on several factors, viz., water relation components like transpiration, stomatal conductance and leaf water potential as well as agro-meteorological factors like solar radiation, rainfall and humidity, which may interplay to facilitate over all drought tolerance mechanism (Passos and da Silva 1991). Epicuticular wax (ECW). ECW is one of the important parameters, which influence the energy balance of leaf. In coconut, the ECW on leaves did not differ significantly among cultivars and hybrids under favourable conditions. However, almost three to four fold increases in ECW was noticed during dry season in some of the coconut hybrids viz., WCT x GBGD, WCT X COD, LCT x COD and LCT x GBGD (Voleti and Rajagopal 1991; Kurup et al. 1993). The physiological age of palms and of leaves influenced the formation of wax on leaf surface. Leaves of coconut seedlings have almost 50% less ECW than those of adult palms even at the same degree of stress. Osmotic adjustment. Coconut palms accumulate organic solutes such as sugars and amino acid during stress period. Accumulation of these solutes was more in the tolerant types than the susceptible types during severe stress condition (Kasturi Bai and Rajagopal 2000). This implies that osmotic adjustment plays an important role in the drought tolerance mechanism in coconut. Root-shoot signals. Roots in drying soil are known to over produce abscisic acid (ABA) thus providing signals to shoot for closure of stomata for water regulation in plant (Zhang and Davies 1989). Root-shoot relationship was also reported to be an effective indicator of soil compaction and water stress for coconut seedlings (Nainanayake et al. 287 CHAPTER 5: Germplasm use 2000). High ABA/cytokinin ratio in leaf has positive influence on water use efficiency (WUE), whereas a high ABA/ cytokinin ratio in root has a negative influence on WUE in coconut seedlings (Kasturi Bai 2003). Biochemical responses to drought stress The biochemical responses of coconut palm to drought stress include regulation or synthesis of scavenging enzymes to maintain cell membrane integrity thus enabling cells to tolerate stress. Effect of water deficits on enzyme activity. Concomitant with the decrease in the leaf water status during drought stress, the activities of stress sensitive enzymes differ depending upon the nature and function of the enzyme in question. Drought stress caused an increase in the activities of some of the stress sensitive enzymes viz., peroxidase (PO), polyphenol oxidase (PPO), superoxide dismutase (SOD), acid phosphatase (APh) and L-aspartate: 2-oxoglutarate amino transferase (AAT) in adult WCT palms, while activities of Malic dehydrogenase (MDH) and nitrate reductase (NR) were decreased (Shivashankar et al. 1991; Kasturi Bai et al. 1996 b, Kasturi Bai et al. 2003). Drought tolerant varieties are endowed with a biochemical mechanism in adapting the adverse effects of drought through appropriate regulation of enzyme activities. Many enzymes exist in multiple molecular forms called isozymes and changes in the activity or appearance of isozymes represent the relative tolerance of coconut cultivars to water stress (Shivashankar 1988). Increased intensity of APh isozyme II shows the susceptibility of the genotype to water deficits since APh is a hydrolytic enzyme (Shivashankar and Nagaraja 1996). Two additional fast moving bands of PPO were located in the drought susceptible varieties under stress, while the drought tolerant cultivars showed no change (Shivashankar 1988). The variability in the isozyme patterns of enzymes like esterase, peroxidase, phosphoglucoisomerase, alcohol dehydrogenase, glutamate oxaloacetate transaminase and acid phosphatase were also reported in coconut germplasm (Fernando and Gajanayake 1997). Membrane stability in relation to drought stress. At the cellular level, the impact of stress is generally seen on the integrity of membranes and extent of solute leakage, which is regulated by the cell membrane stability. Normal cell functions are affected due to changes in peroxidation of cell wall lipids (LP) during stress resulting in increased cell permeability and solute leakage. In coconut, lipid peroxidation was high in drought susceptible cultivars as compared to tolerant ones (Chempakam et al. 1993). Drought tolerance is thus characterized by higher activities of the 288 COCONUT GENETIC RESOURCES protective enzymes like SOD, catalase and peroxidase and consequently coupled with lower levels of lipid peroxidation and higher membrane integrity. Coconut seedlings of the tolerant group maintained lower water loss and lipid peroxidation than the susceptible group and a negative correlation between leaf water potential and lipid peroxidation was observed (Kasturi Bai et al. 2001). Repellin et al. (1997) observed marked reduction in total leaf lipid and chloroplastic major lipid (monogalactosyl diacylglycerol) contents in drought susceptible cultivars. Role of K+ and Cl- nutrition in relation to drought tolerance in coconut The role of K+ and Cl- nutrition in relation to drought tolerance in coconut has been explained on the basis of stomatal regulation (Braconnier and D‘Auzac 1990; Braconnier and Bonneau 1998). Unlike in most of the crops where malate serves as a balancing ion for K+, in coconut the absence of chloroplasts in the guard cells deprives the availability of malate (Braconnier and D ‘Auzac 1985). Increase in drought tolerance of palms under dry conditions with the addition of KCl was reported in Ivory Coast (Ollagnier et al. 1983; Rajagopal and Naresh Kumar 2001). Chlorine is important in coconut nutrition and for resistance to water stress; the critical level of Cl was identified as 0.7% in 14th leaf (Bonneau et al. 1993 and 1997). Potassium nutrition also plays an important role in drought tolerance in coconut (Quencez and de Taffin 1981; Bopaiah et al. 1996). Screening for drought tolerance The cell size and number, sub-stomatal cavity size, stomatal frequency, epicuticular wax content and thickness, leaf thickness, stomatal resistance water potential components, cell membrane stability are the essential anatomical and physiological traits for assessing moisture stress in plants (Rajagopal et al. 1991; Repellin et al. 1994; Naresh Kumar et al. 2000). Based on these, coconut germplasm collections comprising of Talls, Dwarfs and hybrids were screened under field conditions for drought tolerance (Rajagopal et al. 1990). Genetic variation in leaflet anatomy in relation to drought tolerance The anatomical basis for physiological efficiency for drought tolerance in coconut was delineated (Naresh Kumar et al. 2000). The study revealed that the leaf anatomical features which favour high photosynthetic rates are favourable for high transpiration rates as well. Thicker leaflets and thick cuticle are some of the xeromorphic characters observed in WCT and FMS. Correlations between anatomical features and physiological parameters also indicated that thick cuticle lowers the cuticular 289 CHAPTER 5: Germplasm use transpiration. The WCT and FMS, which are tolerant to water stress, had thick leaflets, thick cuticle on both surfaces, larger parenchyma, hypodermal and water cells compared to less tolerant ones (COD x WCT, GBGD and MYD). Drought tolerant types had also more scalariform thickening on xylem trachieds in vascular bundles and large sub-stomatal cavities. These traits are lesser in size in moderately tolerant cultivar like PHOT and WCT x COD. The values for these traits were least in susceptible types like COD x WCT, GBGD and MYD. However, the differences for these traits between moderately tolerant and susceptible cultivars were not great. Certain parameters like epidermal cell size (upper and lower) and guard cell size are related to the drought tolerance character of a cultivar. It is possible that the cumulative effects of all these traits contribute to drought tolerance (Naresh Kumar et al. 2000). Ranking of cultivars for drought stress tolerance All the aforesaid parameters showed clear differences between the groups and among the cultivars within the group. A significant negative correlation between stomatal resistance and transpiration rate was found in Talls, Dwarfs and hybrids. Ranking for drought tolerance was done based on all stress sensitive parameters (viz., stomatal regulation, leaf water potential, lipid peroxidation, ECW content, polyphenol oxidase, super oxide dismutase, catalase and peroxidase) using parametric relationships (Rajagopal et al. 1990; Chempakam et al. 1993). All Dwarfs performed badly ranking ranks between 11 and 20, whereas all hybrids (except COD x WCT) and all Talls (except the SS Apricot, Andaman Ordinary and Laccadive Micro) were within rank 10. Based on anatomical features such as thicker leaflets, thick cuticle on both surfaces, larger palisade and spongy parenchyma cells, larger hypodermal cells, water cells and sub-stomatal cavity, genotypes like WCT, FMS and PHOT and WCT x COD hybrid were identified as relatively tolerant to drought stress (Naresh Kumar et al. 2000). Thus, coconut cultivars with different levels of drought tolerance could be identified based on the desirable traits, which reflect on the overall water relations of palms. Presence or absence of desirable traits imparts higher degree of drought tolerance (e.g. WCT x WCT; FMST; LCT; WCT x COD, LCT x GBGD and LCT x COD) or drought susceptibility (e.g. MYD) (Rajagopal et al. 2000a). Two cultivars - San Ramon and Ambakelle Special - were identified as drought tolerant in Sri Lanka (Wikremaratne 1987 and Fernando 1987). In Cote d’Ivoire, PB–121 was identified as tolerant while WAT was classified as moderately tolerant and Rennell Tall x West African Tall as the most sensitive to drought based on drought tolerance index 290 COCONUT GENETIC RESOURCES and effect of edaphic drought on the leaf water status, gas exchange and membrane lipids (Pomier and de Taffin 1982; de Nuce de Lamothe and Benard 1985; Repellin et al. 1994). Genetic variability for photosynthetic efficiency and water use effi- ciency of coconut under drought conditions The photosynthetic rates (Pn) were reduced by water stress mainly due to increase in stomatal or mesophyll resistance, with higher reduction noticed in susceptible types than in tolerant types (Kasturi Bai et al. 1998). Drought tolerant hybrids such as WCT x COD, LCT x GBGD and LCT x COD exhibited higher increase in Pn/gs ratio as well as higher WUE than that of susceptible types during stress period. The potential of palms for higher dry matter (DM) production is reflected on WUE. WUE can be determined based on dry matter accumulation (g DM mm water-1 used) as well as by gas exchange measurements (µmol CO2 mmol -1 H2O). Significant difference in WUE has been observed between the cultivars and hybrids. The hybrids WCT x COD, LCT x GBGD and LCT x COD, and cultivar WCT had higher WUE than the other cultivars and hybrids. Under mild stress conditions, the WUE improved in coconut juvenile palms (Rajagopal et al. 2000b). Recently, efforts are on to find high WUE types in coconut germplasm using carbon and oxygen isotope discrimination method at CPCRI, India; Coconut Research Institute (CRI), Sri Lanka; and Essex University, United Kingdom. Field trials: Nut yield in relation to intensity and length of drought stress The intricate relationship between dry spell and stages of nut development right from inflorescence initiation to the nut maturity as well as annual nut yield in different agroclimatic zones have been well described in literature (Rajagopal et al. 1996; Rajagopal et al. 2000a). Physiological traits responsible for drought tolerance correlated with yield performance under stress conditions and some of the cultivars identified as drought tolerant also proved to be good yielders (Bhaskara Rao et al. 1991; Rajagopal et al. 1992). There were genotypic variations for drought index (Pomier and de Taffin 1982) in coconut. Naresh Kumar et al. (2003) worked out the influence of soil moisture conservation practices on source-sink relationship in coconut. Drought tolerance mechanism in coconut All the above-mentioned research results helped in deciphering the mechanism of drought tolerance and stability in yield of coconut under water stress conditions (see Fig. 1). To sum up, drought tolerance in 291 CHAPTER 5: Germplasm use coconut is the cumulative effect of several inductive morphological, anatomical, physiological and biochemical mechanisms (Rajagopal and Kasturi Bai 2002, Naresh Kumar et al. 2000). The genotypes possessing the above traits of drought tolerance can be used in breeding programmes. The genetics of these important traits are being looked into for developing future coconut improvement strategies. Genetics of drought tolerance related to physiological and biochemical traits To understand the genetics of drought responsive physiological traits in coconut (Rajagopal et al. 2000a), cultivars with desirable characters were selected and crossed in a 2 x 4 line x tester mating design to study their combining ability and gene action. Physiological parameters like, leaf water potential, transpiration rate, net photosynthetic rate (Pn) and lipid peroxidation were recorded in seedlings under non-stress, water stress and recovery conditions. Analysis of variance for combining ability revealed significant differences among parents and hybrids for all characters except transpiration rate on recovery and leaf water potential under stress. Seedling transpiration rate showed higher specific combining ability (SCA) effects than general combining ability (GCA) effects due to predominance of non-additive gene action indicating heterosis for this character. Leaf water potential showed a similar trend. The Pn under stress was additive with good combining ability, while the Pn during non-stress and recovery were governed by non-additive gene action that could be exploited for heterosis. In case of lipid peroxidation, gene action was unpredictable in non-stress with additive gene action being nil with low dominance. These indicate that the nature of gene actions governing drought sensitive traits can be exploited by selecting proper breeding strategies for drought tolerance. Methodology for screening drought tolerance in coconut Drought tolerant coconut palm can be selected at seedling stage in nursery and at maturity stage. Apart from these, one can use in vitro screening technique as well. The screening of coconut germplasm can be done using morphological, anatomical, physiological and biochemical traits (see Fig. 2). Molecular marker-assisted selection could also be used once developed. It is essential to note that one has to develop the threshold levels for development of stress in a given climatic and soil conditions. 292 COCONUT GENETIC RESOURCES Fi gu re 1 . M ec ha ni sm o f d ro ug ht to le ra nc e in c oc on ut „ L ea f w at er p ot en ti al + „ St om at al re si st an ce + „„„„ „ T ra ns pi ra ti on al r at e- „ E pi cu ti cu la r w ax + „ L ea f l et th ic kn es s+ „ C ut ic le th ic kn es s+ „ Su b- st om at al c av it y+ „ Pa lis ad e ce ll si ze + „ Sp on gy c el l s iz e+ „ N et a ss im ila ti on ra te + „ W at er u se e ff ic ie nc y+ „„„„ „ Fv /F m + - y ie ld o f c hl or op hy ll fl uo re sc en ce „ Su pe ro xi d e d is m ut as e+ „ Pe ro xi d as e+ „„„„ „ C at al as e+ „ Po ly ph en ol o xi d as e- „ A ci d p ho sp ha ta se - „ L a sp ar ta te 2 -o xo gl ut ar at e am in o tr an sf er as e+ „„„„ „ N it ra te re d uc ta se + „ L ip id p er ox id at io n- „ T ot al d ry m at te r+ „ V eg et at iv e dr y m at te r+ „„„„ „ R ep ro du ct iv e dr y m at te r+ „ H ar ve st in d ex + Po si ti on in d ro ug ht -t ol er an t t yp es : + hi gh /i nc re as ed ; - lo w /d ec re as ed So lu te a cc um ul at io n „ T ot al s ol ub le s ug ar + „„„„ „ A m in o ac id + W at er b al an ce Le af a na to m y Ph ot os yn th et ic e ff ic ie nc y T ur gi d it y m ai nt en an ce D R O U G H T T O L E R A N C E D ry m at te r p ro du ct io n O sm ot ic a dj us tm en t C el l m em br an e in te gr ity ÏÏÏÏ Ï ÏÏÏÏ Ï ÏÏÏÏ Ï ÏÏÏÏ Ï ÏÏ ÏÏÏÏÏ ÏÏÏ ÏÏ ÏÏÏ ÏÏ ÏÏÏÏÏ ÏÏÏ Ï Ï Ï Ï 293 CHAPTER 5: Germplasm use Fi gu re 2 . Sc he m at ic d ia gr am fo r s cr ee ni ng d ro ug ht to le ra nt c oc on ut g er m pl as m Se ed lin g sc re en in g G er m p la sm s ow n in n u rs er y (m ai nt ai ne d u nd er n on -w at er st re ss c on d it io ns ) S cr ee n in g fo r d ro u gh t t ol er an ce in c oc on u t A d u lt p al m s cr ee ni ng G er m p la sm b lo ck s (U nd er ra in fe d c on d it io n) O bs er va ti on s on p ar am et er s lik e st om at al co nd uc ta nc e, le af w at er p ot en ti al s, E C W , l ip id p er ox id at io n, p ho to sy nt he ti c ra te s, tr an sp ir at io n ra te s, W U E , e nz ym es li ke S O D , P P O , c at al as e, P O , N R , l ea f a na to m ic al tr ai ts , c ro w n m or p ho lo gy , n u t yi el d (o nl y in a d u lt p al m s) , s oi l m oi st u re , m et eo ro lo gi ca l d at e Su rv ey fo r d ro u gh t- to le ra nt pa lm s (G ar d en s u nd er n o m an ag em en t f or d ec ad es ; ra in fe d ; d ro ug ht p ro ne a re as ) Se le ct p al m s ba se d o n ob se rv at io ns o n cr ow n m or p ho lo gy (f ro nd n u m be r sh ou ld b e > 3 2, r eg u la r be ar in g w it h 12 b un ch es o n cr ow n, m or e nu m be r of fe m al e fl ow er s, n u t y ie ld ). R ec or d in g of p ar am et er s lik e st om at al c on d u ct an ce , l ea f w at er p ot en ti al s, E C W , ph ot os yn th et ic r at es , tr an sp ir at io n ra te s, W U E , so il m oi st ur e, m et eo ro lo gi ca l d at e W it hh ol d th e ir ri ga ti on in nu rs er y fo r ~ 2 0 d ay s O bs er va ti on s on p ar am et er s lis te d a bo ve In v it ro s cr ee ni ng (U si ng P E G o r N aC l a s os m ot ic um ) Su rv iv al c ap ac it y, p hy si ol og ic al ac cl im at iz at io n an d vi go u r of p la nt le ts Ir ri ga te n u rs er y fo r ab ou t 1 5 d ay s R an ki ng o f c ul ti va rs fo r su rv iv al c ap ac ity an d s ta bi lit y O bs er va ti on s on ab ov e p ar am et er s d ur in g p os t m on so on an d d ry p er io d s In s it u- to le ra nt p al m s U se to le ra nt o ne s as p ar en ta l m at er ia l f or b re ed in g d ro u gh t t ol er an t v ar ie ty D ro u gh t- to le ra nt c oc on u t h yb ri d /v ar ie ty Id en ti fi ca ti on o f to le ra nt li ne s E st ab lis hm en t o f th re sh ol d le ve l f or w at er s tr es s d ev el op m en t; p ro gr es si ve in cr ea se in w at er E va lu at io n fo r yi el d a nd to le ra nc e st ab ili ty 294 COCONUT GENETIC RESOURCES Identification and characterization of in situ tolerant palms The plants that can withstand the natural occurrence of drought and other stresses and still produce good yields are of premium value, as they are likely to possess desirable genes. Surveys in hotspot areas were conducted to identify the palms yielding very high compared to others in their vicinity. Palms at different agroclimatic regions were identified in farmers’ plots with desirable canopy shape and leaf number with good yields. The physiological WUE of these palms was also found to be high (Naresh Kumar et al. 2002b). This type of in situ tolerant plants with desirable traits should be used in breeding programmes, to reduce the time gap in breeding for drought tolerant cultivars in coconut (Naresh Kumar et al. 2002b). Molecular markers for drought tolerance traits In coconut, the development of molecular markers for drought tolerance is in its infancy. The work on stress responsive proteins is being carried out at CPCRI (Naresh Kumar 2003). Efforts are on to link the drought tolerance to molecular diversity to find putative molecular markers, which can be useful for marker-assisted selection (MAS). Although the lack of a viable regeneration technique is a bottleneck for genetic engineering of coconut palm, the molecular markers should be identified for use in large- scale rapid screening of germplasm. This will not only increase the efficiency for selection of parental material but will also reduce the gestation period for breeding improved varieties with drought tolerance (Batugal 1999). The RFLP analysis indicated that Tall and Dwarf ecotypes from Pacific and Far East Asia were different from those from India, Sri Lanka and West Africa (Lebrun et al. 1998, 1999). An in vitro screening technique was developed using NaCl as the osmoticum at different concentrations in coconut embryo culture medium (Karunaratne et al. 1991). It is possible to link the in vitro and nursery screening techniques to molecular techniques for development of molecular markers. Once the markers are established, they will be of prime importance to identify the parental material in breeding for drought tolerance. At the same time, it is essential that the stability of drought tolerance through pheno-phases should also be established. Thus, the development of molecular markers and application of biotechnological tools for the improvement of drought tolerant coconut varieties need more emphasis and concerted efforts. The future challenge is in overcoming the bottlenecks in the use of genetic engineering for the development of drought tolerant coconut variety. 295 CHAPTER 5: Germplasm use Constraints and opportunities Drought is a major constraint for coconut productivity in the entire coconut growing area at global level. The realization of the impact of drought on coconut yield increased attention towards this problem. A methodical research approach led to understanding the drought tolerance mechanism in coconut. So far, conventional breeding strategies were applied for the development of drought tolerant varieties/hybrids. However, this takes a lot of time and so is testing for yield stability under stressful conditions. The lack of regeneration techniques handicapped the genetic engineering approach to impart drought tolerance in high yielding cultivars. Hence, it is very important that a globally-coordinated breeding programme for drought tolerance be set in place, as studies indicated that hybrids with Talls as parents can perform better under water stress conditions (Pomier and de Taffin 1982; Rajagopal et al. 1990). It is essential to conserve the natural desirable gene pools present in farmers’ fields before they become extinct. These materials are highly valuable for crop improvement programme. Molecular markers need to be developed for rapid screening of coconut germplasm for drought tolerance at global level. Further, it is important to characterize the nature and intensity of drought in different coconut growing areas in order to develop suitable drought management strategies. Conclusion The results obtained so far indicate that variation exists among the Talls, Dwarfs and hybrids for drought tolerant traits. Generally, Talls and hybrids with Tall as mother palm have higher drought tolerance compared to Dwarfs and hybrids with Dwarf as mother palm. The heterosis for desirable traits can be exploited for breeding drought tolerant varieties. Further, in situ tolerant palms should be identified and used in breeding programme. These experiments can be extrapolated to other germplasm sources, which were not studied so far, and for making promising cross combinations. Since this requires a comprehensive study, a global research network on this topic will facilitate the development of varieties/hybrids with high drought tolerance and stable yield. References Batugal, PA. 1999. The role of international cooperation in the development of biotechnology in coconut. Pp.19-30. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardena and JM Santamaria (eds). Current advances in coconut biotechnology. Kluwer Academic Pub., The Netherlands. 296 COCONUT GENETIC RESOURCES Bhaskara Rao, EVVB, RV Pillai and Jacob Mathew. 1991. Relative drought tolerance and productivity of released coconut hybrids. Pp. 44 -149. In: EG Silas, M Aravindhakshan and AI Jose (eds). Coconut breeding and management. KAU, Vellanikkara 680 654. Thrissur, India. Bonneau, X and K Subagio. 1999. Coconut growing in zones at risk of drought. Plantations Recherche Développement 6: 432-442. Bonneau, X, R Ochs, WT Kitu and Yuswohadi. 1993. Le chlore: Un element essential de la nutrition minerale des cocotiers hybrids dans le Lampung (Indonesie). Oleagineux 48: 179-189. Bonneau, X, D Boutin, R Bourgoing and J Sugarianto. 1997. Le chlorure de sodium, frtilisant ideal du cocotier en Indonesie. Plantations, recherché, development 4: 336-346. Bopaiah, MG, CC Biddappa and V Rajagopal. 1996. Is common salt an alternative to potassium nutrition in coconut? Indian Coconut Journal 26 (11): 5-7. Braconnier, S and X Bonneau. 1998. Effects of chlorine deficiency in the field on leaf gas exchanges in the PB 121 coconut hybrid. Agronomie 18: 563-572. Braconnier, S and JD Auzec. 1985. Anatomical study and cytological demonstration of potassium and chlorine fluxes associated with oil palm and coconut stomatal opening. Oleagineux 40: 547-551. Braconnier, S and JD Auzec. 1990. Chloride and stomatal conductance in coconut. Oleagineux 45: 259-265. Chempakam, B, KV Kasturi Bai and V Rajagopal. 1993. Lipid peroxidation in relation to drought tolerance in coconut (Cocos nucifera L.) Plant Physiology & Biochemistry 20(1): 5-10. Child, R. 1974. Coconut 2nd ed. Longman, London. Pp. 335. Coomans, P. 1975. Influence des facteurs climatiques sur les fluctuations saisonnieres et annuelles de la production du cocotier. Oleagineux 30: 153-159. de Nuce de Lamothe, MW and G Benard. 1985. L’hybride de cocotier PB 121 (ou MAWA ) (NJM x GOA). Oleagineux 40: 255-266. Fernando, WMU. 1987. San Ramon – Big with promise. Coconut Bulletin 4: 15. Fernando, WMU and G Gajanayake.1997. Patterns of isozyme variations in coconut (Cocos nucifera L.) populations used for breeding improved varieties. Plantations, recherché, développement 4: 256-261. Jayasekara, C, CS Ranasinghe and DT Mathes. 1993. Screening for high yield and drought tolerance in coconut. Pp. 209-218. In: MK Nair, HH Khan, P Gopalasundaram and EVV Bhaskara Rao (eds). Advances in coconut research and development. Oxford & IBH Publishing Co. Pvt. Ltd., New Delhi. 297 CHAPTER 5: Germplasm use Juma, MA and RKW Hornung. 1997. Effects of induced water stress on coconut leaf stomata. Pp. 320-325. In: CP Topper, PDS Caligari, AK Kullaya, SH Shomari, LJ Kasuga, PAL Masawe and AA Mpunami (eds). Proceedings of the International Cashew and Coconut Conference: Trees for Life - The key to development, Dar es Salaam, Tanzania, 17-21 February 1998. BioHybrids International Ltd, Reading, UK. Juma, M and R Fordham. 1998. The effect of environmental stress on coconut (Cocos nucifera L.) growth in Zanzibar. Pp. 342-347. In: CP Topper, PDS Caligari, AK Kullaya, SH Shomari, LJ Kasuga, PAL Masawe and AA Mpunami (eds). Proceedings of the International Cashew and Coconut Conference: Trees for Life - The key to development, Dar es Salaam, Tanzania, 17-21 February 1998. BioHybrids International Ltd, Reading, UK. Karunaratne, S, S Santha and A Kovoor. 1991. An in vitro assay for drought tolerant coconut germplasm. Euphytica 53: 25-30. Kasturi Bai, KV. 1993. Evaluation of coconut germplasm for drought tolerance. Ph.D. Thesis, Mangalore University, Mangalore, India. 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Genetic Diversity in coconut (Cocos nucifera L.) revealed by restriction fragment length polymorphism (RFLP) markers. Euphytica 101: 103-108. Lebrun, P, L Grivet and L Baudouin 1999. Use of RFLP markers to study the diversity of the coconut palm. Pp 73-87. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardena and JM Santamaria (eds). Current advances in coconut biotechnology. Kluwer Academic Pub, The Netherlands. Mathes, DT. 1988. Influence of weather and climate on coconut yield. Coconut Bulletin 5 (1): 8-10. Milburn, JA and MH Zimmermann. 1977. Preliminary studies on salt flow in Cocos nucifera, L. 1. Water relations and xylem transport. New Phytology 79: 535-541. Murray, DV. 1977. Coconut palm. Pp. 384- 407. In: TA Alvim and TT Kozlowski (eds). Ecophysiology of tropical crops. Academic Press, New York. Nainanayake, NPAD and DC Bandara. 1998. Effect of water stress on coconut (Cocos nucifera L.) seedlings under different soil types and compaction levels. Tropical Agricultural Research 10: 12-26. Nainanayake, NPAD, DC Bandara and SP Nissanka. 2000. Root shoot relationships: An effective indicator of soil compaction and water stress for coconut (Cocos nucifera L.) seedlings. Tropical Agricultural Research 12: 151-162. Naresh Kumar, S. 2002. Agro-meteorology and drought management in coconut. In: Training manual on drought stress and its management in plantation crops. Central Plantation Crops Research Institute, Kasaragod, India. Naresh Kumar, S. 2003. CPCRI, Annual Report, 2002-2003. CPCRI Pub., Kasaragod, India. Pp. 90. Naresh Kumar, S, V Rajagopal and Anitha Karun. 2000. Leaflet anatomical adaptations in coconut cultivars for drought tolerance. Pp. 225-229. In: N Muralidharan and R Raj Kumar (eds). Recent advances in plantation crops research. Allied Pub. Ltd., New Delhi, India. Naresh Kumar, S, V Rajagopal, RH Laxman and HP Maheswarappa. 299 CHAPTER 5: Germplasm use 2002a. Photosynthetic characteristics and water relations in coconut palm under drip irrigation on sandy and laterite soils. Pp. 116-120. In: P Rethinam, HH Khan, VM Reddy, PK Mandal and KSuresh (eds). Plantation crops research and development in the new millennium. Coconut Development Board, Kochi. Naresh Kumar, S, V Rajagopal., T Siju Thomas, Vinu K Cherian, M Hanumanthappa, B Anil Kumar, Srinivasulu and D Nagvekar. 2002b. Identification and characterization of in situ drought tolerant coconut palms in farmers’ fields in different agro-climatic zones. In: Abstracts of PLACROSYM XV, 10-13 December 2002, Mysore, India. Naresh Kumar, S, V Rajagopal., T Siju Thomas and Vinu K Cherian. 2003. Influence of soil moisture conservation on coconut (Cocos nucifera L.) under different agro-climatic conditions. Pp. 572. In: Proceedings of the 2nd International Congress on Plant Physiology, 8-12 January 2003, IARI, New Delhi, India. Abs S13-P25. Ollagnier, M, R Ochs, M Pomier and G de Taffin. 1983. Effect of chlorine on the hybrid coconut PB 121- in the Ivory Coast and Indonesia- Growth, tolerance to drought, yield. Oleagineux 38 (5): 309-321. Passioura, JB. 1977. Grain yield, harvest index and water use of wheat. Journal of Australian Institute of Agricultural Science 43: 117-121 Passioura, JB. 1986. Resistance to drought and salinity: avenues for improvement. Australian Journal of Plant Physiology 62: 199-217. Passos, EEM and JV da Silva. 1991. Determination de I’ etat hydrique du cocotier par la methode dendrometrique. Oleagineux 46 (6): 233- 237. Peiris, TSG and RRA Peries. 1993. Effects of bimonthly rainfall on coconut yield in the low country intermediate zone (IL1 ) of Sri Lanka. Cocos (1991-1993) 9. 1-11. Peiris, TSG, RO Thattil. 1988. The study of climate effects on the nut yield of coconut using parsimonious models. Journal of Experimental Agriculture 34: 189-206. Peiris, TSG, RO Thattil and R Mahindrapala. 1995. An analysis of effect of climate and weather on coconut (Cocos nucifera). Journal of Experimental Agriculture 31 (4) 451-460. Pomier, M and G de Taffin. 1982. The tolerance to drought of some coconut hybrids. Oleagineux 37 (2): 55-62. Quencez, P and G de Taffin. 1981. Relation between potassic nutrition and rainfall in oil palm and coconut growing. Oleagineux 36: 1-7. Rajagopal, V, KD Patil and B Sumathykutty Amma. 1986. Abnormal stomatal opening in coconut palms affected by root (wilt) disease. Journal of Experimental Botany 37: 1398-1405. Rajagopal, V, S Shivashankar, KV Kasturi Bai and SR Voleti. 1988. Leaf 300 COCONUT GENETIC RESOURCES water potential as an index of drought tolerance in coconut (Cocos nucifera L.). Plant Physiology and Biochemistry 15 (1): 80-86. Rajagopal, V, A Ramadasan, KV Kasturi Bai and D Balasimha. 1989. Influence of irrigation on leaf water relations and dry matter production in coconut palms. Irrigation Science 10: 73-81. Rajagopal, V, KV Kasturi Bai and SR Voleti. 1990. Screening of coconut genotypes for drought tolerance. Oleagineux 45 (5): 215-223. Rajagopal, V, S Shivashankar and KV Kasturi Bai. 1991. Physiological and biochemical basis of coconut production. Cord Journal VII (2): 12-30. Rajagopal, V, KV Kasturi Bai, RV Pillai and K Vijayakumar. 1992. Relationship between physiological characters and nut yield in coconut genotypes under rainfed condition. Journal of Plantation Crops 20 (Suppl): 277-283. Rajagopal, V, S Shivashankar and Jacob Mathew. 1996. Impact of dry spells on the ontogeny of coconut fruits and its relation to yield. Plantation Research and Development. 3 (4): 251-255. Rajagopal, V, KV Kasturi Bai and S Naresh Kumar. 2000a. Drought management in plantation crops. Pp 30-35. In: P Rethinam, HH Khan, VM Reddy, PK Mandal and K Suresh (eds). Plantation crops research and development in the new millennium. Abstracts of PLACROSYM XV, 10-13 December 2002, Mysore, India. Rajagopal, V, S Naresh Kumar, KV Kasturi Bai and RH Laxman. 2000b. Day time fluctuations in juvenile coconut palms grown under rainfed and irrigated conditions. Journal of Plant Biology 27(1): 27-32. Rajagopal, V and S Naresh Kumar. 2001. Avenues to improve productivity potential under drought condition- A case study on coconut. Pp. 31-36. In: MB Chetti (ed). Proceedings of the national seminar on the Role of Plant Physiology for Sustaining Quantity and Quality of Food Production in Relation to Environment. Indian Society for Plant Physiology, IARI, New Delhi, India. Rajagopal, V and KV Kasturi Bai. 2002. Drought tolerance mechanism in coconut. BuroTrop 17: 21-22. Ramadasan, A, TK Balakrishnan and V Rajagopal. 1991. Response of coconut genotypes to drought. Indian Coconut Journal 21(2): 2-5. Rao, GSLHVP. 1985. Drought and coconut palm. Indian Coconut Journal 15 (12): 3-6. Repellin, A, C Daniel and Y Zuily Fodil. 1994. Merits of physiological tests for characterizing the performance of different coconut varieties subjected to drought. Oleagineux 49 (4): 155-169. Repellin, A, A D’Arcy Lameta, AT Pham Thi, A Tashakorie and Y Zuily Fodil. 1994. Physiological parameters as screening tools for drought- 301 CHAPTER 5: Germplasm use stress resistant varieties of coconut palm (Cocos nucifera L.). Pp. 299. In: Proceedings of the Societe Francaise Physiologie Vegetale, Colloque Sciences Vegetales, Saint-Malo, 12-14 October 1994, Paris, France. Repellin, A, AT Pham Thi, A Tashakorie, Y Sahsah, C Daniel and Y Zuily Fodil. 1997. Leaf membrane lipids and drought tolerance in young coconut palms (Cocos nucifera L.). European Journal of Agriculture 6: 25-33. Shivashankar, S. 1988. Polyphenoloxidase isozymes in coconut genotypes under water stress. Plant Physiology and Biochemistry 15:87-92. Shivashankar, S, KV Kasturi Bai and V Rajagopal. 1991. Leaf water potential, stomatal resistance and activity of enzymes during the development of moisture stress in coconut palm. Tropical Agriculture 68 (2): 106-110. Shivashankar, S and KV Nagaraja. 1996. Water stress induces kinetic changes in the properties of an acid phosphatase isozyme from coconut leaves. Plant Physiology and Biochemistry 23 (1):21-26. Voleti, SR and V Rajagopal. 1991. Extraction and identification of epicuticular wax in coconut. Plant Physiology and Biochemistry 18(2); 88-90. Voleti, SR, KV Kasturi Bai., CKB Nambiar and V Rajagopal. 1993 a. Influence of soil type on the development of moisture stress in coconut (Cocos nucifera L.). Oleagineux 46: 505 - 509. Voleti, SR, KV Kasturi Bai and V Rajagopal 1993 b. Water potential in the leaves of coconut (Cocos nucifera L.) under rainfed and irrigated conditions. Pp. 243-245. In: MK Nair, HH Khan, P Gopalasundaram and EVV Bhaskara Rao (eds). Advances in coconut research and development. Oxford & IBH Publishing Co Pvt. Ltd., New Delhi, India. Wickremaratne, MRT. 1987. Breeding coconuts for adaptation to drought. Coconut Bulletin 4: 16-23. Zhang, J and WJ Davies. 1989. Abscisic acid produced in dehydrating roots may enable the plant to measure the water status of the soil. Plant Cell and Environment 12: 73-81. 302 COCONUT GENETIC RESOURCES Performance of coconut hybrids in some countries of Asia, Africa and Latin America P Batugal COGENT Coordinator and Senior Scientist, International Plant Genetic Resources Institute- Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction Coconut (Cocos nucifera L.) varieties grown worldwide are popularly classified as Tall, Dwarf or Hybrid. The Talls (T) and the Dwarfs (D) are mostly selected races of economic importance among the local farmers. Most of them evolved from continuing natural or mass selection. The hybrids are mostly produced from intercrossing these selected races or traditional varieties (i.e., D x T, T x D, T x T) to develop the desired ideotypes, which for most breeders meant varieties with broad adaptability, pests and disease resistance, and high yield. Promising hybrids A few capable national coconut breeding programmes in coconut growing countries, either on their own or through foreign-assisted projects, have been in the forefront of collecting, conserving, evaluating and breeding coconut germplasm since the early 1980s. Each of these country programmes has produced their own set of recommended or promising hybrids. A survey of the performance of some of these hybrids was conducted by the International Coconut Genetic Resources Network (COGENT) (Batugal 2004), and the results of this survey are summarized and analyzed below. China The Wenchang Coconut Research Institute’s sole recommended hybrid is a cross between Malayan Yellow Dwarf (MYD) and the local Hainan Tall (HAT) variety. This MYD x HAT hybrid (WY78F1) exhibited early flowering (3-4 years) and 3-4 fold increase in terms of harvested nuts (80/palm/year) and copra (4 t/ha/year), compared to the Tall parent. The Philippines The Philippine Coconut Authority (PCA) recommended nine hybrids derived from single crosses involving the local cultivars, Catigan Green Dwarf (CAT), Tagnanan Tall (TAG), Baybay Tall (BAY), Laguna Tall (LAG), Bago-Oshiro Tall (BAO), and the introduced varieties, Malaysian Red Dwarf (MRD) and Polynesian Tall (PYT). Most of these recommended 303 CHAPTER 5: Germplasm use hybrids started flowering on the 3rd to 4th year. The average number of nuts per palm ranged from 117 to 155 and copra yield per hectare, from 4-6 tonnes. The local tall BAY was comparatively good producing 114 nuts/palm with a copra yield of 5t/ha. Among the nine hybrids, MRD x TAG (PCA 15-2) and MRD x BAY (PCA15-3) were outstanding giving the highest number of nuts (144-155/palm) and copra yield (6t/ha). Thailand The Chumphon Horticulture Research Centre (CHRC) of the Horticulture Research Institute of Thailand recommends three high-yielding hybrids: Sawi Hybrid No.1 (an introduced hybrid known as PB 121 or MAWA), and the locally developed hybrids Chumphon Hybrid No.60 (Maphrao Yai or Thai Tall x West African Tall) and Chumphon Hybrid No. 2 ( MYD x Thai Tall). A trial comparing the locally developed hybrids with the local Thai Tall (THT) in 1975 showed that THT yielded the least. The recommended hybrids exhibited nut and copra yields ranging from 80- 126/palm and 3.4 4.2t/ha, respectively. Vietnam The Oil Plant Institute (OPI) of Vietnam recommends seven introduced high-yielding hybrids in the country which have significantly outyielded the local Tall (Ta). The introduced hybrids were PB111, PB121, PB 132, PB 141, JVA 1, JVA2 and CRIC 65 with nut production ranging 48-69/ palm in 1996. The local variety Ta yielded 31-35 nuts on the same year. OPI is currently testing six local hybrids in Dong Go Experimental Center (Eo x Ta; Tam Quan x Ta; Tam Quan x BAOT); and in Binh Thanh Experimental Station (MYD x Renell Island Tall; MYD x Palu Tall; and MYD x Ta). Bangladesh The Agricultural Research Institute (BARI) has developed two high- yielding coconut varieties: BARI Narikel –1 and BARI Narikel-2. These varieties are broadly adapted and capable of producing 65-70 nuts/palm throughout Bangladesh. In addition, BARI is recommending two introduced varieties to the country’s coconut growing communities, namely: Sri Lanka Tall (SLT) and Malaysian Yellow Dwarf (MYD). India The Central Plantation Crops Research Institute (CPCRI) has released the largest number (12) of single-cross hybrids among the surveyed countries, involving Chowgat Orange Dwarf (COD), West Coast Tall (WCT), Laccadive Ordinary (LCT), Gangabondam (GBGD), MYD, SS Apricot by KAU (SSAT) and East Coast Tall (ECT). All the hybrids 304 COCONUT GENETIC RESOURCES performed better than the traditional cultivar WCT. The recommended hybrids have reported average nut yields of 98– 156/palm while the WCT has only 80 nuts/palm of recorded yield. COD x WCT (Chandra Sankara), WCT x SSAT (Kera Sowbagya) and WCT x MYD (Kera Sree) produced the highest copra yields (i.e., more than 4 t /ha/year). Sri Lanka The Coconut Research Institute (CRI-SL) has developed two hybrids, [(Sri Lanka Green Dwarf (SLGD) x Sri Lanka Tall (SLT)] and SLT x SR, and a first generation inbred (SLT x SLT) for its national replanting programme. Their yields ranged from 80-125 nuts/palm and 3.6 – 4.0t copra/ha. Hybrids’ nut yields are double that of the usual yield of the local cultivar, Sri Lanka Tall, but their copra content/yields are similar. Vanuatu The Vanuatu Research and Training Centre have produced hybrids involving the local cultivars Vanuatu Tall (VTT) and Vanuatu Red Dwarf (VRD), and the introduced varieties Renell Island Tall (RIT) and Brazilian Green Dwarf (BGD). The Malaysian Red Dwarf (MRD) was also used as a mother palm for crossing with RIT but the resulting hybrids only performed slightly better (in terms of copra yield) compared to the local VTT and were very susceptible to coconut foliar decay (CFD). The BGD crossed with either RIT or VTT produced the best copra yields of 4.4-5.2 t/ha but they were also found to be very susceptible to CFD. The VRD x VTT hybrids had lower copra yields (3.3-3.7t/ha) but were found to be more tolerant against CFD. Both the traditional and improved VTT types had the lowest reported copra yields of 2.6-2.8 t/ha, but comparable with the hybrid MRD x RIT. Côte d’Ivoire The CNRA Marc Delorme Research Station has initially identified seven outstanding hybrids: PB 213 (WAT x RIT), PB 214 (WAT x VTT), PB121 (MYD x WAT), PB 132 (MRD x TAT or Tahitian Tall), PB123 (MYD x RIT) and PB111 (CRD or Cameroon Red Dwarf x WAT).These hybrids flower very early (40-57 months after field planting) under Côte d’Ivoire conditions. Despite early flowering, they produced from 100 to 132 nuts/ palm/year, which is 34% to 138% higher than the population control West African Tall (WAT). Further, their copra yields ranged from 3.15- 4.8t/ha or 86-135% more compared with WAT. Ghana All coconut cultivars in Ghana are considered to be at risk from the Cape St. Paul Wilt disease (CSPWD), a lethal yellowing type of disease. Hence, 305 CHAPTER 5: Germplasm use the coconut breeding programme in the country is geared towards developing hybrids resistant or highly tolerant to CSPWD. There are six cultivars and 21 hybrids being tested in four locations: Cape Three Points, Discove, Agona Junction and Akwidae. These varietal resistance trials are still under observation although some of the test materials were already totally infected by the CSPWD. Tanzania The Mikocheni Agricultural Research Institute (MARI) is currently testing six hybrids with the local East African Tall (EAT) as sole pollinator. Mother palms included the Malayan Green Dwarf (MGD), CRD, Pemba Red Dwarf (PRD), MYD, MRD and improved EAT populations. In addition to determining their yield performance, the F1 progenies are also being evaluated for their resistance to lethal disease and tolerance to drought stress. Mexico Coconut research at the Instituto Nacional de Investigacion Agropecuaria Y Forestal is focused on developing hybrids resistant to lethal yellowing disease (LYD). Initial hybrids were mainly derived from crosses between MYD and improved Pacific Tall populations. Intra population crosses of selected Pacific Tall were also done and these are currently being tested. COGENT, through its CFC-funded multilocation trials, is in the process of determining the suitability of selected hybrids across its member countries (see Batugal et al., this chapter). The inclusion of all promising hybrids, however, is constraint by financial and material resources limiting the number of hybrid entries and location trials. Conditions favouring coconut hybrid performance and use Agroclimatic In a comprehensive hybrid performance assessment study (Rethinam et. al 2004) initiated in 1998 in 10 countries, most of the participating countries reported that, with few exceptions, hybrids generally came into early bearing and exhibited better productivity in the wet zones than in intermediate and dry zones. The result of the study suggested that to maximize the potential of most hybrids, they should be planted under favourable soil and moisture conditions. 306 COCONUT GENETIC RESOURCES Farmers’ preferences As part of the Asian and Pacific Coconut Community (APCC)/COGENT study, farmer respondents in the surveyed countries were asked to indicate their varietal preference and their reasons for their selection. Of the total 381 responses, 55.6% were in favour of hybrids and 28% preferred planting the local and/or selected Talls (Table 1). However, individual countries showed diverse rates of preference for hybrids. In Samoa, all the farmers covered in the survey stated they would grow hybrids, given a second chance. In Thailand, 70% of the farmers remained satisfied with the hybrids and the rest preferred to plant Tall variety. In Indonesia, where hybrids have already spread to a large extent, only 5.56% wanted to plant the same coconut hybrids while 99.44% opted for selected local Talls and locally produced hybrids for planting the next time. High yield, early bearing and good nut size were cited as the main reasons for satisfaction with the hybrids. And the major reasons made known by the farmers for their dissatisfaction with hybrids are their being vulnerable to moisture stress, high input requirement and susceptibility to pests and diseases. Table 1. Farmer’s preferences of cultivars, given a second chance (number of responses) Source: (P Rethinam, P Batugal and F Rognon, 2004) Zone Tall Local/ Selected Hybrids Dwarf Total Wet 31 91 43 165 Intermediate 59 75 8 142 Dry 18 46 10 74 TOTAL 108 212 61 381 Narrowing the technology gap Although hybrids are generally known to perform better than the traditional varieties, they are currently being grown in limited areas, less than 0.1 (or even nil) to 14% of cultivated coconut farms in various countries (Table 2). The poor adoptions of hybrids are commonly attributed to inadequate information dissemination on the availability of improved hybrids/varieties, lack of adequate supply and affordability of planting materials, and inadequate management and cultural practices. These factors resulted to failure in narrowing down the productivity gap between the farmers’ fields and research stations. Comparing the national yield average in farmers’ fields and those of research centers in 15 coconut growing countries, the estimated technology gap in terms of either nuts or copra yield ranged from 33 to 84% (Table 2). To maximize the potentials of using hybrids to increase the income of resource-poor farmers and the 307 CHAPTER 5: Germplasm use total national coconut productivity, an effective campaign to disseminate suitable planting materials should address the reasons cited earlier for the poor adoptions of hybrids. Table 2. Coconut productivity in farmers’ field and research stations, and area planted to hybrids Source: (P Batugal and J Oliver, 2003) Annual yield (A) Farmers' Fields/National Average (B) Research Station/Hybrids Country Nuts Copra (t ha-1) Nuts Copra (t ha-1) Technology gap [100-(A/B x100)] Area grown to hybrids (% of production area ) South Asia Bangladesh 21/palm 69/palm 70 nil India 6892/ha 23 700/ha 71 14 Sri Lanka 42/palm 63/palm 33 11 Southeast/East Asia Indonesia 1.1 3.5 69 5 Malaysia 10 000/ha 23 000/ha 57 n.d. Philippines 0.78 4-6 84 n.d. Thailand 1.2-1.5 3.0 55 10 Vietnam 38-40/palm 55-80/palm 42 <0.1 China 1.27 3.6 65 1.5 South Pacific Fiji 0.3-0.5 2.0 80 <5 PNG 0.66 2.8-3.6 80 n.d. Africa Ghana 20/palm n.d. 3 Tanzania 40/palm 80/palm 50 n.d. LAC Jamaica 0.8 3.7 78 n.d. Mexico 0.65 4.0 84 1 Conclusion The country reports on recommended hybrids (Batugal 2004) and the APCC surveys on the performance of high-yielding hybrids and farmers’ varietal preferences indicated that there is no universal hybrid and that, generally, hybrids perform better than traditional varieties under good rainfall and soil conditions. Based on these analyses, national breeding programmes should be designed to develop and provide either varieties or hybrids that suit specific agroecological conditions and small-scale farmers’ needs. In the end, each national coconut breeding programme should be able to propose to farmers a set of well-evaluated varieties including Dwarfs, Talls, and Hybrids. COGENT is proposing a global breeding programme to address the collective needs of COGENT member countries instead of merely those of individual countries and the adoption of participatory plant breeding approach to incorporate farmers’ varietal preference. The programme aims to significantly increase the choice of hybrid cultivars among coconut growing countries, by maximizing the use of available genetic resources 308 COCONUT GENETIC RESOURCES for breeding purposes, facilitating the development of efficient breeding tools and varietal selection, and improving the quality of the planting materials for distribution to users or farmers. References Batugal, P. 2004. Country survey (2001-2003): Proposed globally coordinated breeding programme. COGENT. IPGRI-APO, Serdang, Selangor, Malaysia. (Unpublished). Batugal, P and V Ramanatha Rao (eds). 1998. Coconut breeding. Papers presented at a workshop on Standardization of Coconut Breeding Research Techniques, 20-25 June 1994, Port Bouet, Cote d’Ivoire. IPGRI-APO, Serdang, Selangor, Malaysia. 150p. Batugal, P and J Oliver (eds). 2003. Poverty reduction in coconut growing communities Volume I: The framework and project plan. IPGRI-APO, Serdang, Selangor, Malaysia. 337p. Rethinam, P, P Batugal and F Rognon. 2004. Performance evaluation of coconut varieties and farmers’ varietal preferences. APCC, Jakarta, Indonesia and COGENT, IPGRI-APO, Serdang, Selangor, Malaysia. (Unpublished). 309 CHAPTER 5: Germplasm use Performance evaluation of coconut varieties and farmers’ varietal preferences P Rethinam1, P Batugal2 and F Rognon3 1Executive Director, Asian and Pacific Coconut Community (APCC), Jakarta, Indonesia 2COGENT Coordinator and Senior Scientist, International Plant Genetic Resources Institute, Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia 3Director, Bureau for the Development of Research on Tropical Perennial Oil Crops (BUROTROP), Montpellier, France Introduction Coconut, (Cocos nucifera L.) has two forms, the Talls and Dwarfs. Predominantly, in all coconut growing countries, Tall varieties are commercially grown for copra and oil. Dwarfs are primarily grown in a limited area for ornamental purpose as well as for its sweet tendernut water for drinking. Talls are highly cross pollinated and hence, the variations in nuts are spectacular. Dwarfs are mostly self pollinated. Studies on varietal improvement using the existing germplasm were taken up in many countries over a period of more than seven decades. This has resulted in the identification of many high yielding varieties and hybrids of different Tall and Dwarf combinations. Inter and intra-varietal crosses were made to develop progenies with combined desirable characteristics of parents and over-dominant traits particularly on yield performance. In India, the first report on hybrid vigour in progenies resulting from the crosses between Tall and Dwarf varieties was available in 1932. In Sri Lanka, the results of exploratory crosses became available in 1948. In both these countries, organized production and distribution of hybrid planting material began in the early fifties. Similar programmes were taken up in the seventies by other countries especially Indonesia, Côte d’Ivoire, Malaysia, Philippines and Vanuatu. Although many high yielding varieties and hybrids were developed and commercial seed production programmes were started, there were always some reservations by small holders of coconut in using them as planting materials. In order to assess the extent of adoption of these varieties and hybrids by farmers, identify the constraints experienced by the farmers in adopting them under field conditions and further assess the yield performance of these hybrids and varieties, three studies were conducted during the year 1980, 1988 and 1998 in different coconut growing countries. 310 COCONUT GENETIC RESOURCES High-yielding varieties and hybrids Among the traditionally cultivated Tall (T) varieties, there are genetically superior palms possessing intrinsic traits for high yield. Such elite palms have been identified in some countries and used for seedling production and for intravarietal crosses. Similarly, these palms have also been used either as pollen parents or female parents in intervarietal crosses with Dwarf (D) varieties. The resulting progenies were therefore, either T x D hybrids or D x T hybrids depending on the pistillate parent involved. Sri Lanka was the first country to produce T x T hybrids from isolated seed gardens planted with progenies of selected T x T crosses. Natural cross pollination was permitted to take place between the planted progenies without resorting to emasculation. The T x T hybrid so produced is known by the name CRIC 60 which is superior to the local Tall cultivars. The hybrid palms commence flowering at 5-8 years from planting and yield about 100 nuts per palm per annum. The copra content per nut is about 200g. They are generally hardy palms and tolerant to drought, pests and diseases and are recommended for planting in all districts in Sri Lanka. The hybrid CRIC 65 has been released to the farmers in the sixties. This hybrid flowers in 3-4 years from planting, yields about 120 nuts per palm per year with a copra content of 200-215g per nut. As it is sensitive to environmental changes, it has been recommended for planting in home gardens (Wickramaratne 1989). In India, the production is mostly T x D hybrids and the different forms of Dwarf variety used are the local Orange Dwarf (COD), Green Dwarf (CGD) Gangabondam (GB) and the Malayan Yellow Dwarf (MYD) which had been earlier introduced in the country. The Tall parents belong to West Coast Tall (WCT), Lakshadweep Ordinary (LO), Andaman Ordinary (AO) and East Coast Tall (ECT). T x T hybrids and D x T hybrids are presently produced only in small numbers. The performance of the released coconut hybrids in India were significantly better than the West Coast Tall in terms of nut yield per palm per year ( 98-140), and except for ECT x MYD, copra yield per palm (16-25 kg) (Nair et al. 1996). In Indonesia, four improved intravarietal hybrids (Kelapa Baru or KB1, KB2, KB3 and KB4) produced by crossing selected Tall cultivars in Kima Atas Experimental Garden have been released. The hybrids started flowering on the 6th year, with total bunches of 16/year, nuts 96-124 palm/year, copra yield 3.88-4.66 t/ha and oil content of 67-71percent. Another three D x T hybrids evolved from the local material: Khina 1 (Nias Yellow Dwarf x Tenga Tall), Khina 2 (Nias Yellow Dwarf x Bali Tall) and Khina 3 (Nias Yellow Dwarf x Palu Tall) have been released. These hybrids have been found to be superior to the parental types with respect to precocity for bearing and high production of copra (Liyanage et al. 1986). 311 CHAPTER 5: Germplasm use In Cote d’Ivoire, a number of promising hybrids were developed since 1965. Some of the promising D x T combinations produced are Port- Bouet or PB 121 (Malayan Yellow Dwarf x West African Tall), PB 111 (Cameroon Red Dwarf x West African Tall), PB 132 (Malayan Red Dwarf x Polynesian Tall) and PB 122 (Malayan Yellow Dwarf x Polynesian Tall). In the case of PB 121, a yield level of 5.5 t/ha of copra has been recorded. The studies on the comparative performance of the different hybrids conducted at the Marc-Delorme Research Station in Ivory Coast have shown that the productivity of PB 132 is equivalent to that of PB 121 and twice that of West African Tall (Sangare et al. 1988). The other promising hybrids PB 122 and PB 111have also been reported to be more precocious and higher yielding than the other local hybrids. The best PB hybrids initiated first flowering 44-57 months after planting, with number of bunches per palm ranging from 12-16, 101-132 nuts/palm, 212-311 g copra/nut and 3.15-4.8 t copra/ha ( Bourdeix et al. 1993). The T x T combinations such as PB 214 (West African Tall x Vanuatu Tall) and PB213 (West African Tall x Rennel Tall) were also found to be higher yielding than the West African Tall (De Nuce 1989). In Malaysia, varietal studies commenced in 1920s with Malayan Dwarf forms which showed that Green Dwarfs are robust, resistant to adverse conditions and produce the best quality copra (Jack and Sands 1929 and Jack 1937). The Malayan Green was capable of producing 11.3 kg copra per palm per year, nearly as much as the average Tall palm. MAWA hybrids ( Malayan Yellow, Orange or Red Dwarf x West African Tall combinations) were less variable in terms of nuts/palm, fruit weight and copra weight and, the copra produced has been higher than other hybrids. These MAWA hybrids have been used in the National Planting Programme since 1978. The Malayan Red and Yellow Dwarf x West African Tall have been found to be high yielding (25.82 and 24.98 kg copra/palm, respectively) and the most suitable planting material particularly in the imperfectly drained and highly fertile coastal plains (Chan 1983). In Papua New Guinea, hybrid seed production was started in 1973. Maren (Malayan Red Dwarf x Rennel Tall ) hybrid was released to the farmers in the seventies (Turner 1989). The cumulative yield up to 1982 for Maren was estimated at 11.08 t/ha. However, its field performance has been disappointing to the farmers. It is highly susceptible to the local forms of Rhynchophorus weevil, Scapanes and Rhinoceros beetles. Moreover, the genetic base is also very narrow. In the Philippines, the hybrids that evolved from the local parental material, namely Catigan Green Dwarf x Laguna Tall and Catigan Green Dwarf x Tagnanan Tall, have been found to be as productive as PB 121 312 COCONUT GENETIC RESOURCES introduced from Côte d’Ivoire. The local hybrids produce bigger sized nuts than PB 121 and also exhibit buffering ability against environmental stress. The local hybrid Catigan Green Dwarf x Laguna Tall has been released as PCA- 15-1 for mass adoption (Santos 1989). In Jamaica, crop improvement studies were started since 1950 with more emphasis on evolving resistant strains for lethal yellowing disease. All the three Malayan Dwarfs were found to possess a high degree of resistance and only a very small percentage of MYD has been affected by lethal yellowing. The D x T hybrids involving the Malayan Yellow, Green or Red Orange Dwarf and Panama Tall as parents have been found to inherit sufficient immunity to the lethal yellowing disease and also have the capacity for high yield. This hybrid is locally called Maypan (Harries 1971). In Samoa, Malayan Red Dwarf x Rennel Tall hybrid was produced on a large scale in a coconut seed garden established in 1977 (Efu 1989). Under the FAO Project, the production of open-pollinated MRD x Samoa local Tall was undertaken at Aela where Dwarf palms were planted with local Talls in the surrounding areas. The natural hybrids found in Aela were planted in three locations along with Samoan Tall in 1980. In Thailand, among the different hybrid combinations tested, PB 121 (MYD x WAT) has displayed greater precocity and has also been found to give more uniform yields than selected Thai Talls. PB 121 was released as Sawi 1 to farmers for general cultivation. Both MYD x WAT and Thai Tall x WAT have been found superior to Thai Tall variety. Two more hybrids, Chumphon Hybrid No. 60 (Maphrao Yai x WAT) and Chumphon Hybrid No. 2 (MYD x Maphrao Yai), were released in the country in 1987 and 1995, respectively. In Vanuatu, a coconut improvement programme was started during 1962. A number of combinations involving different Talls like Vanuatu Tall, Samoan Tall, Rennel Tall and Rotuman Tall were produced and their field performance was studied under different locations (Calvez 1989). One of the promising combinations, Vanuatu Tall (VTT) x Rennel Tall (RLT), is under field test for tolerance to Foliar Decay disease. Vanuatu Red Dwarf (VRD) x VTT has also been planted in 1984. Initial trials have indicated that T x T hybrids might be superior. In Tanzania, production of the hybrids MAWA (MYD x WAT) and Camwa (CRD x WAT) was initiated in 1981-1982. However, these hybrids were not different from local East African Tall with respect to tolerance to lethal yellowing and drought. In Vietnam, high yielding hybrid seednuts were imported in 1985 and the seedlings raised from them were planted in seven different adaptability trial sites in various agroecological zones. The objectives were 313 CHAPTER 5: Germplasm use to evaluate the performance of the imported hybrids in comparison with local ones and to establish trial sites that could also serve as demonstration centres before the hybrids are finally selected and planted on a large scale. The Oil Plant Institute (OPI) subsequently imported four hybrids from Côte d’Ivoire and three more hybrids, JVA I and JVA 2 from the Philippines and CRI 65 from Sri Lanka in 1986 and 1987. These hybrids were planted in different agroecological zones. The hybrids planted in 1985 had shown higher economic efficiency than the local varieties under the same planting and maintenance conditions. They always had significantly higher values for total nuts, copra and oil than the local Ta and Dau cultivars. However, higher efficiency was recorded only under favourable conditions of soil, water and maintenance. For the hybrids planted in 1987, flowering and fruiting rates were better than the local varieties (Linh and Long 2000). Among the local varieties, Ta, Dau and Giay have been identified as the best for copra making. A seed garden of 120 ha has also been established at Trang Bang, Tay Ninh Province for producing hybrids and seed material of local varieties. Other countries In Fiji, a seed garden is now producing high yielding hybrids using Malayan Yellow Dwarf or Malayan Orange Dwarf with Rennel Tall or Rotuma Tall. In Tonga, a seed garden has been established to produce hybrids. Similarly in Tuvalu, a small seed garden has been operated to produce hybrids of Malayan Yellow Dwarf and Malayan Orange Dwarf with Rennel Tall or with local coconuts. In the Solomon Islands, where controlled pollination began in the 1960’s, a replanting programme is being carried out with the Malayan Red Dwarf x Rennel Tall hybrid. Findings of 1980 and 1988 assessment of new varieties The Asian and Pacific Coconut Community (APCC) organized two studies in 1980 and 1988. The objective of the first study was to identify the constraints faced by small-scale farmers in the adoption of high yielding varieties with a view to assisting countries to formulate remedial counter measures. The study covered India, Indonesia, Malaysia, Philippines, Papua New Guinea, Solomon Islands, Sri Lanka, Thailand and Western Samoa. The second study in 1988 covered India, Indonesia, Malaysia, Sri Lanka, Papua New Guinea, Philippines and Western Samoa. The objectives of this study were: (1) to identify new technology packages for coconut, (2) to review the state of art of adoption, and (3) to identify reasons for adoption and non-adoption. The findings of these two studies were compiled and released by the APCC (Sumith de Silva 1989). 314 COCONUT GENETIC RESOURCES In India, Indonesia, Philippines, Sri Lanka and Thailand, the number of holdings studied was 223 occupying a total area of 720.9 ha. The average size of a holding in the sample was 3.23 ha. The holdings of less than 2 ha amounted to 58% of the total holdings but occupied only 20.3% of the total area. Holdings above 2 ha and up to 10 ha accounted for 34% of the total holdings with a higher share of 48.4% in the total area. Although holdings above 10 ha formed only 8% of the total holdings, their share in the area was 31%. Among the cultivars studied, Tall varieties accounted for 27.8%, T x D and D x T hybrids comprised 62.6%, T x T hybrids formed 6.2% and the balance of 3.4% was constituted by miscellaneous other strains. Between T x D and D x T, the latter accounted for 86% of the hybrids spread. The average copra yield during the first 8 years of bearing was 907 kg for Tall, 1352 kg for T x D and 1634 kg for D x T. The unit recovery of copra was, however, high in the Tall which was 4764 nuts to a tonne of copra whereas the corresponding numbers for T x D and D x T hybrids were 7651 and 5825 nuts, respectively. Although the hybrids commenced bearing in the third year as against five years for the Tall varieties, the capital cost and also the recurrent cost were much higher for the hybrids. However, the net returns generated by both the hybrids were higher than that by the Tall variety. Based on their field experience, the farmer participants of the study expressed their opinion about each test material. The study recorded 740 responses of satisfaction for varieties and 457 responses of dissatisfaction. While 28.5% of the respondents expressed preference for Tall varieties, around 43% favoured the existing hybrids while another 28.5% desired to have new varieties possessing better qualities. Higher yield, early bearing, ease to harvest, vigour, uniformity, superior quality copra, higher income, etc. were the notable benefits observed by the farmers who were satisfied with the performance of hybrids, particularly the D x T.The reasons expressed for dissatisfaction were: less than expected yield, vulnerability to diseases and pests, small nuts, low prices, low income, etc., in case of hybrids; and low yield, non- uniformity, late bearing and low income in case of Talls. The farmers’ views on new varieties were not significantly different in most countries. In India, of the two hybrids, farmers favoured only D x T. Even in this case, one hundred percent acceptance was not reported by the farmers. Only 50% of the farmers were satisfied with the performance of T x D. More than 50% of the farmer participants of the study preferred the traditional Tall variety for future planting. About 30% preferred D x T and other tested hybrids and the balance desired to have new (not yet tried) varieties. In Indonesia, field survey revealed that 78% of the farmer participants were satisfied with the performance of hybrids mainly due to their early 315 CHAPTER 5: Germplasm use bearing and high yielding characteristics. The remaining respondents were not satisfied because of the higher cost involved in the use of fertilizers and other inputs, the yields realized were less than expected, small size of nuts and immature nut fall. In Malaysia, survey results indicated that the yield increase in terms of number of nuts obtained by the farmers from hybrids was more than 200%. But the corresponding increase in net income was not materialized due to escalating cost of production. Although the Mawa hybrids yielded larger number of nuts, the nut size was small causing higher cost of producing copra on per unit basis. As such, increase in the number of nuts has not produced a proportionate increase in the farmers’ income. In Papua New Guinea, the hybrids were highly susceptible to the local strains of Scapanes and Rhinoceros beetles and Rhynchophorus weevil. The hybrid programme in PNG had to be suspended pending a solution to the pest problem. In the Philippines, about 55% of the farmer respondents expressed satisfaction with hybrids due mainly to early bearing characteristic and compatibility with the environment. The dissatisfied farmers (45%) cited high fertilizer cost and lower realization of yield than expected, as the reasons. In Sri Lanka, both CRIC 65 and CRIC 60 were found suitable only to certain parts of the country. However, the planting material of the hybrids was distributed indiscriminately with the result that the performance was not up to expectation in unsuitable locations. In the field survey, 66.7% of the farmer participants were satisfied with T x T hybrids and only 16.7% were satisfied with the intervarietal hybrids or D x T hybrids. The study conducted in Thailand showed that the majority of the farmer participants favoured the hybrid due to its vigorous growth, uniformity and consistency in yield, early bearing characteristics and its adaptability to a wide range of environment. The dissatisfaction expressed by others was due to smaller fruit size which, due to its low market price, results in low income to the farmers. The hybrid Sawi 1 gave the lowest direct income while the top earner, Thai Tall, generated income double that of Sawi 1. The difference was due to the higher unit income for mature nuts of Talls in the local market. Summary of major findings of the 1998 varietal assessment study A more comprehensive varietal assessment study was initiated in 1998 in the member countries of the Asian and Pacific Coconut Community (APCC) and of the International Coconut Genetic Resources Network (COGENT) with the financial support of COGENT and the Bureau for 316 COCONUT GENETIC RESOURCES the Development of Research on Tropical Perennial Oil Crops (BUROTROP) and APCC. In several countries, either the study was not completed or the final reports were not prepared. Consequently, this paper has been prepared based on the completed reports received from the following 10 countries (examinant countries): Brazil, Cote d’Ivoire, Federal States of Micronesia, Fiji, Indonesia, Jamaica, Malaysia, Mexico, Papua New Guinea, Philippines, Samoa, Sri Lanka, Tanzania, Thailand, Vanuatu and Vietnam. The samples for the study were drawn in each country from three distinct agroclimatic zones: wet, intermediate and dry. Size class distribution of sample holdings The details of the distribution of the sample farms according to size in India, Philippines, Samoa, Sri Lanka, Thailand and Vietnam are given in Table 1. Table 1. Size and class distribution of the sample farms* The average size of the sample farms in the six countries was 3.5 ha with a range of 0.44 to 20 ha. The total area covered by all the farms was 1079.62 ha. The farmers possessing below 2 ha formed over 60% of the sample although their share in the total area was only below 14%. On the other hand, farmers with area above 2 ha but below 8 ha formed around 30% but commanded over 36% of the total area. At the same time, the farmers who had farms of the size between 8 and 20 ha had a share of over 42% in the total area although they formed only 9% of the total number of farmers. Of the total 308 farms in the sample, 124 or 40% were in the wet zone, 125 or 41% fell in the intermediate zone and 59 or 19% represented the dry zone. Age at first fruiting and productivity The age at first fruiting and the annual production of nuts and copra varied between varieties and the agroclimatic zones as shown in Table 2. * In India, Philippines, Samoa, Sri Lanka, Thailand and Vietnam Size Class (ha) No. of Farms Extent (ha) Average Size of Farm (ha) % of Farmers % of Area No. of Farms by Zone Wet Intermediate Dry < 1.0 113 49.29 0.44 23.70 4.57 50 49 14 1-2 73 100.29 1.37 36.69 9.29 30 30 13 2-4 52 162.43 3.12 16.88 15.05 18 17 17 4-6 28 144.39 5.16 9.09 13.37 14 6 8 6-8 13 88.62 6.80 4.22 8.20 4 5 4 8-10 9 82.60 9.17 2.92 7.65 4 4 1 10-20 19 380.00 20.00 6.18 35.20 4 13 2 Above 20 1 72.00 20.00 0.32 6.67 - 1 - Total / Average 308 1079.62 3.50 100.00 100.00 124 125 59 317 CHAPTER 5: Germplasm use The general trend was that the hybrids came into early bearing in the wet zone although exceptions were observed in some countries. Irrespective of the growing zones, the average pre-bearing period for D x T and T X D hybrids observed in India, Sri Lanka and Vanuatu were 4.4 and 4.7 years, respectively. In the case of T x T hybrids in Sri Lanka and Vanuatu, the average fruit initiation was 5.8 years. Selected local Talls in the Philippines, Vanuatu and Vietnam, started bearing fruits at an average of 5.5 years while the Malayan Dwarfs of Jamaica and selected Dwarfs of Vietnam were more precocious with an average bearing age of 3.7 years. In India, the production of nuts and copra was highest for T x D hybrid in the wet zone, but was such only in terms of nuts in the intermediate zone. For D x T hybrid, the production of nuts as well as copra output was much higher in the intermediate and dry zones than in the wet zone. The difference in the production between T x D and D x T revealed the preference of the former for more favourable soil moisture relations. In Indonesia, the performance of Mawa hybrid was recorded in the wet zone and that of Nias Yellow Dwarf hybrid (NYD x WAT) in the intermediate zone. Since their agronomic performances were observed under different zone conditions, their optimum growing conditions for maximum productivity has not been elucidated. The Malayan Dwarf variety and the Maypan hybrid were compared under the different zones in Jamaica. In all the three zones, the Malayan Dwarf outperformed the hybrid in precocity of bearing, as well as in the production of nuts and copra. Productivity in terms of nuts and copra output was also higher for the Malayan Dwarf compared with the Maypan hybrid through all the zones. The Dwarf variety and hybrid showed preference for wet agroclimatic zone to maximize production. In the Philippines, the two Tall types (Laguna and Tagnanan) , and two hybrids (Mawa and PCA 15-1) exhibited better productivity in the wet zone than in the other two zones with the lowest in the dry zone indicating the sensitivity to soil moisture stress. Between the two hybrids, PCA 15-1 was found to be superior to Mawa in all the three zones. One significant observation recorded in the Philippines studies was the better productivity of Laguna Tall compared to the two hybrids in all the three zones. Hence, the Laguna Tall cultivar is a better option for planting in the wet, intermediate and dry zones. In Sri Lanka, the D X T showed preference for the dry zone. When compared with San Ramon x Dwarf, the latter yielded better in the intermediate zone but production figures for the other two zones were not available for this hybrid. The data for T x D hybrid were available 318 COCONUT GENETIC RESOURCES INDIA Wet T x D 4.1 14 832 5962 2.49 D x T 5.0 13 366 6300 2.12 Intermediate T x D 4.4 18 187 7802 2.33 D x T 4.6 17 627 5861 3.00 Dry T x D 4.8 10 111 5925 1.71 D x T 4.5 11 844 5440 2.17 INDONESIA Wet Mawa hybrid 3.8 12 444 5447 2.28 Intermediate NYD x WAT 4.0 11 277 6200 1.82 JAMAICA Wet Tall - 5850 3500 1.67 Malayan Dwarf 3.3 20 588 6277 3.28 Maypan hybrid 4.1 12 775 4125 3.10 Intermediate Tall - - - - Malayan Dwarf 3.3 13 013 5733 2.27 Maypan hybrid 4.5 6230 3785 1.65 Dry Tall - - - - Malayan Dwarf 3.3 18 545 6440 2.88 Maypan hybrid 4.4 11 527 4388 2.63 PHILIPPINES Wet Laguna Tall 6.0 11 214 3528 3.18 Tagnanan Tall 6.0 7342 2738 2.68 Mawa hybrid 4.0 11 351 4087 2.78 PCA15-1 hybrid 4.0 10 624 3368 3.15 Intermediate Laguna Tall 6.0 11 214 4151 2.70 Tagnanan Tall 6.0 6465 3221 2.00 Mawa hybrid 4.0 9849 4809 2.05 PCA15-1 hybrid 4.0 9276 3963 2.34 Dry Laguna Tall 5.0 9528 4744 2.00 Tangnanan Tall 5.0 5460 3704 1.47 Mawa hybrid 4.0 7029 5530 1.27 PCA 15-1 hybrid 4.0 8366 4557 1.84 SRI LANKA Wet T x T 6.2 5 4492 5500 0.82 D x T 4.70 8415 6000 1.40 Intermediate T x T - 10 620 - - San Ramon x D 6.0 12 883 - - D x T 4.0 7952 4360 1.82 T x D 5.5 10 166 4400 2.31 Dry T x T 5.0 9575 5000 1.92 D x T 4.0 9106 - - SAMOA Wet Tall 6.0 4006 5053 0.79 Hybrid 3.5 6694 4687 1.43 Intermediate Tall 5.6 3860 5053 0.76 Hybrid 3.5 7649 4687 1.63 VANUATU Local Tall 5.0 9620 6013 1.60 Improved Tall 5.0 11 840 5382 2.20 T x T 6.0 12 580 4341 2.90 D x T 4.0 23 200 6629 3.50 VIETNAM Wet Tall 5.4 4903 4064 1.21 Dwarf 3.7 19 456 - - Hybrid 4.0 5364 5000 1.07 Intermediate Tall 5.25 7375 4265 1.73 Dwarf 5.0 26 667 - - Hybrid 3.5 6195 5500 1.13 Dry Tall 5.0 5360 5188 1.03 Country & Variety Years to Nuts Nuts/t Copra/ zone bearing per ha copra ha (t) Table 2. Average production and bearing time 319 CHAPTER 5: Germplasm use only for the intermediate zone which showed higher productivity than those of D x T hybrid. The T x T performed well under the intermediate and dry conditions but had the least number of nuts and copra yield among the test materials when planted under the wet zone. In Samoa, the hybrid yield was more than that of Tall in both the wet and intermediate zones. The average productivity of hybrid through the two zones was 7172 nuts or a copra equivalent of 1.53t against 3933 nuts or 0.78 t of copra/ha/year of the Tall. Differential performance of local Tall, improved Tall, T x T and D x T was recorded in Vanuatu for only one zone. The performance of D x T was found to be far superior compared with all the other types in terms of precocity, number of nuts and copra output. The Dwarf types produced significantly more nuts/ha compared with the hybrid and Talls in Vietnam. The Dwarf variety yielded profusely in the wet and intermediate zones but its production figures for the dry zone were not recorded. Further, its fruits were mostly consumed as tendernut. Farmers’ evaluation of coconut cultivars The farmers covered in this study were individually contacted and their observations on each cultivar including their satisfaction or otherwise and varietal preferences were recorded. A total of 851 responses of satisfaction were recorded with 599 or 70% in favour of T x D and D x T hybrids. High yield, early bearing and good nut size, were the major reasons for satisfaction with the hybrids. This was followed by the Dwarf with 128 or 15% favourable responses and the most preferred traits were better resistance to pests and diseases, high yield, easy marketability and early bearing. The Tall cultivars including the T x T hybrids were preferred for thick kernel and cream, high yield and miscellaneous other traits. The number of responses in their favour was 124 or 14.5%. The specific reasons for satisfaction of farmers with each cultivar are shown in Table 3. The study also recorded 339 responses of dissatisfaction with the varieties already experienced by the farmers. Out of the total, 235 or nearly 70% responses of dissatisfaction were with T x D and D x T hybrids and the major reasons made known by the farmers were small size of nuts, bunch buckling, high input requirement, vulnerable to moisture stress, vulnerable to pests and diseases, low yield, less creamy kernel, alternate bearing, etc. There were 67 responses of dissatisfaction with Dwarf variety but only 36 such responses were recorded in the case of Tall variety. The major reasons for dissatisfaction with Dwarf variety were small nuts, difficult to market, pilferage, etc. The major reason expressed against the Tall variety was its late bearing tendency. 320 COCONUT GENETIC RESOURCES Reasons Tall TxD/ TxT Dwarf Total DxT High yield 31 153 8 30 222 Early bearing 11 137 1 23 172 Good nut size - 119 - - 119 Thick kernel/high copra content 40 28 - - 68 Ease of scraping - 11 - - 11 Thick cream with good flavour 10 9 - - 19 High oil content - 8 - - 8 Good toddy yield 6 4 - - 10 Sweet tendernut water - 20 - - - 20 Low incidence of bunch buckling 6 - - - 6 Short & strong fronds - - - - - - Better resistance to pests and diseases 2 48 - 38 88 Medium/short stature- 12 - - - 12 Long life span 6 - - - 6 Low maintenance costs- 1 - - - 1 Easy marketability - 29 - 23 52 Good timber 1 - - - 1 High frequency of harvest - - - 11 11 Others 2 20 - 3 25 Total 115 599 9 128 851 *The Philippines and Vanuatu were not included due to lack of quantified data Varietal preference of farmers The farmers in the surveyed farms were asked to indicate the varieties preferred by them for planting, if another chance arises. To this inquiry, 381 quantified responses were received from India, Indonesia, Jamaica, Samoa, Thailand and Vietnam. The results are given in Table 4. Of the total 381 responses, 55.6% were in favour of hybrids and 28% in favour of the local and / or selected Talls. In Indonesia, where hybrids have already spread to a large extent, only 5.56% of the farmers wanted to plant the same coconut hybrids while 94.44% of the farmers opted for selected local Talls and locally produced hybrids for planting the next time. In India, 59% of the farmers preferred selected Talls and only 41% expressed preference for hybrids. In Sri Lanka, only 53% of the farmers having experience with D x T expressed the desire to plant the same hybrid whereas, 55% of the T x T growers were completely satisfied with the variety and were willing to plant the same if given a second chance. Table 3. Reasons for satisfaction by cultivar (no. of responses)* 321 CHAPTER 5: Germplasm use In Samoa, all the farmers covered in the survey stated that they will grow the hybrid, given a second chance. Out of these farmers, 47% also wanted to grow the local Tall besides the hybrid. In Thailand, 70% of the farmers remained satisfied with the hybrids and the rest preferred to plant Tall variety. The variety preferred by 54% of the farmers in Vietnam for planting, given a second chance, was the local Tall. Only 26% of the farmers wanted to grow hybrid and the balance 20% stated that they would grow the local Dwarf cultivar, Xiem. Table 4. Farmer’s preference of cultivars, given a second chance (no. of responses) Conclusion and recommendations Hybrid seed production has a history of 3-6 decades in most of the coconut growing countries. In countries like India and Sri Lanka, the programme has been continuing since 1940’s and in Côte d’Ivoire it was commenced in 1951. Hybrid combinations were evolved and planted in the research institutes at the first stage and at farmers’ fields later under diverse agroclimatic conditions. The performance of hybrids was good in terms of nut and copra yield besides early bearing, vigorous growth and uniformity in yield, particularly in D x T. The farmers’ response was varied since they are not very clear about the required input management to maximize yields for coconut in general and high yielding varieties and hybrids in particular. The APCC studies of 1988 and 1998 have, however, revealed that the level of acceptance of T x D and D x T hybrids has been lagging below expectation despite their potential for early bearing and higher production. These hybrids exhibited certain undesirable traits which were not appreciated by the farmers. By and large the hybrids under favourable weather and optimum management conditions have performed better than the local Tall. But experience in some countries showed that even under intensive management, the hybrids do not perform better than the local Tall cultivars growing under the same conditions. In Vietnam, the performance of hybrids in terms of copra production under the intensive management category was not as much as the local Tall cultivars by the Zone Local /Selected Hybrids Dwarf Total Tall Wet 31 91 43 165 Intermediate 59 75 8 142 Dry 18 46 10 74 Total 108 212 61 381 322 COCONUT GENETIC RESOURCES eight year after coming into production. The same trend was observed in Sri Lanka. These results highlight the fact that the productivity of some of the hybrids, even under intensive management, is not significantly different than those of selected Tall varieties; and that the production technology of hybrids has to be improved if farmers are to be benefited from their cultivation. Except, perhaps, in Côte d’Ivoire, the production of hybrids in most other countries has been organized without developing adequate populations of strains or ecotypes tested for specific combining ability. The general combining ability of the existing varietal forms is presently the yardstick for the production of hybrid combinations, which invariably show differential performance and exhibit undesirable traits when grown under field conditions. There is a scope for identifying ecotypes possessing superior traits from among the available cultivars and testing them for both general and specific combining abilities with other strains or ecotypes. The populations comprising selfed progenies of those ecotypes, proven to be the best combiners when established, can serve as the basic material for evolving better hybrids. It is not difficult to identify special strains or ecotypes in each country through participatory approach. Among the countries where coconut hybrids have been introduced, Indonesia appears to have taken the lead with having 244 310 ha under hybrids in 1996 with the estimated production of 121 729 t of copra per year, which is only 50% of the average productivity of the Tall variety in that country. Therefore, when farmers in that country are given the opportunity to plant coconuts again, only 5.56% of them wanted to plant the introduced hybrids, while 94.44% of the farmers wanted to plant selected local Talls, T x T hybrids and the locally produced Khina hybrids. The experience in most of the countries is that the intervarietal hybrids perform well only under intensive management hence, their poor adoption by the small- and medium-scale farmers who are not used to high external input farming in coconut. As long as the new hybrids are not amenable to low external input farming, the farmers may not opt for them either for new planting or replanting. Appropriate breeding technology has to be perfected for producing hybrids possessing potential for higher productivity under average management without exhibiting undesirable traits. Based on the results of the trials, the following recommendations are made: 1. Ecotypes possessing favourable traits should be identified in each country through participatory approach. These ecotypes should be crossed with selected cultivars and the best combiners identified through progeny testing. The parents of the promising crosses 323 CHAPTER 5: Germplasm use should be multiplied by selfing to raise a sizeable population of selfed progenies for large-scale reproduction of the best crosses. 2. The performance of high yielding varieties should be satisfying to small and medium-scale farmers under low external input situations. T x T combinations amenable to such situations should be produced by crossing genetically superior Tall palms which normally constitute 3-5% of the palm population in any holding. 3. The Malayan Green Dwarf, which under Malaysian conditions proved to be better than the yellow and red colour forms in productivity, resistance to pests, diseases, and adverse conditions as well as in vigour and robustness, should be utilized as one of the parents in cross breeding. The resulting progenies should be tested for their yield performance. 4. Coconut farmers should be trained on the appropriate care and management of high yielding varieties and hybrids in coconut plantations, and should be educated on the benefits they could get in terms of increased yield and returns. 5. Production of quality planting materials of selected Talls, Dwarfs and hybrids should be augmented and made available to the farmers in adequate quantity. References Balakrishnan, PC, S Sukumaran Nair and K Kumaran. 1988. Coconut improvement: Six decades of coconut research. Kerala Agricultural University. Pp. 7-37. Balingasa, EN and CB Carpio. 1976. Genetic potential of some coconut populations of the Philippines. Pp. 72-81. In: NM Nayar (ed). Coconut research and development. Wiley Eastern Ltd. Bourdeix, R, A Sangare, JP Le Saint, J Meunier, JP Gascon, F Rognon and MW de Nuce de Lamothe. 1993. Coconut breeding at IRHO and its application in seed production. Pp. 79-94. In: MK Nair et al. (eds). Advances in coconut research and development. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Calvez, C. 1989. Assessment of experience with high yielding varieties, Vanuatu’s experience. Pp. 1-22. In Proceedings of the APCC/ COCOTECH/XXVI/S2-07. Chan, E. 1983. Progress in coconut breeding in United Plantations Bhd, Malaysia. Oleagineaux 38 : 6, 371-376. Darwis, Michellia. 1996. Coconut research in Indonesia. Pp. 207-214. In: PK Thampan (ed). Coconut for prosperity. Peekay Tree Crops Development Foundation, Kerala, India. Davis, TA, H Sundasrip and SN Darwis. 1985. An overview of research 324 COCONUT GENETIC RESOURCES activities. Coconut Research Institute, Manado, Indonesia. De nuce de Lamothe, MW and A Sangare. 1993. Current status of coconut genetic resources research in Ivory Coast. Pp. 9-14. In: MK Nair et al. (eds). Advances in coconut research and development. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Dhamodaran, S, MJ Ratnambal, B Chempakam, RV Pillai and BC Viraktamath. 1993. Evaluation of tender nut water in coconut cultivars. Pp. 123-128. In: MK Nair et al. (eds). Advances in coconut research and development. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Dootson, J, A Thirakul, C Petchpiroon and M Rattanapruk. 1988. Early yields of a number of coconut varieties in Thailand. Oleagineux 43 : 12, 445-451. Duhamel, G. 1993. Crop improvement programmes in South Pacific Region. Pp. 95-100. In: MK Nair et al. (eds). Advances in coconut research and development. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Efu, S. 1989. Assessment of experience with high yielding varieties: Western Samoa’s experience. Pp. 1-5. In: Proceedings of the APCC/ COCOTECH/XXVI/S2-08. Gapasin, Dely P. 1986. Coconut research and development in the Philippines. Cord Journal 11 : 2,1-19. Harries, H.C. 1971. Coconut varieties in America. Oleagineaux 26 : 235- 242. Jack, HW. 1937. The dwarf coconut. Fiji Agri. Journ. 8 : 4, 47-48. Jack, HW and WN Sands. 1929. Observations on the dwarf coconut palm in Malaya. Mal. Agri. Jour. 17 : 140-65. Linh, NA and VV Long. 2000. Evaluation of performance for suitable varieties in Vietnam. COGENT Newsletter, May 2000. IPGRI-APO, Serdang, Selangor, Malaysia. Liyanage, DV. 1988. Coconut seed gardens: A review. Cord Journal 4 : 1, 14-21. Liyanage, M de S. 1996. Coconut research in Sri Lanka. Pp. 221-240. In: NM Nayar (ed). Coconut for prosperity. Peekay Tree Crops Development Foundation, Kerala, India. Manthriratna, MAPP. 1983. The mechanics of hybrid seed production through seed garden. Pp. 56-61. In: NM Nayar (ed). Coconut research and development. Wiley Eastern Ltd., New Delhi, India. Nair, MK, MJ Ratnambal and P Gopalasundram. 1996. Coconut research in India. Pp. 181-198. In PK Thampan (ed). Coconut for prosperity. Peekay Tree Crops Development Foundation, Kerala, India. Ovasuru, T, GY Tan and LA Bridgland. 1993. Coconut germplasm 325 CHAPTER 5: Germplasm use collection in Papua New Guinea. Pp. 33-42. In: MK Nair et al. (eds). Advances in coconut research and development. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Rognon, F and MW de Nuce de Lamothe. 1978. Harvesting and conditioning of pollen for the pollination of coconut seed gardens. Oleagineaux 33:1, 22-23. Santos, GA. 1989. Assessment of experience with high yielding varieties, Philippines experience. Pp. 1-23. In: Proceedings of the APCC/ COCOTECH/ XXVI/S2-04. Santos, GA, RT Bahala, SB Cano and BV dela Cruz. 1986. Yield and agronomic trials of four variety hybrids and some local tall coconut plantations in the Philippines. Oleagineux 41 : 6, 269-280. Sumbak, JH. 1976. Trials with some coconut varieties and hybrids in Papua New Guinea. Pp. 82-84. In NM Nayar (ed). Coconut research and development. Wiley Eastern Ltd., New Delhi, India. Sumith de Silva. 1989. High-yielding varieties of coconut: An intra- regional study on small farmer’s experience. APCC, Jakarta, Indonesia. Tarigans, DD. 1989. Assessment of experience with high yielding varieties, Indonesian experience. Pp. 1-29. In: Proceedings of the APCC/ COCOTECH/ XXVI/ S2-02. Thampan, PK. 1993. Handbook on coconut palm. Oxford and IBH Publishing Co. Pvt. Ltd., New Delhi, India. Turner, P. 1989. Assessment of experience with high yielding varieties: Papua New Guinea’s experience. Pp. 35-43. In: Proceedings of the APCC/ COCOTECH/ XXVI/ S2-03. Wickramaratne, MRT. 1989. Assessment of experience with high yielding varieties of coconut in Sri Lanka experience. Pp. 66-75. In Proceedings of the APCC / COCOTECH / XXVI /S2-06. 326 COCONUT GENETIC RESOURCES Multilocation coconut hybrid trials in three African and three LAC countries P Batugal1, JL Konan2, A Sanaoussi3, A Kullaya4, EA Tupinamba5, R Castillo6 and B Been7 1COGENT Coordinator and Senior Scientist, International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia 2Head, Coconut Programme, Centre National de Recherche Agronomique (CNRA), Abidjan, Côte d’Ivoire 3Researcher, Institut National de Recherche Agricoles du Benin (INRAB), Sous- Programme Cocotier, BP 884, Cotonou, Benin 4Director, Mikocheni Agricultural Research Institute (MARI), Dar es Salaam, Tanzania 5Coconut Researcher, Coconut Germplasm Bank, Brazilian Agricultural Research Corporation (EMBRAPA), Coastal Tablelands, Av. Beira Mar, Aracaju, SE, Brazil 6 Investigador del Sistema Producto, Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias (INIFAP), Mexico 7 Director of Research, Coconut Industry Board (CIB), 18 Waterloo Road, Kingston 10, Jamaica Introduction The main objectives of the multilocation trials are: 1) to assist each of the six participating countries in identifying suitable high-yielding varieties/ hybrids with high adaptation to prevailing local conditions; and 2) to estimate genotype x environment (G x E) interaction, which will serve as a guide to the application of the results to other countries with similar growing conditions. Each test country compared the six common multi-site hybrids produced and shipped from Côte d’Ivoire with 4-8 of its best hybrids/ varieties. The imported hybrids are four Dwarf x Tall and two Tall x Talls which have been proven to have good yield potential in previous trials. Each experimental unit or plot consisted of 16 palms, planted in a triangular pattern at 9m in a randomized complete block design (RCBD) with five replications. When sufficient vegetative and reproductive data will have been generated, statistical analysis shall be done at country level to compare the different genetic materials, while a combined data analysis will be conducted to determine the interaction between genotype and environment. Project implementation The project was approved by the Common Fund for Commodities (CFC) Executive Board on 22 October 1996 and funds were released to IPGRI in January 2000. However, the six participating countries and IPGRI pre- 327 CHAPTER 5: Germplasm use financed the project from 1997 to 1999 to fund preparatory activities in support of the project. The implementation of this multilocation trial to evaluate the performance of 30 imported and local coconut hybrids and varieties, and their G x E interaction involved three African countries (Côte d’Ivoire, Benin and Tanzania) and three Latin American and Caribbean or LAC countries (Brazil, Jamaica and Mexico). Six promising hybrids (MYD x WAT, CRD x RIT, VTT x TAGT, MRD x VTT, MRD x TAGT and SLT x TAGT) were produced by Côte d’Ivoire and air-shipped to the five participating countries (one set was retained for the Côte d’Ivoire trial) while the six participating countries produced seednuts of their promising local hybrids/varieties to serve as controls. The 30 initial coconut hybrids/ varieties that were identified to be produced and tested in the multilocation trials are shown in Table 1. The six countries originally planned to produce four local hybrids each to be compared with the six common imported hybrids. However, this target output was exceeded because Mexico produced four additional local hybrids, namely: MYD x MXPT05, MYD x MXPT10, MYD x MXPT11 and MYD x MXAT. All six countries conducted two trials each, the first, a general performance trial using the seedlings from the first batch of hybrid seednuts sent from Côte d’Ivoire and the locally produced hybrids/ varieties; and the second, using the second and third batch (only for Brazil) of seednuts from Côte d’Ivoire. Table 1. List of multi-site and local hybrid/variety trial entries A. Six multi-site hybrids (common for all participating countries) Dwarf x Tall hybrids (4) Tall x Tall hybrids (2) MYD x WAT CRD x RIT VTT x TAGT MRD x VTT MRD x TAGT SLT x TAGT B. Four locally-selected materials per country Côte d'Ivoire Bénin Tanzania Jamaica Mexico Brazil MYD x TKT MYD x PNT EAT o.p. MYD x THT MYD x PNT BGD x VTT MYD x TGT CRD x WAT PRD x EAT MYD x PNT MYD x MXPT14 BGD x BRT MYD x PUT CRD xT AT EAT x RIT CGD x PNT MYD x MXPT09 MYD x BRT PGD x VTT PGD x VTT EAT x VTT CGD x THT MYD x MXPT02 BRT o.p. (+ 4 additional hybrids) List of variety names BGD Brazilian Green Dwarf o.p. open pollinated CRD Cameroon Red Dwarf PYT Polynesia Tall CUD Cuban Dwarf BRT Brazilian Tall FJM Fiji Malayan Dwarf PNT Panama Tall MRD Malayan Red Dwarf RIT Rennell Island Tall MYD Malayan Yellow Dwarf SLT Sri Lanka Tall PRD Pemba Red Dwarf TAG Tagnanan Tall EAT East African Tall THT Thailand Tall WAT West African Tall VTT Vanuatu Tall PGD Pumila Green Dwarf CGD Chowghat Green Dwarf 328 COCONUT GENETIC RESOURCES Noting the potential impact of this project in increasing the yields of poor coconut farmers, the Government of Portugal funded a similar project involving the evaluation of the same six multi-site hybrids and four local hybrids (MYD x MZT, BGD x RLT, MZT x SGD and MZT x VTT) in Mozambique. This brought to 38 the total number of coconut hybrids/ open-pollinated varieties being evaluated, making this project the most comprehensive coconut hybrid trial worldwide. The details of the multi-location trials are presented in CFC Technical Paper No. 42 entitled, ‘Coconut Hybrids for Smallholders’ (Batugal et al., eds 2005). Major achievements The most important result of the project is the identification of 16 early bearing and high-yielding new coconut hybrids (Table 2). The first trial showed that 16 out of the 34 test trials in the CFC-funded project started to flower and produce fruits in Brazil, Jamaica and Mexico in 2.5-3.0 years after planting compared to the seven years it would normally take the traditional Tall varieties to reach fruiting stage. In Brazil, two hybrids from Côte d’Ivoire and two local hybrids flowered; in Jamaica, all six hybrids produced in Côte d’Ivoire flowered but none of the local hybrids; while in Mexico, only one hybrid produced in Côte d’Ivoire and eight locally produced hybrids flowered. On the other hand, flowering was not observed in the hybrids planted in Benin, Côte d’Ivoire and Tanzania during the same period. These results suggest that the drought in Africa and the generally drier conditions in that region compared to the LAC region had a negative effect on early flowering of the hybrids, suggesting a possible G x E interaction. This interaction could be verified with the vegetative and reproductive plant measurements and biotic and abiotic data to be gathered and analyzed in the next five years. Based on the yield projection of the potential of the 16 fruiting hybrids on their fourth year (as observed in preliminary trials), they have the potential to produce up to five tonnes of copra (dried kernel) per hectare per year at the peak of production (at 10-12 years) compared to the one metric tonne of copra generally produced by the traditional Tall cultivars. The impact of the results from this CFC-funded project is significant as it has the potential to increase coconut yields of resource-poor smallholder coconut farmers by up to five-fold if the results are effectively promoted, with good management, in many coconut growing communities and countries. Although the hybrids in the second trial are all growing well in five countries (except Benin), the potential of the hybrids could only be determined when they start to produce fruits three more years after the project termination. 329 CHAPTER 5: Germplasm use In Tanzania, although the few seedlings planted from the first batch of seednuts did not grow well due to drought, fire and termite infestation, the seedlings in the second replicated trial were very robust and were growing very well and are expected to flower within the next 24 months. In Côte d’Ivoire, both the seedlings in the first and second batches of seednuts are growing well. Oryctes beetle infestation is under control through a good integrated pest management system. In Benin, the plants from the first batch of seedlings did not do very well due to severe drought while the plants in the second batch are growing well in the three blocks (replications). The two blocks located in the low-lying area of the experimental field were waterlogged and remedies are being made to construct a drainage system. In Jamaica, the plants in the second trial are growing well, despite some damage by the lethal yellowing disease. In Brazil, the plants in the second batch are growing well except for a few missing hills which were replanted. Capacity building The second most important achievement of the project is capacity building. Based on all project components, 182 coconut researchers participated in 15 training courses and 863 attended various meetings, conferences and workshops for a total of 1045 coconut researchers worldwide whose research capacities have been enhanced (Annex 1). These events allowed coconut researchers and officers worldwide to enhance their skills on coconut genetic resources research and shared expertise, experiences and ideas to address common problems and Reference Hybrids produced in Cote d’Ivoire Brazil Mexico Jamaica MYD x WAT ; ; ; MRD x VTT ; CRD x RIT ; MRD x TAGT ; ; SLT x TAGT ; VTT x TAGT ; Locally produced hybrids MYD x MXPT05 ; MYD x MXAT ; MYD x MXPT10 ; MYD x MXPT11 ; MYD x MXPT02 ; MYD x MXPT09 ; MYD x MXPT14 ; MYD x PNT ; BGD x BRT ; BGD x VTT ; Table 2. Coconut hybrids that started fruiting 2.5 - 3.0 years after planting (with check) 330 COCONUT GENETIC RESOURCES Target output of CFC-funded training component Actual output Excess over target output 40 coconut breeders from 30 countries trained in breeding research techniques 50 coconut breeders from 30 countries trained in breeding research techniques + 10 coconut breeders 30 germplasm workers from 30 countries trained in collecting and conservation 45 germplasm workers from 30 countries trained in collecting and conservation + 15 germplasm workers 15 biotechnologists from 10 countries trained in molecular techniques for diversity assessment 18 biotechnologists from 9 countries trained in molecular techniques for diversity assessment + 3 biotechnologists 30 physiologists from 15 countries trained in embryo culture techniques 42 physiologists from 15 countries trained in embryo culture techniques +12 physiologists 10 researchers from 5 countries trained in cryopreservation techniques 12 researchers from 5 countries trained in cryopreservation techniques + 2 researchers 10 researchers from 5 countries trained in molecular techniques for pathogen characterization Not done but training on Germplasm x Environment Interaction analysis involving 9 participants was substituted - opportunities affecting the farmers and the coconut industry. These capacity building activities have strengthened the research capability of coconut producing countries and promoted inter-country and inter- regional collaboration for conducting research to help resource-poor coconut farmers. The results of training for the CFC-funded components of the project are shown in Table 3. The project exceeded the target outputs by 42 (number of trained researchers) based on the conducted training activities. However, because training on molecular techniques for pathogen characterization was already programmed in the newly approved CFC- funded project on lethal yellowing disease, training on (GxE) interaction analysis was substituted which was identified by the project participants as a priority activity to expand the application of project results in various environments across the world. Table 3. Trained coconut researchers from national programmes based on target outputs of the CFC-funded project Reference Batugal, P, D Benigno and J Oliver (eds). 2005. Coconut hybrids for smallholders. CFC Technical Paper No. 42. CFC/IPGRI. 235p. 331 CHAPTER 5: Germplasm use Training and capacity building activities Description Venue Dates No. of participants A. TRAINING COURSES The Sub-Regional STANTECH Training Course for Africa Training on aspects of coconut germplasm collecting, conservation and documentation, and breeding techniques Station Cocotier Marc Delorme Abidjan, Côte d’ivoire 16-26 June 1997 9 Regional STANTECH Training Course For Latin America And The Caribbean Training on aspects of coconut germplasm collecting, conservation and documentation, and breeding techniques Coconut Industry Board (CIB) Kingston, Jamaica 14-25 July 1997 6 Coconut Germplasm Collecting and Conservation Training Course Training on aspects of coconut germplasm collecting and conservation techniques Philippine Coconut Authority (PCA) Zamboanga, Philippines 1-12 Sep 1997 11 International Coconut Embryo Culture and Acclimatization Workshop Assess and upgrade and standardized embryo culture technology Albay Research Center Philippine Coconut Authority Philippines 27-31 Oct 1997 14 Coconut Multilocation Hybrid/Variety Trials Workshop Workshop to identify suitable hybrids and varieties for Africa, Latin America and the Caribbean Station Cocotier Marc Delorme Abidjan, Côte D’ivoire 10-12 Nov 1997 9 STANTECH Training Course on Collecting and Management Of Coconut Genetic Resources The training course focused on coconut genetic resources collecting strategy, the collecting process and the methods of descriptions of coconut varieties Vanuatu Agricultural Research and Training Center (VARTC) Santo, Vanuatu 29 Jun – 10 Jul 1999 4 Second International Coconut Embryo Culture Workshop Assess and upgrade and standardized embryo culture technology Centro de Investigacion Cientifica de Yucatan (CICY) Merida, Mexico 14-17 Mar 2000 28 Regional Training Course On In Vitro Conservation and Cryopreservation On PGR Training on Coconut Cryopreservation NBPGR, India 12-25 Oct 2000 7 STANTECH Training Course The training course used the standardized techniques as guidelines in coconut breeding and germplasm conservation with the hope that it will help coconut researchers obtain better and comparable results Philippine Coconut Authority-Zamboanga Research Center, San Ramon , Zamboanga City Philippines 2-7 April 2001 6 2nd CFC-Funded Project Workshop and Initial Consultation on a Proposed Globally Coordinated Coconut Breeding Consultations to refine the proposed globally coordinated coconut breeding programme Mikocheni Agricultural Research Institute (MARI) Dar es Salaam, Tanzania 11-12 June 2001 39 Standardized Research Techniques in Coconut Breeding Training on coconut germplasm characterization and seednuts production (controlled and assisted pollination in seed- garden). CNRA Marc-Delorme Côte d’Ivoire 16-28 Jan 2002 2 Workshop on Coconut Genetic Resources Management Using Microsatellite Kit and Dedicated Statistical Software Training on using Microsatellite Kit and Dedicated Statistical Software CIRAD Montpellier, France 15-24 Apr 2002 18 Annex 1. Training and capacity building activities conducted under the CFC-funded multi-location coconut hybrids trial project 332 COCONUT GENETIC RESOURCES Coconut Cryopreservation Training Course Training on Coconut Cryopreservation Institut de Recherche pour le Développement (IRD) Montpellier, France 13-17 Oct 2003 5 Technical writing, seminar presentation, public awareness and proposal preparation Training course on technical writing, seminar presentation, public awareness and proposal preparation Merida, Mexico 6-7 Nov 2003 15 Statistical Design and Germplasm x Environment Interaction Analysis Training Course To acquaint training course participants with the various statistical designs and methods of data analysis that could be used for coconut research and the protocol for G x E interaction Hotel Grand Maya, Kuala Lumpur 25-27 Nov 2004 9 Subtotal for Training 182 B. WORKSHOPS AND MEETINGS WITH INVITED SPEAKERS FROM ADVANCED LABORATORIES AND PRODUCER COUNTRIES Cadang-cadang viroid-like sequences meeting Consultation on the Cadang-cadang viroid-like sequences Serdang, Malaysia 21-23 Apr 1997 14 LAC coconut regional project proposal formulation meeting Initial consultation to refine an LAC coconut regional project proposal Kingston, Jamaica 7-12 July 1997 7 Coconut Genetic Resources Network In Asia and the Pacific Region (CGRNAP) Phase 1/Phase 2 annual Review and Planning Meeting Review the progress of the conservation and utilization of the coconut genetic resources project and work plans Bogor, Indonesia 15-17 Sep 1997 28 Annual meeting, Coconut multi- purpose uses project Review the progress of the coconut multipurpose uses project and work plans Bogor, Indonesia 18-20 Sep 1997 28 6th COGENT Steering Committee meeting Review the progress of COGENT and work plans Port Bouet, Côte d’Ivoire 13-15 Nov 1997 14 ADB-Funded Projects Annual Meeting Review the progress of the germplasm collecting and conservation project and work plans Kuala Lumpur, Malaysia 29-31 Oct 1998 29 IFAD-Funded Projects Annual Meeting Review the progress of the coconut multipurpose uses project and work plans Kuala Lumpur, Malaysia 2-4 Nov 1998 21 International Coconut Genebank Workshop Review the progress of the field and regional genebanks project and work plans Madang, Papua New Guinea 6-7 Nov 1998 20 7th COGENT Steering Committee meeting Review the progress of COGENT and work plans Madang, PNG 9-11 Nov 1998 20 IFAD-funded project meeting Review the progress of the coconut multipurpose uses project and work plans Ho Chi Minh city, Vietnam 13-15 Sep 1999 27 ADB Phase 2 project meeting Review the progress of the germplasm collecting and conservation project and work plans Ho Chi Minh City Vietnam 16-17 Sep 1999 27 COGENT Steering Committee meeting Review the progress of COGENT and work plans Ho Chi Minh City , Vietnam 20-22 Sep 1999 16 2nd International Coconut Embryo Culture Workshop Review the progress of the project and work plans Merida, Mexico 14-17 Mar 2000 28 333 CHAPTER 5: Germplasm use Meetings of ADB/IFAD Funded Coconut Research Projects and Future Directions of the Coconut Industry in the South Pacific Review the progress of the collecting/ conservation and multipurpose uses project and work plans Apia, Samoa 26-30 Jun 2000 20 ADB & IFAD Funded Projects Joint Annual Meeting for Asia Review the progress of the collecting/ conservation and multipurpose uses project and work plans Manila, Philippines 10-15 July 2000 21 International Coconut Genebank Workshop Review the progress of the project and work plans Chennai, India 17-19 July 2000 24 9th COGENT Steering Committee Meeting Review the progress of COGENT and work plans Chennai, India 20-22 July 2000 24 International Coconut Conference Review the progress of the coconut genetic resources and work plans Chennai, India 24-28 July 2000 167 CFC project workshop Review the progress of the multilocation coconut hybrid trials project and work plans Dar es Salaam, Tanzania 11-12 Jun 2001 22 10th COGENT Steering Committee meeting Review the progress of COGENT and work plans Dar es Salaam, Tanzania 13-15 Jun 2001 22 Poverty Reduction in Coconut Growing Communities, Project Inception and Stakeholders’ Meeting Initial consultation on a proposed project proposal Ho Chi Minh City, Vietnam 25 Feb-1 Mar 2002 25 11th COGENT Steering Committee meeting Review the progress of COGENT and work plans Bangkok, Thailand 25-28 Jun 2002 20 CFC Mid Term Evaluation Project Meeting Review the mid-term progress of the CFC- funded project and work plans Kingston, Jamaica 25-27 July 2002 15 2nd International Coconut Genebank (ICG) Meeting and Consultation on Proposed Globally Coordinated Coconut Breeding Review status of the ICGs and consultations on a proposed project proposal Kasaragod, India 30 Oct-1 Nov 2002 31 2nd Annual ADB funded Project Meeting Review the progress of the poverty reduction in coconut growing communities project and work plans Davao, Philippines 21-23 Aug 2003 80 12th COGENT Steering Committee meeting Review the progress of COGENT and work plans Merida, Mexico 10-12 Nov 2003 16 4th Annual CFC-funded project meeting: Coconut Germplasm Utilization and Conservation to Promote Sustainable Coconut Production Review the progress of the CFC-funded project and work plans Merida, Mexico 13-17 Nov 2003 21 Poverty Reduction in Coconut Growing Communities (PRCGC): The Final ADB funded Project Meeting Evaluate the progress and outputs of the project in eight Asia Pacific countries and discuss initial development impact and strategies for upscaling and replication Ho Chi Minh City, Vietnam 27-30 Sep 2004 24 Final CFC funded Project Meeting Review the progress of the CFC-funded multilocation coconut hybrid trials and other project components, and discuss the findings on the final evaluation of the project and issues and recommendations Kuala Lumpur, Malaysia 17-19 Nov 2004 26 13th COGENT Steering Committee Meeting Review the progress of ongoing and proposed projects and activities of COGENT; status of PROCORD; finalize COGENT’s Strategic Plan and refine COGENT’s coconut conservation strategy Kuala Lumpur, Malaysia 22-24 Nov 2004 26 Subtotal for Workshops and Meetings 863 GRAND TOTAL 1045 334 COCONUT GENETIC RESOURCES Coconut micropropagation C Oropeza1, E Rillo2, V Hocher3 and JL Verdeil4 1Coconut Researcher, Centro de Investigación Científica de Yucatán (CICY), Mérida, México 2Scientist, Philippine Coconut Authority - Albay Research Centre (PCA-ARC), Guinobatan, Albay, Philippines 3Scientist, L’Institut de Recherche pour le Développement (IRD), Montpellier, France 4Researcher, Centre de Cooperation Internationale en Recherche Agronomique pour le Developpement (CIRAD), Cedex 5, Montpellier, France Introduction The coconut palm (Cocos nucifera L.) is a very important crop providing cash and subsistence to millions of smallholders in 86 countries where about 12 million ha are planted with this palm (Santos 1999). However, most coconut groves require replanting because of loss due either to palm senescence or to diseases such as lethal yellowing in America (Arellano and Oropeza 1995), the lethal diseases in Africa (Eden-Green 1995) and cadang-cadang in Asia, in particular the Philippines (Hanold and Randles 1991). Research on genotype selection for disease resistance or other traits of interest, such as high yield, are being carried out worldwide with positive results (Santos 1999; Zizumbo et al. 1999). However, propagation of selected genotypes, or even more conveniently, individuals within these genotypes to satisfy the rapidly growing demands will be very hard to fulfill through natural coconut propagation that occurs only sexually, producing very few seeds per palm within its long life cycle. Therefore, alternative approaches for rapid propagation of improved planting materials must be considered. In this respect, in vitro cloning via somatic embryogenesis seems to provide a convenient alternative for the future due to its potential for massive propagation. Unfortunately, coconut is a species that responds very poorly to in vitro culture, being one of the most recalcitrant species to regenerate in vitro (George 1996). This paper summarizes the efforts that have been carried out to develop protocols for the micropropagation of coconut through somatic embryogenesis, presenting the first work carried out during the 20th century and the research advances obtained during the past five years. The paper focuses particularly on research leading to sustained developments such as those related to the use of inflorescence and embryo explants. Research from the 1970’s to the 1990’s Initial developments started with the work of Eeuwens (1976) on improving callus formation and growth by optimizing the mineral composition of the culture media following factorial design experiments. 335 CHAPTER 5: Germplasm use Testing of the different types of explants followed: young roots of mature palms (Justin 1978), stem and leaf (Pannetier and Buffard-Morel 1982; Gupta et al. 1984; Raju et al. 1984), embryos (Karunaratne and Periyapperuma 1989) and inflorescences. The first promising results involving somatic embryogenesis were obtained at Wye College (UK) with the first clonal plants produced in 1983 (Branton and Blake 1983) and similar findings were obtained with young leaf explants (Buffard- Morel et al. 1992). Low percentage of callus formation and the development of abnormal plants were common occurrences (Branton and Blake 1984; Dublin et al. 1991). However, these studies demonstrated that coconut regeneration by somatic embryogenesis was possible. The use of inflorescence explants Coconut tissue culturists were initially interested in the use of immature coconut inflorescences as explants because they contain meristematic tissue, which was encouraged to form callus tissue with the addition of an auxin to the culture medium. Immature inflorescences from mature palms could be excised non-destructively from the palms (Rillo 1989). Initial callus formation started at about three months after culture initiation and was observed until about nine months (Hornung and Verdeil 1999). The most commonly used auxin was 2,4-D at varying concentrations, depending on the amount and type of activated charcoal employed. Cytokinins were usually not added to the medium for callus initiation. For instance in Montpellier, the Eeuwens Y3 mineral solution (Eeuwens 1976) was used with the addition of Morel and Wetmore’s vitamins (1951), 2.5 g l-1 activated charcoal, sucrose at 40 g l-1 and agar at 7.5 g l-1, 2,4-D between 13 and 36 mM, due to the variable sensitivity of different palms and the developmental stage of inflorescences to the auxin. Murashige and Skoog medium (1962) with the addition of sucrose, activated charcoal and auxin was also employed for callus production. Callus grown on media with gradually reduced auxin levels (Blake 1990), or by an increase followed by a reduction (Verdeil et al. 1992), produced nodular somatic structures that subsequently developed into proembryos and finally into embryos (Verdeil et al. 1994). However, the production of good somatic embryos still presented a problem due to the development of fused embryos, fused leafy structures with or without roots and haustorial type tissues (Sugimura and Salvaña 1989; Blake 1990; Verdeil et al. 1992). The main difficulties encountered in coconut regeneration by somatic embryogenesis from inflorescence explants were: intense browning of tissue linked to the oxidation of polyphenols, considerable heterogeneity in tissue response, a strong tendency to produce roots only, low 336 COCONUT GENETIC RESOURCES embryogenic potential, poorly developed embryo formation, poor shoot formation and slow growth rate in vitro. The first one could be overcome to a great extent by the use of activated charcoal. The others were more difficult to overcome. However, in 1993, several groups (from the University of Hanover, Wye College, L’Institut de Recherche pour le Developpement-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (IRD-CIRAD), Philippine Coconut Authority (PCA) and Centro de Investigación Científica de Yucatán (CICY)) involved in coconut regeneration research gathered together for the first time to start a joint effort to overcome the major difficulties encountered in coconut regeneration. This was made possible through a project funded by the European Commission within the Science and Technology for the Development 3rd Period Programme (STD-3) (ERBTS3*CT940298) that started in January 1993. This project increased the fundamental knowledge on the different aspects of somatic embryogenesis in coconut: morpho-histological development ( and infrastructural changes (Verdeil et al. 2001); hormone studies (Verdeil et al. 1994; Hocher 2003); nutritional studies (Dussert et al. 1995a,b; Magnaval et al. 1995; 1997); protein phosphorylation during somatic embryogenesis (Islas-Flores et al. 2000); and plantlet photosynthesis characterization (Triques et al. 1997a,b; Rival et al. 1999). These studies increased the understanding of the regeneration process and helped to overcome some of the difficulties encountered in coconut regeneration and improvement of protocols using either inflorescence or plumule explants. Several experimental protocols using inflorescences or plumules are now available. For a more detailed account on coconut regeneration from inflorescence explants, see Hornung and Verdeil (1999). Studies using plumule are described below. The use of embryo explants As mentioned above, whole coconut embryos had been used as explants to induce embryogenic calli and somatic embryo formation without success. As an alternative, researchers from Wye College and CICY tested embryo isolated parts: plumules, root poles and cotyledonary tissues. Plumules were the only explants that readily formed embryogenic callus and embryos that developed shoots (Hornung 1995; Oropeza and Chan 1995). Further studies within the EC-STD3 project (reported in Chan et al. 1998) were carried out to improve somatic embryogenesis and shoot formation from plumule explants, and ultimately, ex vitro plantlet establishment. Plumules from the embryos of nuts harvested 12-14 months after pollination were used and cultured on Y3 medium (Eeuwens 1976) containing activated charcoal and gelrite. Different parameters were 337 CHAPTER 5: Germplasm use tested to optimize callus and somatic embryo formation: concentration of phytohormones, subculture frequency and light conditions. Callus formation required auxin (2,4-D) at an optimum concentration of 0.1 mM 2,4-D. These calli developed meristematic centers and when kept at the same auxin concentration (0.1 mM), the calli developed embryogenic structures. A greater proportion of plumule explants developed into calli bearing embryogenic structures when cultures were kept undisturbed and no subculturing was practised. Histological observations showed that formation of somatic embryos had already started in calli bearing embryogenic structures, but development of embryos occurred when the auxin concentration was reduced by a hundred-fold and cytokinin was added (50 µM 6-BAP), performing better under illumination (12 h photoperiod) than in the dark. Keeping cultures in these conditions and subculturing every three months, allowed embryos to germinate and the resulting shoots eventually developed into plantlets that, after acclimatization, grew successfully under ex vitro conditions. Following this protocol, different batches of cultures were tested and the performance was found to be reproducible. As reported by Chan et al. (1998), with plumule explants shorter times were required to obtain somatic embryos (7-9 months) than those previously reported with inflorescence explants (14-20 months, Verdeil et al. 1994), and the yields were higher (nearly two-fold for calli and over ten-fold for calli bearing somatic embryos) than those reported with inflorescences (Verdeil et al. 1994). Acclimatization has been successful and plantlets did well in open environmental conditions since they continued producing new leaves, sexual organs and fruits. This protocol by Chan et al. (1998) reported that 60% of the explants formed initial calli using a local brand charcoal. In Montpellier, by using a different brand (Sigma acid washed charcoal), an improvement in callus induction of nearly 100% was obtained (J.L. Verdeil and V. Hocher, Montpellier, unpublished). Similar results were obtained afterwards at CICY. Therefore, careful selection of charcoal is very important. Clonal plantlets are now produced in most of the participating laboratories within the STD-3 project. The use of embryo slices as explants and different treatments were evaluated at Queensland University. Polyethyleneglycol and ABA were tested for somatic embryo maturation and had very little or no effect when these chemicals were added separately, but when added simultaneously, they inhibited the growth of non-embryogenic calli and improved the maturation of somatic embryos (Samosir et al. 1999). Conditions that increased ethylene concentrations in the above coconut cultures were found to affect callus growth and somatic embryogenesis 338 COCONUT GENETIC RESOURCES (Adkins et al. 1999). Accordingly, embryogenic calli were incubated with a number of additives that could reduce ethylene production (aminoethoxyvinylglycine), protect from ethylene (silver thiosulphate) or help combat ethylene-induced stress (polyamines). Coconut somatic embryogenesis was promoted (100%) by the addition of the polyamines putrescine and spermidine to the medium (Adkins et al. 1999). Unfortunately, when tested with plumule explants, no promotion of somatic embryo formation was obtained (CICY, unpublished data). Recent advances The use of plumule explants During the past five years, research on the plumule system has continued in order to further improve its performance and different approaches have been tested. The compound 22(S), 23(S)-homobrassinolide was found to increase embryogenic calli and somatic embryo formation (1.5 and 2 fold respectively, compared with the controls) when applied as a pretreatment to plumule explants (Azpeitia et al. 2003). Cytokinins have been found to decrease embryogenic callus formation in plumule explants and therefore, the anticytokinin 8-azaadenine was tested. It increased somatic embryo formation 1.5 fold in relation to the control treatment (Azpeitia, 2003). There are other two approaches that resulted in even larger increases of yields: secondary embryogenesis and multiplication of embryogenic calli (CICY, unpublished results). Secondary embryogenesis is based on the use of somatic embryos as explants to produce more embryos. This process can be repeated several times. Therefore, it can be useful to increase the total somatic embryo yield obtained per original explant. Embryogenic calli multiplication allows increasing the yield of this type of calli several fold. Unpublished results (CICY) presently show that by combining these two approaches, thousands of embryogenic calli and tens of thousands of somatic embryos can be obtained from one plumule and the amounts depends on the number of multiplying cycles carried out. Furthermore, if these two approaches were combined with the use of 22(S),23(S)-homobrassinolide (Azpeitia et al. 2003), yields could be potentially increased even more. This three-approach strategy is being tested in collaboration with COGENT. Regarding germination and post-germinative development of somatic embryos, studies had been limited by the low yields obtained. Therefore, these studies were approached using the coconut zygotic embryo as a model system. At CICY, this system showed that aerobic respiration was required for embryos to germinate (Pech y Ake et al. 2004). Percentage of 339 CHAPTER 5: Germplasm use germination increased from 66% in liquid medium where embryos were submerged to 93% on solid medium where embryos could be placed with their micropylar end facing upwards exposed to the ambient atmosphere of the vial (Pech y Ake et al. 2004). This also resulted in increased conversion from 46% to 89% for liquid and solid media, respectively. In addition, the use of gibberelic acid (GA3) further promoted both germination and conversion into plantlets (Pech y Ake et al. 2002). The use of ventilated vessels (with filter paper windows) when compared with sealed vessels, improved the leaf water loss control of plantlets formed from zygotic embryos cultured in these vessels (Talavera et al. 2001). Ex vitro survival of plantlets was found to be over 90% if proper development was allowed, plantlets should have a minimum of three bifid leaves and three main roots when transferred from in vitro conditions to ex vitro acclimatization conditions (Pech y Ake 2004). Some of the information obtained using zygotic embryos has been used to help define the optimal germination and post-germination development conditions for somatic embryos. This way, when plantlets derived from somatic embryos were allowed to develop three bifid leaves and three main roots, ex vitro survival was over 90% (CICY, unpublished results). Micropropagated palms established in permanent field conditions over four years ago have done well under and some are already bearing fruits. Similar (unpublished) observations have been noted at the Coconut Research Institute in Sri Lanka. The use of other explants (leaf and inflorescence) The information obtained on coconut regeneration using plumular explants can be useful to for research on the use of other explants. Research on micropropagation based on inflorescence or leaf explants has not been abandoned and it is one of the main objectives of a project supported by the Australian Centre for International Agricultural Research (ACIAR) involving laboratories at the University of Queensland (Australia), the Research Institute for Coconut and other Palmae (Indonesia), the Philippine Coconut Authority (Philippines), the University of Philippines at Los Baños (Philippines), the Cocoa and Coconut Research Institute (Papua New Guinea) and the Oil Plant Institute (Vietnam). It would be interesting to test the combined use of secondary embryogenesis and embryogenic callus multiplication with inflorescence and leaf explants. Genetic engineering In addition, there are new areas of research intended to open new avenues for coconut micropropagation improvement and probably applications. These studies are based on molecular techniques and presented below. 340 COCONUT GENETIC RESOURCES A different approach to increase coconut micropropagation efficiency, not tried before, is to improve the embryogenic capacity of coconut tissues by inserting genes related to this capacity. Hence, the genes and the protocols for their insertion through transformation techniques are needed. Therefore, through a collaborative effort, researchers from Max Planck and Fraunhofer Institutes (Germany) and CICY are attempting to develop transformation protocols for coconut tissues. Agrobacterium tumefaciens-mediated transformation and particle gun (Biobalistic) DNA delivery methods have been applied to transform coconut cells using the reporter genes uidA that codify for b-glucuronidase (GUS), gfp for the green fluorescent protein (GFP) and rfp for the red fluorescent protein (RFP) under the control of constitutive promoters such as 35S CaMv and Ubiquitin from Maize. Transient transformation was successfully obtained with both transformation methods and the three reporter genes. However, the use of uidA was hampered by the finding of endogenous GUS activity in coconut calli. Stable transformation has been confirmed for gfp in A. tumefaciens-mediated transformed calli. In addition, the effect of hygromicin and bialaphos has been evaluated as selective agents for transformed cells. The former was shown to be useful, whereas the latter was not. These results have yet to be published. In vitro manipulation of coconut tissues is limited by the scarce knowledge of their cellular behaviour. A major problem is the difficulty to maintain the meristematic potential of tissues and to further control their capacity for cell division. Therefore, the L’Institut de Recherche pour le Développement - Centre de Cooperation Internationale en Recherche Agronomique pour le Developpement (IRD-CIRAD) started a study on the cell cycle status of in vitro coconut cells. Using flow cytometry, most of the cells were found to be in G0/G1 phase (around 90% in nodular calli and shoot meristems), with a low mitotic index (less than 0.5%) (Sandoval et al. 2003). These results are in agreement with those obtained by Jesty and Francis (1992) with microdensitometry. Adding aphidicolin (a synchronisator of cell cycle) to the media, around 80% of cells were blocked in G0/G1 and only 20% of meristematic cells were cycling cells (Sandoval et al. 2003). Using immature inflorescences and immature leaves, the study showed that flow cytometry methods could be used to rapidly assess the ability of tissues cultured in vitro to divide. It appears to be a useful tool for a more effective monitoring of the meristematic potential of tissues cultured in vitro, in relation to culture conditions. The basic components of the cell cycle machinery appear to be conserved in all eukaryotes and particularly those controlling the re-entry of cells in the cell cycle (transition between G0 and G, transition between G1 and S). Screening a coconut shoot meristem cDNA library with 341 CHAPTER 5: Germplasm use heterologous probes (from Maize, Arabidopsis and mouse) allowed to isolate cDNA (Cyclin D, cyclin dependant kinase, E2F and retinoblastoma) involved in the retinoblastoma pathway known to control the re-entry of cells into cell cycle and the early cell cycle phases. Among the genes isolated from coconut, those encoding D-type cyclins are of great interest because they are known as favourable candidates for linking the perception of the environment (culture conditions) with cell cycle activity in plants. The study of the accumulation of these cDNA in in vitro coconut tissue is now on the way through collaboration between IRD-CIRAD and CICY. It should help to understand the mechanisms controlling the switch from non-cycling cells to cycling cells. Perspectives and conclusion The account presented here of the research for the development of efficient coconut micropropagation protocols via somatic embryogenesis shows that solid progress has been made and that this has been possible because there has been collaboration among institutions all over the world, particularly in the last ten years. Rapid progress has been made using plumule explants, but there is still work to be done. For instance, there is a need for improving embryo development, mastering germination and post-germination development and continuing genetic stability/integrity testing. Interest in plumule micropropagation started because this could be a useful model system, thus developments obtained with this system should now be tested with other explants such as immature inflorescences and young leaves. From a practical point of view, plumule micropropagation cannot be used for elite plant propagation. However, it can be applied for superior population propagation such as genotypes that are disease resistant. Countries affected by the phytoplasma- associated diseases need urgently at least hundreds of thousands of resistant palms. In the near future, plumule micropropagation could be the way to obtain them. Another application for the plumule system is the multiplication of the Makapuno coconut currently produced by rescuing the embryo of the non-germinating Makapuno nut. On the other hand, research work on coconut somatic embryogenesis should also incorporate the latest trends in developmental biology as they become available and in particular those concerning the control of embryogenesis and shoot meristem differentiation and functioning. As mentioned above, transformation protocol development is already under way based on plumule micropropagation. Regarding the search for genes, there are recent reports on interesting genes that have been isolated from Arabidopsis such as BAYBY BOOM (Boutilier et al. 2002) and LEC1 (Lotan et al. 1998; Stone et al. 2001) encoding transcription factors involved in 342 COCONUT GENETIC RESOURCES the conversion from vegetative to embryonic growth. The over-expression of these genes in Arabidopsis led to the formation of somatic embryos at the surface of the leaves with a high rate. Such genes are attractive and are promising tools for improving somatic embryogenesis and clonal propagation in coconut. Finally, it is very important that for future research efforts, collaboration among institutions in different countries is intensified, not only to sustain current progress in coconut micropropagation research, but also to allow it to take place rapidly. To successfully achieve this, the continuing support of Asian Pacific Coconut Community (APCC), Bureau for the development of research on tropical perennial oil crops (BUROTROP) and the International Coconut Genetic Resources Network (COGENT), is absolutely necessary. Acknowledgement The authors would like to thank GTZ, UE, CONACYT, UNESCO, ACIAR, BUROTROP, COGENT and The French Ministry for Teaching and Research for the funding and help in general provided to support the research collaborative efforts reported here. References Adkins, SW, YM Samosir and ID Godwin. 1999. 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Biotech. 76:569-574. 346 COCONUT GENETIC RESOURCES Triques, K, A Rival, T Beulé, M Puard, J Roy, A Nato, D Lavergne, M Havaux, JL Verdeil, A Sangare and S Hamon. 1997a. Photosynthetic ability of in vitro grown coconut (Cocos nucifera L.) plantlets derived from zygotic embryos. Plant Sci. 127:39-51. Triques, K, A Rival, T Beulé, S Dussert, V Hocher, JL Verdeil and S Hamon. 1997b. Developmental changes in carboxylase activities in in vitro cultured coconut zygotic embryos: comparison with corresponding activities in seedlings. Plant Cell. Tiss. Org. Cult. 49:227-231. Verdeil, JL, C Huet, F Grosdemanges, A Rival and J Buffard-Morel. 1992. Coconut (Cocos nucifera L.) somatic embryogenesis: Obtention of several clone ramets. Oléagineux 47:465-469. Verdeil, JL, C Huet, F Grosdemange and J Buffard-Morel. 1994. Plant regeneration from cultured immature inflorescences of coconut (Cocos nucifera L.): Evidence for somatic embryogenesis. Plant Cell Rep. 13:218-221. Verdeil, JL, V Hocher, C Huet, F Grosdemange, N Michaux-Ferrrier and M Nicole. 2001. Infrastructural changes in coconut calli associated with the acquisition of embryogenic competence. Annals Bot. 88:9- 18. Zizumbo, D, M Fernández, N Torres and R Cardeña. 1999. Lethal yellowing resistance in coconut germplasm from México. Pp 131-144. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardeña and J Santamaría (eds). Current advances in coconut biotechnology. Kluwer Academic Publishers, Dordrecht, The Netherlands. 347 CHAPTER 5: Germplasm use Chapter 6 Major pests and safe movement of germplasm 348 COCONUT GENETIC RESOURCES 349 CHAPTER 6: Major pests and safe movement of germplasm Coconut lethal yellowing C Oropeza1, JA Escamilla1, G Mora2, D Zizumbo1 and NA Harrison3 1Coconut Researchers, Centro de Investigación Científica de Yucatán (CICY), Mérida, México 2Epidemiologist, Colegio de Posgraduados (CP), Montecillos, México 3Coconut Researcher, University of Florida, Fort Lauderdale, USA Introduction Lethal yellowing (LY) is a devastating disease that affects more than 38 species of palms (Harrison et al. 1999) throughout the Caribbean Region (see McCoy et al. 1983) where its effects have been more conspicuous on coconut (Cocos nucifera L.) than other palm species because of their abundance. In the last 50 years, LY epidemics in this region have killed millions of palms. The purpose of this paper is to summarize the current status of study and knowledge on LY. For more extensive treatment of particular issues discussed in this section, the reader will be referred to other related publications. Geographic distribution Reports of dying coconut palms exhibiting LY-type symptoms date back to 19th century in the Caribbean region (Eden-Green 1997). During the last three decades, epiphytotics of LY in Jamaica and Florida have been characterized by rapid spread and high losses (McCoy et al. 1983). LY was first recorded in the Yucatan Peninsula of southern Mexico during 1977 (Oropeza and Zizumbo 1997) and has since spread to Belize (Eden- Green 1997), Honduras (Ashburner et al. 1996) and Guatemala (Mejía et al. 2004). Similar diseases known as lethal yellowing-type diseases (LYD) have also been described in Africa in Ghana, Togo, Nigeria, Cameroon, Tanzania, Kenya and Mozambique (Eden-Green 1997; Tymon et al. 1998; Eden-Green and Mpunami this volume). LY had destroyed millions of palms causing great losses, particularly in Ghana and Tanzania (Schuiling et al. 1992). Symptoms The first visual symptom of the disease on infected bearing coconut palms is the premature drop of most of the fruit regardless of their developmental stage. The next symptom to appear is the blackening of new inflorescences. This symptom is most apparent as the inflorescence emerges from the spathe. The first affected inflorescences usually show partial necrosis but as the disease progresses, newer inflorescences show more extensive necrosis. Most of the male flowers are dead and no fruit are set on those affected inflorescences. Yellowing of the leaves usually 350 COCONUT GENETIC RESOURCES starts after necrosis has developed in more than two inflorescences. The pattern of leaf discoloration due to LY is more rapid than that for normal leaf senescence. The first leaves to turn yellow are the oldest (lower) ones, then yellowing advances upwards; affecting the younger middle and finally the upper leaves. Yellow leaves turn brown, desiccate and die. They remain hanging for a few days before falling. Eventually, the whole crown perishes, leaving a bare trunk or ‘telephone pole’. On the other hand, the syndrome does not always follow the same sequence of events. In some LY affected palms the spear leaf or a midcrown leaf occasionally shows yellowing prematurely (McCoy et al. 1983). Sometimes inflorescence necrosis becomes noticeable only after leaf yellowing has appeared as observed in Guatemala (Mejia et al. 2004). The estimated time lag from the probable time of initial infection by the pathogen to the appearance of first symptom has been variously reported as follows: in mature bearing palms, 230-450 days (Romney 1972) and 210-450 days (Heinze et al. 1972); for young non-bearing palms, at least 240-270 days (Dabek 1974). The time between probable initial infection and death of mature palms has been reported as 3-6 months (Grylls and Hunt 1971) or 4-5 months (McCoy 1973). In addition to these symptoms in above ground parts, roots also show necrosis, which becomes more extensive as the disease progresses (Eden-Green 1979). The growth is also affected by LY. Detailed studies on LY affected coconut palms have revealed physiological and biochemical symptoms (Oropeza et al. 1995; Islas-Flores et al. 1999; Martínez et al. 2000; Maust et al. 2003). In general, LY symptoms in other palms are similar but there are some differences (see McCoy et al. 1983). Symptoms of coconuts affected by lethal yellowing-like diseases (LYD) in West Africa and Tanzania are similar to those described here for LY in the Americas (Mpunami et al. 1999). Causal agent Phytoplasmas (previously known as mycoplasma-like organisms or MLO) were first found to be associated with some plant yellows diseases during the 1960s (Doi et al. 1967; lshiie et al. 1967). These results then sparked the search for phytoplasmas in LY-affected palms. In 1972, three groups independently reported their occurrence in the phloem of coconut palms showing LY symptoms (Beakbane et al. 1972; Heinze et al. 1972; Plavsic- Banjac et al. 1972). A cause-effect relationship between phytoplasmas and LY was supported by the differential response of LY diseased palms to antibiotics. LY palms treated with penicillin showed no beneficial response whereas symptom remission occurred when they were treated with oxytetracycline (Hunt et al. 1974; McCoy 1972). Genotypic characterization of coconut-infecting phytoplasmas was possible when 351 CHAPTER 6: Major pests and safe movement of germplasm pathogen-specific PCR and RFLP-typing or sequence analysis of PCR- amplified rDNA becomes available (Harrison et al. 1994a). In this way a Florida strain of the LY agent was assigned as a sole representative member to 16Sr group IV (coconut lethal yellows group), subgroup A (16SrIV-A) (Harrison et al. 1994b). Phytoplasmas have also been associated with LYD in West Africa (Dollet et al. 1977; Epko and Ojomo 1992) and East Africa (Schuiling et al. 1992; Eden-Green and Mpunami, this chapter). Phytoplasmas associated with Caribbean LY and both West and East African LYD were analyzed using molecular techniques and found to be different but closely genetically related (Harrison et al. 1994a). Further studies on phytoplasmas associated with LY-diseased palms in the Americas have shown that there are genetic differences when different locations were compared. Differences were found between the LY phytoplasmas in Florida, México and Jamaica based on the analysis of rDNA (Harrison et al. 2002b); and between Cuba and Mexico based on the analysis of non-ribosomal DNA (Llauger et al. 2002). In addition, strain diversity within a country has been found in Cuba (Llauger et al. 2002), México (Harrison et al. 2002a) and Florida (Harrison et al. 2002b). Also further studies on the East Africa LYD using rRNA and RFLP analysis showed no differences between the phytoplasmas associated with Tanzania and Kenya LYD, whereas those associated with Mozambique LYD were found to be different and closely related to those from West Africa, Cape St. Paul Wilt (CSPW) in Ghana, and Awka or bronze leaf wilt in Nigeria (Mpunami et al. 1999; Eden-Green and Mpunami, this chapter). Since the 1980’s in Florida and Jamaica, unusually high losses of resistant Malayan Dwarfs and MayPan hybrid coconuts have been reported (Howard et al. 1987). Comparative analyses of the LY phytoplasma 16S rRNA gene by PCR, RFLP and base sequencing have shown that LY phytoplasma population in Jamaica was homogeneous and varied from the Florida strains (Harrison et al. 2002b). The authors conclude that phytoplasma rDNA heterogeneity is probably not correlated with the strain variation in aggressiveness. Transmission According to the pattern of spread of LY, it was first hypothesised that it was probably transmitted by flying insects (Johnston 1912). When the causal agent of LY was discovered to be a phytoplasma, the search concentrated on species of Auchenorrhyncha, the sub-order of the Homoptera to which most vectors of phytoplasma-associated diseases belong (Tsai 1979). Surveys conducted in LY-affected areas in Jamaica yielded five species of fulgorids (Schuiling 1976; Schuiling et al. 1976) 352 COCONUT GENETIC RESOURCES and in Florida two fulgorids and one membracid (Howard 1980a; Howard and McCoy 1980; Howard and Mead 1980). The only common species found in coconut palms in both locations was the cixiid Myndus crudus. In addition, the apparent rate of spread of LY decreased in areas where M. crudus populations were reduced by insecticide treatment (Howard and McCoy 1980) and populations of M. crudus were 40 times higher in heavily affected areas than in LY-free areas in Florida (Howard 1980b). As a result of these reports, M. crudus has been extensively tested as a vector for LY (Howard 1995). Successful transmission was achieved in Florida using caged palms (C. nucifera, Veitchia merrillii and Pritchardia thurstonii) exposed to large numbers of insects and long incubation times (Howard et al. 1982; Howard and Thomas 1980; Howard et al. 1984). Every month, approximately 850 wild M. crudus captured from landscape palms were introduced into each cage over a 34-month period. Palms in cages where insects were not introduced remained healthy. More recently, detection by PCR of LY-phytoplasma infection on native M. crudus in Florida was reported (Harrison and Oropeza 1997). Taken together, these studies indicate the importance of this planthopper as a vector of LY in Florida, but its role as a vector of the disease elsewhere in the Americas remains uncertain. At LY-active sites in southern Mexico, the number of M. crudus on coconut palms were found to be several-fold lower than those of many other potential vectors (Escamilla et al. 1994). The possibility of an indirect transmission path, through embryos or asymptomatic alternative host plants, has been considered. DNA of the LY phytoplasma has been detected in embryos from fruits of diseased Atlantic Tall coconut palms by DNA hybridization (Cordova 1994), and PCR analysis (Cordova et al. 2003). Phytoplasma distribution in sectioned tissues from PCR-positive embryos determined by in situ PCR and digoxigenin-11-deoxy-UTP (Dig) labelling of amplification products was limited to areas corresponding to the plumule and cells ensheathing it (Cordova et al. 2003). By comparison, similarly treated embryo sections derived from fruits of a symptomless Atlantic Tall coconut palm were consistently devoid of any label. Occurrence of LY phytoplasma DNA has been shown most recently in embryos from fruits at different stages of development (Chumba 2003). Presence of phytoplasma DNA in coconut embryo tissues suggests a potential for seed transmission which remains to be demonstrated. The palms Thrinax radiata and Cocothrinax redii that have been listed as not susceptible by McCoy et al. (1983) and were not listed as susceptible by Harrison et al. (1999) are very common on the coastal areas of Yucatan where most of the coconut palms have been killed by LY. Analysis by PCR with LY-specific, non-ribosomal primers (Harrison et al. 1994a) of symptomless palms of these species 353 CHAPTER 6: Major pests and safe movement of germplasm resulted in positive detection of the LY phytoplasma in most of them (CICY, 2004, Mexico, unpublished results). Therefore, these palms could serve as potential reservoirs of LY phytoplasma for acquisition by vector insects. Diagnosis and detection Visual symptom progression on palm hosts allows for a tentative diagnosis of LY disease. However, confirmation of disease requires detection of the LY phytoplasmas in host tissues. This can be achieved directly by their observation in preparations of diseased tissue using transmission electron microscopy or indirectly by observation of their DNA, using DAPI staining and epifluorescence microscopy. These techniques are not specific since they cannot differentiate one phytoplasma from another. However, diagnosis can also be supported by the differential responses of diseased palms to the antibiotics penicillin and oxytetracyline, which provide evidence of a cause-effect relationship of phytoplasma infection and LY. Recent progress in the development of molecular diagnostic assays based upon DNA probe hybridisation and PCR has significantly enhanced detection of phytoplasmas, especially in woody perennial plant hosts such as coconut palm which usually contain low pathogen concentrations. These techniques are also highly sensitive and specific, and well suited for assessing large numbers of samples. LY-specific DNA probes were developed by Harrison et al. (1992) and have been used for detection and identification of the LY phytoplasma in symptomatic coconut and several other species (Harrison et al. 1992; Harrison et al. 1994b; Escamilla et al. 1995). PCR assays have been developed for the amplification of rDNA and non-ribosomal DNA for the detection of the LY phytoplasmas (Harrison et al. 1994a, b). Assays can be coupled with RFLP-typing or sequence analysis of PCR-amplified rDNA for genotypic characterization of LY phytoplasmas as detailed above. These techniques, particularly PCR, have been used for studies on the plant-pathogen- vector-environment relations also. One such study investigated the time- space distribution of phytoplasmas throughout the coconut palm. The results confirmed that they are detectable in all growing parts except mature leaves, which are actively exporting photosynthates (Cordova 2000). These results have helped determine what parts of palms are most useful for sampling for diagnostic purposes. The trunk was found to support detectable phytoplasma concentrations even before symptoms appear. Currently, this is the most common sampled tissue because of its convenience (Harrison et al. 1999). 354 COCONUT GENETIC RESOURCES Spread Two types of spread of LY have been reported in Jamaica and Florida (see McCoy et al. 1983). One involves a local centre of infection that appears in one or two palms only, followed by new cases appearing at random around the initial centre, thereby extending local spread. The second type is a jump spread followed by local spread. The jump distance varies from a few to 70 km or more (Carter, 1964). McCoy (1976) noted that the rate of long distance spread of LY in Jamaica appeared to be slower than in Florida. It took more than 60 years to cover the distance between the west and east end of the island, whereas in Florida it jumped from Miami to Palm Beach and Naples and to Nassau in the Bahamas within three years. McCoy (1976) considered that the mountainous terrain of Jamaica probably contributed to the slower rate of long distance dispersal whereas Florida has no barriers to air-borne dispersal. In Mexico, LY spread about 900 km westward from the Cozumel-Cancun area, where it was first observed in 1979, to the Campeche-Tabasco border in 15 years (Escamilla et al. 1995). The account by McCoy (1976) of a survey of LY spread in Dade County, where it first appeared in mainland Florida in late 1971, illustrates patterns of spread for a locality. Of the estimated original coconut palm population of 350 000, 0.015% of the palms were already diseased when the survey began; 0.6% by the end of 1972; nearly 6% by autumn of 1973; 50% by the end of 1974; and 75% by the end of 1975. Regarding LY tree-to-tree spread, in Dade County in the first eight months after arrival of the disease when only a small portion of the area was affected, each infected palm served to inoculate an average of 4.6 new palms according to McCoy et al. (1983). Two years later, when the logarithmic stage of spread was well underway, each infected palm served to infect 9.3 new palms. The author considered this increase as a result of the greater availability of inoculum in relation to the remaining uninfected palms. The type of locality was also found to affect the rate of the spread of LY. The highest rate was found in inland groups of palms receiving regular irrigation and fertilization; the lowest rate occurred adjacent to the ocean, even with high maintenance; and intermediate rates in inland sites receiving minimal maintenance (McCoy 1976). Studies carried out in Yucatan determined the LY spread gradients within a coconut grove and between coconut groves, as well as the palm to palm spread pattern. It was found that within a grove as the LY incidence or proportion of infected palms in an outbreak grows, the greater is the distance the disease spreads from the outbreak; and that it does so as a symmetrical radial gradient (Gongora et al. 2001). For longer distances, dispersal between groves gradients were asymmetrical and depended on the prevailing wind direction. Since the prevailing direction 355 CHAPTER 6: Major pests and safe movement of germplasm is east-west in this part of Mexico, LY spread was greater to the west than to the east (Mora and Escamilla 2001). Regarding the pattern of palm to palm spread, when LY was studied by following visual symptoms, disease was randomly distributed within the first 10 months, started to form aggregates after 12 months and eventually it was found to be uniformly distributed throughout the study area (Perez et al. 2000). However, use of PCR detection shows aggregate formation when, according to symptoms, distribution was random (Canché 2002). Moreover, spatial autocorrelation analysis based on visual symptoms indicated that a diseased palm can infect adjacent palms situated as far as eight lags (a lag is the separation between two palms, in this case 8 m) (Escamilla and Mora 2003). This was also confirmed by PCR analysis (J. Escamilla 2004, personal communication). Studies on CSPW disease in coconut plantations in Ghana have also revealed that in almost every case the disease first occurs randomly on isolated palms, spreading to the entire plot in patches and then little by little to all the coconut plantations in a given region (Dery and Philippe 1997). It can also spread in jumps of varying distances (Dery and Philippe 1997). Control methods Despite decades of research, a cure for LY is not yet available, but measures may be taken to attempt to reduce its rate of spread. Current and potential methods include quarantine, chemotherapy, vector control, sanitation and the use of resistant varieties. Control of the vector has been approached using insecticides. In tests carried out by Howard and McCoy (1980), and Reinert (1977), M. crudus populations were reduced by insecticide treatment, but not sufficiently to be recommended for practical purposes (McCoy et al. 1983). Due to the phytoplasma nature of the causal agent of LY, antibiotics were tested in Florida (see McCoy et al. 1983) and Jamaica (Hunt et al. 1974). It was found that tetracycline group antibiotics suppressed symptom development if applied before expression of systemic foliar yellowing. Chemotherapy has been successfully used for treating host palms used for ornamental purposes, but is not feasible for commercial plantations because of its high cost and perceived health risks. According to McCoy et al. (1983), eradication of diseased palms could be useful in slowing the spread of LY if practised in the early stages of the outbreak. He noted that a major drawback of this practise is that LY has a long latent period. However, according to the current epidemiological knowledge for LY as commented in the previous section, if eradication is rigorously practised very early when an outbreak starts, its contribution to delaying disease spread could be substantial. On the other hand, although LY spreads rapidly in any locality where it has 356 COCONUT GENETIC RESOURCES become established and jumps large distances to establish new infection sites, quarantine of infested areas could retard disease spread since the greatest majority of new cases occur within 100 meters of any established case of disease (McCoy et al. 1976). Nevertheless, replanting with resistant palms has proven to be the most efficient way to deal with LY. Trials evaluating LY resistance have been performed in Jamaica (Been 1991). Resistance was found in some ecotypes, including the Malayan Dwarf varieties, of which the yellow variety (MYD) was subsequently used as a parent for F1 hybrid production. Hybrids produced with MYD as one of the parents had a level of resistance sufficient to be used for commercial planting. Based on these findings, replanting in Jamaica using MYD and the Maypan hybrids (MYD x Panama Tall) has proved successful (Been and Myrie, this chapter). From a commercial perspective, Maypan offers several advantages: it is resistant to LY, precocious and highly productive (Been 1991). Unfortunately unusually high losses of resistant MYD and Maypan hybrid coconuts have been reported (Howard et al. 1987) and recent outbreaks in northern Jamaica have recently killed up to two thirds of MYD and Maypans (see Harrison et al. 2002a). After LY arrived in Mexico, a search for resistant germplasm began. CICY collected 18 coconut populations mostly from the Pacific Coastal areas of Mexico in 1989, when LY was not present there. Resistance trials were established in Yucatan in an LY affected area. These populations were grouped into five ecotypes: Atlantic Tall, MYD, Pacific Tall 1, Pacific Tall 2, and Pacific Tall 3. After more than ten years of testing, new LY highly resistant germplasm have been identified, namely Pacific Tall 1 and Pacific Tall 2 (Zizumbo et al. 1999). They are also surviving in their original planting locations (D Zizumbo 2004, personal communication) co-existing with different LY phytoplasma strains identified there (see Harrison et al. 2002a). They are currently being used for improvement and replanting programmes in Mexico and Honduras. Testing of these ecotypes and Tall x Tall hybrids produced with them started in Jamaica three years ago to determine if they can survive the ongoing resurgence of LY outbreaks there. Future searches for additional sources of resistance could be facilitated by using microsatellite markers (Baudouin and Lebrun 2002). Conclusion Despite decades of efforts to deal with LY and LYD, these diseases are still spreading and killing palms in the Americas and Africa. In the past 50 years, LY has moved to Mexico, Belize, Guatemala and Honduras. New outbreaks in Jamaica, in particular, are very worrisome because 357 CHAPTER 6: Major pests and safe movement of germplasm resistant MYD and Maypans varieties are dying in unusually high proportions. The reasons for this newest development are as yet unknown. However, sustained research efforts on these diseases have been steadily generating novel and important information. PCR-based detection methodologies have provided more sensitive, and specific diagnostic capabilities that are well suited for assessing large numbers of samples than the previous ones. They have enabled studies toward a better understanding of the pathogen and its interactions with host palms and should facilitate studies focusing on vectors. We know now that there is strain diversity among LY phytoplasma populations and several strains have been identified and classified. From a practical stand point, PCR and improved sampling protocols have allowed more efficient monitoring of disease spread within countries and local regions. This timely information has been very important for updating quarantine programmes in affected countries. Epidemiological studies indicate that prompt eradication has potential to limit disease spread and molecular diagnostics are setting the bases for its improvement. Once again, resistant germplasm is a priority, and fortunately, new sources of resistant coconuts have been recognized in the Pacific coastal areas of Mexico. These materials are already being exploited in Mexico and Honduras and hopefully, trials in Jamaica to determine their response to the current outbreaks there will prove successful. For the future, it is necessary to extend the quest for resistant coconut genotypes to encompass the entire Pacific coast of Central America. We need a better understanding of the pathogen and to clarify the vector identity to elucidate vector-host relationships. However, in order to achieve any further advancement, it will be very important to use new techniques as they become available, for example, the microsatellite technology for the characterization of coconut germplasm. Other new avenues have already been opened. A study with the goal of sequencing the LY phytoplasma genome is underway and should, in the near future, yield invaluable information on the molecular basis of phytoplasma-palm- vector interactions and pathogen virulence mechanisms. Coupled with transformation techniques already under development, collectively, these efforts should provide a means for molecular improvement of coconut. Acknowledgement The authors would like to thank CONACYT- SISIERRA, Mexico for the partial funding of the research presented here. 358 COCONUT GENETIC RESOURCES References Ashburner, GR, II Cordova, C Oropeza, R Illingworth and NA Harrison. 1996. First report of coconut lethal yellowing disease in Honduras. Plant Disease 80:960. Baudouin, L and P Lebrun. 2002. The development of a microsatellite kit and dedicated software for use with coconuts. BuroTrop Bulletin 17:16-20 Beakbane, AB, CHW Slater and AF Posnette. 1972. Mycoplasmas in the phloem of coconut, Cocos nucifera L., with lethal yellowing disease. Journal of Horticultural Science 47:265. Been, BO. 1991. Observations on field resistance to lethal yellowing in coconut varieties and hybrids in Jamaica. Oléagineux 36:9-11. Canché, J. 2002. 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Journal of Phytopathology 1550:390-395. Martínez, S, I Cordova, B Maust, C Oropeza and JM Santamaría. 2000. Is abscisic acid responsible for abnormal stomatal closure in lethal yellowing of coconut palms? Journal of Plant Physiology 156:319- 323. Maust, BE, F Espadas, C Talavera, M Aguilar, JM Santamaría and C Oropeza. 2003. Changes in carbohydrate metabolism in coconut palms infected with the lethal yellowing phytoplasma. Phytopathology 93:976-981. McCoy, RE. 1976. Comparative epidemiology of the lethal yellowing, kaincope and cadang-cadang diseases of coconut palm. Plant Disease Reporter 60:498-502. McCoy, RE, FW Howard, M Tsai, HM Donselman, DL Thomas, HG Basham, RA Atilano, FM Eskafi, L Britt and ME Collins. 1983. Lethal yellowing of palms. University of Florida Agricultural Experiment Station Technical Bulletin No 834. Mpunami, A, A Tymon, P Jones and MJ Dickinson. 1999. Genetic diversity in the coconut lethal yellowing disease phytoplasmas of East Africa. Plant Pathology 48:109-114. Mora, G and JA Escamilla. 2001. Potencial de dispersión del amarillamiento lethal del cocotero. Una enfermedad de importancia cuarentenaria en México [Potencial dispersión of coconut lethal yellowing. A disease of quarantine importance in Mexico]. Pp 221- 225. In: EA Bolaños, H Osada and C Mendoza (eds). Memorias del XXVII Simposio Nacional de Parasitología Agrícola. IAP-UMSNH, Uruapan, México. Oropeza, C and D Zizumbo. 1997. The history of lethal yellowing in México. Pp 69-76. In: SJ Eden-Green and F Ofori (eds). Proceedings of the International Workshop on Lethal Yellowing-like Diseases of Coconut, Elmina, Ghana, November 1995. Natural Resources Institute, Chatham, UK. Oropeza, C, L Alpízar, I Islas, A Escamilla and J Santamaría. 1995. Physiology and biochemistry of lethal yellowing affected palms. Pp 65-77. In: C Oropeza, FW Howard and GR Ashburner (eds). Lethal yellowing research and practical aspects. Kluwer Academic Publishers, Dordrecht. Pérez, HO, G Mora, JA Escamilla, CC Góngora and C Oropeza. 2000. Patrón espacio-temporal del amarillamiento lethal del cocotero (Cocos nucifera L.) en Yucatán [Time-space pattern of lethal yellowing spread in coconut (Cocos nucifera L.) palms in Yucatan]. Pp 78. In: Memorias del XXVII Congreso Nacional de la Sociedad Mexicana de Fitopatología. SMF, Montecillo, Mexico. 363 CHAPTER 6: Major pests and safe movement of germplasm Plavsic-Banjac, B, P Hunt and K Maramorosch. 1972. Mycoplasma-like bodies associated with lethal yellowing disease of coconut palms. Phytopathology 58:298-299. Romney, DH. 1972. Past studies on and present status of lethal yellowing disease of coconuts. PANS 18:386-395. Reinert, JA. 1977. Field biology and control of Haplaxius crudus on St. Augustine grass and Christmas palm. Journal of Economic Entomology 70:54-56. Schuiling, M. 1976. A survey of insect populations on Cocos nucifera. (Abstr.) Principes 20:67. Schuiling, M, CG Johnson, SJ Eden-Green and H Waters. 1976. Recent attempts to find a vector associated with lethal yellowing of coconut (Cocos nucifera L.). (Abstr.) Principes 20:65. Schuiling, M, DA Kaiza and A Mpunami. 1992. Lethal disease of coconut palm in Tanzania. I. Comparison with other diseases in East Africa. Oléagineux 47:511-515. Tsai, JH. 1979. Vector transmission of mycoplasmal agents of plant diseases. Pp 266-307. In: RF Whitcomb and JG Tully (eds). The mycoplasmas Vol III. Academic Press, New York. Tymon, A, P Jones and NA Harrison. 1998. Phylogenetic relationships of coconut phytoplasmas and development of specific oligonucleotide PCR primers. Annals of Applied Biology 132:437-452. Zizumbo, D, M Fernandez, N Torres and R Cardeña. 1999. Lethal yellowing resistance in coconut germplasm from México. Pp. 183- 196. In: C Oropeza, JL Verdeil, GR Ashburner, R Cardeña and JM Santamaria (eds). Current advances in coconut biotechnology. Kluwer Academic Publishers, Dordrecht, The Netherlands. 364 COCONUT GENETIC RESOURCES Status of coconut lethal yellowing in Jamaica B Been1 and W Myrie2 1Director of Research and 2Plant Pathologist, Coconut Industry Board (CIB), Kingston, Jamaica Introduction Lethal yellowing disease (LY), probably the most devastating of diseases which affect the coconut palm (Cocos nucifera L.), is one of the greatest threats to coconut cultivation not only in the Caribbean and the Americas but worldwide. It is associated with phytoplasmas, and Myndus crudus Van Duzee is a probable vector (Beakbane et al. 1972; Plavsic-Banjac et al. 1972; Howard et al. 1983). Despite the considerable research, which has been carried out on the disease, no permanent cure has yet been found. History of LY in Jamaica LY was first reported from the Cayman Islands in 1834. In Jamaica, the disease was first observed in the south western section of the island in 1884 (Fawcett 1891). Nevertheless, it is possible that in 1872 it, or a similar disease, wiped out coconut palms along a forty-mile coastal strip in the south west. The disease continued to be endemic in the western region for decades and by 1952 had spread over the western half of the island (Martyn 1949; Nutman and Roberts 1955). The greatest damage was done in the coconut belt bordering the north west coast but the disease was not found a few miles inland, but there were not many plantations inland in that area. In 1961, LY appeared suddenly in the north eastern section of the island, over 100 km from the nearest case in the west, and subsequently spread rapidly throughout the main coconut growing region destroying existing coconut plantations. Of the estimated six million coconut palms growing in Jamaica in 1961, 90% was lost to LY by 1981. By 1981 when LY was active island wide, mortality levels of the Malayan Dwarf and Maypan were 5% and 10%, respectively (Been 1981). During the early 1980s, LY was largely confined to surviving Jamaica Tall palms and materials of uncertain origin. About the mid-1980s, at certain coastal locations in the north western region there were reports of higher than anticipated levels of LY mortality among Malayan Dwarf and Maypan populations. At some places mortalities were as high as 40%. Following a disastrous hurricane in 1988 the incidence of LY increased significantly and new outbreaks were reported in eastern Jamaica. At 365 CHAPTER 6: Major pests and safe movement of germplasm various points along the coastal areas of the northern region mortality levels among stands of the Malayan Dwarf and Maypan were found to be consistently higher than those observed in 1981. The trend continued during the 1990s and in certain places the disease began to move inland. The mortality level varies; in the western section it is generally lower than in the east where a field of 747 Malayan Dwarfs died over the period 1993 to 1999, and a population of 792 Maypan hybrids between 1997 and 2000 (Myrie 2002). In the western section, mortality rate is generally slower, about 2% per annum, than in the east, which is about 14-25% per annum (Myrie 2002). The LY disease of the 1990s had the usual characteristics of the LY of the 1970s; however, there was an interesting difference in that it attacked non-bearing palms with greater frequency. At present, the disease is largely confined to the coastal areas and its incidence on most of the inland farms is low. The main germplasm collections having survived LY of the 1960s and 1970s are still to be exposed fully to the ‘new’ LY. However, to date no variety currently being cultivated in the areas where the disease is active has shown any sign of possessing a high or any level of resistance. At two experimental sites, the following F1 hybrids – Indian Green Dwarf x Panama Tall, Ceylon Green Dwarf x Panama Tall, Ceylon Yellow Dwarf x Panama Tall and Maypan – have all failed to stand up to the disease (Wallace 2002). Indian Green and Ceylon Yellow and Green Dwarfs had shown high levels of LY resistance, almost as high as that of the Malayan Dwarf. It has been estimated that over the past decade about 800 000 palms have been destroyed by LY in the eastern section of the island and the disease continues its advance. The coconut is not indigenous to Jamaica and when LY first appeared in the 18th century, the Jamaica Tall was the principal variety being cultivated – a situation, which remained unchanged until the 1970’s. Unfortunately, it is highly susceptible to the disease. There is no record of LY affecting other palm species in Jamaica during the 18th century. Searching for the cause From the late 1880s, attempts were made to determine the cause of LY and ways of controlling its spread. Considerable research was done over the years by many workers, especially in Jamaica and Florida (Romney 1983; McKoy et al. 1983). Failure to obtain evidence that fungi, bacteria, nematodes, soil and other environmental factors were the cause led to the conclusion that LY had a viral aetiology (Bruner and Boucle 1943; Nutman and Roberts 1955). This led to attempts to transmit the disease to healthy palms and the search for an insect vector (Carter 1966; Grylls et al. 1968; Heinze 1971). 366 COCONUT GENETIC RESOURCES The discovery of phytoplasmas on LY-infected coconut palms in 1971 changed the direction of the research effort. The search for an insect vector was narrowed down to leaf hoppers and, in Florida, Myndus crudus Van Duzee was found to be a vector of the phytoplasma associated with LY (Howard et al. 1983). Living with LY before 1961 Early attempts were made to control the spread of the disease by isolation and elimination of outbreaks. Felling and burning of palms were carried out, but these failed to halt the spread of the disease. Government legislation was enacted to control the movement of plant parts and soil eastwards into the disease-free area, but the boundary was an open one, which was impossible to maintain in accordance with any proposed quarantine requirements. In disease affected areas, replanting was done with whatever planting material was available. In the early part of the century, the north western coastal strip was replanted at least four times with remnants of the fourth replanting having all but died out by 1954 (Nutman and Roberts 1955). Many growers in the eastern part of the island believed that the Jamaica Tall palms they were cultivating were of a different, resistant type. Living with LY after 1961 The sudden appearance of LY in the main coconut-growing area in the eastern end of Jamaica in 1961 posed an enormous problem for the Coconut Industry Board (CIB), which had been established in 1945 to promote the interests and efficiency of the coconut industry. Before 1945, research and extension for coconuts were done by the Ministry of Agriculture. It was realized from the outset that the additional costs of research on LY could not be provided by the CIB and, therefore, external assistance was sought and obtained. Initially, the United States Agency for International Development (USAID) provided collaborating scientists and later the Food and Agriculture Organization (FAO) of the United Nations supported the research effort. Researchers from Australia, Germany and the Netherlands came to Jamaica for extended periods to work with local staff. At about the time that the FAO project was about to end, the United Kingdom (UK) government through its Overseas Development Administration (ODA) supplied a research team and an electron microscope. Researchers from the University of Florida also worked in Jamaica. Research institutions in the UK and United States of America (USA) were involved in the research effort. The Ministry of Agriculture, University of the West 367 CHAPTER 6: Major pests and safe movement of germplasm Indies, Institut de Recherches pour Huiles et Oléagineux (IRHO) and Unilever also cooperated with the CIB. External funding ceased in the early 1980s but before then, considerable work was done and valuable information obtained. Research on the nature of LY was done largely by visiting scientists, while local staff concentrated mainly on plant improvement and management. The original local research programme had to be modified with emphasis being placed on selection and breeding for disease resistance, and the management methods needed were those suited to resistant varieties. A programme to monitor the disease was put in place to determine which palms were dying from LY as opposed to other causes. It also provided useful information on varietal resistance and data for replanting programmes. Felling of affected palms to check the spread of the disease was as ineffective as it had been in the west, and was abandoned. Pesticides were used in an attempt to control new outbreaks before they could spread, but without success. The concept was to treat all palms around a single diseased palm with insecticides so that the vectors would be killed before they could pick up the phytoplasmas from infected palms that had no symptoms. Once phytoplasmas had been found in diseased palms, it was realized that tetracycline could be used to suppress LY symptoms and keep palms alive for years, but it was not a permanent cure. Chemotherapy was rejected as a means of controlling the spread of LY in Jamaica because of the health hazard and high costs. In 1961, the vast majority of the commercially-grown palms in Jamaica consisted of the LY-susceptible Jamaica Tall variety but there were also populations of the Malayan Dwarf and Panama Tall which were grown from seednuts imported after hurricanes destroyed coconut stands earlier in the century. Realization by the mid-1950s that the Malayan Dwarf palms (all three colour forms) were highly resistant to LY led to the search for other resistant material and the introduction and establishment of a large germplasm collection in Jamaica by the CIB during the 1960s. These introductions were screened for resistance in field trials. When it became obvious that none of the introductions was more resistant than the Malayan Dwarf, a hybridization programme was started in an attempt to combine in the F1 the high disease resistance of the Dwarf with the large fruit size and hardiness of the Talls. One of the early crosses (Malayan Dwarf x Panama Tall or Maypan) was found to be productive and resistant, and a system was devised to produce it commercially. Once the resistance of the Malayan Dwarf had been sufficiently 368 COCONUT GENETIC RESOURCES proven, thousands of mother palms were selected for regular seed production. From then on, even in areas not yet attacked by the disease, the Malayan Dwarf was planted instead of the Jamaica Tall. By 1974, the Maypan had shown sufficient disease resistance and productivity and was released to growers. In an attempt to speed up replanting, the Advisory Section of the CIB was expanded and seed gardens and nurseries established at numerous sites for the effective and efficient distribution of seedlings. In addition, assistance programmes under which growers received free planting material and fertilizer, and cash grants to help with weed control were instituted. As a consequence of these efforts by the CIB, three million resistant seedlings were distributed by 1979 and by 1988, ten million. Current status of LY and future prospects The coconut industry has remained viable through the use of varieties with good disease resistance, but it now appears as if this resistance is being overcome. The main varieties being cultivated – the Maypan F1 hybrid and the Malayan Dwarf – are showing little resistance to the resurgent LY, and preliminary observations suggest that other locally developed F1 hybrids and germplasm recently introduced may not be any better. It is possible that the causative agent of LY has mutated and/or exceptional environmental conditions may be combining to produce situations favourable to the development and spread of the disease. The CIB realizes that if the local industry is to remain viable and survive, it is imperative that some ways be found to cope with LY. In response to the resurgence of LY, assistance was sought from many sources including the Common Fund for Commodities (CFC) and FAO. As a consequence, the CFC funded an ‘Expert Consultation on Sustainable Coconut Production through Control of Lethal Yellowing’ which was held in Jamaica in 2002, and a project entitled ‘Sustainable Coconut Production through Control of Lethal Yellowing Disease’ was submitted to the CFC and, following its approval, is now being implemented. The current activities of the CIB related to LY include: • Monitoring the disease and studying its epidemiology; • Characterization of the pathogen and host; • Identification and characterization of insect vector(s) of LY (in collaboration with the University of the West Indies); • Screening of existing local populations and new F1 hybrids, and introduced germplasm for LY resistance; and • Encouraging the planting of coconut palms in areas not currently affected by LY and intensify intercropping. 369 CHAPTER 6: Major pests and safe movement of germplasm At present, the CIB does not have any variety which has proven resistance to the LY of the 1990s, but seedlings of the Maypan and Malayan Dwarf are being made available to farmers free or at subsidized prices. Farmers are given regular updates of the disease situation and told that it might not be advisable to replant until disease activity in their areas has abated. The answer to LY may well lie in the realm of genetic engineering but, in the meantime, conventional methods of plant breeding will have to be used and every effort made to develop an integrated approach to disease control. References Beakbane, AB, CHW Slater and AF Posnette. 1972. Mycoplasma in the phloem of coconuts, Cocos nucifera L., with lethal yellowing disease. Journal of Horticultural Science 47: 265. Been, BO. 1980. Observations on field resistance to lethal yellowing in Jamaica. Oléagineux 36:9-11. Bruner, SC and L Boucle. 1943. La enfermedad conocida como “enfermedad del cogollo del cocotero Cuba.” Rev. Agric. Cuba 26:132- 141. Carter, W. 1966. Lethal yellowing disease of coconuts (Report to Govt. of Jamaica). FAO Rome, TA 2158. Fawcett, W. 1891. Report on the coconut disease of Montego Bay. Bulletin of the Botany Department Jamaica 23:2. Grylls, NE, P Hunt and NA Bor. 1968. Investigations on the aetiology of lethal yellowing disease of coconuts in Jamaica 1. Preliminary results of virus transmission tests and bacterial inoculations. Pp. 15-20. In: Proceedings of the 3rd Session of the FAO TWP on Coconut Production and Processing, Indonesia. Jakarta, Indonesia. Heinze, KG. 1971. Report to the Government of Jamaica and to FAO on lethal yellowing disease of coconut. Howard, FW, RC Norris and DL Thomas. 1983. Evidence of transmission of palm lethal yellowing agent by a plant hopper Myndus crudus (Homoptera: Cixildae). Tropical Agriculture 60: 168-171. Martyn, EB. 1949. Further observations on the “unknown” disease of coconuts. Tropical Agriculture (Trinidad) 26:110-112. Myrie, WA. 2002. Current outbreaks of lethal yellowing in Jamaica and the use of molecular diagnostic techniques in phytoplasma detection. Pp 107-115. In: Proceedings of the expert consultation on sustainable coconut production through control of lethal yellowing disease. CFC Technical Paper No. 18. Amsterdam, The Netherlands. McKoy, RE, FW Howard, JH Tsai, HM Donzleman, DL Thomas, HG 370 COCONUT GENETIC RESOURCES Basham, RA Atilano, FM Eskafi, L Britt and ME Collins. 1983. Lethal yellowing of palms. University of Florida Agricultural Experimental Bulletin No. 834. Pp. 60-65. Nutman, FJ and FM Roberts. 1955. Lethal yellowing: The unknown disease of coconut palms in Jamaica. Empirical Journal of Experimental Agriculture 23: 257-267. Plavsic-Banjac, B, P Hunt and K Maramorosch. 1972. Mycoplasma-like bodies associated with lethal yellowing of coconut palms. Phytopathology 63:298-299. Romney, DH. 1983. Brief review of coconut lethal yellowing. Indian Coconut Journal 13:1-8. Wallace, M. 2002. Coconut breeding programme for lethal yellowing resistance in Jamaica. Pp. 118-127. In: Proceedings of the expert consultation on sustainable coconut production through control of lethal yellowing disease. CFC Technical Paper No. 18. Amsterdam, The Netherlands. 371 CHAPTER 6: Major pests and safe movement of germplasm Indexing and pathogen characterization SJ Eden-Green1 and AA Mpunami2 1Manager, Crop Protection Programme, University of Greenwich Natural Resources International (NRI), United Kingdom 2Principal Agricultural Research Officer, Mikocheni Agricultural Research Institute (MARI), Dar es Salaam, Tanzania Introduction Safe movement means movement of coconut germplasm without introducing pests (taken here to include diseases), or at least those pests for which there are perceived risks of harmful or undesirable effects following introduction to a previously unaffected region. This section is concerned primarily with pests of quarantine significance (mostly diseases) that are likely to be spread through planting materials derived directly from parent plants which appear to be healthy to the naked eye but might be infected without showing symptoms. These include several intractable and lethal diseases which are difficult to diagnose or characterize; some of unknown or uncertain aetiology. Detection of the causal agents may require the use of laboratory diagnostic tests or indexing either of the planting material itself or of the parent population from which it is derived. As yet, most lethal diseases have a limited geographic distribution although different strains of the pathogen may be present both within and between regions that are considered to be affected by the same disease. This emphasises the need for careful and responsible attention to phytosanitary issues when moving germplasm and, in particular, for robust and sensitive diagnostic techniques and accurate characterization procedures to distinguish between strains that could be spread in planting materials. Pests of concern to the safe movement of coconut germplasm Pests of quarantine concern are reviewed in the FAO/IBPGR technical guidelines for the safe movement of coconut germplasm (Frison and Putter 1993; Frison et al. 1997). Adherence to the general phytosanitary principles recommended for the movement of coconut germplasm as seednuts, embryo cultures and pollen eliminates the risk of spread of most arthropod pests and fungal pathogens, although alternatives to fumigation of seednuts by methyl bromide will have to be found as the use of this chemical is phased out under the Montreal Protocol on substances that deplete the ozone layer. This section summarises the more sophisticated diagnostic or characterization techniques that are required to implement the guidelines for diseases associated with phytoplasmas, viroids, viruses, 372 COCONUT GENETIC RESOURCES protozoa, nematodes and draws attention to the risks posed by diseases of yet unknown cause. Phytoplasma diseases Phytoplasmas (formerly known as mycoplasma-like organisms, MLO) have been associated with diseases of coconut and certain other palms in most of the continental coconut growing regions. Lethal yellowing (LY) was originally recognised over 100 years ago in the northern Caribbean (Jamaica, Cayman, Cuba, Hispaniola and southern Bahamas) but is now present in southeastern USA (Florida, Texas), Mexico, Belize and Honduras. Similar lethal yellowing-like diseases (LYD) have been reported from Africa since the early 1900s where they have become known by various local names: Cape St Paul wilt disease (CSPWD) in Ghana, Kaincopé disease in Togo, Awka or bronze leaf wilt in Nigeria, Kribi disease in Cameroon, lethal disease in Tanzania, Kenya and Mozambique (Eden-Green 1997a). All of these show symptoms similar to LY. In Asia, phytoplasmas have recently been detected in coconut palms affected by lethal diseases, but not in symptomless palms, particularly in Indonesia (Kalimantan wilt in Central Kalimantan, Natuna wilt in Natuna Islands) (Allorerung et al. 1999). Some of the symptoms of these conditions differ from those of LY and LYD but resemble those reported elsewhere in Southeast Asia (Sumatra, Malaysia, Socorro wilt in the Philippines). All of these diseases may prove to have similar phytoplasma aetiology (Eden-Green 1997b). In India, electron microscope observations showed of an association of phytoplasmas with coconut root wilt and Tatipaka wilt diseases, and application of tetracycline antibiotics reportedly caused remission of root (wilt) symptoms (Solomon 1997). However, attempts to confirm the presence of phytoplasmas by PCR have not been successful (Harrison and Jones 2003). In West Africa, phytoplasmas have also been implicated in a blast disease of seedling coconut palms that appears to have no association with LYD (Julia 1979). Earlier diagnoses were based on electron microscope examination of ultrathin sections, supported by observation of remission of symptoms following application of tetracycline antibiotics. These remain as valuable diagnostic tools but molecular methods, based on PCR amplification of DNA with specific primers and characterization of the products, now provide more practical and sensitive means to detect the pathogen. These techniques have revealed a considerable genetic diversity of putative strains of phytoplasmas that has not yet been related to phenotypic characteristics. However, differences in the field susceptibility of coconut varieties to diseases in different regions have been known for some time, suggesting that these genetic differences may be of quarantine significance. 373 CHAPTER 6: Major pests and safe movement of germplasm Indexing and pathogen characterization Until molecular properties can be associated with phenotypic characteristics such as pathogenicity and host range, observations on symptoms, varietal susceptibility and alternative hosts remain important characteristic features and should not be neglected. However, molecular techniques are the methods of choice for sensitive and specific diagnosis and characterization of phytoplasmas. DNA probes have been developed for Caribbean LY and East African LYD phytoplasmas (Harrison et al. 1992; Mpunami et al. 1997; Mpunami 1997) and can be used in dot blot hybridization tests to detect phytoplasmas in palm tissues. However, these probes suffer from problems of background hybridization to healthy coconut DNA and have been generally superseded by DNA amplification by polymerase chain reaction (PCR) using oligonucleotide primers (Harrison et al. 1994a). Specific primers for the detection of LYD phytoplasmas in East Africa were subsequently developed (Rohde et al. 1993). As well, Mollicute-specific PCR primers were optimized for amplification of LYD DNA from palms infected by East and West African coconut yellowing diseases (Tymon et al. 1997; Mpunami et al. 1997). The PCR technique has been used to confirm the phytoplasma aetiology of the LYD diseases in Kenya and Mozambique, for routine detection of incubating infections in palm tissue, in coconut embryos and in insects suspected to be potential vectors of the disease in Tanzania (Mpunami 1997; Mpunami et al. 2000;). Similarly, LYD phytoplasma DNA has been detected in the embryos of nuts harvested from diseased palms, but carry over to palm sprouts has not been established. Higher specificity for detection of the LYD diseases in East and West Africa has been achieved by the use of primers based on the nucleotide sequence of the intergenic region between 16S and 23S rRNA genes for each LYD isolate (Tymon and Jones 1997). Specific detection of the West African LYD has since been routinely carried out by using specific primers in PCR reactions (Quaicoe et al. 2000). Similarly, specific primers for the LYD phytoplasmas in East Africa have been useful for detection and for differentiation of LYD strains within the region. By these means, it has been shown that the phytoplasma strains responsible for disease in Kenya and Tanzania are identical, but the isolate from Mozambique is different, and is more closely related to the isolates from West Africa (Mpunami 1997; Mpunami et al. 1999). Procedures that enhance the PCR technique have also offered increased specificity and sensitivity for detection of the pathogen. For example the nested PCR technique (Haqqi et al. 1988; Steffan and Atlas 1991) utilizing Mollicute-specific primers (Deng and Hiruki 1991; Namba et al. 1993) in the first reaction, and phytoplasma specific primers 374 COCONUT GENETIC RESOURCES (Gundersen and Lee 1996) in the second reaction is used for routine detection of the LYD phytoplasma in Tanzania. The PCR assays have been particularly useful for determining the genetic relationships between the LYD isolates from East and West Africa, and their relationship to the LY phytoplasma. By analyzing the restriction fragment polymorphisms of amplified PCR products, it has been shown that the isolates from West Africa are similar but genetically different from those causing disease in East Africa and the Caribbean region (Harrison et al. 1994b; Tymon et al. 1998). The technique has also demonstrated that the LYD strains in Ghana and Nigeria are genetically identical, but different though similar to the LYD strains in East Africa (Tymon et al. 1997). The Caribbean diseases appear to exist as a group of closely related strains that are most closely related to a phytoplasma associated with declines of coconut and Carludovica palmata in southern Mexico (Cordova et al. 2000) and Phoenix canariensis in Texas (Harrison et al. 2002). The relationship of phytoplasmas recently associated with Porroca disease in southern Panama and northern Colombia (http://review.ucsc.edu/winter-03/ panamas.html) has not yet been reported. The significance of these techniques lies in the ability to determine for what materials, and between which countries there are potential quarantine risks and, potentially, to assess whether host resistance observed in one region is likely transferable to another. Sensitive detection procedures also provide a means to monitor the persistence of the presumed pathogen in seedlings derived from diseased palms to resolve the question of whether or not there is a risk of spread of the disease in seednuts. As knowledge on the variability and distribution of LYD phytoplasma strains improves, it should become possible to base quarantine decisions on the local strains present and to facilitate safe movement of germplasm between regions affected by the same strains, avoiding the need for decentralised diagnostic facilities within importing countries. However, recent experience in Jamaica suggests that new strains of the pathogen can arise that are able to invade previously- resistant hosts, and molecular characterization tests do not yet allow strains to be differentiated on the basis of host specificity. Problems and research needs Lethal yellowing type diseases are recognized as the biggest threat to coconut production. Disease resistance is the only feasible method of control and has been used with great effect in the Caribbean region although there is evidence that this has now broken down in Jamaica (CFC 2002). Although specific pathogen characterization techniques have been developed, the mode of transmission has not been established in 375 CHAPTER 6: Major pests and safe movement of germplasm several regions and the suspected insect vectors have not been confirmed. One of the biggest uncertainties is whether seed transmission is possible. Available (and largely circumstantial) evidence suggests that if this can occur at all then it is extremely rare, and provided the recommended guidelines are observed then any risk should be eliminated. However, recent reports of persistence of phytoplasmas, or at least phytoplasma DNA, in coconut embryos collected from diseased palms make this an important topic for research. Virus disease: Vanuatu wilt or coconut foliar decay Coconut foliar decay (CFD) affects introduced coconut palm cultivars in Vanuatu. It is caused by a single stranded DNA (ssDNA) virus (Randles et al. 1986, 1987) transmitted by the plant hopper Myndus taffini (Homoptera: Cixiidae; Julia 1982). Affected palms typically show a normal apex, several yellowish fronds, then several, young dead fronds hanging through green older fronds. The trunk generally narrows and may thicken again if remission occurs, as happens in tolerant palm varieties. Susceptible cultivars die between one and two years after symptoms appear. The Malayan Red Dwarf (MRD) is highly susceptible to CFD but the local Vanuatu Tall is highly tolerant (i.e. can be infected without showing symptoms) and its progenies show only mild symptoms. The use of symptoms alone for CFD diagnosis is thus unreliable (Calvez et al. 1980). Indexing The MRD can be used as an indicator plant as it is highly susceptible to CFD and shows characteristic symptoms. However, detection and diagnosis are usually based on detection and partial characterization of viral ssDNA by gel electrophoresis, cDNA probes or DNA amplification and sequencing. The CFD virus ssDNA has characteristically low electrophoretic mobility in 5% polyacrylamide gels (PAGE) (Randles et al. 1986) and migrates as a single band in denaturing polyacrylamide gels, but generally as two bands in non-denaturing gels (Randles et al. 1987). A two-dimensional PAGE technique has been used to show that the DNA molecules are circular in nature (Randles et al. 1987). PAGE analysis provides presumptive diagnosis but confirmation requires DNA hybridization and/or sequencing. Pathogen characterization Purification by isopycnic density gradient centrifugation [30%–60% Nycodenz (Nyegaard, Oslo) gradient] results in the co-purification of CFD-associated DNA (CFDV DNA) and unusual, 20 nm, icosahedral particles (Nycodenz density range 1.27–1.30 g ml--1), which are considered to be coconut foliar decay virus (CFDV) particles (Randles 376 COCONUT GENETIC RESOURCES and Hanold 1989). Both CFD DNA and the particles occur in very low amounts in diseased coconut palms. Although the DNA sedimentation coefficient is only 12S to 15S, the virus has been placed in the geminivirus group. Geminiviruses have ssDNA (s20,w = 16S at pH 7.0) and contain either 1 or 2 circular ssDNA molecules of approximately 7–8 × 105 daltons mol. wt, comprising about 2700 nucleotides (Harrison 1985). Examination of purified extracts from CFD-infected palms by transmission electron microscopy shows circular molecules with mean molecular weight of approx. 4.3 × 105 daltons. A molecular hybridization assay has been developed using a P32- labelled cDNA probe synthesized from a 1203 bp DNA fragment, amplified by PCR from circular, single-stranded, 1291-nucleotide CFDV DNA (Rohde et al. 1990; Randles et al. 1992). The high specificity and sensitivity of this assay allows CFDV DNA to be detected reliably, despite its low concentration in coconut tissue. A non-radioactive probe using digoxygenin (DIG)- labelled complementary RNA (cRNA) has also been developed as an alternative detection method (Hanold and Randles 1997). Hybridization assays have been useful in studying the distribution of CFDV DNA in palm tissue in order to establish priority areas for disease diagnosis, for localization of the virus in phloem tissues (Randles et al. 1992; Hanold and Randles 1997), and for detection of the virus in the vector. To obtain sequence data on CFDV DNA, a single-stranded (ss), circular, covalently closed (ccc) DNA associated with coconut foliar decay virus (CFDV) was purified, amplified by PCR and subcloned. Its sequence was established by analysis of overlapping subgenomic cDNA clones (Rohde et al. 1990). The complete sequence comprised 1291 nucleotides and contained open reading frames for six proteins of molecular weight larger than 5 kDa. CFDV can be detected in coconut embryos and husks but not in pollen (Hanold and Randles 1997) emphasizing the need for efficient indexing of mother palms including the use of positive controls to confirm the reliability of diagnostic techniques. This is especially important given that the Vanuatu Tall, some of its hybrids and possibly other varieties are highly tolerant and can be infected without showing symptoms. Problems and research needs Much is known about coconut foliar decay virus. The insect vector is available for use in resistance screening and selection programmes for disease resistant germplasm, and sensitive diagnostic techniques are available for disease indexing. Transmission has not been reported via seed or pollen but detection of the virus in nuts and also in embryos 377 CHAPTER 6: Major pests and safe movement of germplasm collected from infected palms indicates that the risk of spread by this means needs to be investigated more intensively. Viroid diseases: Cadang-cadang and Tinangaja Cadang-cadang is a slow decline disease that results in premature death of coconut palms in the Philippines. The disease was first reported from a plantation in Albay Province on San Miguel Island in 1931. It is estimated that by 1980 it had killed more than 80 million palms on San Miguel and other neighbouring islands (Zelazny et al. 1982) but spread is generally slow (0.5 km a year). It causes yellow leaf spotting, reduced growth and reduced frond production which results in reduced crown size, cessation of nut production and eventual death within 5- 20 years. Cadang-cadang has been the major reason for prohibiting movement of Philippine coconut germplasm to many countries. A similar disease known as Tinangaja occurs in Guam. Symptoms differ slightly from those of cadang-cadang in that nuts are characteristically small, elongated and lack a kernel (Boccardo et al. 1981). The disease was first reported as a destructive disease of coconut palms in Guam in 1917 (Boccardo 1985). It apparently spread slowly and destroyed the coconut industry of Guam over the next 40 years. No commercial coconut industry has existed in Guam since 1946 but the disease is still widespread on the island, with the incidence varying from one location to another. Research on cadang-cadang disease began in the Philippines about 1950, and by 1982, the cadang-cadang viroid or CCCVd was identified as the causal pathogen (Zelazny et al. 1982; Hanold and Randles 1991a). This discovery provided a means of diagnosing the disease, and gave impetus to further research on related diseases (Hanold and Randles 1997), which showed that Tinangaja was also caused by a viroid (Boccardo et al. 1981). Indexing Cadang-cadang symptoms are not reliable means of detection and diagnosis owing to the ease with which they may be confused with the effects of other biotic and abiotic factors including diseases, pests, nutritional deficiencies, typhoons and lightning strikes. Although the disease can be mechanically transmitted to healthy coconut and other test plants (Imperial et al. 1985), indexing is impractical because the latent period between infection and appearance of symptoms is well over a year and symptoms, particularly in pre-bearing palms, tend to be non- specific. Pathogen detection and characterization by biochemical and molecular properties are thus of greatest significance for disease diagnosis 378 COCONUT GENETIC RESOURCES and the safe movement of germplasm, particularly as the CCCVd can be detected in seed husks, embryos and pollen and may be spread by these means (Hanold and Randles 1997). Pathogen characterization Viroid purification Viroids are the smallest known plant pathogens; and each consists solely of a small, circular, single-stranded, naked RNA molecule, which is infectious, and can replicate in the host cell and be transmitted independently of any other microorganism (Diener 1987). Reliable diagnosis is based on identifying the viroid RNA in extracts of coconut palms. Single-stranded RNA is very susceptible to degradation by ribonuclease, thus extraction from plant tissue involves the use of inhibitors such as phenol and SDS to minimize the risk of enzymatic degradation and antioxidants to prevent oxidation. The crude extract is then deproteinised, and viroid RNA precipitated with polyethylene glycol (PEG, mol. wt 6000). The resulting partially purified extract is ready for analysis by gel electrophoresis, or further purification by density gradient centrifugation (Hanold and Randles 1997). Gel electrophoresis Viroids migrate in most gel systems with a mobility less than that expected for their molecular weight. Various forms of CCCVd, which differ in size, are normally resolved in 5-20% polyacrylamide gels (Hanold and Randles 1997). Viroid bands are then visualized in the gel slab after staining with an appropriate stain such as silver, ethidium bromide, or toluidine blue. Increasing the temperature or pH of the gel buffer creates denaturing conditions, and causes viroids to denature from their native rod-like state to open circles and migrate more slowly than their linear forms. Diagnostic tests for viroids have been based on PAGE under both non-denaturing and denaturing conditions, and on the characteristic change of behaviour of these molecules when subjected to both sets of conditions. For example, in two-dimensional PAGE, transfer from a non- denaturing gel to denaturing conditions permits screening for all possible viroids. If used in conjunction with silver staining or molecular hybridization, the two-dimensional PAGE system is a sensitive and definitive test for the presence of small, circular nucleic acids in a preparation, and is thus a powerful tool in the detection of viroids (Hanold and Randles 1997). The procedure is however, lengthy and cannot be used on a routine basis. 379 CHAPTER 6: Major pests and safe movement of germplasm Rapid diagnosis assay For rapid diagnosis of cadang-cadang, an assay has been developed that involves three steps (Hanold and Randles 1997). Initially, sap is extracted from coconut leaf tissue and deproteinised. Then, nucleic acids are recovered by cold ethanol precipitation. Lastly, direct detection of the viroid is achieved by fractionation of the nucleic acids using PAGE and silver staining. The procedure is very sensitive, with a detection end- point of about 600 picograms of viroid. It is reliable and suitable for detecting viroids at the early stage of symptom development. Molecular hybridization After fractionation through PAGE, viroid bands on the gel slab are transferred onto a hybridization membrane and detected by molecular hybridization to a cDNA or cRNA probe (Hanold and Randles 1997). The technique is very sensitive and specific for viroid detection; cRNA probes are preferable to cDNA probes because they bind more strongly to the target RNA, giving stronger signals and thus allowing conditions of higher stringency for hybridization and reduce the non-specific binding of the probe. Electron microscopy Electron microscopy of purified samples spread under denaturing conditions can be used to identify circular viroid molecules and estimate their size (Randles and Hatta 1979). It cannot, however, be used for diagnostic purposes on tissue sections or crude extracts, since the small viroid rods and circles cannot be positively identified when mixed with other nucleic acids. Characterization of viroid RNA This requires a combination of the techniques described above. Isolation, purification, PAGE analysis, and electron microscopy can be used to establish that the nucleic acid is single-stranded RNA, circular in nature and in the size range 246 – 380 nucleotides which is typical of viroids (Keese and Symons 1987). The infectious nature can then be established by inoculating young palms with purified nucleic acids. Partial sequencing has made it possible to develop probes for specific detection of the viroids. Using these techniques, it has been shown that the cadang-cadang viroid is composed of two RNA molecules; the small 246/247 nucleotide (monomeric) and large 287/296/301 nucleotide (dimeric) forms, both of which are naked, single-stranded, circular, infectious by mechanical inoculation into coconut seedlings and are simultaneously isolated from infected palms (Randles and Hatta 1979; Mohamed et al. 1985). The small form is more abundant at the early stage of disease, but as symptom 380 COCONUT GENETIC RESOURCES development progresses, the larger form increases in quantity while the former decreases (Hanold and Randles 1997). The Tinangaja viroid (CTiVd) molecule on the other hand, has 254 nucleotides, and has 64% overall sequence homology with CCCVd 246. CTiVd has not been shown to have the larger molecular forms described for CCCVd, but it does have a dimeric form (Boccardo et al. 1981; Keese et al. 1988). On the basis of their nucleotide sequences, CCCVd and CTiVd fall into the same viroid group. Considerable confusion has been generated by the discovery of viroid- like sequences in a wide range of otherwise symptomless coconut and other monocotyledons from south Asia to French Polynesia (Hanold and Randles 1991b), including oil palms affected with an orange spotting (since shown to be a genetically inherited trait). Although the presence of these nucleic acid sequences could not be associated with any symptoms of disease or other adverse effects on the host plants, the observations led to a recommended embargo on the movement of germplasm from regions where they are found to countries where the viroid-like sequences have yet to be reported (Frison and Putter 1993). It is now considered that the risks were overstated and, in the absence of symptoms of disease, the presence of CCCVd-like sequences should not mitigate against such transfers (Frison et al. 1997). Problems and research needs The mode of natural spread of Cadang-cadang and Tinangaja diseases has not been established. The vector is unknown, transmission through pollen or seed has not been ruled out and eradication is not practical. Similarly, resistance has not been found. Although mild strains of other viroids are known to cross protect against severe strains, little is known about the natural occurrence of the mild strains of CCCVd. While the search for resistance continues, replanting of infected plantations has to be maintained in order to reduce production losses, as new plantings in diseased areas are not affected. The search for mild strains of the viroids should also be intensified. Sensitivity, rapidity, simplicity and portability of diagnostic procedures for CCCVd for use in the field need to be improved so as to assist in studying the epidemiology of CCCVd and determining its mode of spread. Finally, quarantine restrictions on the movement of Philippine germplasm from affected regions have to remain in place until safer methods of exchange can be ensured but this need not be a hindrance to safe movement from other regions of the Philippines. 381 CHAPTER 6: Major pests and safe movement of germplasm Protozoa disease: Phytomonas Uniflagellate protozoa of the genus Phytomonas (Family Trypanosomatidae) are associated with and presumed to cause a group of diseases of coconut, oil palm and a few other palms in Central and South America (Brazil, Colombia, Costa Rica, Ecuador, French Guiana, Guyana, Nicaragua, Peru, Surinam, Venezuela) and the southern Caribbean (Grenada, Trinidad and Tobago) (Parthasarathy et al. 1976; Waters 1978; Dollet 1984). The disease is usually referred to as Hartrot but various local names have been used including Cedros wilt, Coronie wilt, fatal wilt and Marchitez sorpresiva and it is thought that these are all caused by variants of the same pathogen. Affected palms show rapid foliar discolouration and decay and usually die within 1-3 months from the first appearance of symptoms. The pathogen is normally restricted to the phloem but has been found in the husk, calyx, and coconut water in nuts of up to 11 months post fertilization but not in nuts showing advanced decay or in dry nuts (Nanden-Amattaram and Parsadi-Sewkaransing 1989). It is not known to be transmitted through seednuts or pollen but as with lethal yellowing, the risk of movement in seednuts cannot be completely ruled out. Indexing Symptoms usually start with yellowing or bronzing of the oldest leaves, loss of immature nuts and blackening of newly opened inflorescences and thus can easily be confused with those of lethal yellowing. However, the presence of the flagellates, which are 12-27µm x 1-1.5µm, can usually be confirmed in sap expressed from inflorescences or roots examined by light microscopy using phase contrast, dark field or after staining with Giemsa or toluidine blue (Waters 1978). All the coconut varieties that have so far been tested were susceptible to the disease, and other palms including Bentinckia nicobarica, Elaeis oleifera, E. guineensis, Maximiliana maripa and Roystonia regia may serve as alternative hosts (Dollet 1984; Kastelein and Parsadi 1986). The Phytomonas associated with Hartrot can be transmitted by pentatomid bugs of the genera Lincus and Ochlerus (Desmier de Chenon 1984; Dollet 1984) and these are thought to be the natural vectors of the disease. Pathogen characterization Although there are very few instances of protozoa being implicated in plant disease, Phytomonas have long been recognised as apparently harmless parasites of latex cells in laticiferous plants especially in the plant families Euphorbiaceae and Asclepiadaceae. Those associated with and thought to cause disease in coconut and oil palm have been described 382 COCONUT GENETIC RESOURCES as Phytomonas staheli (McGhee and McGhee 1979) but the taxonomy of the genus, and the relationships between types pathogenic and parasitic in plants are not clear. Isolates from palms have been distinguished by means of isozyme profiles (Guerrini et al. 1992), restriction length polymorphisms of kinetoplastid DNA (Muller et al. 1995) and from sequences of small subunit 18S rDNA (Marche et al. 1995). However, in practice, demonstration of the presence of flagellates in palm tissues should be sufficient for presumptive diagnosis of the disease. According to Ohler (1999), in addition to other palms, Musaceae and Zingiberaceae are now thought likely sources of the disease. Precautions to ensure safe movement should thus be extended to these species, particularly to vegetative propagation materials. Nematode disease: Red Ring Red ring disease is caused by the nematode Bursaphelenchus (=Rhadinaphelenchus) cocophilus and is spread mainly by the palm weevil Rhycopophorus palmarum and also the sugarcane weevils Dynamis borassi and Metamasius hemipterus (Giblin-Davis 1993), The disease occurs throughout Central and South America, Mexico (Yucatan peninsula), the Lesser Antilles and the Dominican Republic but has not yet been reported in Cuba and Jamaica despite the presence of the weevil vectors in these countries. Young palms (3-10 years old) are reported to be most frequently affected and die within a few months of infection, which takes place through wounds associated with weevil activity. Symptoms vary according to age, variety and growing conditions but may include yellowing or bronzing of leaves (usually the oldest first but sometimes discontinuously within the crown), nutfall and necrosis or withering of newly opened inflorescences. Production of small and sometimes distorted young leaves (‘little leaf’ syndrome) has reportedly been associated with the disease in parts of South America (Hoof and Seinhorst 1962). The characteristic and diagnostic feature of the disease is an internal red discolouration at the base of the stem in the form of a ring about 2-6 cm wide and about 3-5 cm from the periphery. The red ring usually extends 2-3 m up the stem, breaking up into streaks or discrete spots, which may extend into the rachis and petiole, and into cortical tissues of the roots. The discoloured palm tissues contain abundant nematodes (adult stages are about 1mm long). Juvenile stages of the nematode parasitize palm weevil larvae that develop in infested palms which remain infected throughout metamorphosis. They then transmit the nematode to healthy palms primarily during oviposition in moist leaf bases or freshly wounded tissues (Gerber and Giblin-Davis 1990). 383 CHAPTER 6: Major pests and safe movement of germplasm The disease can usually be diagnosed reliably from the presence of internal symptoms, confirmed by observation of nematodes in palm tissues, and no special indexing or characterization techniques have been described. Although it has been shown that artificially inoculated seednuts supported nematodes for up to 16 weeks after germination, none could be found after 20 weeks and the developing seedlings remained free of disease (Giblin-Davis 1991), suggesting that spread of disease is unlikely to occur in seednuts. However, the movement of germplasm from red-ring affected regions should follow the general recommendations in the FAO/IBPGR Technical Guidelines (Frison and Putter 1993). Conclusion Perhaps the biggest threat to the safe movement of coconut germplasm remains from diseases of as yet unknown or uncertain aetiology, or perhaps not yet recognised as infectious conditions at all. The aetiology of several diseases on the Indian subcontinent remains uncertain or unconfirmed: root wilt and Tatipaka on the west and east coasts of India; leaf scorch decline and premature decline in Sri Lanka. Information on transmission of these diseases through seednuts, embryos or pollen is sorely lacking and experimental investigations are hampered by the lack of reliable diagnostic methods and the obvious difficulties in carrying out large-scale, long-term empirical testing by direct observations on progenies and crosses from diseased palms. Elsewhere, there are several instances where ‘new’ diseases, either previously unrecognised or of only minor local importance, have emerged following the introduction of exotic coconut varieties to new regions. Budrot and premature nutfall cased by Phytophthora palmivora were recognised as only minor problems in indigenous varieties grown in north Sulawesi, Indonesia, but caused widespread losses following the large-scale introduction of exotic Malayan Dwarf x West Africa Tall hybrids in the 1980s. Outbreaks of so-called coconut stem necrosis were also associated with the introduction of exotic varieties in Indonesia (Turner et al. 1979) and the emergence of foliar decay virus as a problem in cultivars introduced to Vanuatu as referred to earlier. In South America, lethal diseases of unknown aetiology, the so-called spear rot-bud rot complex, affect oil palm in several countries (de Franqueville 2002). 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Comparative analysis of coconut phytoplasmas from East and West Africa. Pp. 197 –203. In: SJ Eden- Green and F Ofori (eds). Proceedings of an International Workshop on Lethal Yellowing-Like Diseases of Coconut, Elmina, Ghana, November 1995. Natural Resources Institute, Chatham, UK. Tymon, AM, P Jones and NA Harrison. 1997. Detection and differentiation of African coconut phytoplasmas: RFLP analysis of PCR-amplified 16S rDNA and DNA hybridization. Annals of Applied Biology 131:91-102. Tymon, AM, P Jones and NA Harrison. 1998. Phylogenetic relationships of coconut phytoplasmas and development of specific oligonucleotide primers. Annals of Applied Biology 132:437- 452. Waters, H. 1978. A wilt disease of coconuts from Trinidad associated with Phytomonas sp., a sieve tube-restricted protozoan flagellate. Annals of Applied Biology 90: 293-302. Zelazny, B, JW Randles, G Boccardo and JS Imperial. 1982. The viroid nature of the cadang-cadang disease of coconut palm. Scientia Filipinas 2:45–63. 390 COCONUT GENETIC RESOURCES Strategies for safe movement of coconut germplasm M Diekmann Research Advisor, Deutsche Gesellschaft für Technische Zusammenarbeit - Beratungsgruppe Entwicklungsorientierte Agrarforschung (GTZ-BEAF), Bonn, Germany The drastic effects of plant diseases on yield, on plant growth and on landscapes can be seen in many different agricultural as well as natural habitats. Few diseases have as drastic effects as the so-called lethal ones, such as the lethal yellowing disease (LYD) that killed millions of coconut palms. The coconut research community is therefore very much aware of risks due to pests and pathogens. Like many other plant diseases, LYD is also reported as spreading to new areas, after the Gulf and Caribbean coasts of Mexico, Belize and Honduras now to the Pacific coast of Mexico (Harrison et al. 2002). This disease is caused by a pathogen that belongs to the group called ‘phytoplasmas’, or formerly mycoplasma-like organisms (MLO). The pathogen is transmitted by a planthopper, a small insect. Recently, a yellowing disease associated with phytoplasma was reported to occur on date palms in Kuwait (Al-Awadhi et al. 2002). It seems that the pathogen is not only spreading to new areas, but also to new hosts. Some plant pathogens spread to new areas as wind-blown spores, e.g. the coffee rust fungus Hemileia vastatrix that appeared in 1970 in Brazil, and moved from there to all coffee producing areas in South and Central America as well as in Mexico. Little can be done to prevent the spread of such a pathogen. Other pathogens, for example viruses and phytoplasmas, and many fungi, move only with the help of a vector, such as an insect, or an infected plant to new areas. Here, at least in theory, preventing the movement of insect vectors or infected plants will prevent the spread of the pathogen. Phytosanitary measures can help to stop the introduction of non-indigenous, potentially damaging pests and pathogens into an area or to eradicate them before they spread and cause serious yield losses or the death of plants. There is no doubt that activities to strengthen the conservation and use of plant genetic resources worldwide, with special emphasis on the needs of developing countries, are of utmost importance. However, care must be taken not to introduce new pathogens inadvertently. With the high genetic diversity of plants, a diversity of pathogens is often associated. The diversity of plant genotypes is much needed for the selection of resistant varieties, which maybe the only promising control strategy for 391 CHAPTER 6: Major pests and safe movement of germplasm some plant diseases. The above mentioned lethal yellowing disease of coconut could be effectively controlled using resistant varieties, such as Malayan Dwarf hybrids. Clearly, the movement or plants or plant parts between countries or continents entail the risk of introducing exotic plant pests or pathogens. Less-developed countries often lack adequate plant quarantine and diagnostic facilities, and are especially vulnerable to the damaging effects of newly introduced diseases. It is extremely important that the risk is recognized, and that a minimum risk transfer form of germplasm is chosen, such as in vitro plantlets instead of nuts. Networks, such as International Coconut Genetic Resources Network (COGENT), assist their members with pest risk analysis procedures. An effective phytosanitary system acts as a filter and not a barrier to germplasm exchange. It keeps pathogens out and allows germplasm to pass. As some countries have stronger systems than others, the plant breeders and the germplasm community should also give due attention to pathogens. In addition to the risk of spreading pathogens to new areas (there are numerous examples where this has happened with germplasm), there is also the risk of the collections, or part thereof, being destroyed by disease epidemics. This risk is particularly high in field genebanks. The choice of phytosanitary measures (exclusion, import permit stating certain conditions, certification according to the requirements of the importing country, standard quarantine certificate, post-entry isolation and observation) depends on the risk. The instrument of pest-risk analysis helps to make the correct choice, provided the required data are available (see articles by de Franqueville and Ikin in this chapter). Since phytosanitary measures were established (in some countries almost 100 years ago), the justifications for quarantine measures have not changed: • The pest or pathogen does not occur in the importing country; • The pest or pathogen is capable of surviving and multiplying under the conditions of the importing country; and • The pest or pathogen is likely to cause economic damage. The International Plant Genetic Resources Institute (IPGRI), has published jointly with the Food and Agricultural Organization (FAO) Technical Guidelines for the Safe Movement of Germplasm (e.g. Frison et al. 1993 for coconuts). Table 1 summarizes the FAO/IPGRI Technical Guidelines for the Safe Movement of Coconut Germplasm. The general recommendation is to move embryo cultures or pollen, and not seednuts. If this recommendation is followed, the risk of moving fungi, phytoplasma (MLO) and the red ring nematode with germplasm is greatly reduced. 392 COCONUT GENETIC RESOURCES Indexing would be required only for germplasm from Vanuatu (for coconut foliar decay virus), Guam (for tinangaja viroid), and from parts of the Philippines (for cadang-cadang viroid), unless one decides to exclude material from these areas from germplasm movement. Based on this, the priority should be on supporting embryo culture facilities and training. Since the publication of these guidelines, further research was conducted. To date, no disease symptoms could be linked to the presence of viroid-like sequences in coconuts. They were found to be widely distributed in coconuts and understorey plants. Presumably they do not play a role as pathogens (Diekmann 1997, FAO/IPGRI 1997). It is important not to confuse the cadang-cadang viroid with viroid-like sequences. Cadang-cadang viroid causes premature death of coconut palms in parts of the Philippines (see article by Eden-Green and Mpunami in this chapter). Viroid-like sequences were detected by molecular methods in coconuts and other monocotyledons, but could not be associated with disease symptoms (Hanold and Randles 1991). On the other hand, the reported non-transmission of phytoplasmas (through seed, embryo culture or pollen) may need to be reconsidered. Cordova et al. (2003) reported the presence of phytoplasma DNA in embryos from nuts of palms suffering from lethal yellowing disease. Current opinion among plant pathologists is that phytoplasmas are not transmitted by seed because there is no vascular connection between the tissue of the parent plant and the embryo (Jones 2001). Further investigation is needed to clarify whether the presence of phytoplasma DNA in the embryo incidates the risk of seed transmission or not. Furthermore, the transmission rate under field conditions needs to be studied. As premature nut fall is one of the first signs of the disease, nuts (if at all produced) may not germinate properly, reducing the risk of further transmission through seed/embryos. Germplasm health needs to be considered not only at the point of exchange, but at any stage of germplasm management (collecting, multiplication, evaluation and characterization, storage and distribution). Cooperation among breeders/germplasm curators and regulatory organizations is essential. Consultation should occur regularly, particularly at early planning stages for collecting, establishing field genebanks, etc. Germplasm should be exchanged only for immediate use or for safety duplication. 393 CHAPTER 6: Major pests and safe movement of germplasm Pathogen Specific Recommendation(s) Coconut foliar decay virus (CFDV) Indexing or exclusion of germplasm from Vanuatu Coconut cadang cadang viroid (CCCVd) Indexing or exclusion of germplasm from the Philippines Tinangaja viroid (CTiVd) Indexing or exclusion of germplasm from Guam Viroid-like sequences Indexing recommended for germplasm that is moved from countries where these sequences are known to occur to countries where they have not yet been reported. Lethal yellowing (Phytoplasma, MLO) Kerala wilt (Phytoplasma, MLO) Tatipaka disease (Phytoplasma, MLO) Transmission through seed, embryo culture or pollen not reported Blast (Phytoplasma, MLO) A nursery disease which does not occur on adult trees Marasmiellus spp. (bole rot, shoot rot) Possibly seed-borne, can be eliminated in embryo culture Phomopsis cocoina (leaf spot) Bipolaris incurvata (leaf blight) May be dispersed on husks. Recommendations are: • Embryo and pollen transfer should be carried out • Healthy nuts should be partially de-husked and treated with an appropriate fungicide Phytophthora palmivora, P. katsurae (bud rot, fruit rot) Nuts may be infected internally, but then do not germinate. Recommendations are: • Embryo and pollen transfer should be carried out • Healthy nuts should be partially de-husked and treated with an appropriate fungicide Table 1. Summary of FAO/IBPGR Technical Guidelines for the Safe Movement of Coconut Germplasm Source: (Frison et al. 1993) References Al-Awadhi, HA, A Hanif, P Suleman and MS Montasser. 2002. Molecular and microscopical detection of phytoplasma associated with yellowing disease of date palms Phoenix dactylifera L. in Kuwait. Kuwait Journal of Science and Engineering 29(2): 87-109 Cordova, I, P Jones, NA Harrison and C Oropeza. 2003. In situ PCR detection of phytoplasma DNA in embryos from coconut palms with lethal yellowing disease. Molecular Plant Pathology 4: 99-108. Diekmann, M (ed.) 1997. Proceedings of the Meeting on Viroid-like Sequences of Coconut. 21-23 April, 1997, Kajang (Kuala Lumpur), Malaysia. ACIAR, Canberra /IPGRI Rome. FAO/IPGRI. 1997. Addendum to the FAO / IBPGR Technical Guidelines for the Safe Movement of Coconut Germplasm. Food and Agriculture Organization of the United Nations, Rome/International Plant Genetic Resources Institute, Rome, Italy. 48pp. Frison, EA, CAJ Putter and M Diekmann (eds.). 1993. FAO/IBPGR Technical Guidelines for the Safe Movement of Coconut Germplasm. Food and Agriculture Organization of the United Nations, Rome/ International Board for Plant Genetic Resources, Rome, Italy. 394 COCONUT GENETIC RESOURCES Hanold, D and JW Randles. 1991. Detection of coconut cadang-cadang viroid-like sequences in oil and coconut palm and other monocotyledons in the south-west Pacific. Annals of Applied Biology 118(1): 139-151. Harrison, NA, M Narváez, H Almeyda, I Cordova, ML Carpio and C Oropeza. 2002. First report of Group 16SrIV phytoplasmas infecting coconut palms with leaf yellowing symptoms on the Pacific coast of Mexico. Plant Pathology 51(6): 808. Jones, P. 2001. Phytoplasma plant pathogens. Pp. 126-139. In: JM Waller, JM Lenné and SJ Waller (eds). Plant Pathologists’ Pocketbook, 3rd Edition. CAB International, Wallingford, UK. 395 CHAPTER 6: Major pests and safe movement of germplasm Pest risk assessment of the International Coconut Genebank for Africa and Indian Ocean, and Latin America and the Caribbean H de Franqueville Plant Pathologist, Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD), Oil Palm Programme, UMR BGPI – Campus International de Baillarguet TA 41/K F34398 Montpellier, Cedex 5, France Introduction The International Coconut Genetic Resources Network (COGENT) of the International Plant Genetic Resources Institute (IPGRI) has been assisting the establishment of a multi-site International Coconut Genebank (ICG), with sites in five countries, each representing one of the main coconut ranges. They are Indonesia for Southeast Asia, India for South Asia, Papua New Guinea for the South Pacific, Côte d’Ivoire for Africa and the Indian Ocean, and Brazil for Latin America and the Caribbean region. The pest pressure exerted on coconut throughout its major producing regions, and the consequent phytosanitary risks to which it is exposed, are a threat to its sustainability and sometimes lead to it being dropped from a production system. The risks are rarely the same worldwide, and are therefore important to assess them in order to promote germplasm dissemination and exchange under optimum conditions. Generally, it is the overall phytosanitary constraint that needs to be documented in a given zone, not only to avoid the transfer of pests and diseases but also to guarantee a satisfactory phytosanitary situation in the collections planted at a given site. That means also taking into account fungal diseases and the main coconut pests in the entomofauna that are likely to jeopardise the establishment of a germplasm collection. In order to determine this constraint, a pest risk assessment was conducted in two ICG host countries, Côte d’Ivoire and Brazil. This paper attempts to document the main pests and diseases in the study zones, analyse the corresponding phytosanitary risk, determine their potential as quarantine organisms and identify the phytosanitary risks involved for collecting and exchanging germplasm. Material and methods Documentation This study is based on all the information gathered by conventional bibliographical research, the author’s knowledge of coconut diseases, or 396 COCONUT GENETIC RESOURCES oil palm diseases in some cases that could impact on coconut, consultations and discussions with members of the scientific community, supplemented by information gathered from some particularly useful internet sites. Visits to ICG host countries: Côte d’Ivoire and Brazil The visit to Côte d’Ivoire took place in April 1999, in liaison with the Marc Delorme Research Station (Centre National de Recherche Agronomique, CNRA). Sites visited included the station itself, at Port- Bouët, near Abidjan, with its current collection; the lagoon strip between Assinie and the Ghanaian border, to examine the condition of the coconut groves; and the Grand-Drewin Experimental and Production Station (CNRA, Gagnoa regional management), a potential site for a future collection. This station is located at Sassandra, around 240 km from Port- Bouët, and around 330 km from the Ivorian-Ghanaian coastal border. Port-Bouët is around 95 km from the same border (all the distance are direct, as the crow flies). The Ghanaian border at Elubo is around 170 km by road, via Aboisso, from Port-Bouët and 460 km from Sassandra. These details are important, due to the existence of lethal yellowing disease (LYD) in the neighbouring Ghana. Arrangements will have to be made to duplicate all or part of the Marc Delorme Station collection at the Grand-Drewin Station if the disease gets any closer to the Ivorian border. The visit to Brazil took place also in April 1999, in liaison with the EMBRAPA research station at Aracaju, in Sergipe State (Centro de Pesquisa Agropecuária dos Tabuleiros Costeiros - Empresa Brasileira de Pesquisa Agropecuária). The sites visited that are candidates for receiving the future ICG material were Itaporanga, west of Aracaju, the Neopolis plateau, northeast of Aracaju, and Betume, located between Neopolis and Ilha das Flores. Pest risk assessment Pest risk assessment is a step towards a pest risk analysis (PRA), following the process laid down by FAO (1996a, b). Its purpose is to identify pests and diseases necessitating plant quarantine. It is carried out in a potentially or known pest risk area, usually a country. Ikin (1997) applied these directives to coconut germplasm exchanges for cadang-cadang and cadang-cadang viroid-like sequences. His study led to the revision of the directives governing germplasm movement and the quarantine measures applied to it. PRA could be broken down into three stages: 1. Identification of pests or pathways for which PRA is necessary. Here, the pathway is defined by the form in which germplasm is 397 CHAPTER 6: Major pests and safe movement of germplasm transported: seedlings, seeds, pollen or embryo culture. A pest may or may not be defined as being a quarantine organism depending on the germplasm form; 2. Risk assessment serves to determine whether the identified organism, as such or combined with the pathway, is a quarantine organism, depending on its likelihood of entering the PRA zone, the capability of establishing itself and spreading, and its economic importance (Diekmann 1997). 3. Risk management comprises the development, assessment, comparison and choice of options intended to reduce that risk. PRA can be carried out by considering either the pathway or the pest (i.e., the form in which germplasm should be exchanged to significantly reduce the risks of introducing a given pest). It is primarily the second approach that will be taken, given the inventory of pests existing in the study areas. Results Coconut diseases and pests Almost 30 diseases affect coconut worldwide (Frison et al. 1993; Ikin 1997; Mariau 1999). Most are found on the Asian continent and little is known about most of them. In the study areas, neither identified diseases of viral nor viroid origin have been inventoried. On the other hand, LYD shows a strong presence in Africa, Central America and the Caribbean. The insects listed during the study do not figure in the germplasm transfer pathways, given their nature and their biology, although special attention must be given to the recent outbreaks of white flies (Aleurotrachelus atratus and Paraleyrodes bondari) in Comoros Islands (Baudoin and Ollivier 2003, personal communications). However, it is possible that the insect pests do pose a threat for the installation and development of collections in Côte d’Ivoire and Brazil. Mites, especially Eriophyes guerreronis, can be harboured by nuts, primarily beneath the floral parts, and are therefore, a risk that has to be considered if germplasm is moved as seednuts. However, as they cannot withstand a vacuum (JF Julia 1999, personal communication), there is little risk of them contaminating pollen. Likewise, it should be possible to detect any contamination of embryo cultures very rapidly. For the record, vertebrate pests, birds or mammals do not figure in the germplasm exchange pathways, but the risks they represent to collections, especially on young plants, need to be taken very seriously. In general, cultural practices or special arrangements (ditches, fences) help to reduce their impact. 398 COCONUT GENETIC RESOURCES Situation in Africa/Indian Ocean The following seven diseases were found in Africa and the Indian Ocean: Phytoplasma diseases Blast is the main nursery disease on oil palm in Africa, and is also found on coconut (Quillec et al. 1978). It is attributed to a phytoplasma due to the preventive role played by tetracycline (Dollet 1980; Dollet 1985). Blast is carried by a leafhopper, Recilia mica Kramer (Desmier de Chenon 1979). The insect only seems to be infectious at certain times of the year and incubation lasts a few days (de Franqueville et al. 1991). Blast has never been reported on bearing palms, although it has been observed during the first year after planting. Therefore, it is not a major threat to germplasm movement. LYD first occurred in Africa around 1930 (Bachy and Hoestra 1958), in Togo and was called Kaincopé disease (Dollet and Giannotti 1976), then in Southeast Ghana as Cape St Paul wilt (Dabek et al. 1976), in Cameroon as Kribi disease (Dollet et al. 1977) and in Nigeria asAkwa disease (Ekpo and Ojomo 1990). In East Africa, LYD causes serious damage in Tanzania (Schuiling and Mpunami 1990), Kenya and Mozambique (Mpunami et al. 1996). Analyses by restricted fragment length polymorphism (RFLP) and polymerase chain reaction (PCR) suggest a degree of difference between West African and East African phytoplasmas (Tymon et al. 1997, 1998). The disease vector has not been formally identified, but a plant hopper, Myndus adiopodoumeensis is strongly suspected in Ghana (Dery et al. 1996). Phytoplasma diseases are considered to have little chance of being carried by seeds, pollen, or embryos (Dollet 1995). Phytoplasmas seem to have been detected in embryos, but there is no evidence that these embryos would normally germinate. LYD is not widespread throughout the African and Indian Ocean region; it has not been reported in Côte d’Ivoire, Benin or the Seychelles. Fungal diseases Phytophthora katsurae Ko and Chang causes immature nut fall and lethal bud rots (Quillec and Renard 1984). P. katsurae, which was initially identified as P. heveae, a very closely related species, seems to be the only fungal species, found damaging in Côte d’Ivoire (Blaha et al. 1994). Its incidence is effectively controlled by fungicide injection into the stem (de Franqueville and Renard 1989). Phytophthora rot diseases are not documented in the other African countries, but are suspected in Ghana. Marasmiellus cocophilus Pegler is associated with the so-called lethal bole rot, on seedlings or young palms in Kenya and Tanzania (Bock et al. 399 CHAPTER 6: Major pests and safe movement of germplasm 1970). The fungus can act as a saprophyte, colonizing plant matter, either from coconut palms or from other crops. Phomopsis cocoina (Cooke) Punith. causes leaf spots and husk rot. It is reported in Kenya, the Seychelles (quoted by Frison et al. 1993). It can be borne by nuts. Bipolaris incurvata causes leaf blight in the Seychelles. This symptom is similar to the Helminthosporium leaf spot found in Côte d’Ivoire (Quillec and Renard 1975), caused by Helminthosporium halodes (Dresch.), whose limited economic importance has never warranted any intensive intervention. Diseases of unknown origin Dry bud rot, which is documented in Côte d’Ivoire (Renard et al. 1975), also found on oil palm, is transmitted by two similar species of Delphacidae, namely Sogatella kolophon Kirkaldy and S. cubana Crowford (Julia and Mariau 1982). It is a disease of young palms and primarily occurs in the nursery; damage to adult palms has not been observed. There is no information available on its incidence in the other countries of West Africa. Pest risk assessment in Côte d’Ivoire Ivorian coconut plantings are subject to four of the seven diseases documented in Africa and the Indian Ocean. Two are diseases found in the nursery or on young palms - blast and dry bud rot. The other two are fungal diseases, namely: Helminthosporium leaf spot and Phytophthora. LYD, in neighbouring Ghana, is spreading and threatening the eastern part of Côte d’Ivoire, but it is difficult to establish the speed with which the disease is spreading. In the Western Region, it first occurred in 1964 at Cape Three Points. It was not until 1992 that it reached Axim, around thirty kilometres to the West, after affecting Cape Coast in 1984, 100 km to the East. The situation has been described by Philippe (1997): a large focus developed around 15 km west of Axim, i.e. around 74 km from the Ivorian border. A smaller focus was detected 13 km to the West (61 km from Côte d’Ivoire) and two diseased palms were detected 34 km from the border. Those two palms were immediately eliminated. By 1999, the situation had barely changed (R. Philippe 1999, personal communication). The larger focus, near Axim, has spread at a rate of one to two km per year, the smaller focus at a rate of around a hundred metres in two years, and the situation has remained unchanged at the site where the two diseased palms were eliminated. Visits to the lagoon strip, on the Ivorian side, did not reveal any lethal yellowing infection. There were some yellowing palms, in poor condition, 400 COCONUT GENETIC RESOURCES with few or no bunches but there were no signs of any developments in either symptom intensity or dispersion of the symptoms. They are old coconut palms, which have never received any fertilizer or phytosanitary treatment and have always be subjected to attacks from Oryctes and scale insects (Aspidiotus destructor Signoret), which should not be confused, as emphasized by Dollet (1995) with cases of lethal yellowing. The Grand-Drewin station is one of the sites selected for establishing a coconut germplasm collection. It also has a large population of oil palm. There is no Phytophthora disease. Given its location in a low rainfall zone, the risks run by the collection are linked to drought unless an irrigation system is installed. Diseases affecting young palms may also occur (blast and dry bud rot), which can be effectively controlled by chemical treatments against the vectors, and by cultural practices. A close watch will have to be kept on Oryctes sp. outbreaks in the early years after planting, especially if old oil palm plantings have been felled in the vicinity. During production periods, Aceria (Eriophyes guerreronis) damage is to be feared. Damage caused by the Coreid bug Pseudotheraptus sp. is slight, probably due to the good establishment of Oecophylla ants, which limit its development. Lastly, it is reasonable to assume that if LYD were to spread into Côte d’Ivoire, it would probably only occur on a scale of at least one generation of coconut palms. Situation in Latin America and the Caribbean At least the following ten diseases are documented in Latin America and the Caribbean (LAC): Phytoplasma disease LYD was reported for the first time in the Cayman Islands around 1830, has spread throughout the Caribbean, to Haiti, the Dominican Republic, Cuba, Jamaica and then Florida. It reached the Yucatan peninsula in Mexico in the 1980s (Cardeña et al. 1991) and was reported in Honduras in 1996 by Ashburner et al. Its spread in LAC has been much faster than in West Africa and it is carried by a leafhopper, Myndus crudus Van Duzee (Cixiidae). Molecular techniques (RFLP, PCR) have shown greater similarity between the phytoplasmas in LAC and East Africa than with those in West Africa (Jones et al. 1995; Tymon et al. 1998). LYD occurs in most of the countries in the zone in the COGENT network, but not in Costa Rica, Guiana, Trinidad and Tobago, and Brazil. It also doest not exist in Nicaragua or Venezuela. It should be noted that phytoplasmas are reported to have been detected in the embryos of nuts from diseased palms in Mexico. It has 401 CHAPTER 6: Major pests and safe movement of germplasm not been confirmed, but needs to be checked as soon as possible, using all the appropriate techniques (electron microscopy, PCR), along with their viability. Fungal diseases Phytophthora palmivora and Phytophthora katsurae live side by side in Jamaica (Steer and Coates-Beckford 1990), but P. palmivora is usually the only species found in the zone. It causes bud rot leading to the death of coconut palm. Its incidence can be devastating in some parts of the Caribbean, notably the Dominican Republic. It is known to exist in Cuba and Central America, but there is no precise information. Phytophthora rot diseases are not documented in Brazil. Bud rot symptoms have been observed in the Fortaleza region of Ceara state and have been assimilated in their advanced stage to those caused by this fungus. However, neither the development of the disease, nor the isolations carried out, has confirmed this hypothesis (de Franqueville 1996). Bipolaris incurvata occurs in Central and South America. In particular, it was reported in Brazil by Warwick (1997) where it causes Helminthosporium leafspot, or ‘mancha-foliar’, particularly in the nursery. Lixa pequena, caused by Phyllachora torendiella (Bat.) nov.comb., is a widespread leaf disease in Brazil of varying intensity (Subileau et al. 1993). It is also found in French Guiana. It can cause up to 50% loss of leaf area, immature nut fall, and consequent yield loss of 20 to 40% (Renard 1999). Lixa grande is another leaf disease associated with the previous one but caused by Sphaerodothis acrocomiae (Montagne) von Arx & Muller. Queima das folhas is also a leaf disease of Brazil, caused by Botryosphaeria cocogena Subileau. Lixas generally promote the development of this fungus, for which they represent access routes. This constitutes a perfect parasitic complex (Subileau 1993, 1994; Warwick et al. 1994). Phytomonas disease Hartrot is endemic in northern South America, from Peru or Bahia state in Brazil, to Costa Rica (Renard 1999). It is moving up to Honduras, where infected oil palm marchitez is already found. It has also been identified in Trinidad, under the name of Cedros wilt, where 15 000 coconut palms have been killed in three years. It causes sporadic damage in Colombia, Venezuela, Surinam, Brazil and French Guiana. Smallholdings, which do not have access to regular insecticide treatments, can disappear within five years (M. Dollet 1999, personal communication). The presence of Phytomonas (Trypanosomatids) is associated with any Hartrot syndrome (Dollet et al. 1977a; Dollet and Lopez 1978). Hartrot is carried by bugs of the Lincus genus (Louise et al. 1986) or Ochlerus genus (Mariau 1985). 402 COCONUT GENETIC RESOURCES Nematode diseases Red ring disease is caused by Bursaphelenchus cocophilus (Cobb) Baujard, a nematode carried by an insect, Rhynchophorus palmarum (Curculionidae). It is endemic in Central America, South America and the Caribbean (Warwick et al. 1995). This disease also affects oil palm. Its incidence varies depending on the region. In Venezuela, some oil palm plantations have suffered 70% losses in 15 years. Red ring control consists of limiting the vector populations, notably by using aggregation pheromones. Diseases of unknown origin A dry bud rot occurs in Brazil (Renard 1990). It is not known whether it is linked to the one found in West Africa and/or with the so-called oil palm ring spot disease, which is rife in Latin America, although symptoms are similar to those of oil palm dry bud rot in West Africa. As knowledge stands at the moment, it is classified as a juvenile disease in Brazil (Warwick 1998). Porroca is a disease of unknown origin that seems primarily to affect poorly maintained coconut plantings. Currently, its incidence seems to be limited to Colombia and Panama, countries which are not in the COGENT network. Porroca is not reported in Costa Rica for the time being, but it is worth monitoring closely in Central American countries. Similar symptoms (short leaves) exist in French Guiana. Pest risk assessment in Brazil Seven of the 10 diseases listed above are found in Brazil, but the two most serious diseases have not been detected in the country, i.e. lethal yellowing and Phytophthora. Several thousand kilometres separate Brazil from the most active lethal yellowing foci, whether in the Caribbean zone or in Central America. The Andes, in Colombia and Venezuela, also form a natural barrier between Brazil and the Central American foci. It is therefore unlikely that lethal yellowing will occur in epidemic proportions in Brazil. Brazil may be a favourable zone for Phytophthora palmivora development, as shown by attacks on cocoa plantings in Bahia state (Ortiz Garcia 1996). The Aracaju region in Sergipe is characterized by a substantial water deficit and by extended periods of severe drought. The predominant diseases in the region are leaf blights (lixas and queima das folhas), which occur in varying degrees in each of the plantations visited. Hartrot only seems to occur sporadically in the region (DRN Warwick 1999, personal communication). Wherever the collection is planted, it will run the risk of dry bud rot, which can cause major damage in young plants (Warwick 1998), and 403 CHAPTER 6: Major pests and safe movement of germplasm Helminthosporium leaf spot. However, attacks can be limited by preventive treatments. The list of coconut pests in Brazil is long (Ferreira et al. 1998; Morin 1986). The pests that are likely to be a threat to the collection are primarily Brassolis sophorae L., Hyalopsila ptychis Dyar, Coraliomela brunnea Thumberg, Homalinotus coriaceus, Aspidiotus destructor Signoret and the mite, Eriophyes guerreronis Keifer. Generally speaking, a close watch will have to be kept on the germplasm collection to prevent the risks associated with these different pests. If free of any lethal diseases, drought will remain the main limiting growth factor of the germplasm collections in Brazil. Risks linked to germplasm exchange in the study zones From African/Indian Ocean countries to Côte d’Ivoire Lethal yellowing is a threat to Côte d’Ivoire. The causal agent is capable of surviving in this country, spreading and causing major economic damage. It is therefore a potential quarantine organism. As far as fungal diseases are concerned, the risk of introducing Marasmiellus cocophilus, which has yet to be reported in Côte d’Ivoire, exists from Kenya to Tanzania. Phomopsis cocoina is reported in Kenya and the Seychelles and can be borne by nuts. Bipolaris incurvata is also reported in the Seychelles, but causes only minor damage, except in the nursery. Helminthosporium leaf spot occurs in Côte d’Ivoire. These fungi are not a major threat for coconut cultivation in Côte d’Ivoire and their economic importance does not warrant their being considered as quarantine organisms. Phytophthora palmivora, a species not found on coconut in Côte d’Ivoire, has not been reported in the countries of the zone. From Côte d’Ivoire to African/Indian Ocean countries Phytophthora katsurae may be a threat for the other countries in the zone involved, but only causes immature nut fall at the Marc Delorme Station. Hence, nuts do not germinate. The Grand Drewin Station is free of it. Moreover, it can be effectively controlled by appropriate fungicide treatments. For the other fungal diseases, only Helminthosporium leaf spot could be a threat, but methods of control and prevention exist for this disease. It only significantly affects certain ecotypes and is of no economic importance. Dry bud rot and blast are juvenile diseases that only occur at certain times of the year with virtually non existent risk of transmission. 404 COCONUT GENETIC RESOURCES From Latin American/Caribbean countries to Brazil There is nothing to indicate that the causal agent of lethal yellowing is not capable of settling, developing and causing serious economic damage in Brazil, even in the marginal zone of Aracaju. Myndus crudus, the disease vector in Florida and Mexico, has also been seen on oil palm plantations in Para state (Julia 1990). In any event, the Brazilian government has stepped up its quarantine and surveillance measures for palms to prevent the introduction of lethal yellowing in the country. Phytophthora palmivora probably exists in all the countries in the zone. The recurrent drought in Sergipe and the Aracaju region should hinder the establishment of this pathogen and limit its possible economic impact. From Brazil to the other countries of Latin America/Caribbean The economic importance of leaf fungi, Lixas and Queima, in Brazil means that considerable attention needs to be paid to the movement of the parasites involved. They exist in Sergipe, but also in other much more humid zones, such as Para state, where disease incidence is relatively contained by hyperparasitic fungi, Septofusidium elegantulum or Acremonium alternatum (Warwick et al. 1998). There is nothing to indicate that they are present in the other countries in the zone. Possible transmission of this fungus by pollen has been suggested, although not proven. In any event, based on the available information and given the lack of any effective control method, they have to be considered as quarantine organisms, be it via pollen or seeds. Hartrot disease occurs sporadically in Sergipe. With the exception of Trinidad and Tobago, it has not been reported in the Caribbean zone and therefore could be a threat from Brazil to Cuba, Haiti and Jamaica. Because of its causal agent, which also exists in Grenada on Alpinia, and the economic damage it causes, it should be considered as a potential quarantine organism for those countries and for Mexico. However, its intraphloemic nature ought to limit the risk of transmission by pollen or embryos. Red ring disease, which is endemic throughout Latin America and part of the Caribbean, should not be a major threat provided precautions are taken to eliminate the vector. Dry bud rot, as knowledge stands at the moment, is a juvenile disease with virtually non existent risk of transmission. Recommendations The technical directives drawn up by the FAO impose a few basic measures that govern coconut germplasm movements. Among the measures worth noting in particular, is that such movement must be by embryo cultures or pollen, using the techniques described in the recommendations of the FAO. 405 CHAPTER 6: Major pests and safe movement of germplasm It is common sense that germplasm should only be collected from healthy palms (not from the ground) at sites free of serious diseases. In that way, movement of partially dehusked and carefully decontaminated nuts, as recommended by the FAO for most fungal diseases, should not involve any major risks of spreading lethal diseases. It is all the more important in view of the fact that very few countries in the COGENT network have embryo culture facilities as yet. The two ICG host countries, Côte d’Ivoire and Brazil, are countries free from LYD, unlike most countries in the zone they represent. Any germplasm movement to these two countries must be by pollen or embryo cultures, seeing that LYD propagation is not possible by those pathways. Occasionally, seednuts could also be used provided they are collected from zones free of LYD or Hartrot, as certified by the exporting countries through a phytosanitary certificate after the evaluation of the collection site. If movement is by seednut, particular care must be paid to mites, with fumigation where necessary. It cannot be ruled out that Phytophthora may have an airborne phase during its cycle and contaminate pollen. This hypothesis is difficult to confirm or refute. Pollen preparation does not eliminate the fungi, but the measures recommended by the FAO (inspection and search for fungi on leaving the exporting country and on entering the importing country) should enable checks to be made notably by using specific Phytophthora culture media. Germplasm movements from Brazil to the other countries in the LAC zone must take into account the risks of propagating leaf diseases that are widespread in Brazil. As knowledge stands at the moment, it is not possible to say that the causal agents are not conveyed by seednuts or pollen. Moreover, they are difficult to isolate and culture, which does not argue in favour of the phytosanitary inspection recommended for Phytophthora. Embryo culture is therefore recommended for germplasm exported from Brazil. References Ashburner, R, I Córdova and C Oropeza. 1996. First report of coconut lethal yellowing in Honduras. Plant Disease 80 (8): 960. Bachy, A and H Hoestra. 1958. Contribution à l’étude de la maladie de “Kaïncopé” du cocotier au Togo. Oléagineux 13 (10): 721-729. Blaha, G, G Hall, JS Warokka, E Concibido and C Ortiz-Garcia.1994. Phytophthora isolates from coconut plantations in Indonesia and Ivory Coast: characterization and identification by morphology and isozyme analysis. Mycological Research 98 (12): 1379-1389. 406 COCONUT GENETIC RESOURCES Bock, KR, MH Ivory and BR Adams. 1970. Lethal bole rot disease of coconut in East Africa. Annals of Applied Biology 66: 453-464. Cardeña, R, MAVillanueva, JM Santamaria and C Oropeza. 1991. Presence in Yucatan of mycoplasma-like organisms in Cocos nucifera L., palms showing yellowing disease symptoms. Canadian Journal of Plant Pathology 13:135-138. Dabek, AJ, CG Johnson and HC Harries. 1976. Mycoplasma-like organisms associated with Kaincopé and Cape St.Paul Wilt Disaese of coconut palms in West Africa. PANS 22: 354-358. Dery, SK, R Philippe and D Mariau.1996. Auchenorrhyncha (Homoptera), suspected vectors of Coconut Lethal Yellowing disease in Ghana. Plantations, Recherche, Développement 3(5): 355-363. Desmier, de Chenon R. 1979. Mise en évidence du rôle de Recilia mica Kramer (Homoptera, Cicadellidae, Deltocephalinae) dans la maladie du blast des pépinières du palmier à huile en Côte d’Ivoire. Oléagineux 34:107-112. Diekmann, M. 1997. Safe movement of coconut germplasm. Pp. 9-11. In: M Diekmann (ed). Proceedings of the ACIAR meeting to discuss viroids and viroid-like sequences in coconut, 21-23 April 1997, Kuala Lumpur, Malaysia. IPGRI, Rome, Italy. Dollet, M. 1980. Research on the etiology of blast of oil and coconut palms. Pp. 19. In: DL Thomas, FW Howard and HM Donselman (eds). Proceedings of the International Council on Lethal Yellowing. ARS IFAS University of Florida, Fort Lauderdale, USA. Dollet, M. 1984. Plant diseases caused by flagellate protozoa (Phytomonas). Annual Review of Phytopathology 22:115-132. Dollet, M. 1985. Recherches étiologiques sur les syndromes pathologiques des oléagineux tropicaux pérennes (cocotier et palmier à huile). Thèse d’état, université des sciences et techniques du Languedoc, Montpellier, France, vol I et II. 520p. Dollet, M. 1995. Safe movement of coconut germplasm. Pp. 139-147. In: C Oropeza, FW Howard and GR Ashburner (eds). Lethal yellowing: Research and practical aspects. Kluwer Academic Publishers, Dordrecht, The Netherlands. Dollet, M and J Giannotti. 1976. Maladie de Kaïncopé: présence de mycoplasmes dans le phloème des cocotiers malades. Oléagineux 31(4):169-171. Dollet, M, J Giannotti and M Ollagnier. 1977a. Observation de protozoaires flagellés dans les tubes criblés de palmiers à huile malades. C.R. Acad. Sci. Paris, Série D 284:643-645. Dollet, M, J Giannotti, JL Renard and SK Ghosh. 1977b. Etude d’un Jaunissement létal des cocotiers au Cameroun: la maladie de Kribi. 407 CHAPTER 6: Major pests and safe movement of germplasm Observations d’organismes de type mycoplasmes. Oléagineux 32 (7): 317-322. Dollet, M and G Lopez. 1978. Etude sur l’association de protozoaires flagellés à la Marchitez sorpresiva du palmier à huile en Amérique du Sud. Oléagineux 33(5):209-213. Ekpo, EN and EE Ojomo. 1990. The spread of lethal coconut diseases in West Africa: incidence of Akwa disease (or Bronze leaf wilt) in the Ishan area of Bendel State, Nigeria. Principes 34: 143-146. FAO. 1995. 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Oléagineux 46(6): 223-231. de Franqueville, H. 1996. Frutop Produtora de Alimentos S.A. - Coopération technique pour le suivi des plantations de cocotier. Mission Phytopathologie, 1-15 mai 1996. DOC CP N° 688. 23 p. + annexes. Frison, EA, CAJ Putter and M Diekmann (eds). 1993. FAO/IBPGR Technical Guidelines for the Safe Movement of Coconut Germplasm. FAO/IPGRI, Rome. 48p. Ikin, R. 1997. Pest Risk Analysis - Technical justification of quarantine application to the exchange of coconut germplasm material. Pp. 12- 17. In: M Diekmann (ed). Proceedings of the ACIAR meeting to discuss viroids and viroid-like sequences in coconut, 21-23 April 1997, Kuala Lumpur, Malaysia. IPGRI, Rome, Italy. Jones, P, A Mpunami and A Tymon. 1995. Mycoplasma-like organisms as pathogens of coconut palms. Pp. 35-42. In: Lethal yellowing: Research and practical aspects. Kluwer Academic Publishers, Dordrecht, The Netherlands. 408 COCONUT GENETIC RESOURCES Julia, JF. 1990. Recherches sur les possibles insectes vecteurs de la maladie “Amarelecimento Fatal” du palmier à huile au Brésil. Embrapa - CIRAD/IRHO. Doc IRHO n° 2237. 8p. Julia, JF and D Mariau. 1982. Deux espèces de Sogatella (Homoptère Delphacidae) vectrices de la maladie de la pourriture sèche du coeur des jeunes cocotiers en Côte d’Ivoire. Oléagineux 37(11): 517-520. Louise, C, M Dollet and D Mariau. 1986. Recherches sur le hartrot du cocotier, maladie à Phytomonas (Trypanosomatidae) et sur son vecteur Lincus sp. (Pentatomidae) en Guyane. Oléagineux 41(10): 437-449. Mariau, D. 1985. Rapport entomologique de mission au Brésil. - Le Cocotier. Doc IRHO n° 1996. 34p. Mariau, D (ed). 1999. Les maladies des cultures pérennes tropicales. CIRAD, collection Repères. 287p. Morin, JP. 1986. Les ravageurs du cocotier (au Brésil): état des connaissances et des recherches en cours. Embrapa - Cirad/IRHO. Doc IRHO n° 1995. 13p. with annexes. Mpunami, A, P Jones, A Tymon and M Dickinson. 1996. The use of DNA probes and PCR for detection of coconut lethal disease (LD) in Tanzania. Pp. 577-582. In: Proceedings of the Brighton Crop Protection Conference - Pests and Diseases. BCPC publishers, Brighton. Ortiz Garcia, CF. 1996. Etude de la diversité génétique de populations de Phytophthora pathogènes du cacaoyer (Theobroma cacao L) et du cocotier (Cocos nucifera L). Thèse de doctorat, Université Paul Sabatier, Toulouse, France. 85p. + annexes. Philippe, R. 1997. Studies on vector insects and the epidemiology of the coconut phytoplasma disease in Ghana. Contract TS3*-CT92-0055. Mission Report, DOC CP 778 bis. 28pp. + annexes. Quillec, G and JL Renard. 1975. L’helminthosporiose du cocotier. Etudes préliminaires. Oléagineux 30(5):209-213. Quillec, G, JP Morin, JL Renard and D Mariau. 1978. Les maladies du cocotier dans le jeune âge; causes, méthodes de lutte. Oléagineux 33: 495-500. Quillec, G and JL Renard.1984. La pourriture à Phytophthora du cocotier. Conseil de l’IRHO n° 242. Oléagineux 39 (3): 143-147. Quillec, G, JL Renard and H Ghesquière. 1984. Le Phytophthora hevea du cocotier: son rôle dans la pourriture du coeur et dans la chute des noix. Oléagineux 39(10):477-485. Renard, JL. 1990. Mission Défense des cultures au Brésil: les problèmes sanitaires sur cocotier. EMBRAP - Cirad/IRHO. Doc IRHO n° 2266, 52p. Renard, JL. 1999. Symptomatologie et incidence économique. Pp.19-75. In: D Mariau (ed). Les maladies des cultures pérennes tropicales. 409 CHAPTER 6: Major pests and safe movement of germplasm CIRAD, Collection Repères. Renard, JL, G Quillec and F Arnaud. 1975. Une nouvelle maladie du cocotier en pépinière. Symptômes, moyens de lutte. Oléagineux 30 (3):109-112. Schuiling, M and A Mpunami. 1990. Lethal disease of coconut palm in Tanzania: review of research up to date and preliminary result of resistance trials. Pp. 171-183. In: ML Robert and D Zizumbo (eds). La Problematica del Amarillamiento Letal del Cocotero en Mexico, Centro de Investigacion Cientifica de Yucatan, Merida, Mexico, 1989. CICY Publisher, Merida, Mexico. Schuiling, M and A Mpunami.1992. Lethal disease of coconut palm in Tanzania. I.- Comparison with other coconut diseases in East Africa. Oléagineux 47(8-9):511-515. Schuiling, M, A Mpunami and DA Kaiza. 1992. Lethal disease of coconut palm in Tanzania. II.- History, distribution and epidemiology. Oléagineux 47 (8-9):516-522. Schuiling, M, A Mpunami, DA Kaiza and HC Harries. 1992. Lethal disease of coconut palm in Tanzania. III.- Low resistance of imported germplasm. Oléagineux 47(12): 693-698. Steer, J and L Coates-Beckford. 1990. Role of Phytophthora katsurae, P. palmivora, Thielaviopsis paradoxa and Enterobacter sp. in budrot disease of coconuts in Jamaica. Oléagineux 45(12): 539-545. Subileau, C. 1993. Systématique et biologie du complexe parasitaire du Phyllachora torrendiella (Bat.) nov comb. et du Botryosphaeria cocogena nov.sp., agents fongiques du dessèchement foliaire du cocotier au Brésil. Thèse de doctorat, Université Paris VI, Paris, France. 121p. Subileau, C, JL Renard and B Dennetière. 1993. Phyllachora torrendiella (Batista) comb. nov. responsable de la maladie verruqueuse du cocotier. Mycotaxon 49: 175-185. Subileau, C, JL Renard and L Lacoste. 1994. Botryosphaeria cocogena nov.sp., agent causal du dessèchement foliaire du cocotier au Brésil. Mycotaxon 51:5-14. Tymon, AM, P Jones and NA Harrison. 1997. Detection and differentiation of African coconut phytoplasmas: RFLP analysis of PCR-amplified 16S rDNA and DNA hybridisation. Annals of Applied Biology 131: 091-102. Tymon, AM, P Jones and NA Harrison. 1998. Phylogenetic relationships of coconut phytoplasmas and the development of specific oligonucleotide PCR primers. Annals of Applied Biology 132: 437- 452. Warwick, DRN. 1997. Principais doenças do coqueiro (Cocos nucifera L.) no Brasil. 2. ed. rev.ampl., Aracaju: Embrapa - CPATC. 34p. 410 COCONUT GENETIC RESOURCES Warwick, DRN. 1998. Ocorrência e medidas de controle da Podridão- seca do coqueiro no Platô de Neópolis, Sergipe. Agrotrópica 10(1): 43-46. Warwick, DRN, JL Renard and G Blaha. 1994. La “Queima das folhas” du cocotier. Plantations, Recherche, Développement 1(2): 57-64. Warwick, DRN, DL de Q Santana and ERC Donald. 1995. Anel vermelho do coqueiro. Aspectos gerais e medidas de controle. Comunicado Técnico N° 05, Embrapa -CPATC: 1-7. Warwick, DRN, EC Leal and C Ram. 1998 . Doenças do coqueiro. Pp 269-292. In: JMS Ferreira, DRN Warwick and LA Siqueira (eds). A cultura de coqueiro no Brasil, 2nd Ed. rev.e. ampl.Brasilia. Embrapa- SPI; Aracaju: Embrapa-CPATC. 292p. 411 CHAPTER 6: Major pests and safe movement of germplasm Pest risk analysis and guidelines for the safe movement of germplasm in the International Coconut Genebank of Asia and the Pacific R Ikin Biosecurity Consultant, 25 Mayfair Place, Boondall, QLD 4034 Queensland, Australia Introduction Pest risk analysis (PRA) or Import risk analysis (IRA) is the process that is used to technically justify phytosanitary measures that are imposed on the importation of plants and plant products. Although the process is primarily used to determine import conditions for commercial quantities of traded commodities, it must be applied also to importations of small quantities of germplasm because in both cases there must be technical justification for the phytosanitary measures imposed. International guidelines for the pest risk analysis methodology have been developed by the Interim Commission on Phytosanitary Measures (ICPM) under its mandate to harmonise plant quarantine/phytosanitary procedures at a global level under the World Trade Organisation’s Sanitary and Phytosanitary Agreement (SPS Agreement). The two main International Standards for Phytosanitary Measures (ISPMs) are #2 Guidelines for pest risk analysis (FAO 1996a) and #11 Pest Risk Analysis for quarantine pests (FAO 2001). The identification of pests of concern in the movement of germplasm and the formulation of phytosanitary conditions for the management of these pests were initially developed at an international level by FAO/IPGRI (Frison et al. 1993; Diekmann, this chapter). Later, this publication was modified for one key pest using the ISPM Guideline for PRA methodology at the ACIAR/COGENT/IPGRI meeting in Malaysia (Diekmann 1997). The purpose of this article is to examine the process of PRA in more detail, to identify major pests of concern at a global level and to suggest pest management options that are available that could be adopted at national or regional level to reduce pest risk to an acceptable level. In this case, the task is to devise pest management strategies for the exchange of seednuts, embryo cultures and pollen among the International Coconut Genebanks (ICGs) of the International Coconut Genetic Resources Network (COGENT), and between the ICGs and their country members. 412 COCONUT GENETIC RESOURCES The Pest Risk Analysis process Phase 1: Initiating the PRA Initiating the PRA process involves the identification of the range of pests that are likely to be in the pathway (carried by the propagule) such as seednuts, pollen or embryos. In the case of movement within and between the ICG centres, information on national pest status is taken from the available international technical literature. International pest data can be taken from sources such as the FAO/IPGRI Guidelines for the exchange of coconut germplasm (Frison et al. 1993), the CABI Crop Protection Compendium (CABI 2003) and other sources. Such a literature search initially compiles information on all pests associated with the coconut crop worldwide, irrespective of the type of material that is to be moved as germplasm. Therefore, this primary information gathering activity is non-selective but forms the basis for decision-making. Crucial to the correct progression of the PRA process is the determination of the pest status of the respective countries in accordance with ISPM #8 Pest status in an area (FAO 1996b). In this case, the status of many pests is uncertain and, without conducting extensive in-country surveys, will have to rely upon the literature citations presently available. In the Asia Pacific region, there have been a number of useful compilations of pests and diseases of economic importance. These include data on the coconut crop, but these have been obtained in consultation with agencies that have not provided primary technical references as is generally the case of other compendia (APPPC 1987; Waterhouse 1993; Waterhouse 1997; Li et al. 1997). This information has been included at face value by CABI in its Compendium, but it has not been possible to further investigate the specific impact these pests have on the coconut palm, particularly the affected plant part, or to completely validate the records by further cross references. Therefore, it is essential that the relevant plant protection organization of a country verify the lists of pests compiled from literature sources with responsibility for research and extension in coconuts. For each of the pests identified in the primary pest list, the technical data for pests of potential quarantine concern are compiled in a pest datasheet. The data compiled include information on pest biology, in particular that which relates to the capacity of the pest to be in the pathway and to enter, establish and spread in the importing area. When available, information on the economic importance of the pest is also gathered in order to support the classification of the pest as a quarantine pest in accordance with the International Plant Protection Convention’s definition of a ‘quarantine pest’ and Phase 2 of the PRA process. At the end of this first phase of the PRA process, a list of pests in the country of export is compiled together with a list of pests in the importing 413 CHAPTER 6: Major pests and safe movement of germplasm Pests that are recorded as being associated with coconut growing but are not on the germplasm pathway such as larger mammals (vertebrates), nematodes and weeds are eliminated from the analysis for the following reasons: Vertebrate pests Rats (Rattus sp) and the plantain squirrel (Callsciurus notatus) are too large to be in any pathway considered for germplasm exchange. Nematodes Nematodes can be serious pests of coconuts, but are root pests and would not be in the pathway and are not considered further in this analysis. If nuts were harvested from the ground and could be contaminated with soil, nematodes could be in the pathway. country. By subtracting the second (importing) country list from the first (exporting) country list, a list of potential quarantine pests that move into Phase 2 of the procedure are determined. Phase 2: Pest risk assessment The second of final stage in the classification of the potential quarantine pests include those that would be in the pathway for tissue cultured embryos, seednuts or pollen and is undertaken as the final component of the completion of the datasheet for each pest. The assumptions made concerning economic impact are included in this classification. In each datasheet, the quarantine status for exchange of seednuts, embryo culture and pollen is also recorded. For each quarantine pest or pest of potential quarantine threat, on each datasheet, an assessment of the risk of the pest in each of the three pathways is made using the following table: Key biological information PRA* Seednuts Embryo culture Pollen Risk of entry Risk of establishment Risk of spread Economic impact Quarantine status Overall risk Risk management required *Legend: L=low M=medium H= high NA=not applicable Y=yes N= no Q=quarantine pest NR= non-regulated pest 414 COCONUT GENETIC RESOURCES Weeds A large number of weed species are recorded in association with the cultivation of coconuts as an economic crop. Many are economically significant. However, none would be considered in the pathway, as it would be expected that only seednuts from the palm would be used for germplasm exchange and they would be cleaned of any material prior to partial de-husking. Weeds would only be a problem if nuts were harvested from the ground and could be contaminated with soil. Arthropod pests Seednuts. Leaf and trunk pests are not considered to pose a serious risk with the import of nuts. A number of pests are recorded on flower heads and young nuts and are considered in the pathway. A number of general pests such as scales and mites are found on all parts of the coconut plant and have been considered as hitchhikers. These pests will require risk management options. Embryo cultures. Arthropod pests are not considered to enter the pathway for imports that are made as embryo cultured material because of the technique that is used and the sterile conditions under which the material is extracted and cultured. Pollen. A number of pests infest the floral parts of the coconut such as Tirathaba rufivena and Unaspis citri and could be a problem if care is not taken during harvest. However, they are large enough to be able to be detected by visual inspection. Mites are sometimes a problem with contamination of pollen harvested in the field, and would require examination using a hand lens or binocular microscope to detect infestation. Diseases Seednuts. Pests are only considered of quarantine significance if they are known to be seed borne, such as Marasmius palmivorus (oil palm bunch rot). Many pests are systemic but not seed transmitted such as Foliar decay and Anomola pallida. Embryo cultures. Pests that are systemic may not necessarily be present in the embryo. However, one pathogen cadang-cadang, has been detected in the embryo, but is not proven to be seed borne. Pollen. Most pests do not infect pollen although cadang-cadang has been detected in pollen. Whether it is pollen borne is not proven. 415 CHAPTER 6: Major pests and safe movement of germplasm Diseases and pests of unknown etiology Frison et al. (1993) lists a number of diseases/disorders of unknown etiology that are present in Asia and the Pacific regions. Those included are Finschhafen disease, Frond rot, Malaysia wilt, Natuna wilt, Socorro wilt and Stem necrosis. These diseases are restricted in distribution and the precautionary principle is implemented by requiring that material of all types moved from areas where they occur should be from areas free of the pests in accordance with international standards for pest free areas (ISPM # 4 FAO 1996b). Since the causes of the diseases are not known since no tests are available, this is the only phytosanitary management option. In accordance with ISPM #2 at the end of Phase 2 of the PRA process, the quarantine pests in the pathways should have been identified. These pests would now require that phytosanitary management procedures be identified to address the phytosanitary risk identified. Phase 3: Pest risk management It is assumed that all COGENT ICGs have the capacity to handle germplasm as seednuts, tissue cultures and pollen, and that they have the equipment to undertake the risk management operational procedures that are recommended. If the genebanks do not have the capability to undertake the required treatments then they must be undertaken before export at the point of exit or if post entry quarantine is not possible, then third country, intermediate quarantine would be required. Management options for the movement of germplasm between the Asian and Pacific COGENT ICGs in India, Indonesia and Papua New Guinea are given in Annex 1. Management of quarantine pest groups Although a number of different pests have been identified as quarantine pests in Annex 1, these need not be dealt with on an individual basis. Quarantine pests management strategies are developed to deal with the risks of pest groups of ‘like minded’ pests, rather than individual quarantine pests. Arthropod pests Seednuts. The accepted method of managing arthropod pests has been fumigation with an appropriate broad-spectrum chemical (Frison et al. 1993). Currently, the practice is to remove part of the husk of the coconut, thereby removing some of the pests, and to fumigate with methyl bromide (MeBr) at the rate of 32g per cubic metre for three hours at 21oC. This treatment will effectively deal with all arthropod pests that have casually 416 COCONUT GENETIC RESOURCES moved to the coconut fruit, such as the leaf feeders, scales, thrips, bugs and mites. Methyl bromide is known to have some phytotoxic effect on coconuts and care should be exercised in undertaking the treatment. The treatment at 32g per cubic metre for 24 hours at 20oC is used for devitalisation treatment in Australia (treatment A7.b. in FAO 1984).Temperatures for the treatment should not be high, water should be placed in the chamber in trays before the fumigation begins to increase humidity. The dehusked nuts should be removed from the chamber as soon as the treatment is completed and placed in a cool, ventilated area to allow the fumigant to disperse from around the coir. If MeBr is not available as a fumigant, then aluminium phosphide is an alternative at the rate of 225 ppm of phosphine gas for 120 hrs at 20oC (treatment B4h.(5)(e) in FAO 1984) or 2-3 tablets per cubic metre for 24-72 hours (treatment C13 (30) in FAO 1984). Embryo cultures. Arthropod pests are not considered to be in the pathway when movement as tissue cultures is correctly undertaken. However, there have been instances where small mite pests have contaminated cultures, so imports should be carefully inspected for these pests on arrival by microscopic examination. Pollen. Established methods for collecting pollen have been described (Balingasa and Santos 1978 and Frison et al. 1993). These methods would prevent pollen contamination from neighbouring palms and also prevent contamination by airborne pests if carefully applied. Treatment of pollen is not possible, other than sieving out the larger contaminating pests, so all consignments should be carefully, visually inspected using a low power microscope, before dispatch and again at point of entry. Fungal diseases Seednuts. A number of fungal diseases have been recorded on seednuts and flower clusters and therefore have the potential to be in the pathway. Nevertheless, whether all of these are seedborne has not been determined, although the risk exists. Invoking the precautionary principle, it is recommended that where these diseases are identified by the PRA as of concern, the nuts should be grown in post-entry quarantine (PEQ). Where a disease occurs generally in an area, only healthy nuts should be selected for exchange. Where diseases are not widespread and do not occur in specific and defined areas, then nuts should be sourced from these pest free areas. Seednuts should be treated with an acceptable and registered fungicide before sowing in PEQ. 417 CHAPTER 6: Major pests and safe movement of germplasm Embryo cultures. Embryo cultures free of contamination would not present a pathway for the introduction of fungal diseases. Pollen. Pollen should be visually inspected after gathering for fungal spores and again at point of entry. Pollen found infected should be destroyed. Viruses, viroids, mollicutes and phytoplasmas These systemic diseases have to be managed either through sourcing from pest free areas, or by active testing where the disease occurs generally and is not controlled. The causal organism for some of these diseases has not been determined and the precautionary principle is invoked to ensure that risk of incursions with exchange is negligible. As a general principle for these diseases, material should only be collected from palms showing no symptoms. Although this in no way guarantees freedom from these diseases, it does reduce the possibility of the disease being in the pathway. Seednuts. They should never be moved directly from areas where non- cultivable mollicutes or Phytomonas occur, to areas not affected with these pathogens (Frison et al. 1993). This is recommended despite the fact that there is no firm evidence that any of these systemic diseases are transmitted by seed. The research on cadang-cadang in controlled non- infected areas has not been completed so material from the infected area should not be exchanged, or only made from palms indexed free of the viroid. Embryo cultures. The presence of some systemic diseases has been detected in the embryo of coconuts. Therefore, material must only be taken from plants that are known to be free of these diseases, or the material taken as tissue must be indexed before release for growing in a propagation nursery. Pollen. Cadang-cadang has been detected in pollen. However, the evidence of the disease being transmitted this way to seed is not yet available. Nevertheless, as a precautionary measure, pollen should be tested or sourced from areas free of cadang-cadang. General phytosanitary measures for the movement of coconut germplasm Administration (i.e., permits, etc.) As well as compliance with countries’ phytosanitary requirements, the collecting and exchange of germplasm should be undertaken with the 418 COCONUT GENETIC RESOURCES full participation of the stakeholders, which could be collectors, breeders, other scientists and farmers. In the case of exchange between national and regional centres, it can be assumed that formal approval is sought for the movement at a bilateral level. Nevertheless, with the possibility of the movement from national sources outside the collections into other centres, compliance with good collecting practices should be iterated, particularly if a standard procedure is being developed and adopted worldwide, such as the International Code of Conduct for Plant Germplasm Collecting and Transfer. This Code “aims to promote the rational collection and sustainable use of genetic resources, to prevent genetic erosion, and to protect the interests of both donors and collectors of germplasm. The Code, a voluntary one, has been developed by FAO and negotiated by its Member Nations through the Organization’s Commission on Plant Genetic Resources. The Code is based on the principle of national sovereignty over plant genetic resources and sets out standards and principles to be observed by those countries and institutions that adhere to it. The Code proposes procedures to request and/or to issue licences for collecting missions, provides guidelines for collectors themselves, and extends responsibilities and obligations to the sponsors of missions, the curators of genebanks, and the users of genetic material”(FAO 1993). The Code outlines the arrangements that should be made prior to collecting missions. In particular, import permits should be obtained that clearly indicate the phytosanitary conditions that must be met prior to the material being exported. With the increasing reliance on the concept of area freedom and the indexing of source plants, these phytosanitary requirements must be fulfilled otherwise, the material will most likely be destroyed on arrival. Specifically the Code requires collectors or curators of collections to – “(c) make arrangements with quarantine officials, seed storage managers and curators to ensure that the samples are transferred as quickly as possible to conditions which optimize their viability; (d) obtain, in accordance with the importing countries’ requirements, the phytosanitary certificate(s) and other documentation needed for transferring the material collected. Treatments Because of the uncertainty about the distribution of many of these pests and lack of precise information on their biology, particularly the parts of the plant affected, it is prudent to require a set of general measures to address overall pest risk as well as requirements for specific regulated pests. These general recommendations are as follows: 419 CHAPTER 6: Major pests and safe movement of germplasm • Germplasm should only be collected from apparently healthy palms • Seednuts should be collected from the palm, not from the ground • Seednuts partially dehusked and fumigated at port of exit • Seednuts be treated with an approved fungicide and grown in post-entry quarantine for at least one growing season (in the tropics, the duration of a wet season and at least three months of a dry season), for release only after examination and certification of pest freedom by a plant pathologist. References APPPC. 1987. Insect pests of economic significance affecting major crops of the countries in Asia and the Pacific region. Technical Document No. 135. Regional FAO Office for Asia and the Pacific (RAPA), Bangkok, Thailand. Batugal, P. 1997. Implications of restricted coconut germplasm movement. Pp. 4-7. In: M Diekmann (ed). Proceedings of the meeting on viroid- like sequences of coconut, 21-23 April 1997, Kajang, Kuala Lumpur, Malaysia. ACIAR, Australia/IPGRI, Rome, Italy. Balingasa, EN and GA Santos, 1978. Manual for coconut hand pollination technique. Breeding and Genetics Division, Philippine Coconut Authority, Bago-Oshiro, Davao City, Philippines. CABI. 2003. Crop Protection Compendium. CDRom. Commonwealth Agriculture Bureau International, Wallingford, UK. Diekmann, M (ed). 1997. Proceedings of the meeting on viroid-like sequences in coconut. 21-23 April 1997, Kajang, Kuala Lumpur, Malaysia. ACIAR, Australia/IPGRI, Rome, Italy. FAO. 1984. International plant quarantine treatment manual, FAO Plant production and protection paper, 50. FAO, Rome, Italy. 220pp. FAO. 1996a. Guidelines for pest risk analysis. ISPM #2. FAO, Rome, Italy FAO. 1996b. Guidelines for the establishment of pest free areas. ISPM # 4. FAO. Rome, Italy. FAO. 2001. Pest risk analysis of quarantine pests. ISPM #11. FAO Rome. Italy Frison, EA, CAJ Putter and M Diekmann.1993. Technical guidelines for the safe movement of coconut germplasm [and addendum (1997)]. FAO/IPGRI, Rome, Italy. Foale, M and PW Lynch. 1994. Coconut improvement in the South Pacific. Proceedings of a workshop in Taveuni, Fiji. ACIAR Proceedings #53. ACIAR, Canberra, Australia. 420 COCONUT GENETIC RESOURCES Li Li-ying, R Wang and DF Waterhouse. 1997. The distribution and importance of arthropod pests and weeds of agriculture and forestry plantations in Southern China. ACIAR, Canberra, Australia. 185pp. Waterhouse, DF. 1993. The major arthropod pests and weeds of agriculture in Southeast Asia. ACIAR, Canberra, Australia. 141pp. Waterhouse, DF and KR Norris. 1987. Biological control: Pacific prospects. Inkata Press, Melbourne, Australia. 454pp. 421 CHAPTER 6: Major pests and safe movement of germplasm Annex 1. Specific phytosanitary measures for the movement of coconut germplasm between and among the ICG host coun- tries in the Asia and the Pacific regions (India, Indonesia and Papua New Guinea) India to Indonesia Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Coccus hesperidum (brown soft scale) • Nipaecoccus nipae (spiked mealybug) • Oligonychus biharensis • Raoiella indica • Tetranychus ludeni (red spider mite) Fumigation Not applicable Inspection Diseases • Hypocrea rufa (fruit rot: Citrus spp.) Fungicide and PEQ Not applicable Not applicable • Kerala wilt (root wilt) Area freedom* Area freedom Area freedom Note: *Pest-free Area Indonesia to India Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Hidari irava (coconut skipper) • Mahasena corbetti (coconut case caterpillar) • Rhabdoscelus obscurus (New Guinea sugarcane weevil) • Unaspis citri (citrus snow scale) Fumigation Not applicable Inspection Disease • Natuna wilt Area freedom Area freedom Area freedom India to Papua New Guinea Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Coccus hesperidum (brown soft scale) • Nipaecoccus nipae (spiked mealybug) • Oligonychus biharensis • Raoiella indica Mites • Tetranychus cinnabarinus (carmine spider mite) • Tetranychus ludeni (red spider mite) Fumigation Not applicable Inspection Diseases • Hypocrea rufa (fruit rot: Citrus spp.) Fungicide and PEQ Not applicable Not applicable • Kerala wilt (root wilt) Area freedom Area freedom Area freedom 422 COCONUT GENETIC RESOURCES Papua New Guinea to India Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Amblypelta cocophaga (coconut bug) • Amblypelta theobromae (coconut bug) • Axiagastus cambelli • Mahasena corbetti (coconut case caterpillar) • Rhabdoscelus obscurus (New Guinea sugarcane weevil) • Unaspis citri (citrus snow scale) Fumigation Not applicable Not applicable Diseases • Phytophthora katsurae (chestnut downy mildew) Fungicide & PEQ Not applicable Not applicable • Finschhafen disease Area freedom Area freedom Area freedom Indonesia to Papua New Guinea Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Hidari irava (coconut skipper) • Icerya pulchra • Tetranychus cinnabarinus (carmine spider mite) Fumigation Not applicable Inspection Disease • Natuna wilt Area freedom Area freedom Area freedom Papua New Guinea to Indonesia Management Options Quarantine Pests Nuts Embryo Pollen Arthropods • Amblypelta cocophaga (coconut bug) • Amblypelta theobromae (coconut bug) • Axiagastus cambelli Fumigation Not applicable Inspection Diseases • Phytophthora katsurae (chestnut downy mildew) Fungicide and PEQ Not applicable Not applicable • Finschhafen disease Area freedom Area freedom Area freedom 423 CHAPTER 6: Major pests and safe movement of germplasm Treatment recommendations Fumigation: As recommended with methyl bromide (MeBr) or phosphine gas. Dip: As recommended in Frison et al. (1993). The fungicide must be registered for use in particular circumstances and it is not therefore possible to make a specific recommendation here. Post-entry quarantine: The nuts after treatment should be germinated in an enclosed area, preferably a greenhouse or a screenhouse depending on the capacity to control temperature. The plants should be grown for at least one growing season (in the tropics, the duration of a wet season and at least three months of a dry season) and during this time regularly examined by a qualified plant pathologist with experience in palm pathology. If required, samples of leaf and other material should be taken for specific diagnostic testing. Diagnostic tests: Tests for specific disorders and diseases should be conducted as prescribed by Frison et al. (1993). Area freedom: Some quarantine pests are not distributed throughout the country of origin of the germplasm. It is therefore possible to obtain certification from the national plant protection organization of the exporting country a declaration that the material (seednuts, embryo or pollen) have been obtained from an area where the specific pests of concern have not been detected. This declaration should be made on a Phytosanitary certificate that accompanies the consignment. If this is not obtained then testing must be done for the diseases, or if the risk is considered too great, then the material is destroyed. 424 COCONUT GENETIC RESOURCES 425 CHAPTER 6: Major pests and safe movement of germplasm Chapter 7 Information, public awareness, institutional support and partnerships 426 COCONUT GENETIC RESOURCES 427 CHAPTER 7: Information, public awareness,institutional support and partnerships The international coconut genetic resources database C Hamelin1, R Bourdeix2 and L Baudouin3 1Scientist, 2Coconut Breeder and 3Geneticist Centre de Coopération Internationale en Re- cherche Agronomique pour le Développement (CIRAD), Montpellier, Cedex 5, France Background During the International Workshop on Coconut Genetic Resources, held in Cipanas (Indonesia) in October 1991, two major recommendations were made for coconut: the first was to set up the International Coconut Genetic Resources Network (COGENT), and the second, concerning network information and documentation, stated as follows: “Participants agree on the need for a single central coconut database to be developed for the initial stage of the network. The offer by CIRAD Montpellier to act as a host for this database was gratefully accepted. […]” A meeting was then organized in Montpellier (France) from 19 to 22 May 1992, with representatives from national collections to clarify the status of existing collections, define how the database was to be organized and draw up the list of descriptors to be taken into account, along with precise standardized ways of observing those descriptors. It was decided that the database would be developed in several stages. The development of an IT (Information Technology) application was initiated in 1992 with the presentation of a database mock-up called the Coconut Genetic Resources Database (CGRD), which eventually became the formal title for the database. It has continued since then on an annual basis. Each stage involved adding/improving management system functionalities, and increasing the number of accession sites contained in the database. Furthermore, in order to take into account the progress made in IT techniques, the application, which initially functioned under the MS-DOS system, was completely rewritten for the Microsoft Windows system, so that photos and graphs could be displayed, and to provide a more user-friendly interface. Lastly, the progress made in molecular techniques, and their use on coconut to characterize genetic diversity, revealed the need to add the possibility of storing molecular information from these techniques in the database. Table 1 shows how the database developed over the years in terms of its functions and content. It can be seen that the successive increases in the number of accessions over time has been irregular. The growth rates that can be calculated vary between 1 and 34%. From 1995 to 2002, 428 COCONUT GENETIC RESOURCES Brazil, China, India, Indonesia, Ivory Coast, Jamaica, Mexico, Papua New Guinea, Philippines, Sri Lanka, Thailand, Vanuatu and Vietnam (Bourdeix 1996; 1997a; 1997b; 1998; Bourdeix et al. 1999; Baudouin 2002) were visited and local researchers were trained in gathering and inputting data into the database. Such visits also provided an excellent opportunity to test the software in real situations and to detect items that needed improvements. Objectives The work on CGRD was initiated for the main purpose of providing the COGENT members with an easily consultable computerized catalogue of accessions representing a large number of cultivars spread throughout the coconut growing zone, in order to gain a clearer picture of coconut genetic diversity and thereby promote exchange of germplasm. This continues to be the main objective. Another purpose of this database is for the COGENT country members to establish a list of passport descriptors and standardize characterization/assessment descriptors specific to coconut to be used by all partners. In addition, the database was created to speed up cultivar characterization and evaluation. The members of COGENT supplying the information contained in the database are regularly encouraged to add new information or complete the information already recorded requiring them to make a considerable effort to gather information and thereby improve their knowledge of the cultivars planted in their genebanks. Table 1. Stages in the development of CGRD Date Version Functions added/Improved No. of accessions 1993 Mock-up Version 1.0 Passport data Functions to query the database and to create reports 0 500 1994 Version 2.0 Characterization and evaluation data Backup function 669 1996 Version 2.1 DIP (Data Interchange Protocol) format for data export Restoration function 738 1997 Version 2.2 New structure Improved software 1998 Version 2.2 improved New DIP (Data Interchange Protocol) format for data export 936 1999 Version 2.2 New reports 1225 2000 Version 3.0 for Windows Migration to Windows 2001 Version 4.0 for Windows Improved functions Export in delimited ASCII files (to use in statistical software) 1352 2002 Version 5.0 for Windows Introduction of coconut molecular data 1369 2003 Version 5.1 for Windows Improved backup function 1416 429 CHAPTER 7: Information, public awareness,institutional support and partnerships Organization The data entered into the base are the values of passport and characterization descriptors for coconut accessions defined by COGENT. It also includes photos of the palms, along with results and diagrams from microsatellite molecular analyses. Based on an analysis of these data, and of the relations and constraints existing between them, a conceptual model of database organization was established. It provided a formal description of the database (Figure 1) using entities (symbolized by rectangles) corresponding to the natural objects identified in the system (sites, accessions, cultivars, photos, etc.), and relations (symbolized by circles) between those entities. This model was modified during development, when molecular data or photos were integrated. In the model, accessions and cultivars form the core of the database. The database structure respects certain management rules, such as: • It is compulsory for an accession to belong to a collection site and its number is unique in the database; • A cultivar can be represented by several accessions; and • Photographs and molecular data are attached to the cultivars. Figure 1. Simplified conceptual model of the CGRD The conceptual model was translated into a relational type logical model (Figure 2) consisting of tables (symbolized by rectangles) linked to each other, derived from the entities and relations of the conceptual model. A relational type organization was chosen because it is a widely used 430 COCONUT GENETIC RESOURCES The CGRD was designed to function in a semi-centralized way. All COGENT members have a version of the database on their machine, which can be used to consult the entire catalogue, and to add or update data on the accessions at their site. In the latter case, a basic functionality enables data backup per site, for transmission to CIRAD (Centre de Coopération Internationale en Recherche Agronomique pour le Développement), the organization that centralizes the data from all the collection sites. CIRAD checks data coherence before entering them in the database, which is distributed annually to COGENT members. Functions The CGRD has a management system endowed with functions that can be used to carry out all the necessary operations on the data it contains. Among the functions available, which are listed below, there are those that are found in conventional database management systems, but there are also specific functions, given the nature of the database: • Entering/consulting information on collection sites • Entering/consulting data on individual accessions • Selections in the database using criteria • Creation of various types of reports Figure 2. Simplified relational logical model of CGRD database organization, which has proven its worth and enables the use of a very powerful query language. Lastly, a very large number of commercial database management systems function with this type of organization. This logical model was implemented in the chosen relational database management system. 431 CHAPTER 7: Information, public awareness,institutional support and partnerships Figure 3. The tab picture of the accession screen in CGRD • Backing up of the database on an external medium • Restoring database from a previous backup to the computer hard disk (when the database files are damaged on the hard disk) • Exporting accession data in D.I.P. format (Data Interchange Pro- tocol, for introduction into generalized genetic resources databases accepting this format) • Consulting pictures of coconut cultivars on various topics • Consulting the molecular group-based coconut classification • Obtaining information on microsatellite markers and viewing the electrophoretic profile • Online help The database has a Microsoft Windows type graphic interface with a menu from which operations to be implemented on the data can be chosen. Figures 3 to 5 show how data are displayed in CGRD. The screen displayed in Figure 3 is a part of the main screen of CGRD where pictures can be seen and from which descriptors data can be entered by clicking on the Passport or Characterization and evaluation data tabs. 432 COCONUT GENETIC RESOURCES Figure 4 shows a list of accessions retrieved after a successful search in the database. By clicking on the Accessions details tab when an accession is selected, all previously entered information on passport or on characterization data will be displayed on the screen. Figure 4. The screen resulting from a successful selection of accessions in CGRD Figure 5 shows the Classification Tree of the coconut cultivars annotated with the molecular groups identified in the species. A click on a letter of the molecular group, displays the list of cultivars of the same group on the right of the screen. Figure 5. The Classification Tree in CGRD 433 CHAPTER 7: Information, public awareness,institutional support and partnerships Contents Each accession in the database is characterized by 145 descriptors, of which there are 72 passport descriptors and 73 characterization and evaluation descriptors. Passport descriptors are divided into two groups: the accession data group, which contains accession characteristics (category, colour, cultivar, parental origin, etc.) and the collection data group, which contains data on the site and the original population from which the accession was sampled. The characterization and evaluation descriptors are generally quantitative traits, whose values are means, calculated from values measured on several palms. A standard deviation is associated with each of the means, to have an idea of the variability of the trait within the accessions. To obtain these values, a data management system has to be put in place for every palm in the field. This is not trouble-free, because it has to be carried out over a long period following a regular schedule. In order to help researchers to follow this management protocol, COGENT has committed CIRAD in 1996 to develop dedicated software called CDM (Coconut Data Management). This software was designed to manage experimental data observed on collections and experimental fields of coconut and other crops. The version 3 delivered in March 2000 is able to manage the palm identification characters along with data on observations during the vegetative phase, leaf morphology, stem measurements and state of the palms. It is possible to execute powerful queries on the database, to export data into external file, and to make statistical analysis of widely used experimental designs. The CGRD characterization and evaluation descriptors are divided into: • Site descriptors, information about the site at which the acces- sion is to be found and about the people assessing it; • Germination descriptors, germination rates and percentages; • Stem descriptors, stem morphology (height, circumference, number of internodes, etc.); • Leaf descriptors, leaf morphology (petiole, rachis, leaflets, etc.); • Inflorescence descriptors, to characterize inflorescences (pedun- cle, spikelets, number of female flowers, etc.); • Flowering descriptors, such as the length of male and female phases, information on overlapping of these phases, spathe emis- sion date, inflorescence opening date, etc.; • Fruit descriptors, nut characteristics (shape, weight of different compartments, dry matter weight, etc.); • Yield descriptors, number of bunches, number of nuts, quantity of copra; and 434 COCONUT GENETIC RESOURCES • Oil descriptors, oil/nut characteristics (quantity of oil in fresh matter and in dry matter form). In 2003, the main statistics about the database were as follows (Table 2a and 2b): • The accessions of 28 genebanks located in South Asia, Southeast Asia, the South Pacific, Africa, and the Caribbean−Central America zone figure in the database. • Some countries, such as Indonesia, Malaysia and Papua New Guinea, have at least two sites where collections are maintained, with a maximum of four for Indonesia. • A little over 60% of the accessions are in the South and Southeast Asia, primarily in India, the Philippines and Indonesia. • Not all the descriptors are filled in; a little over half of the acces- sions (all sites combined) have values for 25% to 50% of their passport and evaluation descriptors. • Very few accessions have values for 100% of their passport descriptors or evaluation descriptors. • The database lists 599 Tall cultivars, 111 Dwarf cultivars, 1 semi- Tall cultivar plus a few others of small size but cross-fertilizing. • Some cultivars such as the Malayan Tall, Pakistan Tall are de- scribed very often in the database, represented by 49 and 32 ac- cessions, respectively, whereas more than 70% of the cultivars are only represented by one accession. • The database contains 754 photos representing a little over 20% of the cultivars. The aspects illustrated are the whole plant, the crown, the inflorescence, the fruit, genetic diversity, and cultural aspects of the coconut palm. • Photos of inflorescence, bunch and fruit are the most numerous. • The molecular data contained in the database come from microsatellite electrophoresis studies carried out on cultivars. On average, 14 microsatellite loci have been analyzed per cultivar. • For each cultivar and each locus, the frequencies of the different alleles found at the locus are indicated. • Based on the allelic frequencies of the microsatellite loci studied, cultivars have been assigned to different molecular affiliation groups. The molecular group of each cultivar is also recorded in the database. A cultivar classification tree based on the molecular groups is proposed in one of the database modules. It is possible to move along the different branches of the tree, bringing up a list of the cultivars attached to the selected molecular group in each case. The microsatellite data of 119 435 CHAPTER 7: Information, public awareness,institutional support and partnerships Site Number of accessions P=0 02000 mm), the BYD, BGD and CRD flower well at 1.7, 1.8 and 2.0 years, respectively. Hybrids BYD x BRT and BRD x BRT, on the other hand, flowered at 2.5 years; BGD x BRT at 2.6 years; and the giant BRT at 3.4 years. In the regions with annual rainfall of only about 1200 mm and unevenly distributed as in the coastal state of Piauí, BRT, BGD, BGD x BRT, BYD x BRT and BRD x BRT started to produce in the second and third year after blooming, registering 17.2 and 44.8; 109.1 and 91.9; 68.5 and 76.0; 64.2 and 98.1; 92.7 and 75.8 fruits/plant/year, respectively. The research activities described above are also being carried out under different agroecosystems in the states of Alagoas, Pernambuco, Rio Grande do Norte, Pará, Goiás, Mato Grosso, São Paulo, Minas Gerais, Espirito Santo, Paraná and Distrito Federal, being part of the National Net of Evaluation of Cultivars of Coconut Palm (RENAC). A second RENAC is being implemented with other cultivars in the states of Sergipe, Bahia, São Paulo, Paraná, Brasilia and, possibly Alagoas and Mato Grosso. Future plans The following would be undertaken by Embrapa in the near future to further develop the coconut genetic resources in Brazil: 1. Establish the International Coconut Genebank for Latin America and the Caribbean in Itaporanga-SE; and 2. Maintain the experiments of the international multilocation hy- brids trial and disseminate their results. References FAO. 2002. Statistical databases: Agriculture 2002. Rome, Italy. IBGE. 2001. Produção Agrícola Municipal. Rio de Janeiro. http:// www.sidra.ibge.gov.br Moretzsohn, M de C, PJA. Coelho, ZP de S Amaral, A Hercos and EA Tupinambá. 2001. Desenvolvimento e uso de marcadores microssatélites na análise da variabilidade genética de ecótipos de 703 CHAPTER 9: Country reports on status of coconut genetic resources research coqueiro (Cocos nucifera L.). Embrapa Recursos Genéticos e Biotecnologia, Brasília (Boletim de Pesquisa e Desenvolvimento, 16). Ribeiro, FE and ER de Siqueira. 1995. Introdução, coleta e conservação de germoplasma de coqueiro no Brasil. Embrapa-CPATC, Aracaju. (Documentos, 3). Ribeiro, FE, ER de Siqueira, WM Aragão and EA Tupinambá. 2000. Ecótipos de coqueiro gigante no Brasil. Embrapa Tabuleiros Costeiros, 704 COCONUT GENETIC RESOURCES Latin America and the Caribbean Status of coconut genetic resources research in Mexico R Castillo1 and C Oropeza2 1Investigador del Sistema Producto, Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias (INIFAP), Mexico 2Coconut Researcher, Centro de Investigacion Cientifica de Yucatan (CICY), Mexico Introduction Mexico is the major coconut producing country in Latin America after Brazil with 168 000 ha planted to coconut. Since 1998, coconut farmers in the Latin America and Caribbean (LAC) region have been facing serious problems that have diminished their incomes due to the low prices of copra and the decrease in the yield of old coconut palms. In Mexico, in the Gulf–Caribbean region, there are about 30 000 ha of the Atlantic Tall coconut whose average yields have fallen to 550 kg/ha due to the prevalence of Lethal Yellowing Disease (LYD). In the last 16 years, LYD has devastated around 17 000 ha of coconut stands in the country. Coconut palms affected by LYD die within a couple of months after infection. The common symptoms showed by an LYD-infected palm are as follows: 1. Fruits fall prematurely, the ovaries miscarry and also fall; 2. Open inflorescences exhibit partial necrosis while developing ones (after nutfall) exhibit total necrosis; 3. Mature leaves yellow and then turn brown. They later dry com- pletely and hang limp from the shaft of the palm; and 4. The youngest leaves in the middle of the crown also die and fall usually after the mature leaves fall off, leaving the tree bald and with a telephone pole appearance. Generally, it takes only about six months from the onset of the disease until the tree dies and assumes the characteristic telephone pole appearance (Figure 1). Mortality rate is 100% for palms infected by the disease. At the moment, LYD is the most serious problem affecting not only Mexico but also Honduras, Guatemala, El Salvador and Belize in Latin America; Jamaica, Haiti, Dominican Republic and Trinidad-Tobago in the Caribbean, as well as Tanzania, Ghana and Mozambique in Africa. In some of these countries, it is suspected that the causal agent of the disease could be a different strain of the Mycoplasm-Type Organism (MTO), although this is yet to be proven. Due to its extensive and rapid 705 CHAPTER 9: Country reports on status of coconut genetic resources research spread, it is estimated that LYD would eventually infect and devastate about 500 000 ha of coconut stands in these 12 countries. Currently, cooperative efforts are underway among these countries to combat the disease and to develop coconut varieties which are resistant to LYD. Coconut research activities Mexico and Jamaica are currently carrying out combined efforts to develop technology to overcome LYD, which include projects on genetic improvement, microsatellite analysis, in vitro multiplication and seed production. So far, genetic improvement seems to be the most feasible and effective measure against LYD. This involves the evaluation, selection and recombination of coconut germplasm to identify resistant sources which could be used in breeding programmes for LYD-resistant varieties. Other significant coconut R&D projects in LAC are briefly described below. Multilocation trials to identify suitable coconut hybrids and varieties for Africa, Latin America and the Caribbean Starting from year 2000, Mexico, Brazil and Jamaica participated in this COGENT-supported project funded by Common Fund for Commodities (CFC) to evaluate two groups of coconut hybrids where six were common in the experiments of all participating countries and four are local hybrids Figure 1. Atlantic Tall coconuts in South Mexico affected by LYD 706 COCONUT GENETIC RESOURCES or varieties. It is envisioned that results obtained may be applied in other coconut countries with similar environmental conditions. In case of Mexico, eight local hybrids were evaluated: • Malayan Red Dwarf x Panama Tall Aguadulce(MRD x PNT01)• Malayan Red Dwarf x Panama Tall Monagre(MRD x PNT02)• Malayan Yellow Dwarf x Mexican Pacific Tall 14 (MYD x MXPT14)• Malayan Yellow Dwarf x Mexican Pacific Tall 9 (MYD x MXPT09)• Malayan Yellow Dwarf x Mexican Pacific Tall 2 (MYD x MXPT02)• Malayan Yellow Dwarf x Panama Tall Aguadulce (MYD x PNT01)• Malayan Yellow Dwarf x Panama Tall Monagre (MYD x PNT02)• Malayan Yellow Dwarf x Mexican Pacific Tall 10 (MYD x MXPT10 ) These hybrids have progenitors that demonstrated resistance to Lethal Yellowing. The experiment was established in October 2002 in the coast of Tabasco, Mexico. The results revealed that hybrids flowered in 2.5- 3.0 years after planting and have potential to benefit smallholders. Establishing a framework and selecting project sites for the nation- wide deployment of coconut-based poverty reduction interventions in coconut growing communities using COGENT’s 3-pronged strat- egy in Mexico A socioeconomic study in five coconut growing communities in Mexico was carried out to identify suitable pilot communities and groups that could undertake poverty intervention projects. COGENT’s 3-pronged strategy in helping poor coconut farmers, especially women, improve their incomes and living standard, will be tested under this project in one community. The project aims to:: 1. Increase yields and incomes by deploying high-yielding; high- value multipurpose and adapted coconut varieties and hybrids using locally produced seednuts; 2. Increase incomes by promoting the production and marketing of high-value products from the meat, husk, shell, water, wood and leaves and identifying suitable varieties for these uses; and 3. Increase food security and incomes per unit area per unit time through intercropping and livestock/fodder production. Use of biofertilizers for sustainable production Technology has been developed on biofertilizer application for Atlantic Tall coconuts in Gulf of Mexico. The advantages of using biofertilizer are its low cost and ease of application. This biofertilizer is being promoted to help farmers increase yields and farm profitability. 707 CHAPTER 9: Country reports on status of coconut genetic resources research Results/outputs and their significance Zizumbo et al. (1998) conducted a 7-year evaluation of Tall-type coconuts from the Atlantic and the Pacific in Mexico. The Tall cultivars Guerrero 2, Jalisco 1, Michoacan 2, Colima 1, Colima 2 and Colima 3 exhibited some form of resistance to LYD. These genotypes were used as male parents to develop new Dwarf x Tall hybrids, some of which are being tested under the CFC-funded ‘Multilocation trials to identify suitable coconut hybrids in Africa, Latin America and the Caribbean’. Other hybrids from these genotypes are being evaluated under two different experiments with the same purpose of finding resistance to LYD. Carrillo (1998) and Dominguez (1994), after nine years of evaluating the coconut hybrid ‘Chactemal’, found that it has high resistance to LYD in trials established in two Mexican towns. The hybrid is now being propagated in four seed gardens for dissemination and planting in LYD-devastated areas. However, as the current seednuts production can only cater to the planting needs of approximately 700 ha per annum, it cannot keep up with the rate of spread of the disease. Related to this, Dominguez et al. (2003) developed a technology to produce Dwarf x Tall hybrids with a productivity of 150 seednuts per palm per year. This technology, developed under the project entitled, ‘Mixed Seed Garden Production of Coconut Hybrids’ consists of using male and female parents in the same field, cauterizing the spikelets after the emasculation and applying Gibberellic acid at 100 ppm when concluding the pollination using bees. Currently, about 3 200 ha in Mexico have been planted with LYD-resistant hybrids and growing at a rate of 900 ha per year. With regards to the rehabilitation of old Atlantic Tall stands, which were abandoned when world copra prices dropped, Castillo et al. (2003) found that with two applications per year of Micorriza + Azospirillum + 45-35-60 yield could be increased to 872 kg/ha, compared with the yield of the plantation before the application of the treatments that was of 542 kg/ha. This represents a yield increment of 61%. The application of Azospirillum with 816 kg/ha gave an increment of 50% yield. This technology is very economical and could allow coconut farmers to increase incomes as well as to conserve their precious genetic resources. Suggested next steps In Mexico, the following are the priority activities to develop the country’s coconut genetic resources: 1. Identification, development and propagation of LYD-resistant Tall coconut varieties using open pollination; and 2. Establishment of a coconut genebank in Tabasco dedicated to the conservation and propagation of LYD-resistant hybrids/varieties. 708 COCONUT GENETIC RESOURCES References Carrillo, RH. 1998. Lethal yellowing coconut in Mexico. Pp. 49-80. In Memoirs of Masterful Conferences. First National Meeting of Coco- nut Palm. Acapulco, Mexico. (In Spanish). Castillo, GR, CE Dominguez and BP Ruiz. 2003. Evaluation of biofertilizers and fertilizers of slow liberation in Atlantic Tall Coco- nut. Final report. FOSIGOLFO, INIFAP. (In Spanish) Been, BO. 1981. Observation on field resistance to lethal yellowing in coconut varieties and hybrids in Jamaica. Oleagineux 36: 9-12. Domínguez, CE, AJI Lopez and BP Ruiz. 1994. Preliminary evaluation of the coconut hybrid Malayan Yellow Dwarf x Lazaro Cardenas Tall in Tabasco. Pp. 14-16. In: Proceedings of the 7th Scientific Meet- ing. SAGAR-INIFAP. Villahermosa, Tabasco. (In Spanish). Domínguez, CE, GR Castillo and BP Ruiz. 2003. Mixed seed garden production of coconut hybrids. Final report. FOSIGOLFO. INIFAP. (In Spanish). Harries, HC. 1973. Selection and breeding of coconuts for resistance to diseases such as lethal yellowing. Oleagineux 28 8-9: 395-398. Zizumbo, VD, BM Fernandez and LR Cardeña. 1998. Evaluation of the resistance to lethal yellowing of the coconut germplasm in Mexico. Pp. 87-98. In: Memoirs of Masterful conferences. First National Meeting of Coconut Palm, Acapulco, Mexico. (In Spanish). 709 CHAPTER 9: Country reports on status of coconut genetic resources research Latin America and the Caribbean Status of coconut genetic resources research in Guyana O Homenauth Director, National Agriculture Research Institute (NARI), Mon Repos, East Coast Demerara, Guyana Introduction In Guyana, coconut (Cocos nucifera L.) ranks third, next to rice and sugar, among the most economically important crops. In spite of this, the potential of the crop has been largely underexploited and poorly developed. Coconuts contribute only approximately 1% to the total gross domestic product (GDP) of the country, an under achievement, considering its priority ranking in the agricultural sector. It is estimated that there are 24 000 ha under coconut production, with an average annual production of 92 million nuts. In order to increase and sustain the current levels of production to meet market demand for greater economic efficiency, it is imperative that the issue of increasing coconut productivity be urgently addressed. According to Paul (1999), varietal improvement is the most expedient approach to resolving low productivity and consequent economic inefficiency. He further stated that varietal enhancement necessitates an analysis of the genetic structure (population level) and production potential of the diverse types, forms, strains and varieties grown in Guyana. This article describes the coconut varieties and forms, and pests and diseases of economic importance in Guyana. A summary of the proposed coconut R&D development project for Guyana is also discussed. Coconut varieties and forms Coconut is grown widely on the coastal regions of Guyana, primarily along the Pomeroon River, in the Essequibo Coast, East Demerara, and West Berbice and on the Corentyne Coast. Coconut is mainly processed into cooking oil. Average copra yield from 100 nuts ranges from 13 to 16 kg. The use of tendernut as a nutritive beverage is very popular in Guyana. Commercial holdings of coconut are mainly planted with two types of the Tall variety and two types of the Dwarf variety. The Tall types are the predominant source of copra, while the Dwarf variety is specially grown for their sweet water. One variant of intermediate height, known as ‘Bastard Nut’, is grown in the Pomeroon River area and is cultivated for both copra production and for its sweet water, although its copra yield is inferior to the Tall types. 710 COCONUT GENETIC RESOURCES Tall types The most common Tall types existing in the country are the Jamaica Tall and the Panama Tall, each consisting of two basic colour forms: green and bronze. The Jamaica Tall bears long, angular nuts with distinct ridges and a thick mesocarp. Dehusked, its nuts are also angular and pointed at the end. On the other hand, the Panama Tall fruits are much more spherical with thinner mesocarp. These two types may be considered the ‘original’ Tall types in Guyana. Another variant of the Tall type found on the Coastal Corentyne (No. 60 Village), as reported by Manthriratoa (1980), is a type with spherical, medium-sized nuts but with a pronounced dark pink mesocarp. Several variations in epicarp colour forms have also been observed. Generally, commercial stands could not be classified on this criterion alone. Farmers, however, could distinguish between the two Tall types known as Clara Nut and Cocrit Nut. Clara Nut is similar in character to the Panama Tall. Cocrit Nut, on the other hand, seems to combine the nut characteristics of the Jamaican Tall and the Panama Tall. The Cocrit Nut is regarded as a ‘nut number’ type rather than a ‘nut size’ type. Fruits of the Cocrit Nut are more spherical than oblong, with a thin mesocarp and thick kernel. Nut size ranges from small to large, with trees of larger nuts being less prolific. Commercial copra producers prefer the ‘5-year’ (five years to begin production) nuts with an intermediate nut size, high yield and precocity. The Clara Nut is a ‘nut size’ type. Husked nuts are large and spherical, with a high water capacity but thin kernel. The coconut water of this type is described as sweet and is favoured over all the other types grown for their coconut water. The hectarage devoted to Clara Nut, however, is minuscule compared to those variants preferred for copra. A preponderance of Tall types can generally be found in all commercial holdings. However, in the Pomeroon River area, there is a higher frequency of Dwarf types in commercial holdings. In all commercial plantings, demand for new planting materials is generally for the Cocrit Nut type. Dwarf types Commercial Dwarf types are mainly of the green and yellow ‘3-year’ (three years to begin production) variants. Manthriratoa (1980) described the Green Dwarf as being similar to the Brazilian Green Dwarf in growth habit, number of nut per bunch and size of nuts. The Yellow Dwarf, however, is different from the Malaysian Yellow Dwarf, in having a larger nut size and a less intense yellow colour in petioles and epicarps of the nuts. A third Dwarf type variant is the Red Dwarf (Orange Dwarf), restricted mainly to the Pomeroon River area. The Red Dwarf is similar 711 CHAPTER 9: Country reports on status of coconut genetic resources research to the Red Dwarf of India and Sri Lanka and the Malaysian Dwarf. Manthriratoa (1980) suggested that this variant may be a recent introduction from a Caribbean Country. Another variant, the Bronze Dwarf (so-called because of its bronze epicarp), is a relatively new find of about four years ago. The Bronze Dwarf was reportedly introduced from Surinam. Currently, there are only a few homesteads with this variety, but the current demand suggests it has the potential to spread rapidly to commercial holdings. Bastard nut Bastard Nut is predominately recognized in the Pomeroon River area. Manthriratoa (1980) attributed the origin of this variant to natural cross pollination between Dwarf and Tall types, being an apparent Dwarf x Tall hybrid. Bastard Nuts show marked hybrid vigour in trunk and leaf size, number of bunches produced per year and number of nuts per bunch. Pests and diseases The major pests of coconuts in Guyana are the coconut caterpillar (Brassolis sophorae L.); moth borer (Castnia daedalus Crammer); and Azteca ant-scale complex. Of the three predominant coconut pests identified in Guyana, the coconut caterpillar is considered the most destructive. On the other hand, the major diseases of coconuts in Guyana are: cedros wilt, red ring disease; and bud rot. None of these diseases poses serious threats to the coconut industry in Guyana. It is also worthwhile to note that there is no reported case of Lethal Yellowing Disease (LYD) in Guyana. Proposed coconut R & D projects for Guyana Varietal improvement Projects on varietal improvement shall address the immediate breeding objectives of increased yields, early maturity and disease resistance, especially against LYD. Breeding strategies shall include the following: Standardization of evaluation techniques. Not much information is documented in Guyana on the evaluation procedures for coconut. In order to ensure that evaluation results be comparable with those obtained by evaluation programmes in other coconut-producing countries, it would be necessary to adopt the standard protocols for such evaluation programmes. 712 COCONUT GENETIC RESOURCES Characterization of locally-adapted germplasm resources. To date, monitoring of coconut germplasm resources in farmers’ fields has established the location and prevalence of four Dwarf varieties. There are obvious variants of the Tall types, especially those of the intermediate or ‘Bastard’ types. However, unlike other crops cultivated in Guyana, it is not the habit of farmers to assign names to prominent variants of coconut. For this reason, assembling local coconut collections and characterizing them through conventional agronomic and morphometric methodologies could prove to be quite difficult. Faced with a wide range of cultivated variants, it will be most appropriate to focus on analysis relating the observed pattern of traditional cultivation practices with the prevalence of coconut genetic diversity. This is where the application of DNA marker technology, like AFLP, becomes relevant. Elucidating the structure of molecular diversity, for example of ‘Bastard Nuts’, will assist in establishing their genetic origin and structure. Hybrid coconut production. High-yielding varieties are the best way to mitigate low crop production efficiencies; low productivity being the most serious limitation to the viability of the coconut industry in the country. Despite the relatively wide range of germplasm resources available locally, it is apparent that the current rate of varietal improvement is still inadequate to sustain the development of the coconut industry. The solution is basically one of utilization. The performance of non- conventional and conventional Dwarf x Tall hybrids, produced locally, has been reported to be satisfactory. The potential genetic diversity of locally available Tall, Dwarf and ‘Bastard Nut’ germplasm opens up the possibility for testing a number of combinations. In addition, the introduction of refined germplasm and superior accessions of known performance suitable to Guyana’s growing conditions and production circumstances will not only expand the genetic base of existing germplasm but also greatly magnify the spectrum of possible hybrid combinations. Networking with regional and international coconut R&D programmes is therefore necessary. In this regard, three options for hybrid research may be exploited: • Introduction and evaluation of elite hybrids to identify hybrids suitable to nut production in Guyana. It is anticipated that many hybrids are presently available from the existing breeding pro- grammes of other coconut growing countries that could be widely tested, probably in a regional setting. This is intended to have a continuous, medium-term impact; • Development of hybrids using proven progenitors, the objec- 713 CHAPTER 9: Country reports on status of coconut genetic resources research tive being to deliver potentially superior hybrids in the shortest time possible. This option will require the application of DNA markers; and • Hybrid coconut development using combinations of superior inbred Dwarf lines and Tall non-inbred progenitors. Some of the progenitors will include the parental lines presented in the first option. The objectives, in addition to developing hybrids, are to determine the combining ability and heterotic pattern cum grouping of adapted progenitors using DNA analysis; and to ob- tain information on genetic diversity and structure of the local coconut germplasm. This then will set the stage to better man- aged and maintained high level on-farm coconut genetic resources diversity. Specifically, the project will determine the level of in- ter- and intra-varietal diversity, with the objective of establishing a breeding population which ultimately will have to be a com- prehensive national coconut breeding programme. A further ben- efit of DNA typing of local coconut germplasm is that it will en- able the determination of the requirements for infusing exotic germplasm material. However, until DNA typing techniques be- come accessible, in situ maintenance and monitoring will con- tinue to be in the front line of the country’s coconut genetic re- sources conservation strategy. Development of disease indexing potential In order to adopt a pre-emptive strategy to control LYD, the exchange of exotic germplasm and development of disease indexing capability need to be expedited. With minimum staff training and provision of the necessary equipment and materials, the Tissue Culture Laboratory, in conjunction with the Plant Protection Laboratories of the National Agriculture Research Institute (NARI), can accommodate a LYD screening facility. Of specific value is diagnosing the early occurrence of LYD so that its potential damage can be mitigated. Coconut tissue culture As a useful complement to the conventional nursery method, embryo culture technique has the potential of shortening the generation time to establish hybrids and their progenitors. Moreover, this technique, combined with disease indexing, will expedite the introduction of exotic germplasm and in the long term establish the capability for cloning superior palms through tissue culture. With adequate training, the tissue culture facility of NARI can also facilitate this procedure. 714 COCONUT GENETIC RESOURCES Collaborative coconut quality evaluation The importance of copra quality should not be neglected. Quality adds value to copra. The experience of large-scale copra processors will be invaluable in this aspect of research. The processing companies may see it necessary to jointly finance this sector of coconut research in order that they also fully profit from generated technologies. References Manthriratoa, MAPP. 1980. Rehabilitation of the coconut industry of Guyana. Report to the Government of the Cooperative Republic of Guyana and the FAO. Paul, CR. 1999. Coconut growing ecologies and the status of coconut germplasm resources in Guyana. Paper presented at the Laboratory Course on the Application of Biotechnology to Plant Breeding and Crop Potection in Coconut. CICY, Merida, Yucatan, Mexico. 715 CHAPTER 9: Country reports on status of coconut genetic resources research Latin America and the Caribbean Status of coconut genetic resources research in Jamaica B Been Director for Research, Coconut Industry Board (CIB), Kingston, Jamaica Introduction The coconut is not indigenous to Jamaica. The prevailing opinion is that it was first introduced to the Caribbean and Atlantic coasts of South America about the middle of the 16th century (Purseglove 1968). Initially, coconuts seemed to have been planted near harbours and coastal settlements but later, with the expansion of plantation agriculture, the crop was grown inland and by 1681, when Hans Sloane visited Jamaica, coconuts were widespread. By the end of the 19th century, the coconut had become a plantation crop and flourished in many parts of Jamaica especially on hillsides, which are not suitable for sugarcane, located mostly in the wetter eastern section of the island. At the beginning of the 20th century, the majority of the coconuts grown commercially were of the Atlantic Tall (Jamaica Tall) variety but old records showed that there were at least four other varieties, including the King Coconut introduced from Sri Lanka in 1869. In 1973, Captain Bligh brought coconuts along with breadfruit to Jamaica from Tahiti. Following periods of severe hurricane activity in 1903, 1904, 1912- 1917, large numbers of nuts of the Panama Tall variety were imported from the San Blas Islands. Again, in 1922, there was further importation of nuts from Panama. In 1921, after the Panama Canal had been opened, a small number of seednuts of the Niu Leka Dwarf variety were brought to Jamaica from Fiji. The Niu Leka gave rise to two local populations: Tulloch Dwarf and Fiji Dwarf. These have not been of much commercial value. In 1933, 12 open-pollinated seednuts from two hybrids of Malayan Dwarf x Niu Leka were introduced. The third and fourth generation selections, locally known as Fiji-Malayans, are still being used in the country’s breeding programme. From 1938 to 1940, a few selected red-fruited Malayan Dwarf seednuts were introduced from the Caribbean island of Trinidad and from these a population of over 10 000 palms was established in the western part of the island. It was in that plantation that resistance to lethal yellowing was first documented. In 1939, about 150 seednuts from red- and yellow- fruited Malayan Dwarfs were introduced from Florida. Large-scale introductions of Malayan Dwarf seednuts (red, green and yellow colour 716 COCONUT GENETIC RESOURCES forms) were made from the island of St. Lucia in 1945 and 1951 following severe hurricanes. A further introduction from the same source was made in 1968 as a consequence of the lethal yellowing resistance possessed by the Malayan Dwarf. During the 1960s, in response to an outbreak of lethal yellowing disease in the main coconut growing region, germplasm from most of the countries in the Asia-Pacific region were introduced with international assistance for screening. These included Bougainville Tall, Chowghat Green Dwarf, Fijian Tall, Indian West Dwarf, King Coconut, Malayan Tall, Malayan Red Dwarf, Malayan Yellow Dwarf, Malayan Green Dwarf, Niu Leka Dwarf, Peru Tall, Rangiroa Red Dwarf, Rangiroa Tall, Rennell Tall, Rotuma Tall, Samoan Tall, Sarawak Tall, Seychelles Tall, Solomon Islands Tall, Spicata Tall, Spicata Red Dwarf, Tahitian Tall, Thailand Tall, Tonga Tall, Vanuatu Tall and Yap Island Tall. In addition, pollens of Cambodia Tall, Tahiti Tall, Mozambique Tall and Cameroon Red Dwarf were obtained from Institut de Recherches pour les Huiles et Oléagineux (IRHO). As a part of the multilocation trial funded by the Common Fund for Commodities (CFC), six F1 hybrids were introduced into Jamaica in 2000 and 2002. In 2001, seednuts from selected Panama Talls in Mexico and of two locally produced F1 hybrids were imported into Jamaica. Germplasm and hurricanes Jamaica is located in the hurricane belt and during the period 1886- 1986, the island experienced 171 ‘events’ (hurricanes) of varying intensities (Gray 1990). Over the years, multipurpose variety trials have yielded useful data on windstorm damage. Following a hurricane in 1944, it was reported that 60% of 30 560 Jamaica Talls were destroyed compared with only 6% of 5120 Panama Talls (Coconut Industry Board 1962). After another hurricane in 1980, it was observed that the Malayan Dwarf was not as resistant to storm damage as the Panama Tall and the F1 hybrids of Malayan Dwarf x Panama Tall and Malayan Dwarf x Jamaica Tall. Yaps Talls, Seychelles Talls and late generation Fiji-Malayans showed high windstorm resistance (Coconut Industry Board 1980). Data collected from variety trials in six sites following a severe hurricane in 1989 showed that of the eleven varieties involved, the Malayan Dwarf was the most susceptible and Malayan Dwarf x Panama Tall (Maypan), the least damaged. Data gathered from wind-thrown palms that were recovered suggested that canopy may be of less importance in determining wind damage than trunk height and diameter of the bole (Johnson et al. 1994). 717 CHAPTER 9: Country reports on status of coconut genetic resources research Germplasm and lethal yellowing When lethal yellowing appeared in the main coconut belt in 1961, all varieties being grown locally were tested for disease resistance. Fortunately, the Malayan Dwarf, a variety introduced earlier, was found to have good resistance. In addition, it was precocious and highly productive under good conditions. Farmers were encouraged to replant the variety in affected areas and in underplant areas not yet affected by the disease. Unfortunately, the Malayan Dwarf had relatively low oil content and did not do well under marginal conditions hence, the search continued for other resistant varieties. With the assistance of the Overseas Development Administration (ODA) and Food and Agriculture Organization of the United Nations (FAO), germplasm was collected in Southeast Asia and the Pacific. These were screened for resistance in field trials. When it became obvious that none of the introductions was more resistant than the Malayan Dwarf, a hybridization programme was started in an attempt to combine in the F1 progeny the high disease resistance of the Malayan Dwarf with the large fruit size and hardiness of the Talls. One of the early crosses, Malayan Dwarf x Panama Tall, called Maypan, was found to be productive and resistant. A system was then devised to produce it commercially. Screening of introductions for lethal yellowing resistance has been ongoing and other F1 hybrids have been produced and tested (Been 1981), but to date the Maypan remains the most popular variety accounting for more than a half of the palms grown commercially. During the mid 1990s it was observed that, in certain places, Malayan Dwarf and Maypan palms were exhibiting lethal yellowing mortalities at a rate higher than previously observed. The trend continued and now the disease is destroying thousands of palms in the main coconut growing areas. Work done on the pathogen suggests that the phytoplasma may have mutated and/or exceptional environmental conditions may be combining to produce situations, which are extremely favourable for the development and spread of the disease. Screening and breeding for lethal yellowing resistance will continue with every effort being made to import new germplasm for further evaluation on lethal yellowing resistance. IPGRI/COGENT activities in Jamaica Since the 1990s, the International Plant Genetic Resources Institute’s International Coconut Genetic Resources Network (IPGRI/COGENT) has been supporting activities related to germplasm conservation and use in Jamaica, especially in the areas of capacity building and research undertakings. 718 COCONUT GENETIC RESOURCES In 1997, Jamaica received expert technical assistance in formulating a coconut regional project proposal for Latin America and the Caribbean. In the area of training and human resource development, a regional STANTECH (Standardized research technique in coconut breeding) training course was held in Jamaica in 1997 and the Botanist/Plant Breeder of the Coconut Industry Board attended a similar regional course in Africa. Two COGENT-sponsored meetings were hosted by Jamaica in 1997 and 2002. Currently, Jamaica is participating in the Common Fund for Commodities (CFC)-funded Multilocation Trials to identify suitable coconut hybrids/varieties for Africa, Latin America and the Caribbean. The Department for International Development (DFID)-funded project entitled ‘Establishing a Framework and Selecting Project Sites for a Nationwide Deployment of Coconut-based Poverty Reduction Interventions in Coconut Growing Communities using COGENT’s 3- pronged strategy in Jamaica’ has also been completed. The Coconut Industry Board has been the implementing agency for both projects. The Board, a statutory body established in 1945, is responsible for and advises the government on matters regarding the Jamaican coconut industry. Prior to the establishment of a research department in the Board, the Ministry of Agriculture was responsible for coconut research and development. Conclusion The use of introduced germplasm has enabled the local coconut industry to survive despite hurricanes and lethal yellowing disease, the two main threats to the industry. The resurgence of lethal yellowing necessitates a renewed search for and use of germplasm with good resistance to this disease. The eventual solution to the problem may lie in the realm of genetic engineering, but until then, conventional breeding procedures would have to be used. References Been, BO. 1981. Observations on field resistance to lethal yellowing in coconut varieties and hybrids in Jamaica. Oleagineux 36 1:9-12. Coconut Industry Board. 1962. Second report of the Research Depart- ment. Jamaica, West Indies. Coconut Industry Board. 1980. Twentieth report of the Research De- partment. Jamaica, West Indies. Gray, CR. 1990. History of topical cyclones in Jamaica 1886-1986. Jamai- can Journal of Science and Technology 1:29-48. 719 CHAPTER 9: Country reports on status of coconut genetic resources research Johnson, CF, WJ Fielding and B Been. 1994. Hurricane damage to differ- ent coconut varieties. Tropical Agriculture (Trinidad) 71 3:239-242. Purseglove, JW. 1972. Tropical crops: Monocotyledons. Longmans, Lon- don. 607p. 720 COCONUT GENETIC RESOURCES South Pacific Status of coconut genetic resources research in the Cook Islands W Wigmore1 and T Mataora2 1Director of Research and 2Senior Research Officer, Ministry of Agriculture, Rarotonga, Cook Islands Introduction The Cook Islands consist of a group of 15 small islands scattered between 167o west and 8-23o south of the equator. Its total land area is 237 sq km, and the country has one of the largest Exclusive Economic Zones in the Pacific Region covering approximately 1.8 million sq km of ocean. The islands are geographically divided into two groups: the Northern Islands and the Southern Islands. The two island groups have marked differences in their agricultural activities. The Northern Islands group remains relatively isolated from the Southern Islands, with the latter continuing to indulge in more diversified agricultural practices. The Southern Islands group has a cooler climate and more fertile soil enabling a wider variety of agricultural production as compared with the Northern Islands where the soil is relatively unfertile and has poor water holding capacity, thereby limiting agriculture-related activities. A census of agriculture in 2000 reports a total cultivated area of 1945 ha or 8.2% of the total land area, a drop of 3.3% from data collected in the previous census in 1988. Coconut occupied 34.5 ha of the cultivated land, which includes intercrops in a coconut-based farming system. The proportion of land under coconuts is somewhat higher on the atolls. Coconut is widely used, especially in the rural communities and the Northern Islands, for food and other numerous purposes. Copra, in previous years, constituted a major export commodity. However, as international demand for the commodity dropped, copra production became a non-viable venture. In April 2000, Dr Roland Bourdeix, coconut palm geneticist and breeder from the Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD) visited the Cook Islands, with the support of COGENT, from 30 March to 13 April 2000 to collect data on coconut genetic resources in Cook Islands. Tiara Mataora, Senior Research Officer with the Ministry of Agriculture acted as the local project leader. The coconut germplasm collecting, characterization and conservation were conducted on the islands of Rarotonga and Aitutaki. The project aimed to survey and collect available data on diversity of 721 CHAPTER 9: Country reports on status of coconut genetic resources research local coconut populations which will be used for breeding and to mitigate genetic erosion due to population pressure, palm ageing and natural hazards. Collecting and characterization of coconut germplasm The project’s specific objectives are as follows: 1. Train coconut researchers on germplasm collecting, and on coco- nut breeding research techniques using the International Coco- nut Genetic Resources Network’s (COGENT) standardized re- search techniques in coconut breeding (STANTECH) Manual; 2. Collect germplasm initially from one or two sites then to another three to seven sites; 3. Characterize germplasm collected from the sites to identify popu- lations and conserve desirable germplasm; 4. Plant and manage collected germplasm in a genebank; 5. Characterize the collected germplasm and submit passport and characterization data to COGENT’s International Coconut Ge- netic Resources Database (CGRD); and 6. Send embryos of selected populations to the Secretariat of the Pacific Community (SPC), which will culture the embryos and subsequently transfer the resulting in vitro seedlings to the Inter- national Coconut Genebank for the South Pacific (ICG-SP) in Papua New Guinea. Problems and opportunities addressed by the project The Cook Islands lacks the necessary human resources, particularly a coconut expert or a full-time coconut research staff, to carry out coconut genetic resources research, limiting the exploitation of the potential of coconuts and its products. It is anticipated that further survey activities shall include populations in the other islands of the country, such as the island of Pukapuka. The inclusion of other islands should provide a better understanding of coconut genetic diversity in the Cook Islands. Training activities sponsored by COGENT A regional training course was organized on standardized research techniques in coconut breeding techniques which was held at the Agricultural Research and Training Centre in Vanuatu from 29 June to 9 July 1999. The training course was part of the Asian Development Bank-funded COGENT project entitled, ‘Coconut Genetic Resources Network and Human Resources Strengthening in Asia and the Pacific Region’. A researcher from the Cook Islands participated in this activity. 722 COCONUT GENETIC RESOURCES Results/ outputs of the germplasm survey The islands of Rarotonga and Aitutaki were surveyed. A visit to the island of Pukapuka was initially planned; however, this was impossible as there was no air link with the islands at the time of Dr Bourdeix’ visit. Pukapuka is an atoll island and was chosen for its isolation from most other islands and, therefore, may hold some unique coconut ecotypes. Aitutaki, on the other hand, is both a volcanic and atoll island (usually referred to as ‘almost an atoll’). Seednuts from seven populations were collected, taking into account the results of the previous generalized sampling strategy and results of the participatory survey in Aitutaki Island. Due to time constraints, it was not possible to make all the required characterizations for the seven populations. Description of surveyed populations Cook Islands Tall Seven-in-One (COKT01) In the centre of Avarua, capital of the Island of Rarotonga is a group of seven Tall palm trees growing in a circular and, what appears to be, a singular clump. Old folks in the area believe that the ‘clump’ actually originated from just one palm. However, several historians tell otherwise. To settle the issue once and for all, seednuts and leaflets from the trees were collected for DNA analysis to determine whether these palms are real ‘septuplets’, full-siblings, half-siblings or are entirely unrelated. Some 105 seednuts were collected, but many either were pre-germinated on the tree or are too young to harvest. Sixty albumen cylinders were obtained that finally gave 52 excised embryos, which were cultivated in vitro at SPC’s tissue culture laboratory in Fiji. The resulting in vitro seedlings were later transplanted to the ICG-SP which is hosted by PNG. The summary descriptions of the surveyed coconut population in Cook Islands and related ongoing DNA analysis of the germplasm are presented in Table 1. Significance of the survey and impact on Cook Islands’ coconut ge- netic resources The results of the germplasm survey should assist with the assessment of coconut diversity in Cook Islands. It is anticipated that further exploration and collecting activities will include populations in other islands of the Cook Islands group. The collections made during this study will conserve precious germplasm which could be used to develop better varieties for the atolls and enrich the collection of the ICG-SP. The description and the molecular 723 CHAPTER 9: Country reports on status of coconut genetic resources research markers analysis of these populations will not only generate information on genetic diversity within these collections but also improve the knowledge on the origin and the dissemination of coconut in the Pacific Islands, and help facilitate the exchange of important cultivars with other countries. Suggested next steps The selected or identified populations of Tall and Dwarf coconuts with good characteristics will be continuously observed and may be used for further genetic breeding programmes. Selected populations of coconut materials transferred via tissue culture to PNG where the conserved genetic pool is located would continue to provide such materials for future research, not only to Cook Islands but also to other South Pacific countries. Tree No. Proposed Name and Abbreviation Origin Remarks 1 Cook Islands Tall – Sweet Husk (COKT03) Rarotonga Husk from the nut can be removed by hand. The internal part of the husk is sweet. 2 to 13 Cook Islands Tall – Papaaroa (COKT03) Rarotonga One mature fruit and several leaflets were harvested from each palm for molecular analysis. Palm No.2 had big, elongated brown nuts with a relatively high proportion of husk. 14 Red Dwarf Rarotonga There were no seednuts harvested for embryo extraction as similar cultivars are available in Papua New Guinea. Leaflets were collected for DNA analysis. 15 to 29 Cook Islands Dwarf – Totokoitu (COKD01) Rarotonga Ten palms were sampled for fruits and DNA analysis. 30 to 50 Cook Islands Tall – Vivi (COKT04) Aitutaki Seven or eight palms were sampled for DNA analysis. 51 to 80 Cook Islands Tall – Golf (COKT05) Aitutaki Ten palms were sampled for leaf analysis. 81 Cook Islands Dwarf – Vaikoa COKD02) Aitutaki One palm was sampled for DNA analysis. 82 to 88 Cook Islands Tall – Seven-in-One (COKT01) Rarotonga Seven palms were sampled for DNA analysis. 89 to 91 Cook Islands Tall – Papua River (COKT06) Rarotonga The palms are remnants of an old inland plantation. Sampling was done only for molecular analysis. Table 1. Description of the surveyed coconut populations in Cook Islands 724 COCONUT GENETIC RESOURCES References COGENT. 2003. COGENT Newsletter 7:9-12. IPGRI-APO, Serdang, Selangor, Malaysia. Ministry of Agriculture. 2000. Cook Islands Census of agriculture and fisheries. Cook Islands Government. Drew, OJ. 1985. Report on copra industry and potential copra crushing industry in the Cook Islands. Economic and Social Commission for Asia and the Pacific, United Nations. Foale, MA. 1987. Coconut germplasm in the South Pacific Islands. ACIAR Technical Reports Series No. 4. 23p. Foale, MA. 2003. The coconut odyssey: The bounteous possibilities of the tree of life. Australian Centre for International Agricultural Research, Monograph No. 101. 132p. Labouisse, JP and R Bourdeix. 2003. Coconut germplasm collecting, char- acterization, and conservation in Cook Islands, Kiribati, Marshall Is- lands and Tuvalu. Final report. March 2003. Vanuatu Agriculture Research and Training Centre, Santo, Vanuatu. Lombard, K. 2001. Reviewing the coconut (Cocos nucifera L.): The tree of life. A paper presented for PSS 5326 Advanced Seed Science, Texas Tech University, USA. Taffin, G de. 1993. Report of a fact-finding visit on the coconut industry of the Cook Islands. CIRAD, Montpellier, France. 725 CHAPTER 9: Country reports on status of coconut genetic resources research South Pacific Status of coconut genetic resources research in Fiji V Kumar1 and T Kete2 1Director of Research and 2Senior Research Officer, Ministry of Agriculture, Sugar and Land Resettlement (MASLR), Fiji Introduction Coconut germplasm collecting, conservation and varietal description has been an ongoing collaboration between the Ministry of Agriculture, Sugar and Land Resettlement (MASLR) and the International Plant Genetic Resources Network (COGENT) since1994. Fiji has an established germplasm collection consisting of initial eight accessions at the Taveuni Coconut Center. These accessions have not been fully characterized. To better utilize germplasm held in such collection characterization is vital. Characterization carried out in this activity is limited to morphological description using COGENT’s standardized techniques in coconut breeding (STANTECH) manual. The data collected from the evaluation were sent to the Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD) for inclusion in the International Coconut Genetic Resources (CGRD) database. COGENT consultants assisted our national projects by assessing our R&D capacity and also in helping MASLR to identify projects and training necessary and adapted to our situation. COGENT initially provided training for developing the National Coconut Research staff on using STANTECH. The knowledge and skills acquired through this training was useful and complementary to their existing skills. Also COGENT’s support in the evaluation of Fiji germplasm collection enabled us to know better the performance of the different accessions, which would assist us in recommending to farmers and policy makers the best varieties to be used for planting. Researchable problems and opportunities addressed by research conducted in the country Hybrid evaluation trial The evaluation of Dwarf x Tall hybrids is one of the major activities carried out. Four Dwarf by Tall hybrids consisting of Malayan Red Dwarf as their female parents and Fiji Tall, Niu Leka, Rennell Island Tall and Rotuman Tall as their male parents were used. These hybrids were 726 COCONUT GENETIC RESOURCES compared against Fiji Tall in a randomized complete block experiment. Like the establishment and evaluation of the coconut germplasm, this activity was established with the assistance of the European Community and CIRAD. Through COGENT, we were able to continue the evaluation. On a bi-monthly basis, yield in terms of the number of nuts per palm were collected and a fruit component analysis was carried out. The results of this trial were used in planning Fiji’s breeding and replanting programmes. New collections established Several collections were made for varieties, which existed as non– commercial varieties, but they had value-adding potentials. These potentials were never exploited previously. Hence, under the Asian Development Bank (ADB)-funded project, these varieties were collected and conserved at Taveuni Coconut Center for future research activities. On the other hand, several populations of Fiji Tall were studied for genetic diversity. The results showed that there was not much difference between the populations. Diversification of coconut uses In the farmer participatory research conducted by the Research Division, Ministry of Agriculture, Fisheries and Forests, the constraints facing coconut production in Fiji include lack of knowledge of farmer’s varieties and uses especially by the younger generation. In this survey, 21 varieties were identified and several uses. This reveal the potentials existing in communities for value adding which are not being tapped. Projects coordinated by COGENT in Fiji Evaluation of coconut germplasm collection and hybrids in Fiji Selection and breeding of high-yielding varieties (ADB/ COGENT PHASE 1) The Taveuni Coconut Center was established in 1987 to look into Fiji’s ailing coconut industry. To set up this station, the government purchased 384 ha of land for the construction of infrastructure and establishment of seedgardens and trials. The programme was initiated to address the declining production of coconuts through production of high yielding hybrids seednuts and seedlings for rehabilitation. During this time, COGENT coordinated an ADB-funded project to accomplish the following objectives: 1) evaluate and characterize the existing germplasm collection at Taveuni Coconut Center; 2) maintain and monitor performance of 727 CHAPTER 9: Country reports on status of coconut genetic resources research four hybrids compared to Fiji Tall; and 3) monitor nine on-farm demonstration plots. The evaluation of the hybrids and also the germplasm collection include data on yields in terms of copra per nut and per palm and also cyclone resistance of the different accessions. The results are presented in Table 1. Collecting of coconut genetic resources in Fiji (ADB/COGENT Phase II) The phase II project focused on the study of various populations of Fiji Talls in all provinces of Fiji and collecting other cultivars which were not previously considered and conserved, and were under threat of genetic erosion. Twenty-four populations of Fiji Talls were studied in eight islands and these include the three main islands namely Viti Levu, Vanua Levu and Taveuni and other islands in the eastern division. The objective of the study was to confirm the diversity that exists between the populations of Fiji Tall so that appropriate conservation strategies can be designed and implemented. The results revealed that there is not much difference among populations. On the other hand, the new collections made included Niu ni Magimagi, Niu Buludrau, Uto Gau, Niu Kitu and Stripped variety. The passport data have been included in the CGRD. See Tables 2 and 3 for summary of collecting activities. Sustainable use of coconut genetic resources to enhance incomes and nutrition of coconut smallholders in the Asia and Pacific region Component 1: Farmer participatory surveys Farmer participatory surveys using participatory rapid appraisal (PRA) tools were conducted in five villages selected from four islands in the archipelago. The objective was to identify farmer’s varieties and multipurpose uses of coconuts. The focus of the study went beyond the above two themes (i.e., the general preoccupation of the community in relation to coconut plus the impacts of coconuts in their respective community). The five villages identified several cultivars and uses (Table 4). Component 2: Evaluation and conservation of tendernut varieties Three major activities were carried out: 1. Initial market survey - local and overseas (Australia); 2. Transferred seednuts of tendernut variety to the western division for raising and distribution to farmers; and 3. Characterized tendernut varieties. 728 COCONUT GENETIC RESOURCES Cultivar Average dried Copra / nut (g) Average number of nuts produced per palm per annum Average yield copra / ha (tonnes) 10 x 10 m (triangular spacing) = 115 palms /ha Dwarf Niu Leka 236 25 0.7 Malayan Green 152 43 0.8 Malayan Red 166 36 0.7 Malayan Yellow 179 41 0.8 Hybrids MRD x FJT 213 29 0.7 MRD x NLAD 262 27 0.8 MRD x RIT 279 31 1.0 MRD x RTMT 269 33 1.0 Tall Fiji Tall - Taveuni 240 24 0.7 Fiji Tall - Lakeba 248 29 0.8 Rotuman Tall 261 19 0.6 Renell Island Tall 355 15 0.6 Table 1. Results of germplasm evaluation Table 2. Fiji Tall population study sites Table 3. Data of the new collections carried out nationally Date Locality Province No. of samples characterized Nov-97 Cicia Island Lau 2 Jul-98 Taveuni Island Cakaudrove 2 Jul-98 Nawaca Bua 1 Aug-98 Lakeba Island Lau 1 Sep-98 Rotuma Island Rotuma 2 Sep-98 Navutulevu Serua 1 Sep-98 Navua Serua 1 Sep-98 Toga Rewa 1 Oct-98 Savusavu Cakaudrove 2 Apr-99 Saqani Cakaudrove 1 Apr-99 Levuka Lomaiviti 2 May-99 Natavea Naitasiri 1 May-99 Navunibitu Ra 1 May-99 Nailega Tailevu 2 Jul-99 Vanua Balavu Lau 2 Nov-99 Kadavu Kadavu 2 Date of collection Collected from Variety No. of palms November 1997 Cicia Niu ni Magimagi 30 November 1997 Cicia Niu Buludrau 35 September 1998 Rotuma Uto Gau 12 September 1998 Rotuma Stripped Nuts 15 July 1999 Vanua Balavu Niu ni Magimagi 7 July 1999 Vanua Balavu Niu Buludrau 21 August 1999 Cicia Niu ni Magimagi 142 August 1999 Cicia Niu Buludrau 20 729 CHAPTER 9: Country reports on status of coconut genetic resources research Table 4. Coconut cultivars identified by the communities Komo Bouma Kanacea Namuka- l- Lau Nawaca Elders 10 9 7 11 12 Women 11 8 12 Married Men 11 6 11 9 Youth 11 6 9 6 The market study revealed the following: 1. Fiji Tall variety is the major source of green coconuts (90 – 95%) and one of the problems encountered by farmers is difficulty of harvesting due to the height of the palms; 2. Lack of diversity in the green coconut belt areas (Rakiraki and Serua); 3. The stakeholders are unaware of the varieties of coconut locally available; and 4. A demand that cannot be satisfied is bigger size drinking nuts for overseas markets. A nursery was established in the western division (Legalega Research Station) to raise the first batch of seed nuts from Taveuni Coconut Center. The seednuts were raised and established at the station as source of planting materials for tendernuts. Four Dwarf varieties were characterized for potential tendernut production and data sent to CIRAD for inclusion in CGRD in 1998. Developing sustainable coconut based income-generating technolo- gies in poor rural communities in Fiji To increase incomes of coconut farmers, MASLR participated in COGENT’s diversity-linked ‘Poverty reduction in coconut growing communities’ project, involving eight Asia Pacific countries. The project deployed and tested four income generating technologies: 1) production and marketing of high-value products from all parts of the coconut; 2) intercropping cash and food security crop; 3) livestock raising and feed/ fodder production; and 4) establishment of community-managed seedling nurseries. The project was implemented in three coconut growing communities, namely Belego, Tukavesi and Cicia. The project involved 17 participants in production of high-value coconut products, 454 in intercropping and 32 in animal and feed/fodder production. The project increased the incomes of participants by 2-5 times, enhanced their food security and nutrition, and more than 1000 coconut seedlings have been conserved on farm. 730 COCONUT GENETIC RESOURCES Training and capacity building Two coconut research staff (Tevita N Kete and Vijendra Kumar) were trained in Vanuatu in 1996 on STANTECH and the former attended this training course through funding from French Embassy in Fiji. Apart from the above, Fiji benefited from the visit of COGENT consultants (M Hazelman and G Santos) who assessed the National R&D capability, assisted the national program in identifying common problems, opportunities and projects for network collaboration. COGENT also supported the scholarship of Tevita Kete who obtained his Master’s degree from the University of the Philippines at Los Baños. Interpretation of significance or impact of output 1. The results obtained from the evaluation of the collections will be useful for the coconut industry for rehabilitation programmes, espe- cially the effect of the cyclone on varities. Current recommendation for the rehabilitation programme is to use Fiji Tall, Rotuman Tall and Niu Leka apart from hybrids of Rotuman Tall crossed with Malayan Red Dwarf. 2. Coconut research staff trained to execute and implement coconut research programs for the betterment of Fiji’s coconut industry. 3. Collecting of coconut cultivars were done and collected accessions conserved for future use. Suggested next steps New collections The new collections will be evaluated and characterized and multiplied for future breeding work. Most of the new collections made are varieties suitable for value adding, the potentials of which will be exploited. Coconut rehabilitation The results of the germplasm evaluation work will be used for selecting materials for replanting, as the cyclone of 2003 served as a test in determining the cyclone tolerance of the varieties existing in our collection. References Marechal, H. 1928. Observation and preliminary experiments on the coconut palm with a view to developing improved seed for Fiji. Fiji Agri. Journal 1:16-45. Mc Paul, JW. 1964. Coconut growing in Fiji. 2nd edition. Bulletin # 38, Department of Agriculture, Fiji. 78p. 731 CHAPTER 9: Country reports on status of coconut genetic resources research Parham, RW. 1960. Coconut and Breadfruit surveys of the South Pacific Region. South Pacific Commission, Technical Information # 1, Noumea, New Caledonia. Harries, HC. 1978. Evolution, dissemination and classification of Cocos nucifera L. Botanical Review. 44:265-320. Manciot, R and P Sivan. 1988. Coconut hybrids for the South Pacific Islands. In: Coconut breeding and management. Proceedings of the National Symposium, 23-26 November 1988. Kerala Agricultural University, Vellanikara, Trichur, India. 380p. Vernon AJ, PN Emose and T Mudaliar. 1975. Coconut varietal selection and breeding, Part 2: Recent work in Fiji. Fiji Agric. Journal 37:47-52. Annual Coconut Research Reports. 1986-1999. MASLR, Fiji. 732 COCONUT GENETIC RESOURCES South Pacific Status of coconut genetic resources research in Kiribati M Tenang Chief Agricultural Officer, Ministry of Natural Resources Development, Kiribati Introduction Coconut dominates agricultural production in Kiribati as the crop provides one of the main components of the people’s daily diet as well as drinks, copra for export, timber for construction, leaves for thatching, string and materials for handicrafts (Trewen 1985; Edward 1989; Beenna 2001). Edward (1989) commented that the total number of indigenous coconut plant species in Kiribati is very low, which is a reflection of the isolated location of the islands coupled with the infertility of the soil. Barr (1992) stated that 80% of the land area of the main Gilbert Group, where 93% of the population lives, is covered with coconut. The Agricultural Division has, for the last 30 years, conducted extensive research on coconut with assistance from the British Government. The emphasis on coconut improvement has been in response to the perceived importance of the coconut and its products in the lives of the people of Kiribati. The objective of most of the researches conducted has been confined to increasing coconut yields. Hence, two large-scale coconut plantation improvement schemes were devised; one aimed at improving traditional palm groves and the other targeted at rehabilitation and replanting. These coconut plantation improvement schemes mainly involved improving the quality of existing stands of traditional palm groves with reasonable density (Barr 1992). Thinning of over-dense areas was done by poisoning senile palms and non-productive ‘self-seeded’ younger palms. The scheme was terminated in the early 1970s as it was exhausting resources. In addition, data recording was very poor and could not be used to justify the continuation of the activities. Past coconut replanting schemes focused on replanting areas with less than 49 palms per hectare. The result of the scheme was quite disappointing as the actual production per hectare was far below expectations (Barr 1992). The problem was aggravated by poor or unsuitable planting sites, most of which were hard-pan or waterlogged. 733 CHAPTER 9: Country reports on status of coconut genetic resources research Research activities conducted and results/ outputs Experimental studies Coconut management trials were carried out at different locations to address the problems encountered in implementing the two plantation improvement schemes previously mentioned. It was anticipated that the resulting recommendations would then be demonstrated and transferred to smallholder farmers to help them rehabilitate their old palms, including replanted farms in order to increase yields (Trewen 1986; Barr 1992). Spacing/density. Three triangle pattern spacing were tested and conducted on a number of selected sites with variable rainfall, i.e. high and very low rainfall. The trials aimed to determine the optimum spacing for coconut growing. The results of the trials concluded that a spacing pattern of 9 m and 8 m was ideal especially for areas with high rainfall and good soil fertility. Basic NPK requirements. Two trials were carried out to determine the optimal NPK combinations for coconut growing in the atolls under marginal and normal soil conditions. The treatments were combinations of annual applications of 0, 1, 2, or 3 kg/palm of potassium chloride (KCl) fertilizer; 0 or 1 kg/palm of triple superphosphate fertilizer; and 0, 1, 2, 3 kg/palm of IBDU (N) fertilizer. All palms tested also received a trunk injection of iron sulphate at the start of the trial. The results suggested that potassium was the most important nutrient for marginal soil areas and K deficiency could be corrected within three years by applying 1 kg KCl/palm/year. It was also found that nitrogen application increased coconut yields. Manganese, copper and zinc trials. The trial examined the requirement for Mn (0 or 20 g/palm), Zn (0 or 8 g/palm), and Cu (0 or 3 g/palm) when applied with Fe and their interactions with applications of NPK. All the palms tested received a basal dressing of iron sulphate by trunk injection at the start of the trial. The results showed that there should be an optimum balance of Fe, Mn, Zn and Cu in order to promote good plant growth and production. It was also found out that Cu interacts with N to increase yields, while proper Cu and Mn combination improves N assimilation. Trace elements and application trial for coconut seedlings. The trial investigated the optimum method for giving seedlings a long-term supply of Fe at the time of planting and also examined whether other trace 734 COCONUT GENETIC RESOURCES elements are required other than Fe. The trace element compounds used consisted of iron sulphate (75%), manganese sulphate (15%), zinc sulphate (7%) and copper sulphate (3%). The compounds were applied using two doses of FeSO4 (50 g and 375 g), following two different methods of application (applied to the husk of a seedling or buried near the seedling). For good growth, it was recommended that 50 g of FeSO4 be applied to the husk while 200 g is recommended if the soil-covered method was followed. Iron application trial. The trial compared the effect of four different iron compounds: iron sulphate, chelated iron, fritted iron (iron in glass) and iron fillings. The results showed that chelated iron was the fastest acting compound followed by iron sulphate, iron fillings and fritted iron. Application of iron compounds corrected iron deficiency and improved coconut growth. Coconut demonstration project The main objective of the project was to educate and assist farmers who wish to improve their coconuts. Activities included establishing demonstration plots on each island, organizing field days for farmers, conducting radio programmes about coconut improvement, and providing training on recommend crop management techniques. Collaborative activities with the International Coconut Genetic Resources Network (COGENT) Establishment of genebanks A genebank has been set up in the Central Nursery. The hybrids were collected from different locations (islands) in Kiribati, including South Tarawa. Since studies have shown that Tall varieties in the country are not in danger of genetic erosion, the collecting focused more on Dwarf cultivars found in the country. How Dwarf varieties were introduced into the country remains a mystery. Some people surmise that these varieties were brought in illegally from neighbouring countries, as perceived from the local names given to these varieties. Dwarf varieties are sought after because they flower early (most within three years), they are suitable for making toddy and are high-yielding which can supply tendernut for drinking and are easy to manage. Given these qualities, selected Dwarf varieties are being used in breeding programmes to produce Dwarf x Tall hybrids. It is envisioned that the genebank would continue to supply planting materials to the public and serve as a conservation area for the collected varieties. 735 CHAPTER 9: Country reports on status of coconut genetic resources research Capacity building A coconut expert visited the country to assist in training the national coconut programme coordinator and to characterize existing coconut germplasm collections and set up a coconut germplasm conservation site in the country. Two local research officers have also been sponsored by COGENT to undergo staff development training on the use of the STANTECH (Standardized research techniques in coconut breeding) Manual in 1996 and 1999. Significance of research outputs The result of the replanting and rehabilitation schemes resulted to an increase in the land area for coconut. Various coconut researchers in Kiribati reported that more than 80% of the land was occupied by coconut alone (Trewen 1986; Edwards 1989; Barr 1992). The establishment of a genebank for Dwarf varietal accessions is a very significant step for coconut breeding, germplasm conservation and copra production improvement programmes, which would lead to the enrichment of the resource for the improvement of the culture and lives of I-Kiribati. Suggested next steps The introduction of new Dwarf varieties in vitro that could survive atoll condition should be one of the priorities in developing the coconut industry in Kiribati. Technical assistance from institutions such as Secretariat of the Pacific Community (SPC), International Plant Genetic Resources Institute/International Coconut Genetic Resources Network (IPGRI/COGENT), and the Food and Agriculture Organization of the United Nations (FAO) should be requested. Traditional methods of planting coconuts should be further investigated and adapted to modern technologies to improve and increase the production of coconut and the products derived from it. References Barr, J. 1992. Coconut improvement in Kiribati. A guide to coconut re- search development in Kiribati, 1960-1992. Ministry of Natural Re- sources Development, Kiribati. Beenna, TI. 2001. Studies on composts and the effects of composted or- ganic matter applications on the productivity of a coralline soil. Uni- versity of the South Pacific Library, Suva, Fiji. Edwards, S. 1989. Annual technical report on coconut research in Kiribati 1988-1989. Ministry of Natural Resources Development. Kiribati. Purseglove, JW. 1988. Tropical crops monocotyledons. Longman 736 COCONUT GENETIC RESOURCES Singapore Publishers Pte Ltd., Singapore. Trewen, K. 1985. Crop research in Kiribati. Ministry of Natural Resources Development, Kiribati Trewen, K. 1986. Annual technical report on crop research in Kiribati. Ministry of Natural Resources Development, Kiribati. 737 CHAPTER 9: Country reports on status of coconut genetic resources research South Pacific Status of coconut genetic resources research in Papua New Guinea M Faure Coconut Breeder, Cocoa and Coconut Institute (CCI), Stewart Research Station, Madang Province, Papua New Guinea Introduction The PNG Cocoa and Coconut Research Institute (PNG-CCRI), established in 1986, is the research arm of the cocoa and coconut industries in the country. Priority research is currently focused on breeding, entomology, downstream processing, agronomy and farming system. In the 1970s, a number of exotic coconut populations were brought into PNG, initially as planting material. These include Malayan Red and Yellow Dwarf (MRD and MYD), Renell Island Tall (RIT), West African Tall (WAT) and the Solomon Islands Tall populations. The MRD and RIT populations were used to produce the MAREN hybrid which is known to yield higher copra than either of its parents in the Solomon Islands where it was initially developed (Foale 1987). The MAWA (PB121) hybrid was also introduced for planting because of high yield performance as observed in the Ivory Coast. Unfortunately, these two hybrids did not perform well in PNG. The MAWA hybrid was not accepted by the growers as it produced small nuts compared to the local populations. They believe that large but fewer nuts involve less labour while still giving similar copra yield as that from palm with smaller but more numerous nuts. The local people have always preferred bigger fruits for drinking and they also seem to prefer the solid endosperm of the local types, which were considered sweeter and tastier than the MAWA hybrid. When the MAREN hybrid was introduced to growers during the 1970s, it was found to be susceptible to three insect pests, namely: Scapanes australis (beetle), Oryctes rhinoceros (rhinoceros beetle) and Rhynchophorus bilineatus (black palm weevil). These three species are endemic only to the island provinces of PNG where the bulk of the copra is produced. The hybrid trial in the Gazelle Peninsula was devastated by these insect pests. The growers then reverted to picking seednuts from the best palms among the local open-pollinated Tall palms. Breeding work in the 1970s developed Dwarf x Tall hybrids with local Tall, with Karkar Tall (KKT) and Markham Valley Tall (MVT) as the pollen donors. These hybrids were then planted out in progeny trials throughout the country using the MAREN hybrid and RIT as controls. However, the results of the 738 COCONUT GENETIC RESOURCES progeny testing were inconclusive because the trials were terminated prematurely due to lack of funds. Germplasm collecting, conservation and utilization In 1986, when PNG-CCRI was officially established, the institute took over the research on cocoa and coconut from the Department of Agriculture and Livestock (DAL). The main aim of the coconut breeding programme is to develop better and improved varieties for distribution to farmers. The coconut breeding related activities at that time were focused on germplasm survey, identification of the best populations, germplasm collecting and utilization in a national breeding programme (Faure and Moxon 1998). From 1987 to 1992, the Australian Centre for International Agricultural Research (ACIAR) funded and supported a national ‘Coconut Improvement Project’ main focus of which was on germplasm survey, characterization and collecting (Ovasuru et al. 1993). By 1993, seednuts and pollen of over 50 different Tall and Dwarf populations from surveyed sites, PNG-CCRI’s collection in Rabaul and from DAL research stations have been collected based on fruit component analysis (FCA) data. The seednuts from these collections were raised and planted at the Stewart Research Station (SRS) in Madang. Pollen from various Tall populations was used in crosses with MRD, MYD and PNG Brown Dwarf (PBD). The progenies are currently under test, predominantly for general combining ability (GCA) for yield (kg copra/ha), with the aim of selecting the best hybrids for distribution to farmers and growers. In addition, the programme emphasizes further prospecting, collecting of new germplasm and production and testing of Dwarf x Tall and Tall x Tall hybrids. Development of the regional coconut genetic resources centre: The International Coconut Genebank for the South Pacific (ICG-SP) In November 1998, the International Plant Genetic Resources Institute (IPGRI) and the Government of PNG, through the Department of Agriculture and Livestock, signed a Memorandum of Agreement formalizing the establishment of the International Coconut Genebank for the South Pacific (ICG-SP) with PNG-CCRI as host. The Food and Agriculture Organization (FAO) of the United Nations also signed the agreement in its capacity as trustee as witness. The ICG-SP is located at the SRS, located south of the equator at 5o latitude and 146o longitude. The major roles of the genebank include the collecting, conservation, evaluation and utilization of selected and desirable coconut germplasm in the South Pacific region. In addition, the ICG-SP will eventually become 739 CHAPTER 9: Country reports on status of coconut genetic resources research a service centre to facilitate germplasm exchange and utilization amongst the COGENT network countries, especially among the Pacific Island Countries. Other potential sources of germplasm Research stations of the Department of Agriculture and Livestock The research stations owned by DAL could hold introduced germplasm which may have not been sampled by the current breeding programme. These need to be visited and their germplasm collected for conservation in the ICG-SP. If these accessions found on the research stations have been listed in the designated list for introduction, they would be collected immediately and hence, save costs in importation. Farmers’ varieties The use of farmers’ varieties in the national breeding programme has great potential but is not fully explored, except under an ACIAR-funded project on germplasm survey which was conducted from 1987 to 1992. Potential local varieties will be surveyed and either pooled or catalogued collections conserved on farm for future requirements. However, such system of conservation is unpredictable as the farmer could replace the accession with other varieties. One way to safeguard this would be to enable the farmers to conserve the accession by providing some sort of incentives like production of high-value products from the conserved genetic materials. The scattered coconut palms on the fringes of the mainland and outlying atolls of the country are in danger of being swept away by rising sea levels. These germplasm would be a priority for possible characterization, collecting and conservation. The inhabitants of some atolls depend on coconut for their entire livelihood, e.g., Motlok atolls in North Solomon Province, and they have special varieties cultivated for food and artefacts. The sweet husk variety is one of the varieties. Some innovative farmers or large landholders have or may have done their own selections of good performing varieties that need to be sampled for direct use (pollen) or for conservation and further evaluation and utilization. Such populations need to be considered in the national breeding programme. Synthetic varieties The development of synthetic varieties is being explored to utilize the promising Tall populations to generate Tall composite hybrids. Experience and models for synthetic varieties developed in the Philippines will be used in the PNG breeding programme. This programme will use the 740 COCONUT GENETIC RESOURCES outcome (results) of the germplasm currently being evaluated in the ICG- SP. Dwarf x Tall hybrids The 78 series of Dwarf x Tall hybrids produced during 1992-1993 are being field-tested. Additional four Dwarf and four Tall accessions have been used to develop new progenies for GCA trials. Research projects/activities conducted Capacity building Technical assistance Since 1994, specialists, supported by COGENT/IPGRI, have provided technical assistance missions to help CCI/CCRI enhance coconut research. These missions included assessing the country’s coconut R&D capability, identifying common problems and opportunities for network collaboration, identifying a suitable site for the ICG-SP, evaluating embryo culture laboratories, evaluating germplasm collecting and conservation strategies, assessing the pest risks for the ICG, and assisting in the establishment of ICG-SP. Training and human resources development From 1996 to 2002, five local coconut researchers were trained through COGENT/IPGRI on various topics such as coconut germplasm management, collecting and conservation methods, coconut data analysis, computer use, documentation, coconut embryo in vitro culture techniques, and the use of the microsatellite kit and dedicated statistical software. COGENT meetings/workshops In 1998, a meeting and a workshop were held in PNG, including the 7th COGENT Steering Committee Meeting which was held in Madang. The meeting further endorsed the country, through CCRI, as the host of the ICG-SP. Research projects A total of 12 coconut research and development projects have either been completed or are underway in the country, with CCRI as the lead implementing agency. 741 CHAPTER 9: Country reports on status of coconut genetic resources research Financial support and funding Donor funding support generated by COGENT/IPGRI for the coconut projects in PNG totals US$ 100 475, was mostly from the Asian Development Bank (ADB), and the Department for International Development (DFID). National government counterpart financing for these projects amounted to US$ 295 831, mostly in the form of logistics support by the implementing agency (CCRI). Results/Outputs and benefits Improved coconut embryo culture capability As a result of the COGENT/IPGRI-sponsored coconut embryo culture expert visit and training of CCRI staff, research capacity has been improved. The laboratory facilities have also been upgraded and the culture unit has been improved and now fully-functional. A research officer has also been trained to run the coconut embryo culture laboratory. Other COGENT-funded infrastructure development includes the building of the embryo culture room and procurement of appropriate equipment, acquisition of post-entry quarantine and acclimatization units, and provision of stand-by generator. Genebank personnel have also been trained on the approved embryo culture protocol and now adopted in practice. The first generations of plantlets using the approved protocol are now planted in polybags in the nursery. In addition, other infrastructure funding by COGENT in support of the ICG-SP has greatly benefited PNG. This includes mainly the recognition of country’s role in coconut research and development in the region, and the consequent greater attention from scientific community to invest in PNG coconut programme (e.g. the Centre de Coopération Internationale en Recherche Agronomique pour le Développement or CIRAD, and the Australian Centre for International Agricultural Research or ACIAR) through donor-assisted projects. ACIAR-funded project on coconut embryo quality studies The ACIAR-funded project on ‘Coconut Tissue Culture for Clonal Propagation and Safe Germplasm Exchange’ has been approved enabling PNG to participate in the project. Although the project was originally intended to start in July 2002, project activities, particularly experiments on embryo quality studies, only started in April 2003. The results to date are being reviewed and future directions are being discussed. Current germplasm maintained at the ICG-SP In 2003, the ICG-SP held 38 Tall and 11 Dwarf accessions, most of which are national germplasm collections provided by CCI as stipulated in the 742 COCONUT GENETIC RESOURCES MOA establishing the ICG-SP. Since then, more accessions have been collected and conserved in the regional genebank. In 2000, CIRAD, in collaboration with the Secretariat of the South Pacific (SPC) and COGENT, collected coconut germplasm from atolls of Cook Islands, Marshall Islands, Kiribati and Tuvalu. These were cultured at SPC in Fiji and transferred to ICG-SP for in vitro culture and eventual conservation. If funds are available, PNG-CCRI will organize a collecting team to visit other germplasm centres of the donor-member countries to collect their designated germplasm. Planned activities Site establishment A new area has been identified and being prepared as part of the PNG’s commitment to expand the coconut breeding programme. Major field preparation activities include brushing, felling, drainage and cover crop planting. Germplasm introduction and establishment A total of 200 accessions are scheduled to be planted and conserved in the ICG-SP in the next seven years (Table 1). The 7-year development plan and the budget had been submitted to AusAID for possible funding. Table 1. Schedule of germplasm planting in the ICG-SP for the next seven years Year 0 1 2 3 4 5 6 7 Total No. of germplasm 54 75 96 117 138 159 180 200 200 Budget (US$) 1272 1776 2280 2784 3288 3792 4296 4800 4800 Note: Zero year means germplasm currently conserved in the field genebank at ICG-SP Germplasm management This activity will include in vitro culture of imported germplasm, growth and management in culture room until potting and acclimatization stage. Also the poly bag nursery maintenance until the field planting stage. Field genebank management includes weed control, pest management, drainage and general field upkeep. Income generation from the sale of dry nuts and intercrops is part of the management activities to augment the ICG-Management Fund. The trust account is being organised for the management fund and for future donor funding. Other planned activities of the ICG-SP 1. Coconut leaf analysis to determine soil nutrient level at ICG 2. Germplasm survey in high risk areas, including the atolls 743 CHAPTER 9: Country reports on status of coconut genetic resources research 3. Survey and identification of farmers’ varieties 4. Evaluation of Dwarfs for tender nut drink and food production 5. Research and training for the ICG staff and those from the region 6. Conduct of income generation activities for the maintenance of the genebank References Faure, MG and JE Moxon. 1998. Coconut Breeding. Pp. 73-18. In: PA Batugal and V Ramanatha Rao (eds). Proceedings of a Workshop on the Standardization of Coconut Breeding Research Techniques, 20- 25 June 1994. COGENT, IPGRI-APO, Serdang, Selangor, Malaysia. Faure, MG. 2003. International Coconut Genebank Progress Report 2003. Foale, MA. 1987. Coconut germplasm in the Pacific islands. ACIAR Tech- nical Report Series No. 4. Australian Centre for International Agri- cultural Research, Canberra, Australia. Ovasuru, T, GY Tan and LA Bridgland. 1993. Coconut germplasm collection in Papua New Guinea. Pp. 33-41. In: MK Nair, HH Khan, P Gopalasundaran and EVV Bhaskara Rao (eds). Advances in coconut research and development. Oxford and IBH, New Delhi, India. 744 COCONUT GENETIC RESOURCES South Pacific Status of coconut genetic resources research in Samoa A Peters1 and K Jayashree2 1 Assistant Chief Executive Officer, Crop Division, Ministry of Agriculture, Forests, Fishery and Meteorology, Apia, Samoa 2 Scientific Assistant, International Coconut Genetic Resources Network (COGENT), International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction Coconut is the most predominant crop grown in Samoa. Its traditional value and multipurpose uses make it one of the most important crops in the everyday lives of Samoans as an important source of food and cash. In 1996, Samoa exported coconut food products such as coconut cream, copra, copra meal and coconuts, worth SAT 3.598M (US$ 1.3M). The Agricultural Census (1989) stated that 96% of farmers’ holdings grew coconuts, which bring to a total land area of 27 692 ha. In Samoa, one of the most important crop mixtures being identified is coconut intercropped with cocoa, the others being crops like banana and taro (Agricultural Census 1999). However, due to price fluctuations of coconuts all around the world, there is a need to upgrade and improve approaches in coconut farming and encourage adoption of new production and processing technologies to enhance farmers’ incomes. Project activities conducted and outputs Farmer participatory research on the multipurpose uses of the coco- nut and characterization of farmers’ varieties Under this International Coconut Genetic Resources Network (COGENT)- supported project, farmers’ coconut varieties were characterized, documented and conserved on farm. Farmer participatory survey was conducted in Siufaga Savaii where seven varieties were identified and conserved in farmers’ fields. These varieties include the Samoan Tall (SMOT), Samoan Tall Samatau (SMOT01), Niu Vai Tall (NVIT), Niu Afa Tall Samoa (NAFT), Niu Lea Dwarf Samoa (NLAD02), Samoa Yellow Dwarf (SYD) and Samoan Tall Siufaga Savaii (SMOT03). A database on these farmers’ varieties has also been compiled. 745 CHAPTER 9: Country reports on status of coconut genetic resources research Coconut food recipes A total of 10 coconut food and five cocktail (beverage) recipes have been compiled and submitted to COGENT for inclusion in the International Catalogue of Coconut Food Recipes. Coconut-based Farming System (CBFS) as strategies for enhancing farmers’ incomes and conserving coconut germplasm One experimental plot has been established at Nuu Crop Development Centre where intercrops such as banana, guava, taro and taamu are being grown under coconuts. The favourable initial results were used to formulate new research on coconut-based farming system (CBFS). Television promotional programmes on CBFS and production of post- ers on value-added products from coconuts In collaboration with the Televise Samoa Corporation under the ‘Atinae Samoa’ Programme (Samoan Development Programme), three special episodes on coconut were produced. The episodes, with 15 minutes run time each, are on the following topics: (1) coconut-based farming systems; (2) export potential of coconuts; and 3) the coconut breeding programme at Olomanu Hybrid Seed Garden. A poster was also prepared and printed to promote the potential value-added products made from coconuts. Copies of the poster were distributed to different government offices, schools, manufacturing agencies and during special events such as the Coconut Day and the World Food Day. Compilation of coconut literature A list of publications on coconuts has been compiled from several sources such as the Nelson Memorial Library in Apia, University of the South Pacific, Alafua Campus Library, the National University of Samoa Toomatagi and others, which could be used by researchers in finding relevant information on coconut. The compiled list includes the following publications: 1. Asian Development Bank. 1985. Western Samoa Agriculture Sec- tor Study, Volume II. Background and Sector Review. ADB, Ma- nila, Philippines. 2. Bourdeix, R. 2001. Coconut cultivars in Samoa. Draft mission re- port. Apia, Samoa. 3. Christophersen, E. 1935. Flowering Plants of Samoa. Bull. Bernice P. Bishop Mus. 154. 4. Cumber, RA. 1957. The Rhinoceros Beetle in Western Samoa. SPC Technical Paper, No. 107. Noumea, Fiji. 5. Curry, L. 1955. The physical geography of Western Samoa. NZ 746 COCONUT GENETIC RESOURCES Geographer, Vol. 11. 1:42. 6. Efu, S and SV Tuia. 2001. Fruit measurement and fruit compo- nent analysis of Samoan Tall Siufaga Savaii (SMOT03). Unpub- lished technical paper. 7. Efu, S and Pouono. 1996. Performance Trial of two hybrids and three selected Talls under various agro-ecological zones. Unpub- lished technical paper. 8. Fili, Manaia Lolo. 2000. Farmer participatory research on farm- ers’ coconut varieties, multipurpose uses and evaluation of inter- cropping technologies as strategies to promote germplasm con- servation through increased farmer productivity. Project report submitted to IFAD and COGENT. IPGRI-APO, Serdang, Selangor, Malaysia. 9. Fili, Manaia Lolo. 2000. Collecting and conservation of coconut germplasm from Vava’u Haapai and the Niua. In ADB Phase II Project Report (October 1999-June 2000) submitted to COGENT. IPGRI-APO, Serdang, Selangor, Malaysia. 10. Fox, JW and KB Cumberland. 1962. Western Samoa: Land, Life and Agriculture in tropical Polynesia (Whitcombe and Tombs, editors). Christchurch, New Zealand. 11. Geographer, Vol.11, No.1. Pp. 42 12. Hamilton, WM and LI Grande. 1937. Report on the soils and ag- riculture of Western Samoa. Department of Science and Indus- trial Research Report to the Samoan Administration on New Zealand Reparation Estates. 13. Hunter, David. 1998. Confirmation letter of statistical analysis results for the comparison of coconut oil content of the Nu’u hy- brid and Siumu’s local tall variety. Suva, Fiji. 14. Laufasi Ola. 1957. Field Control of Rhinoceros Beetle. Laufasi Ola. 2,5:4-5. 15. Mendoza and Efu. 1986. Early performance of Hybrid in Samoa. CORD, APCC, Indonesia. 16. Ministry of Agriculture, Forests, Fisheries and Meteorology and the South Pacific Commission. Biological control of the Rhinoc- eros Beetle. Suva, Fiji. 17. Peters, A, U Dkatulla, O Aukuso, S Meleisea and H Hammans. The Coconut Hispid Beetle - Brontispa lingissima. Crop Protection Leaflet No 2. Samoan German Crop Protection Project. Depart- ment of Agriculture, Forests, Fisheries and Meteorology. Nuu, Samoa. 18. Pouono, K and S Efu. 1996. Performance trials of two hybrid and three selected Talls under various agro-ecological zones. Project 747 CHAPTER 9: Country reports on status of coconut genetic resources research Progress Report (August 1994 - July 1996). Project technical re- port submitted to COGENT. IPGRI-APO, Serdang, Selangor, Malaysia. 19. Pouono, K and S Efu. 1998. Coconut germplasm collecting and conservation in Samoa. ADB Phase 2 Project Progress Report (November 1997 - September 1998). Project technical report sub- mitted to COGENT. IPGRI-APO, Serdang, Selangor, Malaysia. 20. Solomon Island Joint Coconut Research Scheme. 1974. Review of the coconut breeding program (1971-1974). 21. Tilialo, Oscar. 1998. Olomanu hybrid coconut seed garden. An- nual Report, 1997-1998. 22. Tuia, Saena Valerie. 2001. Fruit measurement and fruit compo- nent analysis of Samoan Tall Spicata Kogau (SMOT02). (Unpub- lished technical paper). Future research and development strategies The Ministry will continue to conduct surveys to identify and characterize new ecotypes. Nuts of the new ecotypes will be collected for multiplication and conservation in Olomanu Hybrid Seed Garden. In addition, two new CBFS plots will be established on farmers’ field. 748 COCONUT GENETIC RESOURCES South Pacific Status of coconut genetic resources research in Tonga P Taufatofua1 and K Jayashree2 1 Head of Research, Ministry of Agriculture and Forestry, Vainani Research Division, Nuku’alofa, Tonga 2Scientific Assistant, International Coconut Genetic Resources Network (COGENT), International Plant Genetic Resources Institute - Regional Office for Asia, the Pacific and Oceania (IPGRI-APO), Serdang, Selangor, Malaysia Introduction Coconut is an important crop which has supported the livelihoods of the Tongan people for hundred of years. Lately, Tongan coconut farmers have been suffering from declining productivity and unstable markets of copra and coconut oil, traditional products derived from the coconut. Tongan farming systems are basically multi-storied and agro-forestry based, with root crops and other crops as the common intercrops. Farmers in the country are willing to learn and adopt new and more efficient farm management strategies and approaches to improve their existing coconut-based farming system. Research activities conducted and outputs Collecting and conservation of coconut germplasm from Vava’u, Ha’apai and the Niua Islands Tonga has a coconut germplasm collection that was started six years ago. Part of the collecting activity was carried out during Phase I of the Asian Development Bank (ADB)-funded project entitled, ‘Collecting, conserving and characterizing coconut genetic resources in eight Asia Pacific countries’, which was coordinated by the International Plant Genetic Resources Institute’s International Coconut Genetic Resources Network (IPGRI/COGENT). The accessions collected came mainly from the main island of Tongatapu. However, there is a need to explore and collect the coconut genetic resources of the other islands, particularly those in the north such as Vava’u, Ha’apai and the Niua Islands. To date, four ecotypes, namely: Niu Kafa, Niu Vai, Niu ‘Utongau, Niu Matakula and Niu Talokave, have been collected and characterized from Vava’u, Utongau and Ha’apai. Data collected have been submitted to COGENT and entered into the International Coconut Genetic Resources Database (CGRD). These ecotypes have also been planted in 749 CHAPTER 9: Country reports on status of coconut genetic resources research the genebank in Vaini Research Station for conservation and further study. Promoting germplasm conservation of Tonga’s coconut diversity through increased farm productivity This COGENT-assisted project aims to: (1) identify and describe the multipurpose uses of coconut, and the local ecotypes grown by farmers for these uses; (2) develop strategies for product utilization and to add value to products from local varieties; (3) quantify marketable products from coconut and to evaluate and promote technologies for commercializing these; and (4) evaluate coconut-based farming system technologies to enhance germplasm conservation and farmers’ incomes. Farmer participatory survey to identify multipurpose uses and vari- eties suitable for these uses and to promote germplasm conserva- tion The main purpose of this project component was to characterize, document and conserve farmers’ coconut varieties. Farmer participatory survey was conducted in Tongatapu where eight varieties were identified and conserved in farmers’ fields. These varieties include: Niu Vai, Niu Kafa, Niu Matakula, Niu Leka, Niu Ta’okave, Niu Loholohotaha, Niu Mea and Tonga Talls. During the participatory rural appraisal workshop, it was also found out that one of the main problems of smallholder farmers was the limited income-generating opportunities from coconut primarily caused by irregular supply of raw materials and lack of markets (both domestic and overseas). The irregular supply of raw materials was caused by low crop productivity due to ageing palms, animal damage, seasonal productivity, saltwater intrusion (sea spray) and water logging. Another major contributor to low productivity was the close and dense planting of coconut pamls resulting in competition for nutrients and moisture, as well as difficulty in maintenance. Evaluation of coconut-based intercropping technologies Farmers preferred the Tonga Tall variety compared to others because of its capacity to accommodate intercrops due to its greater spacing requirement. Based on COGENT’s recommended strategy, farmers planted squash, kava, watermelon, vanilla and root crops as intercrops with coconut primarily for their export market potential. The project found out that intercropping can be promoted to more farmers to further enhance food security and expand export markets. 750 COCONUT GENETIC RESOURCES South Pacific Status of coconut genetic resources research in Vanuatu JP Labouisse1 and J Lahva2 1Coconut Breeder, Centre de Coopération Internationale en Recherche Agronomique pour le Développement - Vanuatu Agricultural Research and Technical Centre (CIRAD- VARTC), Santo, Vanuatu 2Agronomist, Department of Agriculture and Rural Development, Port-Villa, Vanuatu Introduction Vanuatu, formerly called New Hebrides, is an archipelago located in the Southwest Pacific Ocean between the Solomon and Fiji Islands. It consists of some 80 widely dispersed islands between the Torres Group (13°S) to the uninhabited Matthew and Hunter islets (22°S). As in most of the Pacific Island countries, coconut is widely planted and used by the rural populations for food and for numerous other domestic purposes. The production of copra started in the 1870s and was the mainstay of Vanuatu’s economy until the 20th century. Even when world demand and prices for the product declined, copra remained as the most important export commodity of the country, with around 30 000 metric tonnes exported annually. Coconut is grown in an estimated 90 000 ha, representing nearly 60% of the total cultivated area in the country. On the southeast coast of Espiritu Santo Island, near the village of Saraoutou, a coconut research station was established in 1962. Up to 2001, the station was managed by the French research organization Institut de Recherches pour les Huiles et Oléagineux (IRHO), which became the Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD) in 1985. The Saraoutou Research Centre is now called the Vanuatu Agricultural Research and Technical Centre (VARTC). Researchable problems and opportunities Tolerance to coconut foliar decay When the Saraoutou Station was created, the main objective of its research work was to increase coconut productivity through agronomic and genetic improvement, particularly by developing high-yielding and suitable hybrids to replace the ageing established local varieties. A number of exotic varieties were planted in a field genebank, but they quickly started 751 CHAPTER 9: Country reports on status of coconut genetic resources research to succumb to a previously unseen wilt of unknown aetiology, while the local Vanuatu Tall (VTT) variety, remained unaffected. This new wilt was named ‘coconut foliar decay’ or CFDV, a viral disease transmitted by the insect vector Myndus taffini (Julia 1982). Following this discovery, tolerance to CFDV was decided as the main criterion for selecting and developing coconut planting materials for Vanuatu. Enrichment of VARTC field genebank and conservation of local co- conut genetic resources As the main source of CFDV tolerance, the use of the local Vanuatu Tall was the central strategy for the coconut breeding programme. Before the International Plant Genetic Resources Institute’s International Coconut Genetic Resources Network (COGENT)-sponsored projects started, the genetic diversity of local coconut genetic resources in Vanuatu was not properly assessed and conserved. There was also a need to conduct a survey to identify the different uses of coconut, other than copra. At that time, the in situ management of coconut genetic resources by farmers was still unexplored. Improvement of coconut-based farming system With the recent drop in world copra prices, it is urgent to find ways to improve smallholders’ incomes. The coconut groves are senescent and occupy large areas. The planting of precocious, high-yielding cultivars created through research can optimize the landuse. However, the dissemination of the improved planting material is expensive due to the distant locations of the cultivated areas, and the difficulties and high cost of transport around the archipelago. Diverse associations of coconut with other crops have been observed during farmers’ participatory surveys. The performance, the sustainability and market opportunities of such associations must therefore be assessed. Diversification of coconut uses Even if a wide range of coconut by-products and uses have been observed at rural household level, very few are marketed in the urban areas or exported. The industrial processing for grated coconut, canned coconut milk and other similar products is not profitable in Vanuatu due to the limited domestic market, expensive inputs, high transport and labour costs. Nevertheless, the marketing of fresh products (tender and mature coconuts) and small-scale processed products (e.g. virgin oil) could be developed. The use of copra oil as biofuel for vehicles shows promise. Projects for the electrification of remote areas by using copra oil powered- generators are also being explored. 752 COCONUT GENETIC RESOURCES Collecting, evaluation and characterization of coconut genetic re- sources in Vanuatu The main purpose of this ADB-funded project was the evaluation and the ex situ conservation of the genetic diversity of local coconut genetic resources. Fourteen sites located in 10 different islands of the archipelago were surveyed. Two hundred nuts each of the 12 populations were collected. Eighteen variants showing special characteristics (spicata form, unique nut colour, etc) were also collected but with smaller sample size. They were all established in VARTC field genebank (Labouisse and Sileye 2001). As of 2003, the local germplasm collection of VARTC consists of 20 populations of Vanuatu Tall and the Vanuatu Red Dwarf (VRD). Table 1. Comparative performance of three cultivars in Vanuatu for yield and copra processing Characteristics VTT VTT x RIT VRD x VTT Average annual yield under farmers’ field conditions as evaluated from 1994 to 1997 (tonnes) 2.0 2.6 2.2 Average copra content per nut as measured from 1994 to 2000 (in grams) 199 258 134 Percentage of oil in albumen dry matter 66.2 66.0 65.4 Percentage of water in fresh albumen 47.4 50.8 55.8 Number of nuts needed for one tonne of copra 5555 4360 7600 Time to process these nuts (comparison with VTT x RIT) 26.5 hours (+ 11%) 24 hours 37 hours (+ 55 %) Quantity of copra obtained from one tonne of fresh kernel by hot air drying process (in kg) 467 447 415 Legend: VTT = Vanuatu Tall RIT = Renelle Tall VRD = Vanuatu Red Dwarf Research and training activities conducted in the country During the last 10 years (1994-2003), the Department of Agriculture and Rural Development (DARD) and VARTC have actively participated in the different projects and training activities coordinated by COGENT, which are as follows: Evaluation of selected coconut cultivars planted in farmers’ fields in Vanuatu The agronomic performance of three improved coconut cultivars, distributed to farmers during the implementation of the Coconut Development Project (CDP) between 1982 and 1993, were evaluated through COGENT’s Asian Development Bank (ADB)-funded project. For this purpose, observations were conducted on farmers’ plots and results are presented in Table 1. 753 CHAPTER 9: Country reports on status of coconut genetic resources research Enhancing farmer incomes and germplasm conservation through coconut- based farming system and identification of varieties for multipurpose uses This International Fund and Development (IFAD)-funded project consisted of three components: Component 1: Farmer participatory surveys During the period July 1998 - December 2000, eight participatory rural appraisal surveys (PRAS) were conducted on seven different islands gathering substantial information about local names and uses of coconuts, and different coconut-based farming systems. An average of 11 distinct types (or variants) of coconuts per village were identified by the farmers with numerous by-products and uses documented (Table 2). Some variants are associated to specific uses (Lahva and Labouisse 2000). This component is closely linked with the project on characterization and conservation of local cultivars. Samples of leaves of some collected populations were analyzed with the microsatellite kit developed by CIRAD (Baudouin and Lebrun 2002) to assess the genetic diversity within and between the populations. The results of the PRAs were presented and discussed in a journal article by Labouisse and Caillon (2001). A set of three posters in bislama (Vanuatu’s official language) was prepared and posted in the rural communities in order to make them aware of coconut genetic resources conservation strategies. Component 2: Feasibility of coconut based intercropping systems for promoting coconut germplasm conservation through use During the PRAs, the associations between coconuts and others crops have been documented. The socioeconomic survey on marketable crops produced in association with coconuts shows the opportunities and constraints of the different crop varieties and present some data on costs and returns (Bule 2000). The survey indicated that Xanthosoma sagittifolium and Musa sp. are the most frequently cultivated crops under coconuts, while Kava (Piper methysticum) is shown to be the most profitable. It was also identified that high transportation costs from farms to markets is the main productivity constraint. Component 3: Evaluation of the improved cultivars used by farmers in Vanuatu for processing Under the ADB-funded Project, the agronomic performance of three cultivars, (improved VTT, VRD x VTT hybrid, VTT hybrid x RIT hybrid, were assessed in farmers’ fields (Labouisse and Buletare 1997). Under 754 COCONUT GENETIC RESOURCES Table 2. List of the common uses of and products from the coconut as surveyed in the villages of Vanuatu (Source: Lahva and Labouisse 2000) Coconut part Uses and products Whole palm • Land marking • Garden ornamentation • Cattle shade Roots • Medical uses Trunk • Building material and furniture (post, plank, part of canoe) • Support for plants (yams, vanilla) • Medical uses (bark) Leaves • Handicrafts (hat, mat, fan, broom, baskets, hoop net) • Building material (roof, walls) • Support for plants • Fuel and light (torch) • Filter for kava Whole fruit • Ceremonial uses (wedding present, customary exchanges) Husk • Rope for building and for canoe • Container, support and protection for plants • Fuel • Abrasive Shell • Handicrafts (container, cup, spoon) • Kava cup • Fuel Water • Beverage • Medical and ’magic‘ uses (excipient) Albumen • Food • Copra Milk • Food • Medical uses (excipient) Oil • Food • Oil for human body and hair conditioning • Fuel (for lamp) the IFAD-funded project, the quality of the fruits of these three varieties was also evaluated for copra production and processing (Lahva and Labouisse 2000). Results showed that the hybrid VTT x RIT outperformed the two other varieties in terms of nut yield and copra production (Table 1). Contribution to the Coconut Genetic Resources Database (CGRD) The entries of Vanuatu in the International Coconut Genetic Resources Database (CGRD) comprise of 60 living accessions. In addition, 755 CHAPTER 9: Country reports on status of coconut genetic resources research Other activities carried out within the framework of COGENT 1995 - Participation in the finalization of the STANTECH Manual (Santos et al. 1996) in Manado, North Sulawesi, Indonesia. 1996 - Pacific Projects review of Coconut Genetic Resources Network and Asia Pacific (CGRNAP) at VARTC. 1996 - Participation in the COGENT Steering Committee in Merida, Mexico as a representative of the Pacific region. COGENT training courses organized in Vanuatu No. of trainees/ participating countries Regional STANTECH course for South Pacific (VARTC, 6- 13/8/1996) 9 / PNG, Solomon Islands, Tonga, Fiji, Kiribati, Vanuatu STANTECH training course on collecting and management of coconut genetic resources (VARTC, 29/6-10/7/1999) 4 / Kiribati, Cooks Islands, Marshall Islands, Tuvalu COGENT training courses organized outside Vanuatu Participating Vanuatu researchers Coconut collecting and conservation course (PCA, Philippines, 1-12/9/1997) Godefroy Buletare Farmers participatory research on coconut diversity (Taveuni Coconut Centre, Fiji, 24-28/3/1998) Pierre-Chanel Watas, Jeffrey Lahva, Jean-Pierre Labouisse Computer use, documentation and data analysis course for South Pacific (SPC, Suva, Fiji, 3-7/8/1998) Godefroy Buletare Technical writing, seminar presentation and proposal writing course (PCA, Philippines, 30/8-3/9/1999) Jeffrey Lahva Coconut data analysis training course (PCA, Philippines, 6- 10/9/1999) Jeffrey Lahva Establishment and management of field genebanks for conservation and use (PSGT, Malaysia, 28/9 – 10/10/1999) Tiata Sileye Table 3. List of COGENT-sponsored training courses with participating Vanuatu researchers morphometric and passport data of 12 accessions which do not exist anymore (due to cyclones or felling) have also been recorded and included in the CGRD. Training activities sponsored by COGENT Two regional training courses were held in Vanuatu while five Vanuatu researchers were sponsored by COGENT to undergo staff development training on topics such as standardized research technique in coconut breeding (STANTECH), coconut germplasm collecting and conservation, farmer participatory research, computer use, documentation and data analysis, and technical writing and seminar presentation (Table 3). 756 COCONUT GENETIC RESOURCES 1999 - Participation in the COGENT consultancy on coconut collecting strategy (Bourdeix et al. 1999). 2000/2001- Appointment of VARTC as the implementing agency for the ADB-CGRNAP project on ‘Coconut germplasm col- lecting, characterization and conservation in Cook Is- lands, Kiribati, Marshall Islands and Tuvalu’ (Labouisse and Bourdeix 2003). 2002 - Participation in the preparation of the International Cata- logue of Conserved Coconut Germplasm and Farmers’ Varieties. Activities supported by other donors Conservation and observation of exotic germplasm Exotic varieties which are susceptible to CFDV can be conserved in VARTC field genebank by removing, within a radius of about a hundred meters, all stumps of Hibiscus tiliaceus, the breeding host of the CFDV vector. Through the Pacific Regional Agricultural Programme (PRAP) project, the exotic germplasm of VARTC have been fully rejuvenated by hand pollination between 1992 and 2000. To date, the collection comprises of 14 distinct Tall and 13 Dwarf varieties imported from different countries of Africa, America, Asia and Pacific. Growth characteristics, yield, fruit component analysis are regularly recorded. Observations were also done in 1999 on the tolerance of Dwarfs to strong winds (Figure 1). 0 20 40 60 80 100 TACD VRD SYD CRD NLAD PILD CATD THD MBD BGD MRD AROD MYD Varieties Pe rc en ta ge o f f el t t re es Figure 1. Damage inflicted by cyclone Dani (January 1999) on the different Dwarf varieties (aged 15) in the VARTC genebank. 757 CHAPTER 9: Country reports on status of coconut genetic resources research Improvement of coconut planting materials for Vanuatu Three cultivars were selected for propagation in Vanuatu for their tolerance to CFDV. These include the improved Vanuatu Tall, the VRDxVTT and the VTTxRIT hybrids. The improved Vanuatu Tall, obtained from several cycles of mass selection, has an average copra yield of 2.2 t/ha/year and a copra content of 195 g/nut. It is completely tolerant to CFDV and can be easily multiplied by farmers. The VRD x VTT hybrid line, tolerant to CFDV, is produced in seedgardens at Saraoutou Station and was released to farmers between 1986 and 1996. In spite of its high-yielding potential of 3.4 t/ha/year (as recorded in station trials), its dissemination was discontinued due to its low germination rate in the nursery, a low copra content (154 g) and the frequent dropping of immature bunches. The VTT x RIT hybrid is the most promising with an average yield of 2.6 t/ha/year and a high copra content of 237 g/nut. The first lines of this hybrid showed very slight susceptibility to CFDV. However, the tolerance has been improved by using, as female parent, several self- pollinated progenies of RIT which show no symptoms of the disease. The VTT and the VTT x RIT hybrid, with good nursery and cultivation practices coupled with an ideal climate, start to bear flowers 30 months after planting, which is remarkable for Tall cultivars. PRAP - PDICC Project From 1989 to 1999, with the financial support of the European Union (EU) and the French Ministry of Foreign Affairs, and the technical assistance of CIRAD, VARTC implemented the Production and Dissemination of Improved Coconut Cultivars (PDICC) project in the framework of the PRAP. Eight countries (Fiji, Kiribati, PNG, Tonga, Samoa, Solomon Islands, Tuvalu and Vanuatu) were associated with this regional programme. The objectives of the PDICC project were: 1. To improve the potential of coconut production by increasing the choice of hybrid coconut cultivars in the South Pacific Region. A wide range of hybrids was created and the performance assessed in experimental trials established at VARTC. Before 1999, the project also supported the maintenance and the data collecting of VARTC coconut germplasm; and 2. To improve the quality of planting materials disseminated to farm- ers from the seedgardens of the participating countries. Techni- cal assistance and training were provided to these countries for seedgarden management, coconut breeding and coconut genetic resources management. 758 COCONUT GENETIC RESOURCES Under the project, 39 new hybrid crossings were made by hand- pollination and eight trials were successfully established in Saraoutou Station between 1992 and 1999. This represents a total area of 57 hectares with approximately 9000 palms under individual observation. Each of the first seven trials included hybrids created by crossing diverse Dwarf cultivars with a Tall cultivar native to the region (i.e., Rennell Tall (RIT), Tonga Tall (TONT), Rotuman Tall (RTMT), Kiribati Tall (KIT), Gazelle Peninsula Tall (GPT), Samoan Tall (SMOT) and Markham Valley Tall (MVT)). In the eighth trial, six different Tall cultivars were crossed with RIT. For each trial, the following data were gathered: rate of germination, growth in the nursery, growth in the field (young age), flowering, yield, fruit component analysis, oil content, stem measurements, resistance to cyclone and susceptibility to diseases. For copra yield, the hybrid MRD x RIT showed good performance in all the trials, producing about five tonnes of copra/ha when six years old. The hybrids MYD x RIT, BGD x RIT, MRD x TONT and MRD x RTMT also showed good potential. Some of the varieties also showed some promise on the tolerance to cyclones (Labouisse 2002). Study of in situ management of coconut genetic resources With the support of CIRAD and the Institut de Recherche pour le Développement (IRD), a three-year study in the framework of a PhD thesis was conducted since 2001 on in situ management of coconut genetic resources by farmers in Vanuatu (Caillon 2001). The study aims to further understand the biological and sociological processes that build the diversity of a crop system. The study was performed in Vanua Lava in the north of Vanuatu. The farmers themselves distinguished the existing variants in Vanuatu Tall variety according to some specific morphological traits, production characteristics or particular origin. The average number of variants identified by village in Vanua Lava is 30, far above the number found during the IFAD-funded project, which was attributed to a longer and more intensive survey (Caillon and Malau 2002). Variants are being described according to the STANTECH Manual. Statistical analyses done on 105 individuals showed that the most discriminant characters are those related to the description of fruit components. The results of molecular analysis using 14 microsatellites performed on 69 coconut leaves collected in Vanua Lava from 12 variants revealed that the whole population is distinct from the rest of Vanuatu and other Pacific countries. However, this technique is inadequate to differentiate the variants except for one (a Tall with yellow fruits). As the sampled population is small (two to eight individuals per variant), additional analyses have do be done to confirm these initial results. 759 CHAPTER 9: Country reports on status of coconut genetic resources research Study on coconut-based farming systems Since 2002, a study on the methods of assessment of performances and sustainability of associations of staple crops with old coconuts has been undertaken on the island of Malo in the framework of a PhD thesis sponsored by CIRAD (Lamanda et al. 2003). Interpretation of significance or impact of output The use of participatory approach for the assessment of coconut diversity in Vanuatu was very fruitful. Several coconut variants were identified and traditional uses discovered. VARTC’s collection has been enriched with populations of Vanuatu Tall collected in different environments. The improved VTT represents a significant advancement compared to ordinary VTT, with a better yield but, above all, a higher copra content which reduces the labour needs for copra processing. The hybrid VTT x RIT is also promising but its utilization by farmers is impeded by the high cost of transportation of nuts or seedlings within the archipelago. Contrary to improved VTT, it cannot be reproduced by farmers. The results of the PRAP trials constitute a database of great value for the research and extension services within the region. These results will also benefit other Pacific countries which would be advised to reproduce the best crossings by using their own germplasm and seedgardens. Unfortunately, these hybrids would not be disseminated to Vanuatu farmers because of their susceptibility to CFDV. Future research and development activities in coconut Maintenance and observations of VARTC germplasm The recently collected populations of Vanuatu Tall will be observed and the most interesting and promising ones could be used for further genetic breeding programme. Besides potential copra yield, the characteristics of the fruits for processing will be assessed. Maintenance and dissemination of the results of PRAP-PDICC hy- brids trials Due to the biological cycle of the coconut, the performance assessment of a hybrid could only be undertaken about 9-10 years after field planting. Therefore, the evaluation of the first seven trials established at VARTC would only be done by 2006 although the financial support of the EU ended in 1999. Due to the regional significance of these trials, VARTC needs to source external funds for the maintenance and full evaluation of these trials until 2006. 760 COCONUT GENETIC RESOURCES Improvement of coconut-based farming systems With the objective of improving the effectiveness and profitability of coconut-based farming systems (CBFS), the evolution of the different CBFS in Malo Islands is being studied. Likewise, the assessment of performance of the association of ageing coconut with other crops requires further research on agronomy, plant protection and physiology of coconuts and other crops (fruit trees, rootcrops, kava Piper methysticum, legume trees, etc) and on farm economy. The results of these studies will enrich CBFS technologies and, hopefully, provide increased benefits to resource-poor coconut farmers and their households. References Baudouin, L and P Lebrun. 2002. The development of a microsatellite kit for use with coconuts. Centre de Cooperation Internationale en Re- cherche Agronomique pour le Développement, Montpellier, France. Bourdeix, R, L Baudouin, J Ollivier and JP Labouisse. 1999. COGENT consultancy report on coconut collecting strategy. Centre de coopération Internationale en Recherche Agronomique pour le Développement (CIRAD), Montpellier, France. Bule, F. 2000. Socioeconomic survey on marketable crops and livestock under coconuts. In: J Lavah and JP Labouisse (eds). Enhancing farm- ers’ incomes and germplasm conservation through coconut based farming system and identification of varieties for multipurpose use. Final project report. DARD, Port-Vila, Vanuatu. Caillon, S. 2001. Gestion traditionnelle et conservation in situ: cas du cocotier Cocos nucifera et du taro Colocasia esculenta au nord du Vanuatu. Projet de thèse. Université d’Orléans, France. Caillon, S and EF Malau. 2002. Catalogue of coconuts and taros of the West Coast of Vanua Lava. Julia, JF. 1982. Myndus taffini (Homoptera Ciixidae), vecteur du dépérissement foliaire des cocotiers au Vanuatu. Oléagineux 37 8- 9:409-414. Labouisse, JP. 2002. Hybrids trials of the Pacific Regional Agriculture Programme. Progress report, January 2002. Vanuatu Agricultural Research and Training Centre, Santo Vanuatu. Labouisse, JP and R Bourdeix. 2003. Coconut germplasm collecting, char- acterization and conservation in Cook Islands, Kiribati, Marshall Is- lands and Tuvalu. Final report. March 2003. Vanuatu Agriculture Research and Training Center, Santo, Vanuatu. Labouisse, JP and G Buletare. 1997. Evaluation of selected coconut culti- vars planted in farmers’ fields in Vanuatu: COGENT - CGRNAP 761 CHAPTER 9: Country reports on status of coconut genetic resources research Project. Terminal report. Vanuatu Agriculture Research and Train- ing Center, Santo, Vanuatu. Labouisse, JP and S Caillon. 2001. Une approche de la conservation in situ par l’étude d’un systeme semencier informel: cas du cocotier au Vanuatu (Pacifique Sud). An approach to in situ conservation through the study of an informal system of seed supply: the example of coco- nut in Vanuatu (South Pacific). OCL - Oléagineux, Corps Gras, Lipides 8 (5):534-539. Labouisse, JP and T Sileye. 2001. Collecting, evaluation and character- ization of coconut genetic resources in Vanuatu. Final report. Vanuatu Agricultural Research and Training Centre, Santo, Vanuatu. Lahva, J and JP Labouisse. 2000. Enhancing farmers’ income and germplasm conservation through coconut-based farming system and identification of varieties for multipurpose uses in Vanuatu. Final technical report. Department of Agriculture and Rural Development, Port Vila, Vanuatu. Lamanda, N, E Malezieux and P Martin. 2003. Cocoteraies et jardins vivriers dans les îles mélanésiennes : diversité spatiale et dynamique temporelle des systèmes de culture. Le cas de Malo (Vanuatu). Colloque : Organisation spatiale et gestion des ressources et des territoires ruraux. 25-27 février 2003. CIRAD, Montpellier, France. Santos, GA, PA Batugal, A Othman, L Baudouin and JP Labouisse. 1996. Manual on standardized research techniques in coconut breeding. IPGRI-APO, Singapore. 46p. 762 COCONUT GENETIC RESOURCES Index A Accessions conserved in the Southeast Asian Region 533 AFLP 229. See also Amplified Fragment Length Polymorphism Africa and Indian Ocean, Research on coconut genetic resources in 546 Africa and the Indian Ocean 546 AIO. See Africa and the Indian Ocean Amplified Fragment Length Polymorphism 229 Area freedom 423 Arthropod pests 415 Assessing the completeness of collecting 59. See also Farmer participa- tory approach Assessing the reasons for, extent and danger of genetic erosion 59. See also Farmer participatory approach B Bangladesh, Status of coconut genetic resources research in 596 Biochemical and molecular methods for characterizing coconut diver- sity 225-243 Different genetic markers 225 Biochemical markers 226 Molecular markers 227 Choice of markers to be used 232 Results 236 Biofuels Project, SPC 521 Brazil, Status of coconut genetic resources research in 695 Bursaphelenchus. See Nematode disease C Cadang-cadang 377 Candidate-gene approach 271. See also Molecular markers for coconut improvement Catalogue of coconut food recipes 466 Catalogue of conserved germplasm and farmers’ varieties 456 763 Index Catalogue of high-value coconut products 463 CCCVd. See Cadang-cadang Cedros wilt,. See Phytomonas CFD/CFDV. See Coconut foliar decay CGIAR. See Consultative Group on International Agricultural Re- search CGIAR’s support to coconut research 473 Historical perspective 473 Present status of international coconut research efforts 477 Future perspectives 477 CGRD. See International coconut genetic resources database CGRD, Conceptual model of the 429 CGRD, Stages in the development of 428 Characterization of viroid RNA 379 Characterizing diversity 207 China, Status of coconut genetic resources research in 648 Choice of markers to be used 232-237. See also Biochemical and mo- lecular methods for characterizing coconut diversity The coconut microsatellite kit 232 Geneclass2 software 234. See also Geneclass2 software perfor- mance Use for classification purposes 236 Chronology of international coconut research (1900-2000) 480 Clustering methods and dendrograms 216. See also Morphometric methods of determining diversity in coconuts Coconut breeding 443, 450 Coconut collecting sites and gaps identification 24 Coconut collections, general statistics on 45. See also Status of coconut germplasm collecting Coconut conservation, Integrated approach to 108 Coconut diseases and pests 397 Coconut diversity for nut weight in the Philippines 25 Coconut embryo in vitro culture 445 Coconut embryo in vitro culture: Part II 446 Coconut evolution, key features of 16 Coconut field genebank 91 Coconut foliar decay 375 Coconut genetic diversity: General considerations 15 Coconut genetic resources book 450 Coconut germplasm collecting, Status, gaps and strategy 44 Coconut hybrids performance and use, Conditions favouring 305-306. See also Performance of coconut hybrids in Asia, Africa and Latin America 764 COCONUT GENETIC RESOURCES Agroclimatic 305 Farmers’ preferences 306 Coconut hybrids for smallholders 449 Coconut in Africa, History of 546 Coconut is good for your health, CD-ROM 449 Coconut lethal yellowing 349 Geographic distribution 349 Symptoms 349 Causal agent 350 Transmission 351 Diagnosis and detection 353 Spread 354 Control methods 355 Coconut lethal yellowing in Jamaica 364 History of LY in Jamaica 364 Searching for the cause 365 Living with LY before 1961 366 Living with LY after 1961 366 Current status of LY and future prospects 368 Coconut micropropagation 334-342 Research from the 1970’s to the 1990’s 334 Recent advances 338 Perspectives and conclusion 341 Coconut microsatellite kit 232 Coconut productivity in farmers’ field and research 307 Coconut Research for Development Programme 500 Goal and objectives 502 Priority research areas 503 Organizational structure 504 Programme coordination 505 Cocos 13 COGENT. See International Coconut Genetic Resources Network: Its history and achievements COGENT member countries 106, 190, 483 COGENT Newsletters 450 COGENT publications and other PA materials 443 COGENT publications and other public awareness materials 443 COGENT, Objectives of 107 COGENT’s public awareness strategy 440 Complementary conservation of coconuts 75 Complementary conservation strategy of coconut germplasm 83 Complementary conservation strategy, Considerations for 79-83 Costs and risks 82 765 Index Genetic diversity 79 Infrastructure 80 Network 81 Policy/Legal issues 82 Socioeconomic aspects 81 Stakeholders 80 Complex hybrids 258. See also Conventional breeding schemes Conservation of coconut germplasm in field genebank, Considerations for 92 Agronomic considerations 96 Genetic considerations 92 Germplasm health issues 100 Legal issues 99 Policy and management issues 98 Conservation in the multi-site ICG 109 Conservation methods for coconuts, Advantages and 86 Conservation objective 79 Conservation options for coconuts, Comparison of 85 Conservation strategy of COGENT 108 Conserving coconut germplasm, Methods for 76 Consultative Group on International Agricultural Research 473 Conventional breeding schemes 254-259. See also Conventional coco- nut breeding Mass selection methods 254 Other intra-population breeding methods 255 Conventional coconut breeding 251-264 Population base 252 Conventional breeding schemes 254 Status of coconut improvement 259 Breeding limitations and opportunities 261 Farmers’ varietal preferences 263 Future breeding plans 264 Cook Islands, Status of coconut genetic resources research in the 720 Coronie wilt. See Phytomonas Côte d’Ivoire, Status of coconut genetic resources research in 654 Country reports on status of coconut genetic resources research 571 Cryopreservation, Definition of 143 Cryopreservation research in coconut, Status of 142 Immature embryos 144 Mature embryos 144 Plumules 145 Prospects 146 Cultivar 9 766 COCONUT GENETIC RESOURCES D Deciding what to collect and how 59. See also Farmer participatory approach Definition of terms and nomenclatures 9 Diagnostic tests, Pest risk assessment 423 Dip 423 Diseases of unknown origin 399, 402 Distribution 6 DIVA-GIS 23. See also Geographic Information System (GIS) DIVA-GIS 35 Diversity 2 Diversity indexes 217. See also Morphometric methods of determining diversity in coconuts Documenting and using the collection 59. See also Farmer participa- tory approach Drought stress. See Drought tolerance in coconut Drought tolerance in coconut 282-295 Scientific and theoretical basis 282 How serious is the problem? 282 Environmental factors 283 Impact of drought stress 284 Role of K+ and Cl- nutrition 288 Screening for drought tolerance 288 Drought tolerance mechanism 290 Genetics of drought tolerance in coconut 291 Methodology for screening 291 Constraints and opportunities 295 Dynamis borassi. See Nematode disease E Ecotype 9 ECW. See Epicuticular wax Electron microscopy 379 Embryo explants, Use of 336. See Coconut micropropagation Epicuticular wax 286 Ethnobotany 5 Euclidean distance. See Mahalanobis distance Evaluation descriptors, CGRD 433 Evolution of the coconut palm 1 767 Index F Farmer participatory approach 58 Farmer participatory approach in coconut germplasm 58 Farmer participatory approach, Importance in collecting 59 Farmer participatory research on coconut diversity 446 Fatal wilt. See Phytomonas FGB, Steps in establishing, maintaining and managing 101 FGB. See Field genebanks Field genebanks 91 Fiji, Status of coconut genetic resources research in 725 Framework for complementary conservation strategy 83 Framework for developing a complementary conservation strategy 87 Fumigation: 423 Fungal diseases 416 Fungal diseases of coconut 398, 401 G Gaps in coconut germplasm collecting 46 Gel electrophoresis 378 Geneclass2 software 234. See also Geneclass2 software performance Geneclass2 software performance 234 General consideration, Coconut genetic diversity 15 Genetic diversity 13 Genetic engineering 339. See Coconut micropropagation Genetic erosion. See Gaps in coconut germplasm collecting Genetically-modified organisms 272. See also Molecular breeding in coconut Geographic Information System 23 Geographical gap-filling. See Gaps in coconut germplasm collecting Georeferencing coconut accessions 32 Georeferencing of coconut accessions in CGRD 33 Germplasm conserved in the multi-site ICG 113 Germplasm exchange, Risks linked to 403-404 From African/Indian Ocean countries to Côte d’Ivoire 403 From Brazil to the other countries of Latin America 404 From Côte d’Ivoire to African/Indian Ocean countries 403 From Latin American/Caribbean countries to Brazil 404 Germplasm health 392 Germplasm health management for COGENT’s multi-site International Coconut Genebank 448 768 COCONUT GENETIC RESOURCES Germplasm use 249 Ghana, Status of coconut genetic resources research in 66 GIS. See Geographic Information System GIS, main elements of a 23 Global coconut conservation strategy 190-205 Integrated approach to coconut conservation 191 Scope and status of coconut conservation 191 Priority activities 202 GMOs. See Genetically-modified organisms Guyana, Status of coconut genetic resources research in 709 H Hartrot. See Phytomonas Helminthosporium halodes (Dresch.) 399 Hemileia vastatrix 390 Hurricane tolerance in coconut 561 I ICG, Plan of action 112 ICG. See International Coconut Genebank ICG-AIO, Designated germplasm for 140 ICG-AIO. See International Coconut Genebank for Africa and Indian Ocean ICG-LAC. See International Coconut Genebank for Latin America and Caribbean ICG-SA, Designated germplasm for 137 ICG-SA. See International Coconut Genebank for South Asia ICG-SEEA, Designated germplasm for 138 ICG-SEEA. See International Coconut Genebank for Southeast and East Asia ICG-SP. See International Coconut Genebank for the South Pacific ICG-SP. See International Coconut Genebank for the South Pacific Improving location of diversity 21 In vitro collecting of coconut germplasm 65 In vitro collecting protocols developed by Assy-Bah et al. 67 In vitro culture of embryos 69 In vitro culture, sequence of operations 66 In situ conservation of coconut diversity 149-155 769 Index Basic information needed for conservation programme 152 Benefits of 155 Building and implementing a conservation programme 152 Definition of 150 Importance of 151 Indexing 375, 377, 381 Indexing and pathogen characterization 371, 373 India, Status of coconut genetic resources research in 573 Indonesia, Status of coconut genetic resources research in 608 Inflorescence explants, Use of 335. See Coconut micropropagation Information, public awareness, institutional support and partnerships 425 International Coconut Genebank 106 International Coconut Genebank for Africa and Indian Ocean 121, 552 International Coconut Genebank for Latin America and the Caribbean 123 International Coconut Genebank for South Asia 117, 524 International Coconut Genebank for Southeast and East Asia 119 International Coconut Genebank for the South Pacific 115, 520 International coconut genetic resources database 427-437 Background 427 Objectives 428 Organization 429 Functions 430 Contents 433 Technical references 437 International Coconut Genetic Resources Network: Its history and achievements 482 Goal, objectives and organizational structure 484 Achievements in the last 14 years 484 Introduction to the coconut palm 1 Inverse Sequence-Tagged Repeat 230 Invisible adaptation in coconut 3 ISTR. See Inverse Sequence-Tagged Repeat J Jamaica, Coconut lethal yellowing in. See Coconut lethal yellowing in Jamaica Jamaica, Status of coconut genetic resources research in 715 770 COCONUT GENETIC RESOURCES K Kenya, Status of coconut genetic resources research in 682 Kiribati, Status of coconut genetic resources research in 732 L LAC. See Latin America and the Caribbean Latin America and the Caribbean 557 Latin America and the Caribbean, Research on coconut genetic re- sources 557 LAVPD. See Leaf to air vapour pressure deficit Leaf to air temperature difference 285 Leaf to air vapour pressure deficit 285 Leaf water potential 285 Lethal yellowing disease 390 Lethal yellowing. See Coconut lethal yellowing Lethal yellowing. See Coconut lethal yellowing in Jamaica Lethal yellowing-like disease. See Coconut lethal yellowing Linkage mapping 269. See also Marker-assisted selection in coconut Locating and accessing target areas and material 59 Locating and collecting germplasm 11 Locating coconut genetic diversity 13 Losses from existing ex situ collections. See Gaps in coconut germplasm collecting LY. See Coconut lethal yellowing LYD. See Lethal yellowing disease M Mahalanobis distance 215 See also Morphometric methods of deter- mining diversity in coconuts Main elements of a GIS 23 Major pests and safe movement of germplasm 347 Malaysia, Status of coconut genetic resources research in 634 Manual on Standardized Research Techniques (STANTECH) 444 Manual on technical writing, public awareness, seminar presentation and proposal preparation for coconut researchers 449 Mapping coconut diversity for nut weight in the Ph 25 Mapping collection sites 35 771 Index Mapping for collecting genetic diversity 40 Mapping molecular markers 39 Mapping morphometric characteristics 37 Mapping of coconut genetic diversity 32 Mapping of major coconut cultivation areas, Vietnam 24 Marasmiellus cocophilus Pegler 398 Marchitez sorpresiva. See Phytomonas Marker-assisted selection in coconut 269, 275-276 Usefulness of various markers 275 Linkage mapping 275 Prospects of GMOs to date 276 MAS. See Marker-assisted selection in coconut Matrix summary of COGENT’s public awareness strategy and method- ologies 440 Mechanism of drought tolerance in coconut 292 Metamasius hemipterus. See Nematode disease Mexico, Status of coconut genetic resources research in 704 Microsatellites or Simple Sequence Repeats 230 MLO. See Mycoplasma-like organisms MOA for the establishment of the ICG-SP 124 Molecular breeding in coconut 269-273. See also Molecular markers for coconut improvement Linkage mapping and marker-assisted selection 269 Synteny 270 Candidate-gene approach and physical mapping 271 Genetically-modified organisms 272 Molecular genetics related to coconut improvement 273. See also Molecular markers for coconut improvement Devising markers for coconuts 273 Linkage mapping and QTL identification 274 Devising adapted mapping populations for QTL identification 274 Molecular hybridization 379 Molecular marker kit for COGENT partners, A 21 Molecular markers for coconut improvement 268-279 Potential for success in molecular breeding 269 Research activities in molecular genetics related to coconut im- provement 273 Results and evidence of the success of MAS in coconut 275 Immediate research needs 276 Constraints and opportunities 278 Molecular studies of diversity 20 Molecular techniques, increase in the use of 20 772 COCONUT GENETIC RESOURCES Morphometric methods of determining diversity in coconuts 209-222 Observations 211 Statistical methods 213 Application 217 Morphometric studies of diversity 17 Mozambique, Status of coconut genetic resources research in 688 Multilocation coconut hybrid trials 326-333 Project implementation 326 Multi-site and local hybrid/variety trial entries 327 Major achievements 328 Coconut hybrids that started fruiting 2.5 - 3.0 years after planting 329 Capacity building 329 Training and capacity building activities conducted 331 Mycoplasma-like organisms 372 N National collections in CGRD, List of 193 Nematode disease 382 Nematode disease: Red Ring 382 Nematode diseases 402 Nigeria, Status of coconut genetic resources research in 667 O On-farm conservation programme, Criteria for site selection 152-155 Diverse use values 153 Ecosystems. 153 Farmers and communities 153 Genetic erosion 153 Intra-specific diversity within target species 153 Logistics 153 Partners 153 Specific adaptations 153 Osmotic adjustment 286 Other explants (leaf and inflorescence), Use of 339. See Coconut micropropagation 773 Index P PA. See Public awareness initiatives in coconut. See also Information, public awareness, institutional support and partnerships Pacific Agricultural Plant Genetic Resources Network 521 Pakistan, Status of coconut genetic resources research in 604 PAPGREN. See Pacific Agricultural Plant Genetic Resources Network Papua New Guinea, Status of coconut genetic resources research in 737 Partner institutions in COGENT member countries 497 Pathogen characterization 375, 378, 381 PEQ. See Post-entry quarantine Performance of coconut hybrids in Asia, Africa and Latin America 302-308 Promising hybrids 302 Conditions favouring coconut hybrid performance 305 Narrowing the technology gap 306 Conclusion 307 Performance evaluation of coconut varieties and farmers’ varietal preferences 309-323 High-yielding varieties and hybrids 310 Findings of 1980 and 1988 assessment of new varieties 313 Summary of major findings of the 1998 varietal assessment study 315 Conclusion and recommendations 321 Performance of high-yielding coconut varieties/ hybrids 446 Pest risk analysis 411 Pest risk analysis and guidelines for the safe movement of germplasm in the ICG of Asia and the Pacific 411 Pest Risk Analysis process 412 Management of quarantine pest groups 415 General phytosanitary measures for the movement of coconut germplasm 417 Pest Risk Analysis process 412. See also Pest risk analysis and guide- lines for the safe movement of germplasm in the ICG of Asia and the Pacific Phase 1: Initiating the PRA 412 Phase 2: Pest risk assessment 413 Phase 3: Pest risk management 415 Pest risk assessment 396 Pest risk assessment in Brazil 402 Pest risk assessment in Côte d’Ivoire 399 Pest risk assessment of COGENT’s multi-site ICG 395 774 COCONUT GENETIC RESOURCES Pest risk assessment of ICG-AIO and ICG-LAC 395-405 Material and methods 395 Results 397 Recommendations 404 Pest risk assessment, Situation in Africa/Indian Ocean 398 Pest risk assessment, Situation in Latin America and the Caribbean 400 Pest risk assessment, Visits to ICG host countries 396 Philippines, Status of coconut genetic resources research in the 639 Phomopsis cocoina (Cooke) Punith. 399 Phytomonas 381. See also Protozoa disease Phytoplasma diseases 372 Phytosanitary measures for the movement of coconut germplasm between and among host countries in Asia and the Pacific 421- 423 India to Indonesia 421 India to Papua New Guinea 421 Indonesia to India 421 Indonesia to Papua New Guinea 422 Papua New Guinea to India 422 Papua New Guinea to Indonesia 422 Plumule explants, Use of 338. See Coconut micropropagation Polyethylene glycol 378 Population and variant 9 Populations 10 Post-entry quarantine 416, 423 Poverty reduction in coconut growing communities project 161-189 Activities 162 Constraints and recommendations 176 Lessons learned 173 List of participating organizations 178 Objective 161 Project benefits 165 Project impact 166 Project outputs 163 Summary of achievements 187 Summary of activities and target outputs 180 Sustainability 169 Poverty reduction in coconut growing communities, Volume 1: The framework and project plan 447 Poverty reduction in coconut growing communities, Volume II: Mobi- lizing for Action 447 775 Index Poverty reduction in coconut growing communities, Volume III: Project achievements and impact 448 PRA. See Pest risk analysis PRCGC. See Poverty reduction in coconut growing communities project PRCGC project partners 498 Primer sequences, Microsatellite kit 233 Priority activities of the Global Coconut Conservation Strategy 202- 206 Accelerate the full establishment and upgrading of 203 Additional collecting and conservation of germplasm 203 In situ and on-farm conservation of important farmers’ varieties 204 Upgrading the International Coconut Genetic Resources Database 204 Development of national coconut germplasm conservation strategy 205 Development of Regional Coconut Conservation Strategies 205 Proceedings of the COGENT regional coconut genebank planning workshop 444 PROCORD priority research areas 507 PROCORD. See Coconut Research for Development Programme Project grants provided to COGENT member countries 495 Promising hybrids in Asia, Africa and Latin America 302-305. See also Performance of coconut hybrids in Asia, Africa and Latin America China 302 The Philippines 302 Thailand 303 Vietnam 303 Bangladesh 303 India 303 Sri Lanka 304 Vanuatu 304 Côte d’Ivoire 304 Ghana 304 Tanzania 305 Mexico 305 Promoting conservation through use, Mechanisms for 199-202 Globally-coordinated coconut breeding programme 200 Community-managed coconut seedling nurseries 200 Linking conservation with broader areas of research and develop- ment 201 776 COCONUT GENETIC RESOURCES Developing catalogues and other public awareness materials on coconut 201 Upgrading the International Coconut Genetic Resources Database 202 Promoting multi-purpose uses and competitiveness of the coconut 445 Prospects of GMOs to date 276. See also Marker-assisted selection in coconut Protocol 1 (inoculation of embryos in the laboratory). See In vitro collecting protocols developed by Assy-Bah et al. Protocol 2 (inoculation of embryos in the field). See In vitro collecting protocols developed by Assy-Bah et al. Protozoa disease 381 Protozoa disease: Phytomonas 381 Provisional classification of coconut cultivars 247 Public awareness 439 Public awareness initiatives in coconut 439 Public awareness under COGENT’s ‘Poverty reduction in coconut growing communities’ project 451 Public awareness, institutional support and partnerships 425 Publications and other PA materials of COGENT 443 Q Q factor 37. See also Mapping morphometric characteristics QTL. See Quantitative trait loci Quantitative trait loci 269. See also Molecular genetics related to coconut improvement R Randomly Amplified Polymorphic DNA 229 RAPD. See Randomly Amplified Polymorphic DNA Rapid diagnosis assay 379 Red Ring. See Nematode disease Regional network reports 511 Related palm species. See Gaps in coconut germplasm collecting Restriction Fragment Length Polymorphism 228 RFLP. See Restriction Fragment Length Polymorphism Rhycopophorus palmarum. See Nematode disease Root-shoot signals 286 777 Index S Samoa, Status of coconut genetic resources research in 744 Schematic diagram for screening drought tolerant coconut varieties 293 Scope and status of coconut conservation 191-202 Conservation in COGENT’s multi-site International Coconut Genebank 192 In vitro embryo culture and cryopreservation 196 In situ and on-farm conservation 198 Promoting conservation through use 199 Secretariat of the Pacific Community 514 SEEA Network member countries 543 SEEA Network, COGENT member countries 543 SEEA. See Southeast and East Asian region Seychelles, Status of coconut genetic resources research in 691 Shannon-Weaver index. See Diversity indexes Simpson index. See Diversity indexes Single cross hybrids 255. See also Conventional breeding schemes Single Nucleotide Polymorphism 231 SNP. See Single Nucleotide Polymorphism South Asia sub-regional network 524 South Asia, Research on coconut genetic resources in 524 South Pacific, Research on coconut genetic resources 513 Southeast and East Asia (SEEA) sub-regional network 533 Southeast and East Asia, Area and production of coconut in 533 Southeast and East Asia, Research on coconut genetic resources in 533 Southeast and East Asian region 533 SPC. See Secretariat of the Pacific Community Sri Lanka, Status of coconut genetic resources research in 581 SSR. See Microsatellites or Simple Sequence Repeats Status of coconut germplasm collecting 44 Status of work on in vitro collecting of coconut germplasm 65 Stomatal regulation 285 Strategies for safe movement of coconut germplasm 390 Strategy in coconut germplasm collecting 53 Sustainable livelihoods framework, Five capitals 169-172 Financial capital 172 Human capital 169 Natural capital 170 Physical capital 171 Social capital 170 778 COCONUT GENETIC RESOURCES Synteny 270. See also Molecular breeding in coconut Synthetic varieties 258. See also Conventional breeding schemes T TAC. See Technical Advisory Committee Tanzania, Status of coconut genetic resources research in 670 Targeted surveys and under-represented phenotypes. See Gaps in coconut germplasm collecting Taxonomy 2 Technical Advisory Committee 474 Technical guidelines for the safe movement of coconut germplasm 393 Thailand, Status of coconut genetic resources research in 618 The Coarse Grid Strategy 54. See also Strategy in coconut germplasm collecting The farmer participatory approach. See Strategy in coconut germplasm collecting The International Coconut Genetic Resources Database 443 Tinangaja 377 Tonga, Status of coconut genetic resources research in 748 Treatment recommendations, Pests 423 Treatments, Pest 418 “True” Varieties 10. See also True variety True variety 9. See also True variety U Understanding the origin and distribution of diversity 59. See also Farmer participatory approach Use of Isozymes 18 Use of polyphenols 20 Using GIS tools 23. See also Geographic Information System (GIS) V Vanuatu Agriculture Research and Training Centre 520 Vanuatu, Status of coconut genetic resources research in 750 Vanuatu wilt 375 Variant 9 779 Index Varietal assessment study (1988), Summary of major findings 315- 321. See also Performance evaluation of coconut varieties and farmers’ varietal preferences Size class distribution of sample holdings 316 Age at first fruiting and productivity 316 Farmers’ evaluation of coconut cultivars 319 Varietal preference of farmers 320 Varietal preference of farmers 320. See also Performance evaluation of coconut varieties and farmers’ varietal preferences VARTC. See Vanuatu Agriculture Research and Training Centre Vietnam, Status of coconut genetic resources research in 625 Viroid diseases 377 Viroid diseases: Cadang-cadang and Tinangaja 377 Viroid purification 378 Viroid-like sequences of coconut 444 Virus disease: Vanuatu wilt or coconut foliar decay 375 780 COCONUT GENETIC RESOURCES 781 Index