BP Singh

This publication has been brought out jointly by theDivision of Germplasm Exchange and Division of Plant Quarantine,National Bureau of Plant Genetic Resources, New Delhi, based onthe lectures delivered during the training programme onEXCHANGE AND QUARANTINE OF PLANT GENETICRESOURCES at the NBPGR, New Delhi from 10 to 24 January, 1995conducted under INDO~US PGR Project (386~D513).© National Bureau of Plant Genetic ResourcesNew Delhi-ll0012, Ir~dia.Copies can be obtained from:Director,Nati,onal Bureau of Plant Genetic Resources,Pus a Campus, New Delhi-110012, ,IndiaPrinted by :Ashok Computers, EA-1/75, Indj;lrpuri, New Delhl-110012 (Tel. : 5782(01)

CONTENTSPage. Foreword iContributors1. Conservation of Plant Genetic Resources inIndia -RS. Rana 12. Centres of Origin and Diversity of Crop Plants-Bhag Singh 183. Need for Exchange of Plant Genetic Resources-B.P. Singh 30 .4. Principles of Plant Introduction and Exchange ofPlant Genetic Resources -M. Kazim3S( 5.! Principles and Concepts of Plant Quarantine -V.K. Mathur 406. 'Plant Quarantine Regulations and theIrImplementation in India -Ram Nath . 467. Medium and Long~term Storage of Seed Materialin Gene Bank -R.K. Saxena 548. Promising Introductions and Prioritize Needsfor exotic germplasm -M. Kazim 619. Use of Tissue Culture Techniques inExchange and Utilization of Plant GeneticResources -S.R. Bhat 67to. Promising Introductions in HorticulturalPlants (Fruits, Ornamentals) and Oil Seeds-B.P. Singh 7411. Promising Introductions and Prioritize Needsfor Exotic Germplasm of Grain Legumes, Fibrecrops and Under-utilized Crop Plants-R. V. Singh 8612. Priorities for Promising Introductions ofMedicinal, Aromatic, Spices and Condimentsgermplasm -Deep Chand 9913. Promising Introductions and Prioritize Needs ofExotic Gerrnplasm in different groups ofVegetable Crops -Po Brahmi 110ii

Page14. Genebank Management System (GMS) Software-R.L. Sapra 11515. New Policy on Seed Development and Role ofDesignated Inspection Authorities -RadheyShyant 11916. Symptoms of Fungal al;1d Bacterial Diseases-P.c. Agarwal 13517. Detection procedures for Fungal Plant Pathogensand Salvaging of Infected Germplasm-A. Majumdar 14218. Detection of Bacterial Pathogens and Salvagingof Infected Germplasm -Baleshwar Singh 14919. Symptoms Caused by Plant Viruses.:-D.B. Parakh 15920, Techniques for Detection of Plant Viruses-Shamsher Singh & R.K. Khetarpal 16521. Symptoms of Insect Damage in Crops-Shashi Bhalla 17122, Methods for Detection' of Insects and Mites inIinported Germplasm -B,R, Verma 18323. Plant Quarantine Treatments for SalvagingGermplasm Infested with Insects and Mites-Mnnju Lata Kapur 19324. Symptoms of Nematode Damage -V.K. Mathur& N. Gokte, 20225. Detection Techniques for. Plant ParasiticNematodes in imported Germplasm Materialfor Research -Arjun Lal 20826. Treatment Schedules for Eradication ofnematodes from Germplasm -Rajan 21727. Plant Genetic Resources Activities at IPGRIOffice for South Asia -R.K. Arora 22828 .. Intellectual Property Rights: Protection of PlantVarieties -R.S. Rana 23529. The Indian National Gene Bank -R.S. Rana 24230. List of Participants 249

FOREWORDIntroductions of useful plant materials from abroad have played a significant rolein diversifying Indian agriculture, in stabilising agricultural production in face of biotic ;;Indabiotic stress factors, New Policy on Seed Development introduced by the Governmentof India in 1998 opened up new vistas by enabling farmers and seed growers to importthe best planting materials available anywhere in the world, Incentives are also beingprovided to aid export-oriented imports of agri-horticultural planting materials and toencourage Indian collaboration with foreign agri-business.Equally important Is the role of. plant quarantine in safeguarding the unprotectedIndian farmers by preventing the possible entry of exotic pests, pathogens and weedswhich may sneak in alongwith the incoming seeds and other planting material.Germplasm exchange and rigorous quarantine checks need to be effectively linked andintegrated into one system that must be effective, efficient and'dependable. Our pastexperience reveals that the distribution of several pests and pathQgens has resulted fromthe movement of infested/infected/contaminated germplasm. While strict enforcementof quarantine regulations Is essential, it is equally important that healthy materials arereleased for planting without undue delay.A nine-years Indo-US Project on Plant Genetic Resources (PGR) is beingimplemented by the National Bureau of Plant Genetic Resources since 1988 andprofessional development of scientists as well as technicians engaged in different PGRactivities is an approved item of the project document. This objective is sought to beachieved by conducting in-country training courses and also by arranging advancedtraining at leading plant germplasm sites in USA.Under provisions of this Indo-US project, NBPGR organises short·term trainingcourses on various aspects of PGR activities for the professional development of indianscientists engaged in maintaining active germplasm collections for use in cropimprovement programmes in ICAR institutes, state agricultural universities and non·governmental organisations. A training programme on EXCHANGE AND QUARANTINEOF PLANT GENETIC RESOURCES was conducted at New Delhi Campus -from 10 to24 January 1995. The present publication is based on lecture notes prepared by thefaculty members and invited speakers for the benefit of trainees. It is expected that thecontents will also serve as useful reference material on the selected topics. Readersare welcome to' visit the Bureau for more detailed information and discussions.It gives me great pleasure to acknowledge expert and overwhelming supportprovided by my colleagues in the Bureau and also the financial assistance provided bythe ICAR and USAID for organising this course and bringing out this publication.10 January, 1995R.S. RanaDirectorNBPGR, New Deihl(i)

CONTRIBUTORS(Number in parentheses refers to the page on which authors'contribution begins)P.c. Agarwal (135)R.K. Arora (228)Arjun Lal (208)Shashi Bhalla (171)S.R. Bhat (67)Deep Chand (99)N. Gokte (202)Manju Lata Kapur (193)M. Kazim (35, 61)RK. Khetarpal (165)A. Majumdar (142)V.K. Mathur '(40, 202)Senior Scientist, Division of PlantQuarantine, NBPGR, NewDelhi-110012.Coordinator, IPGRI Office forSouth Asia, NBPGR Ca,mpus,New Delhi-110012.Senior Scientist, Diyision of PlantQuarantine, NBPGR, New Delhi-110012.Scientist (Se!. Gr.), Division ofPlant Quarantine, NBPGR, NewDelhi-110 012.Senior Scientist, NFPTCR,NBPGR, New Delhi-110 012.Scientist, Division of GermplasmExchange, NBPGR, New Delhi-110 012.Scientist (Se!. Gr.), Division ofPlant Quarantine, New Delhi-110 012Scientist (Sel. Gr.), Division ofPlant Quarantine, NBPGR, NewDelhi-110 012.Head, Division of GermplasmExchange, NBPGR, New Delhi-110012.Senior Scientist, Division of PlantQuarantine, NBPGR, New Delhi-110012.Senior Scientist, Division of PlantQuarantine, NBPGR, N.ew Delhi-110012Head, Division of PlantQuarantine, NBPGR, New Delhi-110 012.(ii)

D.B. ParakhPratibha BrahmiRadhey ShyamRajanRam NathItS. RanaRL. SapraR.K. SaxenaB,:,leshwar SinghBhag SinghB.P. SinghRV .. SinghShamsher SinghB.R. Verma(159)(110)(119)(217)(46)(1,235,242)(115.) .(54)(149)(18)(30, 74) \(86)(165)(183)Scientist (Sel. Gr.), Division ofPlant Quarantine, NBPGR, NewDelhi-ll0 012.Scientist (Sel. Gr.), Division ofGermplasro Exchange, NBPGR,New Delhi-110 012.Joint Director, Directorate ofPlant Protection, Quarantine &Storage, N.H. IV, Faridabad.Scientist (Sel. Gr.), Division ofPlant Quarantine, NBPGR, NewDelhi-11O 012.Principal Scientist, Division ofPlant Quarantine, NBPGR, NewDelhi-lID 012.Director, NBPGR, New Delhi-110012.Senior Scientist, Division ofGermplasm Evaluation, NBPGR,New Delhi-ll0 012.Senior Scientist, Division of'Germ p lasro Conservation,NBPGR, New Delhi-110 012.Scientist (Sel. Gr.), Division ofPlant Quarantine, NBPGR, NewDelhi-110 012.Head, Division of GermpJasm. Evaluation, NBPGR, New Delhi-110 012.Principal Scientist, Division ofGermplasm Exchange, NBPGR,New DelhH10 012.Senior Scientist, Division ofGermplasm Exchange, NBPGR,New Delhi-lID 012.Senior Scientist, Division of PlantQuarantine, NBPGR, New Delhi-110 012.Senior Scientist, Division of PlantQuarantine, NBPGR, New Delhi-110 012.(iii)

IIConservation of Plant Genetic Resources in IndiaR.S. RanaIntroductionPlants use solar energy for synthesis of organic products,using carbon dioxide from 'air and water as raw materials andreleasing oxygen into atmosphere. Conservation of plant diversityis thus of utmost importance in ensuring protection of healthyenvironment over the globe and also for meeting basic humanneeds of nutrition, health care, clothing and fuel. Although,around 250,000 plant species have been described so far and amuch larger number still remains to be duly recognised yet onlyabout 3,000 out of them were picked up from the wild by humanbeings for their use and grown on varying scales since thebeginning of agriculture considered to be nearly ten thousandyears ago. .Progressive improvement of these economic species overtime by successive generations of farmers hf\$ led to evolution oftoday's crop plants giving rise to landraces and traditionalcultivars. These prized materials alongwith their wild closerelatives serve as building blocks that are used by plant breedersfor development of improved crop varieties. They also constitutea priceless reservoir that contains genes conferring better adaptationto stress environments and resistance to diseases and pests. Thesevaluable plant genetic resources are, however, getting lost becauseof their replacement by high yielding modern varieties or due toheavy pressure ol;l their natural habitats. India has taken timely, steps for their long term conservation and sustainable utilization.

2 R.S. RanaIndian Centre of Origin and Diversity of Cultivated PlantsIndian subcontinent is well known since long as an importantcentre of" origin and diversity of a large number of agrirrrticulturalcrops. Over 1S,(10() species of flowering plants aren~tive to this region. These include more than 160 domesticated'species of economic importance alongwith over 320 species of theirwild ancestral forms and close relatives and also around 800species of ethnobotanical interest. Agri-biodiversity compriseslandraces and traditional cultivars· of several cereals, millets,legumes, vep;etables, fruits, forages, fibres, sugarcane, spices,condiments and medicinal & aromatic plants.Crops in which rich diversity still occurs in India includerice (Oryza sativa var. indica and several wild species), wheat(Triticum aestivum, T. durum, T, dieoccum), barley (Hordeum vulgare),sugarcane (Saccharum species and allied genera), Echinochloa,Panicum species, Setaria species and Paspalum species, foragegrasses, legumes (Vigna radiata, V. mungo, V. aconitifolia,V. umbellata and V. unguiculata, Cieer arietinunt, Cajallus cajan,Lablab purpureus), several Brassica species, sesame (Sesamumindictlm), okra (Abelmoschus esculentus and other allied species), eggplant (Solanum melongena and related types), Citrus species, bananaand plantains (Musa species belonging to different genomicgroups), jackfruit (Artocarptts heterophyllus, A. illtegl'ijolia), mango(Mangifera indica), Phyllanthus emblica, Carissa carandus, Syzygiu11lspecies, jute (Col'chorus capsularis), tree cotton (Gossypium arboreul1i),ginger (Zingiber officinale), turmeric (Curcuma species), pepper(Piper species), cinnamon and cardamom. Among tuber crops, richvariability exists in sweet potato (Ipomoea batatas), taros (Colocasiaand Alocasia), yams (Dioscorea esculenta, D. alata, D. dcltoidea etc.),Coleus species, swordbean (Canavalia species), velvet bean (Mucunaspecies), elephant foot yams and coconut (Cocos nucifera), particularlyin Nicobar group of Islands. Diversity also occurs in several minor .fruits such as Zizyphus jujube and Z.nummularia, Capparis decidua,Syzygium cumin ii, Myristica fragrans, Aegle ntarmelos and severalspecies of the genera Ribes, Rubus, Juglans, Pyrus and Prunus.Concentration of genetic diversity comprising native speciesand landraces occurs more in Western ghats and NorthweasternHimalayas. The richness of plant diversity is largely due to

Conservation of Plant Genetic Resources in India 3ecological diversity superimposed with tribal and ethnicdiversification, plant usages and religious rituals.Endemic Indian plant wealth is also supplanted with newspecies and forms that have transgressed national boundaries andenriched our flora which got diversified on being isolatedclimatically and spatially. The past linkages with Indo~Chinese­Indonesian, Chinese-Japanese and the Central Asian region helpedconsiderably in augmenting our crop plant resources. The influxof genetic material in distant past from Mediterranean and Africanregions has also resulted in the accumulation and diversificationof enormous genetic variability. The ancient travellers, invadersand religious missionaries have also contributed significantly. to,:",ards enriching the Indian gene centre.Wild Genetic Resources in the Indian Gene CentreWild species and putative ancestral forms of plants containvaluable genes that are of immense genetic value in crop breedingprogrammes using conventional methods or modern biotechnology.These genetic resources are likely to playa unique role in thedevelopment of new cultivars, strains and hybrids and also inrestructuring of the existing ones that lack one or the otherattribute. A comprehensive synthesis of available information onthe distribution, habitat preferences, ecology, utility, diversity etc.of such wild flora enlist wild relatives of crop plants comprisingwell over 325 species of agri-horticultural/ silvi-pastoral importanceoccurring in natural habitats as members of disturbed, bio-edaphiccommunities within the major vegetation types. Disturbedgrassland and scrub vegetation and similar open forest areas areconsiderably rich in such components except fruit trees which arelargely associated with evergreen, sub-humid, humid-tropical andtemperate ecotones.The distributional pattern of the wild plant genetic resourcesin different botanical/phytogeographical regions and the areas oftheir concentration where rich diversity of wild species stillcontinues to perpetuate, are of special significance for undertakingprogrammes on collection as well as for in situ conservation ofbiodiversity. The distribution pattern of wild species in differentphytogeographical regions reveal seven zones as follows:

4R.S. Ranai)ii)iii)iv)v)vi)vii)Western Himalayas (125 species),Eastern Himalayas (82 species),North-eastern region (132 species),Gangetic plains (66 species),Indus plains, North-western plains (45 species),Western peninsular region and Malabar coast (145species), andEastern peninsular region/Deccan plateau (91 species).PGR Conservation in IndiaConservation programmes are being implemented in Indiaat three levels, namely, genotypes, species and ecosystems. In situconservation of wild flora through protection of habitats andecosystems is being implemented by the Ministry of EnvirolUnentand Forests. Fourteen biosphere reserves have been identified onthe basis of survey da ta and seven of them have already been madeoperational (Table 1). Ex situ conservation of genetic variability ofcultivated plants and their wild relatives is the sole responsibilityof the National Bureau of Plant Genetic Resources (NBPGR) thatoperates under the Indian Council of Agricultural Research(ICi\R). Numerous botanical gardens managed by the BotanicalSurvey of India and several other organizations help in ex situconservation of endangered, threatened and rare plant species.Tabl.e 1. Ecosystem Conservation: in situ conservation in IndiaBiogeographic Biosphere Area StatesRegion Reserve (Sq. km.) InvolvedHimalayan Highlands Nanda Devi 1,560 U.P.Burma Monsoon Nokrek 80 MeghalayaBengalian Manas 2,837 AssamRain Forest Sundarbans 9,630 West BengalCorQmandel Gulf of MannaI' 555 Tamil NaduMalabar Nilgiri 5,520 Karnataka, Keralaand Tamil NaduAndaman & Nicobar Great Nicobal' 885 A & N islandsIslands

Conservation of Plant Genetic Resources in India 5National Bureau.of Plant Genetic ResourcesNBPGR was established in its present set up in 1976 althoughPGR activities were initiated in 1946. It is the nodal organisationin India for planning, conducting, promoting, coordinating andleading all activities concerning collection, introduction, exchange,evaluation, documentation, safe conservation and sustainablemanagement of diverse germplasm of crop plants and their wildrelatives with a view to ensuring their continuous availability foruse of breeders and other researchers in India and abroad. IndianNational Plant Genetic Resources System, operated by the NBPGR,comprises National Seed Repository (Base collection), NationalFacility for Plant Tissue Culture Repositories, Cryo-preservationFacility and a chain of Clonal Repositories and Field Gene Banks.This system is suppoited by an active partnership of over 30institutions/ centres designated as the National Active GermplasmSites as well as activities taken up under several bilateral andinternational agreements. Conservation programmes should,however, be justified by the effective use of preserved germplasm.The NBPGR has the following national mandate:* To plan, conduct and coordinate plant explorations forcollection of diversity in germplasm of cultivated plants,"their wild relatives and naturally occurring species ofeconomic importance.* To undertake introduction and exchange of plantgermplasm for research purpose.* To examine seed and plant propagules under exchangefor the presence of associated pests and pathogens andalso to salvage healthy materials from the infected/infested! contaminated samples.* To undertake and promote conservation of plant geneticresources on a long term basis employing in vivo, in vitroand cryopreservation techniques and also to assist in situconservation efforts.* To develop and operate the National Database for storageand retrieval of information on plant genetic resources.* To conduct basic research for prOViding a sound scientificback up to its services.

6R.S. Rana,,.,,,.........f' "."'..... ....

Conservation of Plant Genetic Resources in India7Indian NationalPlant Genetic Resources SystemNODAL ORGANISATIONNBPGRPARTNERSCrop Research Instit~teNational Research CentresAll India Coordinated Crop Improvement ProjectsState Agriculture UniversitiesNBPGRINDIAN NATIONAL GENE BANK(NATIONAL BASE COLLECTIONS)NATIONAL DATABASE ONPLANT GENETIC RESOURCES/' '-NATIONAL ACTIVE ./ GERM PLASM ADVISORYGERMPLASM SITES 7' COMMITTEES'uieRS/

8R.S. Rana***To develop a~d operate the National Herbarium of CropPlants and their Wild Relatives.To organise suitable training programmes at the national,regional and international levels.To develop and implement workplans based onMemoranda of Understanding with InternationalAgriculture Research Centres and bilateral agreementswith other countries.The concept of conservation of PGR in India has undergoneremarkable transformation in recent years. NBPGR has ,nowdeveloped a network of activities based on partnership with otherrelevant institutes and organizations. Its regional stations/ centresrepresent different phytogeographical regions with distinctecological conditions and these are located in the temperate regionat Shimla; arid region at Jodhpur; semi-arid region at Hyderabad,Akola and Amravati; hUmid-tropical region at Thrissur andhumid-subtropical region at Shillong. It also has 10 explorationbase centres; 7 of these located in the existing regional stations and3 located at Cuttack (Orissa), Ranchi (Bihar) and Srinagar (J&K) asindicated in Fig.1.At the main campus, the Bureau is organised into fiveDivisions, namely, Plant Exploration and Collection, GermplasmExchange, Plant Quarantine, Germplasm Evaluation andGermplasm Conservation. A National Facility for plant TissueCulture Repository, funded by the Department of Biotechnology,also operates at NBPGR and complements the Bureau's programmeson genetic conservation of orthodox seed materials. In addition,three All India Coordinated Projects are located at NBPGR,namely, on Medicinal and Aromatic Plants, on Arid Legumes andon Under-utilized and Under-exploited plants. With this set upat the Bureau, India has taken a lead among the developing nationsin establishing a well organized and effective plant geneticresources system.Major efforts are now devoted towards further strengtheningthe national plant genetic resources system in the country withNBPGR as the leading organization linked effectively with over 30lCAR institutes, National Research Centres, All India CoordinatedProjects and State Agricultural Universities (Fig.2). These centresare deSignated as· National Active Germplasm Sites for specific

Conservation of Plant Genetic Resources in India 9crops (Table 2) and have been assigned responsibility formaintaining, evaluating and supplying germplasm out of itscollections of different crops which are also under long termstorage at -20°C in the National Gene Bank (Table 3). In vitrocollections are maintained if). the National Plant Tissue CultureRepository (Table 4). Field collections are maintained by theHeadquarters and the Regional Stations. It has also developedfacilities for cryopreservation of seeds, pollen and in vitro culturesin liquid nitrogen (-1~6°C). A chain of clonal repositories has alsobeen developed under its network. Computerized NationalDatabase on plant genetic resources and National Herbarium ofCultivated Plants and their Wild Relatives are also operative at theBureau. Basic researches related to conservation and utilization ofgermplasm form an integral part of Bureau's research programmesso as to improve quality and effectiveness of its multi-facetedservices for the sustainable management and use of plant geneticresources. Major accomplishments of NBPGR are highlighted inTable 5.Table 2. Directory of National Active Germplasm SitesS1.No.CropNAG SiteNo. of Accessions1,.)~'3.j!567WheatRiceMaizeBarleySorghumPearlmilletSmallMilletsDirectorate of Wheat Research,Kamal 132 001 (Haryana)Central Rice Research Institute,Cuttack 753 006 (Orissa)Directorate of Maize Research,Indian Agriculturai ResearchInstitute, New Delhi 110 012Directorate of Whent Research,Kamal 132,001 (Haryana)18,00042,0002,500National Research Centre for 5,160Sorghum, Rajendranagar, Hyderabad(Andhra Pradesh) 50ll 030All Indin CoordinatedPenrlmillet Improvement Project,College of Agriculture, Shivaji Nagar,rune 411 005 (Maharashtra)All Indin CoordinatedSmnH Millets Improvement Pl'Oject,University of Agri1. Sciences,Bungall1re 560 065 (Karnataka)8,572

10 R.S. RanaContd ..... (Table 2)81. Crop NAG Site No. of AccessionsNo.8 Pulses Indian Institute of Pulses Research, 9,310(ICAR), Kanpur 208 024 (UP)9 Soybean National Research Centre for Soybean, 2,500Indore 425 00 (Madhya Pradesh)10 Oilseeds Directorate of Oilseeds 15,629Reseal'ch (ICAR), Rajendranagar,Hyderabad 500 030 (Andhra Pradesh)11 Rapeseed National Research Centre on 8,082& Mustard Rapeseed & Mustard,Bharatpur (Rajasthan)12 Groundnut National Research Centre 6,432for Groundnut, TimbawadiP.O., Junagarh 362 015 (Gujarat)yti' Sugarcane Sugarcane Breeding Institute, 3,979Coimbatore 641 007 (Tamil Nadu)/,,)4 Cotton Central Institute for Cotton 6,896Research, P.B. No. 125,Nagpur 440 001 (Maharashh'a)15 Jute & Central Institute of Jute 3,226Allied& Allied Fibres,Fibres Barrackpore 743 101 (West Bengal)16 Vegetables Directorate of Vegetable 16,139Research, Varanasi 221 005 (UP)17 Potato Central Potato Research Institute, 2,375Shimla 171 001 (Himachal Pradesh)18 Forages Indian Grassland & Fodder 6,267Research Institute, Jhansi, 284 003(Uttar Pradesh)19 Spices National Research Centre for Spices, 2,847Marikunnu, Calicut 673 012 (Kerala)20 Tobacco Central Tobacco Research Institute, 1,500Rajahmundry 533 105 (Andhra Pradesh)21 Plantation Central Plantation Crops Research 307CropsInstitute, Kasargod 671 024 (Kerala)22 Medicinal All India Coordinated 375& Aromatic M & AP Improvement Project,Plants NBPGR, New Delhi 110012

Conservation of Plant Genetic Resources in India 11Contd ..... (Table 2)51. Crop NAG Site No. of AccessionsNo.23 Agro- National Research Centre for 40Forestry Agro Forestry, Indian Grassland &Plants Fodder Research Institute,Jhansi 284 003 (Uttar Pradesh)24 Fruits National Research Centre on Arid 541(Semi-Arid) Horticulture, Bikaner (Rajasthan)25 Fruits NBPGR Regional Station, Phagli, 454(Subs tropical Shimla 171 004& Temperate) (Himachal Pradesh)26 Fruits Indian Institute of Horticultural, 13,118Research 255, Upper Palace Orchards,Bangalore 560 080 (Karnataka)2.7 Citrns National Research Centre for Citrus, 51Seminary Hills, Nagpur440 006 (Maharashtra)28 Fruits Central Institute for Horticulture 587(NorthernPlains)for Northern Plains,Lucknow 226 016 (Uttar Pradesh)Z9 Tuber Central Tuber Crops Research Institute, 3,586Crops Sreekariyam, Trivandrum695 017 (Kerala)Jd' Pseudo- NBPGR Regional Station, Phagli,cereals Shimla 171 0043,682(Himachal Pradesh)NBPGR is helping the collaborating institutions in developingmedium term seed storage facilities and also computer facilities fordocumentation. Bureau also provides training to their scientistsand technicians.Plant Quarantine ResponsibilityUnder the authority delegated to its Director, the NBPGR isresponsible for quarantine of incoming and outgoing germplasmfor research purposes. Director, NBPGR has also been empoweredto issue import permits and phytosanitary certificates. Introductionof germplasm envisages in~E?':~ o.~ small sampl~s (usnal~y' up to200 .ru~ or limited number of propagules accompanied by--pnytosanitary certificate.

12 R.S. RanaTable 3.Base collections in National Gene Bank at NBPGRkept in long term storage in seed repository maintainedat -2oDe (as on 31 December, 1994)Crop GroupsNo. of accessionsCereals 60,221Pseudo-cereals 560Millets and Minor Millets 15,667Oilseeds 20,491Pulses 25,143Fibre Crops 3,607Vegetables 7,243·Spices & Condiments 78Medicinal & Aromatic Plants 179IOthers 778Improved Varieties 857Duplicate Safety Samples 4,752Total 1,39,576Table 4. Status.of in vitro Conservation at NBPGR (NFPTCR)Crop No. of Storage Temp. Storage periodaccessions (0C) (months)Allium sativunt 64 10 16Allium spp. 15 10 12Ipomoea batatas 230 25 12Dioscorea spp. 31 25 12Zingiber officinale 120 25 12Curcuma spp: 24 25 8Musa spp. 246 25 12Others 35 25 10

Conservation of Plant Genetic Resources in India13Table 5. Major accomplishments at a glanceExplorations conducted in IndiaExplorations undertaken abroadSamples of indigenous germplasm collectedImport permits issued (since July, 1989)Exotic germplasm samples ir~.roduced and distributedSamples supplied to users from germplasm holdingsGermplasm samples exportedPhytosanitary certificates issuedSamples examined for quarantine checkSeed samples stored in the National Gene bankCultures stored in the In vitro RepositorySamples kept in cryopreservationGerni.plasm lines characterized/evaluatedCrop catalogues publishedCrop Inventories brought outIssues of Plant Introduction Reporter brought outIssues of NBPGR Newsletter published CQuarterly)Training programmes organizedRegional/international workshops organizedReference specimens prepared for the Herbarium .Seed samples displayed in the MuseumGermplasm Advisory Committees functioningBooks and monographs published472994,5942,52711,15,9051,44,1263,01,0266,41114,43,2141,36,5427658121,49,7965435693435611,4643,166539Germplasm Advisory CommitteesAnother milestone in the organization of a strong NationalPGR System has been the constitution of Germplasm AdvisoryCommittees, which have been set up recently for specific crops orgroups of crops. They play an important role by advising theBureau regarding the status of its current holdings of differentcrops, shortcomings in storage and management system as wellas gaps in exploration and collection of indigenous geneticvariability of native crops. These committees also suggest

14 R.S. Ranapromising countries/regions in the world that need to be exploredor approached for introduction of new crops/ genetic variability tosustain our crop improvement programmes.International CollaborationsCollaboration with CGIAR crop"based institutesBesides operating the above mentioned network of activegermplasm sites in the country, the Bureau also actively collaborateswith International Agricultural Research Centres in India andabroad. International Plant Genetic Resources Institute (IPGRI)'has contributed significantly to the Bureau's efforts offeringexpertise, training and research support. NBPGR has also an activecollaboration with ICRISAT on joint exploration and multi-locationevaluation programme on five ICRISAT's mandate crops. This hashelped in documentation of germplasm collections in pearlmillet,sorghum, pigeonpea, chickpea and groundnut. Considerable'exchange of germplasm takes place between NBPGR and IRRI(Philippines), and CIMMYT (Mexico), and ICARDA (Syria).IPGRI's office for Sourth Asia is located, in the NBPGRcampus and there is an active collaboration between them basedon biennial work plans. In addition to supporting joint explorationand collection programme in this region, the IPGRI helps indeveloping training programme in conservation and managementof plant genetic resources designed primarily for the Asian region.Collaborations under Bilateral ProgrammesBesides the International Research Centres, many cou.ntrieshave well developed systems for assemblage, enrichment,documentation and conservation of plant genetic resources andalso have computerized database network. ICAR has Memorandaof Understanding as well as bilateral agreements with several suchinternational organizations and national programmes. Exchangeof plant genetic resources activities is carried out through NBPGRwith over 80 countries.Global responsibility for PGRFollowing an assessment of the gene bank standards,infrastrudural facilities and trained manpower available at NBPGR,the Bureau has been identified for responsibility of global and

Conservation of Plant Genetic Resources in India 15regional base collections of more than a dozen crops. The firstInternational Okra Workshop was organized at NBPGR in 1990under the sponsorship of IPGRI. Besides the IPGRI experts,representatives from Belgium, France, Brazil, China, India,Indonesia, Ivory Coast, Nigeria, Papua New Guinea, Philippines,Senegal, Sri Lanka, Sudan and USA participated in the deliberations.An International Workshop on Sesame Genetic Resources wasorganized at Nagpur and at NBPGR Regional Station, Akola(Maharashtra) in 1993.INDO~US PGR ProjectThe Indian National Plant Genetic Resources Programme(IN-PGRP) has gained substantially from the PGR systemsadopted by leading nations in this field yet it has not whollyadopted any of their models. India is panistakingly developingits own indigenous system taking full advantage of its inherentadvantages and incorporating suitable features that have beenevolved over a long period of time by repeated trials in developedcountries. NBPGR is currently operating a 9 years INDO-USProject on Plant Genetic Resources with a budget provision of 27.95million US dollars. This project is being implemented to enhanceNBPGR's national capability and also to enhance its role at theregional and international levels.NBPGR'S National Information SystemThe Bureau is gradually strengthening its computerizeddata-base to back up conservation of genetic resources for not onlyimmediate utilization of already conserved and evaluated/characterized germplasm in the ongoing plant breeding programmesbut also meeting needs of future generations. Success of both theseactivities is dependent upon the availability of descriptive andevaluation information on accessions stored in the gene bank.National database system gathers all relevant data from diversesources that are required by user scientists. Selection ofappropriate descriptors and their descriptor states, softwarecomputer programmes and retrieval system are pre-requisites todevelopment of computerized information on genetic resources.Success of genetic resources database management depends uponeffective cooperation among genetic resources personnel,computer experts and plant breeders/biotechnologists.

16 RS. RanaNBPGR proposes to expand its database through strong regionaland international linkages.Recent Developments and ConcernsSince success of crops improvement programmes depen4slargely upon availability of suitable genotypes having the desiredtraits, it is important that the breeders should continue to haveunrestricted access to crop genetic resources. International. Undertaking on Plant Genetic Resources, developed by the FAOCommission on Plant Genetic Resources in 1983, is anintergovernmental agreement for 'free' supply of plant germplasmfor research purpose. India is among the 112 countries who havesigned this Understanding even though many developed countrieslike USA, Canada and Japan have expressed their reservations. Itis noteworthy that this International Undertaking is based on theconcept that all plant genetic resources are a common heritage ofmankind and this should be made freely available for use in cropimprovement programmes.A turning point came in 1992 when the Convention onBiological Diversity (CBD), signed by 154 countries, recognisedsovereign rights of nations over their genetic resources and alsoover determining terms for access to them. The eBD became. legally binding on 29 December, 1993 and India has ratified itbecoming thereby a Contracting Party. Text of the InternationalUnderstanding is being revised in a step by step process so as toharmonise it with provisions of the CBD. The revised Undertakingis likely to become protocol for implementation of CBD.India has also signed the general Agreement on Tariffs andTrade (1994) that has brought agriculture and its services alsounder the purview of this multilateral trade agreement that willlead to establishment of World Trade Organisation on 1 January,1995. Under the provisions of this agreement, member nations areto provide for Breeders' Rights through a patenting system orthrough an effective sui generis system or a suitable combinationof both. Indian Patents Act 1970 provides for process patentingbut excludes agriculture. Indian Government proposes to developa sui generis system by legislating a Plant Variety Protection Act.Likewise, it is also proposed to regulate the outflow of geneticresources through a suitable legislation.

Conservation of Plant Genetic Resources in India 17To sum up, the Indian Plant Genetic Resources Programmehas taken a lead among the biodiversity-rich developing countriesnot only in collecting and safeguarding crop genetic resources butalso in promoting their utilization in crop improvementprogrammes. Wherever, access to genetic resou;:ces is linked totransfer of appropriate technology, India will opt for obtaininghigh technology in preference to soft and hard technologies sincethe Indian system is fully capable of absorbing such technology.NBPGR is already engaged in characterization of its more valuablegermplasm accessions employing modern techniques like isozymeprofiling, RFLP, RAPD and MSV. Efforts are underway to developand employ multi-locus DNA probes for DNA fingerprinting ofprized native materials and also single-locus DNA probes fordetection of pathogens as well as for screening of availablecollections for identification of genotypes possessing desirabletraits.

•Centres of Origin and Diversity of Crop PlantsBhag SinghIntroductionHuman beings may have existed as a species for about 2million years yet it was only about 10,000 years ago that humanbeings started growing food, rather than merely gathering it fromwild state and hunting wildlife. With the origin of settledagriculture, the interest in environmental assets began. Whereverman has gone, his plants have gone with him. When the transitionfrom 'gathering' to I growing' food took place, there was immediateinterest in the collection and utilization of useful plants, andanimals from native flora and fauna. Most of the importanteconomic plants and animals were domesticated thousands ofyears ago. The pioneer work of de Candolle (1886) and N.I.Vavilov(1926) led to the wide spread recognition that the diversity of cropswas centred in certain geographical regions.Concept of centres of origin and crop plant diversityThe study of origin of agriculture and its spread providesclues to the geographical distribution of centres of domestication.These centres were found to be located in areas of maximumdiversity.In the forefront of contributors to our knowledge of the globaldispersal of crop plants and their wild relatives has been Vavilov(1927). He believed that time was the only factor that influencedthe dispersal of species and their increase in variation. Hesuggested that centres of origin of species coincide with the areas

Centres of Origin and Diversity of Crop Plants 19where greater diversity occurs in species. Based on the presenceof great diversity,he identified 8 centres with three sub-centres asI centre of origin' where agriculture developed independently.Appendix-I lists the crop plant species that originated in thesecentres. These centres were found to be characterized by theaccumulation of dominant genes in the middle portion andrecessive genes in the periphery. This possibly led to the highconcentration of genetic diversity in the secondary centre ratherthan the primary centre of origin. The centres of diversity locatedby Vavilov have proved to be fertile promising collecting areas forfuture explorations. A well known hypothesis for the origin ofagriculture is the Rubbish-Heap hypothesis. It says that earlyhumans gathered roots and seeds for their food. Such plantsactively colonized the base areas around their dwellings, whichwere rich with the discarded rubbish.Vavilov also recognized secondary centres of origin andpainted out that valuable forms are found far removed fromprimary area of origin. A good example is the Washington Navel.~ra!!.g~ .. ~!~~

20 Bhag Singh3. Establish the distribution of genetic diversity anddetermine the geographical centres where this is atmaximum, especially those centres with endemic forms Icharacters.4. Determine the centre where diversity of geneticallyallied species is concentrated.5. Correlate the above centres with the areas ofconcentration of nearest wild relatives.6. Compare centres of origin of group of cultivated plantswith certain specialized parasite(s).7. Support the above with the evidences from archae,ologylinguistics and history.Transformation of concept of originThere has been considerable chan~{n the concept of originever since Vavilov (1951) propounded it.It was considered later that the presence of wild relatives wasessential for designating the centre of origin. Subsequently, severalworkers revised the boundaries supported with the conceptsdeveloped by them.Schiemann(1951) criticised the Vavilovian gene centre theoryspecially in relation to Ethiopean and central Asian centres whichwere considered by Vavilov as the centres of origin of durumwheat (Triticum durunt) and barley (Hordeum vulgare) and that ofhexaploid wheat (T. aestivum) on the other. The criticism isprimarily based on the fact that none of wild progenitors werepresent in these centres, hence termed them as /I Accumul~tiQnCentres". Zohary (1970) also criticized the Vaviiovian"~oncept ofcentres of origin. Similarly Ku.ckuck (1963) criticised the Vavilovianconcept as he did not find the dominant alleles in the centres oforigin and recessives in the periphery, e.g., bread wheat andsorghum as claimed by Vavilov.Megagene centresZhukovsky (1965), a dose associate of Vavilov, proposed 12megagene centres of crop-plant diversity. The new areas addedto Vavilov's eight centres were Australia, Africa, and Siberia

Centres of Origin and Diversity of Crop Plants 21followed by reVISIon of the boundaries to make 12 centres.M!.~:,'?gE;!:l:e:,5:~.11trft,s of wild growing species related to crop plantswhere the cultigens first o.riginated, were also indicated. He alsodistinguished between the 'pr~~~:rI pl,!crpgene centres~ for therestricted areas where the cultigen actually first originated and thesecondary megagene centres which are the areas into which theyha"tr~ now spread. Zeven and Zhukovsky (1975) in "Dictionary ofcultivated plants and their centres of diversity" have listed thespecies for different mega gene centres. Zeven and de Wet (1982)prefer the term region to centre. By and large, these twelve regionshave wider coverage and more acceptability.Centres and non-centresHarlan (1971) recognized only three main centres eachwith more or less connected but large and diffuse non-centres(Table-I).Table 1. Centres and non-centres (Harlan, 1971).CentreNorth Chinese-BINorth East Centre-AIMeso American Centre-CINon centreSouthEastAsian and SouthPacific non-centre-B2African non-centre-A2South American non-centre-C2Harlan also recognized smaller areas/pockets of varietal and/orracial diversity within a Vavilovian Centre and termed these as'Micro centres'. Darlingtan (1973) has increased the number ofcentres·~of' origin to sixteen.N udear Centres and Regions of diversityHawkes (1983) advocated that agriculture began not once butseveral times, more or less simultaneously in different regions ofthe world. He envisaged centre of agricultural origin from whichfarming spread into one or more regions for which he proposedthe name Nuclear centres and regions of diversity (Table-2). Helinked up his Nuclear centres with the archaeological evidences to .provide proofs of the agricultural origins.

22 Bhag SinghTable 2.Nuclear centres and regions of diversity ofdomesticated plants (Hawkes, 1983).Nuclear centresA. Northern ChinaB. The Near EastC. Southern MexicoD. Central to SouthernPeruRegions of diversityI. ChinaII. IndiaIII. South East AsiaIV. Central AsiaV. The Near EastVI. The MediterraneanVII. EthiopiaVIII. West AfricaIX. Meso-AmericaX. Northern Andes(Venezuela to Bolivia)There arp several such regions where crop actually did notoriginate. This argument is based on the wild progenitors, absenceof archaeological remains to suggest antiquity of a crop species.These are regions int which the crops perhaps spread from theNuclear centres in the past in which spatial isolation in time andintensive human selection played a pre~dominant role in theincrease of genetic diversity.Micro centresHawkes (1983) further identified small 'micro' centres forseveral cr, ps.1. New Guinea~Sugarcane (Saccharum officinarum)2. Solamon Islands and Fuji-Mlisa species3. North Western Europe-Avena strigosa, Seeaie cerealc4. United States, Canada-Helianthlls annuus, H. tuberoslts5. Southern Chile-Bromus, mango6. Brazil - Mnnihot esculenta, Ananas comostlsDiversity of crop plants in Indian sub~continentRich genetic diversity occurs in several crop plants (about 166species) and their wild relatives (about 320 species). Antiquity of

Centres of Origin and Diversity of Crop Plants 23agriculture and ethnic diversity in the sub-continent have playeda major role in the diversification of crop resources in this region.In addition, there has been a continuous stream of introductionsof new crops and their cultivars by man since the ancient times.Crops like pear, grape, nuts, datepalm, garlic, opium,maize,potato, sweet potato, tomato, chillies, french bean etc. werebrought by the Mughals, Spaniards, Portuguese and the British.Thus, both indigenous and well adapted exotic set of materialsconstitute a well balanced matrix of crop diversity in Indian subcontinent.Eight regions where diversity in major crop plants isconcentrated are as under.1. Western HimalayasBarley, wheat, maize, buckwheat, amaranth, prosomillet,finger milletFrench bean, soybean, lentil, blackgram, peasPumpkin, cucumber, Allium spp., ginger, BrassicaePome, stone, soft and nut fruitsMedics2. Eastern HimalayasBarley, maize, buckwheat, amaranth, finger millet,foxtail millet.French bean, soybean, cowpea, blackgram, peas, scarletbeanPumpkin, cucumber, Allium spp., ginger, chayote, treetomato, BrassicaePome and stone fruits3. North-Eastern RegionRice, maize, sorghum, finger millet, foxtail millet, job'stearsFrench bean, soybean, pigeonpea(perennial),blackgram,rice bean, dolichos bean, winged beanPumpkin, chayote,· cucumber, okra, eggplant, chilli!Capsicum spp. pointed gourd, ash gourd

24 Bhag SinghTaros, yams4. Gangetic Plains5. Indus PlainsCitrus spp. - Lime/lemon/orange/grape fruit, bananaTea, tree cotton, jute, kenaf and mesta, largecardaIl'l:om, ginger, long pepper, sugarcaneRice, sorghum, barnyard millet, little millet/ Panicumspp.Chickpea, cowpea, mungbeanOkra, eggplant, bittergourd, Cucumis spp., Lulla spp.Jackfruit, mango, lemon/lime, orange, jujube, Indiangooseberry I Emblica spp., jamunl Syzygium spp., melonsLinseed, niger, sesame, BrassicaeSugarcane, mulberryDurum wheat, pearl milletMoth bean, cluster bean, chickpea, blackgramOkra, tucumis spp.Jujube, Khirnil Mimusops sp., Phalsal Grewia sp.Sesame, taramiral Eruca sp.Cotton6. Eastern Peninsular Region/E.Ghats/Deccan pleatueRice, sorghum, finger millet, pearl millet, foxtail millet,little millet, prosomillet, kodo milletBlackgram, greengram, cowpea, horsegram, Mucunaspp., pigeonpea, Dolichos bean, rice beanTaros, yams, elephant-foot yamBanana, mango, lemon/lime, jackfruitNiger, Brassicae, sesameGinger, turmeric, chilli/ Capsicum spp., kenaf, sugarcane, .coconut, cotton.

Centres of Origin and Diversity of Crop Plants 257. Western Peninsular Region/W:Ghats/MalabarRice, sorghum, finger millet, small millet/ Panicum spp.Blackgram, greengram, cowpea, pigeonpea, dolichosbean, horsegiam, sword beanOkra, eggplant, cucumber, chilli/Capsicum spp.Taros, yams, elephant-foot yamJ ackfruit, banana, lime/lemon, orange, jamun/ Syzygium ..'spp.Sugarcane, black pepper, turmeric, ginger, coconut,arecanut, cotton.8. The, Islands RegionsCoconut, bread fruit, chilli, taros, yams, XantilOsomaspp.ReferencesCandolle, A.de. 1886. Origin of cultivated plants. Haffner, NewYork.Darlington, C.D. (1973) Chromosome Botany and the origin ofcultivated plants, Allen and Unwin., London 237 p.Harlan, J.R(1971). Agricultural origins: centres and non-centres.Science 174 : 468-474.Harlan, J.R (1975). Crops and man. American Society of AgronomyMadison, Wisconsin. 295 p.Hawkes, J.G. (1983). The diversity of crop plants. Harvard UniversityPress, Cambridge, Mass. 184 p.Kuckuck, H. (1963). Present views on Vavilov's gene-centretheory. Plant Genet. Resources Newsletter 12 : 8-10.Paroda, RS. and Arora,RK.(1991). Plant genetic reSOlll'ces :Conservation and Management, IBPGR, ROS & SEA,N. Delhi. 'pp. 392Schiemann, E.(1951) New results on the history of cultivatedcereals. Heredity (London) 5 : 305-320.

26 Bhag SinghVavilov, N.I.(1951). Phytogeographical basis of plant breeding.The origin, variation, immunity and breeding ofcultivated plants (K.J. Choster, translator). ChronicaBotanica 13 : 366 p.Zeven, A.C. and Zhukovsky, P.M.(1975). Dictionary of cultivatedplants and their centres of diversity. Centre for AgriculturalPublishing and Documentation, Wageningen. 219 p.Zohary, D.(1970). Centres of diversity and centres of origin. In :O.H. & Benett. E. (Ed.) Genetic Resources in Plants.Oxford, Blackwell, pp. 33~42.Zhukovsky, P.M.1965. Genetic and botanical irregularities in theevolution of cultivated plants. Genetika Mosc. 1 : 41-49(in Russian; English summary).1. China2. IndiaWorld centres of diversity of cultivated plantsAvena nuda, naked oat (Secondary centre of origin)Glycine max, soybeanVigna angularis, adzuki beanPhaseolus vulgaris, common bean(receSSive form; secondary centre)Phyllostachys spp., small ba:rp.boosBrassica juncea, leaf mustard(secondary centre of origin)Prunus persica, peachCitrus sinensisSesamum indicum, sesame (endemic groupof dwarf varietiesj secondary centre)Camellia(Thea) sinensis, China teaOry:z;a; sativa, riceElusine coracana, African milletCieer arietinum, chickpeaVigna aconitifolia, moth beanVigna umbellata, rice bean'Appendix I

Centres of Origin and Diversity of Crop Plants27Macrotyloma uniflorum, horse gramVigna unguiculata, asparagus beanSolanum melongena, eggplantRaphanus caudatus, rat's tail radishColocasia antiquorum, taroCucumis sativus, cucumberGossypium arboreum, tree cotton, 2XPiper nigrum, pepperIndigojera tinctoria, indigo2a. Indo-MalayaDioscorea spp., yamCitrus maxima, pamelaMusa spp., bananaCocos nz/cijera, coconut3. Central ~siaTriticum I aestivum, bread wheatTriticum compactum, club wheatTriticum sphaerococcum, shot wheatSecale cereale, rye (secondary centre)Pisum sativum, peaLens culinaris, lentilCi"'c>r arietinum, chickpea'Se~. m indicum, sesame (a centre of origin)Lint", dsitatisimum, flax (a centre of origin)Carthamus tinctorius, safflower ( a centre of origin)Daucus carata, carrot (basic centre (If Asiatic varieties)Raphanus sativus, radish ( a centre of origin)Pyrus communis, pearPyrus malus, appleJuglans regia, walnut4. The Near EastTriticum monococcum, einkorn wheat­Triticum durum, durum wheatTriticum turgidum, Paulard wheatTriticum aestivum, bread wheat (endemic awnless group; acentre of origin)Hordeum vulgare, cultivated two rowed barley (endemicgroup)

28Bhag SinghSecale cereale, ryeAvena byzantina, red oatCicer arietinum, chickpea (secondary centre)Lens culinaris, lentil (a large endemic group of varieties)Pisum sativum, pea (a large endemic group; secondary centre)Medicago sativa, blue alfalfaSesamum indicum, sesame (a seperate geographic group)Linum usitatissimum, flax (many endemic varieties)Cucumis melo, melonAmygdalus communis, almondFicus carica, figPunica granatum, pomegranateVitis vinifera, grapePrtmus armeniaca, apricot (a centre of origin)Pistacia vera, pistachio (a centre of origin)5. The MediterraneanTriticum durum, durum wheatAvena strigosa, hulled oatsVida Jabal broad beanBrassica oleracea, cabbageOlea europaea, oliveLactuca {3ativa, lettuce6. AbyssiniaTriticum durum, durum wheat (an amazing wealth of forms)Triticum turgidum, Poulard wheat (an exceptional wealth offorms) .Triticum dicoccum, Emmer wheatHordeum vulgare, barley (an exceptional diversity of forms)Cicer arietinum, chickpea ( a centre of origin)Lens cullinaris, lentil ( a centre of origin)Eragrostic abyssinica, teaEleusine abyssinica, tef .Eleusine coracana, finger milletPisum sativum, pea ( a centre of origin)Linum usitatissimum, flax (a centre of.origin)Sesamum indicum, sesame (basic centre)Ricinus communis, castor bean (a centre of origin)Coffea arabica, coffee

Centres of Origin and Diversity of Crop Plants297. Southern Mexico and Central AmericaZea mays, cornPhaseolus vulgaris, common beanCapsicum annUUnl, pepperGossypium hirsutum, upland cottonAgave sisalana, sisal hempCucurbita spp., squash, pumpkinS. South America (Peru, Ecuador, Bolivia)Ipomoea batatas, sweet potatoSolanum tuberosum, potatoPhaseolus lunatus, lima beanLycopersicon esculentum, tomatoGossypium barbadense, sea island cotton (4X)Carica papaya, papayaNicotiana tabacum, tobaccoSa. ChileSolanum tuberosum, potato8b. Brazil and ParaguaySolanum utilissima, maniocArachis hypogaea, peanutTheobroma cacao, cacao (secondary centre)Hevea brasiliensis, rubber treeAnanas comostls, pineapplePassiflora edt-dis, purple grandilla or passion fruit

•Need For Excha~ge of Plant Genetic ResourcesB.P. SinghIntroductionAgriculture evolved some 10,000 years back, with thedomestication of plants of choice and economic importance to man.The result was narrowing down the dependence of human kindfrom an array of wild usable plant resources to a limited anddefined species grown under organised package of agronomicpractices. The era of Green Revolution further confined thevariability in the local cultivars, land races and even withwithdrawal of lesser important crops from particular areas, therebynecessitating the collection and conservation of existing cropspecies, taxa and their relatives.Stages of plant genetic resources for exchangeEvaluation under domestication may be viewed at threedifferent stages(i)(ii)(iii)DomesticationStabilizing semi-domesticated/semi-wild cropsBreeding for improved varieties.The germplasm resources exchange is equally needed at allthe three stages.(i) Domestication: It implies domestication of a crop species innew areas beyond their habitat/home, e.g., Simmondsia chinensis(Jojoba) in area outside southern USA and Mexico.

Need For Exchange of Plant Genetic Resources 31(ii) Stabilizing semiwild/semidomesticated crops : Import ofdesirable germplasm under this category is made in respect of'land races', old and obsolete cultivars etc. for the purpose ofincorporating adaptability / resistance/ quality genes etc.(iii) Breeding for improved varieties : Centres for enhancedgermplasm, International screening nurseries/trials or improvedvarieties from other countries for specific trials I genes.Germplasm diversity in IndiaIndia· is endowed with wide range of agro-ciimatic conditions(tropical, subtropical, temperate) with diverse crop Iplants of agrihorticulture/agro-forestry. Indian gene centre is one of theimportant region of diversity of crop plants. It holds prominentpOSition among the eight Vavilovian centres of crop plants origin.Out of 2,40 J OOO economic plant species recounted by Zeven andDewet (1982) distributed in 12 megagene centres of crop diversity,160 species are enumerated in Hindustani gene centre, distributedin eight diverse phy,togeographical/ agro-ecological regions ofIndia. There are several crops in which rich diversity exists in India.Cereals and millets: Rice (Oryza sativa), little millet (Panicumsumatrense), kodo millet (Paspalum scrobiculatum)Legumes/pulses: Urid or black gram (Vigna mungo), arhar orpigeonpea, (Cajanus cajan), horse gram or kulthi (Macrotylomauniflorum).Fruits: Mango (Mangifera indica), Banana (Musa sp.), Jamun(Syzygium cumini), Jackfruit/kathal (Artocarpon heterophyllus, A.integrifolia), Citrus sp., Karonda (Carissa carundus), Khirni (Manilkarahexandra)., Phalsa (Grewia asiatica/G.maequalis), bael (Aegle marmelos),wood apple (Feronia limonia), kokam (Garcinia indica).Vegetables : Egg plant/brinjal (Solanum melongena), parval/pointed guard (Trichosanthes dioica), tOri/ridgegourd & smoothgourd (Luffa spp.), tinda/round gourd (Citrullus lanatus), taro/arbi(Colocassia esculenta), yams (Dioscorea spp.), jimikand(Amorphophullus campanulatus), Kundru (Coccinia indica) cucumber(Cucumis sativus).Oilseeds : Brassica sp. (rai, sarson and tori types), sesame(Sesamum indicum).

32 B.P. SinghOrnamentals: Rhododenchron, orchids, Hedychium fern and fernallies, Nerium indicum, Polyathea longijolia, Bauhinia, Primula,Camellia and many others.Exchange of Plant Genetic Resources - Past effortsExchange of plant genetic resources in the past has helpedconsiderably number of nations to grow new plants and cropspecies that were not known before. In the process people ofvarious countries were greatly benefited. For example, most of thecrops cultivated in the industrialized countries have their centresof origin and diversification in developing countries. All cropsexcept sunflower and a few :minor species of fruits etc. wereintroduced in USA like wheat ,rice, soybean, which are now beingexported by USA..Wheat, barley, sorghum, maize, groundnut, tea, coffee,potato, tomato, cauliflower, cabbage, pea, cashew, rubber, etc. allwere introduced into India from abroad. Many of these crops havebecome integral part of '~he existing cropping systems in differentparts of the country. This process is still on.Organised Exchange of Plant Genetic ResourcesSystematic activities of introduction and exchange of plantgenetic resources of agri-horticultural crops started as early as 1946at the Indian Agricultural Research Institute (JARI), New Delhiunder a scheme initiated at the Division of Botany. Thus, activitiesfurther accelerated by the creation of a new Division of PlantIntroduction in 1961. In 1976 , this Division was raised into anindependent institute, National Bureau of Plant Genetic Resource(NBPGR), New Delhi under Indian Council of AgriculturalResearch. NBPGR has been functiOning as a nationwide research/service organizations for exchange of plant genetic resourcesstrictly under phytosanitary conditions. It is assisting various cropbased institutes, all India Crop improvement programmes,agricultural universities as well as agri-horticultural/forestrydepartments in the country.It has close linkages with more than 85 countries besides thePlant Introduction Agencies having headquarters at Beltsville(USA), Canberra (Australia), Leningt;ad (USSR/Russia), Ottawa(Canada), San Paulo (Brazil), Buenos Aires (Argentina), Lisbon(Portugal), Peredeniya (Sri Lanka), Dacca (Bangladesh), Islamabad

Need For Exchange of Plant Genetic Resources 33(Pakistan), Addis Ababa (Ethiopia), Tapivizele (Hungary), Sofia(Bulgaria), Manila «Philippines), Tsukuba (Japan) and many alliedagencies, universities, botanical gardens private institutes/organisations. The Bureau has also developed cooperatomgrelationship with the International Agricultural Research Centres(IARC's) under CGIAR viz IRRI (Philippines), CIMMYT (Mexico),CIAT (Colombia)1 CIP (Peru), ICRISAT (India), ICARDA (Syria)IlTA (Nigeria) as well as other centres like AVRDC (Taiwan) andWARDA (Liberia), beside IBPGR (IPGRI).Import of Plant Genetic ResourcesThe import and export of seeds, plants, plant products andplanting materials in India are regulated by the rules andregulations framed under, DIP Act of 1914. NBPGR has beenauthorised to issue Import Permit and import /export of agrihorticulturaland agri-silvicultural plants in small quantities forresearch purposes for public and private sector with research baseas well as for ICRISAT located in Patancheru near Hyderabad.During the period 1976-1994 (ending December) 3,81,982accessions of different agri-horticultural corps and agri-silviculturalcrops/plants were introduced through correspondence.Export of Plant Genetic ResourcesDuring the period 1976-1994, Division of Germplasm Exchangeexported above 1,84,000 accessions of different agri-horticulturalcrops to various foreign collaborating agencies/institutes in 101countries under phytosanitary conditions. These exports werebased on specific requests received .Intensifiction in Introduction EffortsThe Division of Germplasm Exchange of the Bureau has beencontinuously making all out efforts for introduction of diverseplant genetic resources by way of (a) Bibliography Screening ofvarious J oumals /Periodicals/Reports/ Catalogues. Index seminamsetc. in search of useful genetic stocks, latest improved cultivars,donors of useful characters, new crops etc., for their introduction,(b) Indents for seed/ plant materials received from various cropbased institutes/universities scientists l (c) Annual Workshopsrecommendations, Mini-Mission meet on pulses & oilseeds andalso on suggestions from various Crops Advisory Committees.

36 Mansoor KazimThe last five centuries have witnessed much give and takeof economically useful flora of different regions in the world. Thisexchange in the beginning was at the genus and species level andmore in emperical way. These activities were more done by thetravellers in search of new plants for their economic attributes, bythe rulers during Moughal time and by Portuguese, English,French and Dutch during 16th century.The. distribution of plants has been established byVavilov(1951) not to be uniform throughout geographical areas buthave different pattern in different ecological areas. This could beattributed to Gene centre concept of cultivated plants and theirwild progenitors . This has given rise to two basic concepts :Agro-climate (climatically homologous regions)The introduction of a plant species from closely alike climateor ecological similar region could have better possibilities ofadaptation and success in establishment in a new site.Adaptability- introduced crop will have a greater chance ofsurvival provided it has much amount of adaptability allowing itto fit well into wide variety of environment.Zhuk9y~1;:y. (1975) has. further elaborated the principles ofcenter Dtorigin to centres of diversification. This means that aparticular genus or species has diversified from a particular regionto greater areas by way of its wide adaptability.Introduction of a particular genotype, thus from an area ofdiversification to climatically similar region would be mores·uccessful as compared to one which is from a diverse centre. Thisdoes not hold true with all the plant species since the adaptabilityis an important factor in establishment of a species in newenvironment.In recent years, plant scientists have realized that a stage hascome where introduction of new economic species has limitedpossibility but there is much scope in improving agronomicpotential of presently grown species by genetic improvementthrough judicious use of extensive germplasm .Therefore, greateremphasis has been laid on location and assemblage of varietalwealth through correspondence or through specific explorations.

Principles of Plant Introduction and Exchange of Plant Genetic Resources 37This assemblage of diverse germplasm could be achievedfrom the centres of diversity, primary and 'secondary centres;national organisations USDA (USA), IPI (Russia), CSIRO (Australia),CENERGEN /EMBRAPA (Brazil) and other PGR centres;international network of CCIR institutions: IRRI (Philippines),ICARDA (Syria), CIMMYT (Mexico), eIP (Perut IITA (Nigeria),CIAT (Colombia), AVRDC (Taiwan) and many others; FAO/IPGRt GeneBanks; National arboreta/Botanic gardens/privatenurseries etc.India being one of the important centres is known asHindustani centre of diversity of crop plants has much diversityto several crops, rice (OI:!za sativa), finger millet (£leusine coracana),tree cotton (Gossypium arboreum), pepper (Piper nigru'11!-), pulses andcucurbits etc. as an old centre. It has well established linkage withother 58 countries over the world and free exchange of plantgenetic resources in agri-horticultural and silvi-agricultural crops, ,on reciprocal basis. India has not only exchange of mandate cropsof CGIAR institutions but has also equally strong programme ofexchange of plant genetic resources with well over 85 countriesunder different protocols/memoranda of understanding atGovenunentallevels under bilateral programmes.The National Bureau of Plant Genetic Resources (NBPGR),New Delhi owes its birth to the erstwhile Division of PlantIntroduction of Indian Agricultural Research Institute. It has beenengaged since 1940 in introducing elite strains, varieties, geneticstocks and improved varieties/cultivars of various crops fromdifferent sources under strict quarantine control. Some of them asprimaty introductions, Mexican wheats, and Taiwan paddy havebrought revolution in the country. Others were utilized indevelopment of new more adaptable, stress tolerant types in cropimprovement programmes.The NBPGR has been functioning as a national research cur"service organisation for introduction and exchange of plant geneticresources of varied agri-horticultural plants for research purposes.It provides basic raw material in the form of germplasm to thescientists in reAR crop based institutes, agricultural ~niversities,all India crop improvement programmes and private seedindustries.

38 Mansoor KazimIt also monitors the testing of different international trials/nurseries received from CGIAR centres for adaptability studies.The Bureau also does export of plant genetic resources byarranging their quarantine examination, issuance of phytosanitarycertificate to the researchers abroad under different collaborativeprogrammes.Besides the NBPGR, the exchange of seed/planting materialshas also been authorised by the Government of India to Deptt. ofAgriculture and Cooperation for bulk commercial materials, Deptt.of Environment & Forests for plants of botanical interest , andIndian Council of Forestry Research & Education for forestryplants.The Division of Germplasm Exchange at the Bureau has beengiven the responSibility of importation/export of plant geneticresources for research use. It is following two ways of selecting thematerials for introduction into the country. One is by scanning thecurrent world literature for promising varieties developed worldover and selecting them to their suitability to Indian conditions.The second approach is where a scientist makes a request to theBureau for his specific needs of the material relating to his researchprogramme.The enactment of new policy on seed development by. theGovt. of India has made it obligatory for all researchers intendingto import seed/planting materials to meet the following twomandatory requirements of Plants, Fruits & Seeds Order (regulationof import into India) 1989:1. Import permit 'before importation of seed/plantingmaterial and2. Phytosanitary certificate from country of originThe above two documents should invariably accompany theplant material conSignments on arrival at Indian port for customclearance. In case of materials meant for research, the DirectorNBPGR is authorised to issue the required import permit for allICAR institutes/centres" State Agricultural Universities~ ICRISA Tand private seed industries having research and developmentprogramme. Import permits could be obtained in advance fromDirector, NBPGR on an application made in prescribed format.

Principles of Plant Introduction and Exchange of Plant Genetic Resources 39This import permit should be sent by the researcher to the sourcecountry with special request to enclose this import permitalongwith official phytosanitary certificate at the time of despatch.The consignment should invariably be addressed to Director,NBPGR for its quarantine clearance and national accessioning ofthe material. In no case it should be got in onels own name sincethis would not be released by the customs for w

•Principles and Concepts of Plant QuarantineV. K. MathurIntroductionPlant quarantine is based on the principle that prevention isbetter than cure with the objective of reducing the risk factoragainst the introduction of exotic pests and diseases and if alreadypresent to check its further spread and multiplication. No doubtthough it is very important that our plant breeders must continueto obtain all the promising planting material in the form of newcrops, new varieties and germplasm in the form of primitive landraces, wild and weedy relatives of crop plants possessing genes fortolerance/resistance against biotic and abiotic stress situations andother desirable traits for crop improvement programmes but at thesame time it is equally essential to adopt quarantine safeguards toprevent the entry of exotic diseases and weeds into the country.This activity possesses a great potential for transfer of associatedpests and pathogens from one region/ country to another as allkinds of planting materials are affected by diseases or injuriescaused by one or more organisms, viz., insects, nematodes, fungi,bacteria, viruses etc.Need for QuarantineAlthough a good number of pests and pathogens have worldwide distribution but still there are several of these which are notyet reported to be present in many regions/ countries, and ifpresent, have only a limited distribution.It is well known now that the present distribution of severalimportant pests and diseases has resulted from the movement of

Principles and Concepts of Plant Quarantine 41infested/infected/ contaminated planting material from one countryto another, as of their own they are not capable of travelling longdistances. Thus, the accelerated pace of exchange of germplasmthroughout the world these days has exposed the world to greaterrisk of introducing exotic pests and- pathogens in 'clean areas'.There are several such cases reported in literature. This would beevident from the following few examples. European com borer(Ostrinia nubilalis) entered USA from Europe and became moreprolific and polyphagus than in Europe resulting in losses ofmillions of dollars. The introduction of the late blight pathogen(Phytophthora infestans) of potato into Ireland from Centrai Americaabout middle of 19th Century, resulted in almost completedevastation of potato crop and the result was mass migration ofIrish people to all parts of the world. In 1870 Ceylon (now SriLanka) was one of the foremost coffee producing countries in theworld with an industry based on Coffea arabica, an EthIopianspecies, recognis,ed for quality. In 1875, coffee rust appeared inCeylon and devastated the plantations. Production dropped from400 million pounds in 1870 to 5 million pounds by 1889 forciI,lgthe cultivators to abandon coffee plantation and take to teaplantations. It is believed that the rust got introduced from EastAfrica. In 1908, cotton production in Mississippi (USA) was1,800,000 pounds. Four years later, this production was reducedto one-third due to the inroads made by cotton boll weevil(Anthonomus grandis) which moved into USA from Mexico. In,1960, it was estimated that this pest was costing the USA anaverage of $ 375 million per year. Similarly, stem and bulbnematode (Ditylenchus dipsaci) severely attacked red clover insouthern part of Sweden for many years, so lucerne was suggestedas an alternate crop because though a host, it was not attacked bythe nematode. But recently there are reports that this also is beingattacked by this nematode~doubtlessly the 'lucerne race' of D.dipsaci got introduced into that area from somewhere through seedas this nematode is also transmitted through seed. This examplealso illustrates the fact that introduction of a 'race' may be asdisastrous as introducing a new species or an exotic pest in acountry /region. These are a few significant examples of lossesarising from the introduction of a disease or a pest into a country,which could have been avoided had there been an effectivequarantine'in those days.

42 1l.K. MathurWhat is Plant QuarantinePlant Quarantine can be defined as legal enforcement ofmeasures aimed to prevent pests and pathogens from spreading,or to prevent them from multiplying further in case they havealready found entry and have established in a new restricted area.Although it is difficult to predict how an introduced pest willbehave, if it gets established, in a new location and environment,but there is a distinct possibility that freed from its natural enemiesand competitors and with possibly more susceptible hostsavailable, pest in a new environment may have more deleteriouseffect than in its place of origin. Though plant quarantine measuresmay not' guarantee an everlasting protection against the entry ofexotic species but will certainly check or delay the introduction ofthese unwanted organisms and their subsequent establishment inhitherto clean areas.' Therefore, practicing IPlant Quarantine' i.e.protection of plants by observing quarantine measures will go along way in protecting our agriculture from the ravages of exoticpests and diseases and also from those which are not present inthat area.Plant Quarantine is in effect a national service provided bythe Central Govt./State Govts. for the benefit of the farmers. Ashas been rightly stated by Mathys (1975) "Government quarantineoffers services which are beyond the capabilities of individualbeneficiaries or that are difficult to obtain in some other way at alesser cost."Analysis of the pest risk involved in germplasm introductionThis is an important aspect to be taken into consideration bythe quarantine officials in the germplasm introduction as itprovides a sound biological basis to decide quarantine policies. Ifrisks are low, quarantine should be ~ in permitting entry butif the risks are high, the material may even be cl.e:cied entry. Thepest risk analysis should take into account the benefits that arelikely to accrue from the introduction of the planting materialunder reference, cost of quarantine inspection, treatment includingdetention in post-entry quarantine facility as well as cost oferadication, should an exotic pest get established. It should alsoconsider factors such as availability of trained staff, efficaciousdetection techniques, treatments, knowledge about the life cycle of

Principles and Concepts of Plant Quarantine 43the pest, existence of races/strains, mode of transmission, worlddistribution, factors favouring establishment, adequacy of surveyand surveillance programmes. Risk which is higher with theintroduction of vegetative propagules, rooted plants, andunderground plant parts such as rhizomes, tubers, suckers, bulbsetc. than true seed, budwood, scions and unrooted cuttings shouldalso be taken into consideration. In case of bulk imports the risksare maximum apparently because thorough examination andtreatment is very. difficult and the planting area is also far too large.Qua;antine Reg'ulationsPlant quarantine measures will be justified only when thepest has UP natural means of spread and when these are based onbio]ogica!.consideratious alone. The regulations vary from countryto country, but they all have the following c,ommon features.(a)(b)(c)(d)(e)(f)Import permit is required;Phytosanitary Certificate is required;Specific prohibition;Grant exceptions to the prohibition for scientificpurposes;Specify inspectiQn and treatment on arrival;Prescribe post-entry quarantine or other safeguards.Guidelines for promulgating quarantine regulations aregenerally based on the suggestions made by internationalorganisation or intergovernmental agreements or conventions,which are organised within the frame work of the FAO. Whilepromulgating quarantine regulations local conditions like cropspectrum, environmental conditions, presence of alternate hostsshould also be considered. The quarantine regulations shouldhave flexibility within permissible safeguards and undue restrictionson movement of planting materials should be removed. Thiswould help in clearing the doubts in the minds of the scientists andthe farmers as well that quarantine regulations are not bottlenecksto hamper their activities but they are in the best interest of ourcountry / agricultural economy.International/Regional CooperationInternational or regional cooperation is very important forthe success of plant quarantine particularly in the present situation

44 V.K. Mathurbecause crop improvement programmes in almost every countryare dependent on ~xotic germE.!~sm. Cooperation on the followinglines would help in reducing the cost of quarantine operations asalso the pest/pathogen introductiqn risk to a considerable extent.1. Consortium of Plant Quarantine Stations: Under this conceptproposed ~~ K~~_.{t~?7) different quarantine stations wouldundertake the-processing of different materials (several accessionsof the same crop or a group of crops) at each station and then sharethe material. This would avoid duplication of efforts, reduce costof processing and more material would be available with adequatequarantine safeguards. In the same way, scientific information likedetection techniques, treatments, distribution of pests/pathogensand antisera for serological diagnosis of viruses and bacteria couldbe shared by quarantine stations of different countries.2 .. Establishment of Central Seed Health Testing Laboratories:~;N~~g!!.cg:9J!2Z?') proposed establishing a few central seed healthtesting laboratories well equipped with required facilities andtrained personnel. These may be coordinated with regional genebanks or international c~ntres, but should be independent of theorganisation for whom they are working. Such an arrangementwould lower pest risk to a considerable extent and avoidduplication of efforts, thus reducing the costs of processing. Plantquarantine services of different countries will have faith in suchlaboratories and will accept their certification.3. Third Country/Intermediate Quarantine: Under this 'conceptthe material could be grown, tested/indexed for hazardous pestsand pathogens in a temperate country without much risk becauseeither the possible hosts of such pests are not present there or theenvironment is unfavourable for their establishment. Severalstations in France, USA, UK and Netherlands offer their facilitiesfor crops like cocao, rubber, banana, sugarcane, groundnut etc.4. Biogeographical Regions: The concept of biogeographicalregions proposed by: Ma!~ (19~~) is an example of regionalcooperation. The eight biogeographical regions proposed foreffective quarantine are separated by J;latural barriers like sea, highmountains and deserts, thus making pest/pathogen disseminationextremely difficult. Accordingly, all countries in each region musthave common quarantine regulations so that chances of entry of

Principles and Concepts of Plant Quarantine 45exotic pests are greatly minimised. Countries of the ASEAN regionhave come to an official level understanding and formulatedsimilar set of quarantine regulations to protect the region againstexotic pests.Finally, it would be quite appropriate in the present dayscenario to conclude with the following quotation from Hewitt andChiarappa (1977), "with to-day's knowledge, technology andscientific cooperation, there seems little excuse or reason for plant .germplasm to be transferred in anything but a healthy condition.There is no excuse, not even impatience for the boot-legging ofplant germFbsm through or around quarantine./IReferencesHewitt, W.B. and Chiarappa, L. (Eds.) (1977). Plant Health andQuarantine in international transfer of genetic resources. CRCPress, Cleveland, USA.Kahn, RP. (1977). Plant Quarantine: Principles, methodology andsuggested approaches, pp. 209-308. In: Plant Health andquarantine in international transfer of plant genetic resources(Eds., W.B. Hewitt & L. Chiarappa). CRC Press, Cleveland,USA.Mathys, G. (1975). Thoughts on quarantine problems. EPPO Bull.S : 55.Neergaard, P. (1977). Seed Pathology. Macmillan Press Ltd., London,UK, 1187 p.

IIPlant Quarantine Regulations and theirimplementation in IndiaRam NathIntroductionIt can safely be stated that movement of pests and diseasesacross national boundaries has been taking place ever since manstarted transporting seed and other propagating materials fromone country to another for diversification of agriculture andincreasing food production. However, the magnitude of croplosses that could be caused by the introduced .R1?~ts and ctiS.5!_?sgswas realised around the middle of the last century when the potatocrop was completely devastated by the late blight pathogen(Phytophthora infestans) introduced from Central America resultingin the well known Irish famine of 1845. During 1860's powderymildew (Uncinula necatar),root eating aphid (Phylloxera vitifalia)and downy mildew (Plasmopara viticala) got introduced into :&e.:nc;eon the nursery stocks imported from tTor.th .America. The wineindustry of France was virtually anhilated due to the combinedeffect of the three diseases/pests. Although true nature of thedisease was not yet fully understood, farmers were convinced thatthese got introduced from America. Alarmed by these diseaseoutbreaks,governments in European countries started puttingembargo on the import of American products. In the neighbouringcountry Sri Lanka, rust (Hemileia vastatrix) severely mauled coffeeplantation bringing down coffee production from 400 millionpounds in 1870 to just 5 million pounds by 1889. Because of theseand other disease/pest outbreaks in different parts of the world,certain countries like Germany, France, Australia, USA and UK

Plant Quarnatine Regulations and their implementation in India ,47enacted plant quarantine legislations with fl. view to prevent entryof exotic pests! diseases during the first and second decades of thecurrent century.Development of plant quarantine in IndiaLearning from the experience of these countries, Indianauthorities also took early steps in this direction and in '1906"ordered compulsory flLW,g.atio).:1 of all the incoming cotton baies' at'the port of entry to prevent the introduction of the dreadedMexican cotton boll weevil (Antlzonomus grandis). However, it wassoon realised that cotton boll weevil was not the only pest thatneeded to be kept out. Therefore, a comprehensive plant quarantinelegislation was passed on 3 February,12,l!.by the Governor Generalof India which is popularly known as "Qestr,uctive Insect and Pe~ts~t,)91.4" (DIP Act). Since its first promulgation, Govt. of Indiahas issued eight amendments/ adaptation orders, so far. The DIPAct empowers the Central Government to:i) prohibit or regulate the import into India or any partthereof or any place therein, of any article or class ofarticles likely to cause infection to any cropiii)iii)prohibit or regulate the export from a State or thetransport from one State to another in India, of anyarticle or class of articles likely to cause infection to anycrop;make rules prescribing the nature of documents whichshall accompany any article or class of articles, theexport or transport of which is subject to the conditionsimposed or which shall be held by the consignor orconsignee thereof, the authorities which may issuesuch documents and the manner in which thesedocuments shall be employediIt authorises. the State Governments to make rules fordetention, inspection, disinfection or disinfestation of any articleor class of articles in respect of which a notification has been issuedand for regulating the powers of the officers whom it may appointin this behalf.It provides for penalty for persons who knowingly contravenethe rules and regulations issued under the act.

48 Ram NathIt also protects the persons from any suit, prosecution orother legal proceedings for anything done in good faith orintended to be done under the act.Plant quarantine rules and regulations have been f.raroed bythe government under the provisions of the Q}:f ... bet. Theseregulations can be grouped as, (a) foreign quarantine regulationswhich regulate the import into India of seeds, plants and plantproducts from other countries and, (b) domestiC quarantineregulations which regulate the transport/movement of suchmaterials from one State to another within the country.Foreign quarantine regulationsUnder the foreign quarantine, regulations have been issuedfor (a) import of cotton, (b) import of live insects and (c) importof live fungi. Besides these, a comprehensive Plants, Fruits andSeeds (Regulation of Import into India) Order, 1984 waspromulgated which came into force in June 1985. After theannouncement of the 'New Policy on Seed Development' by theGovt. of India in October 1988 liberalising the import of 'se~ds andother planting materials for the benefit of the Indian farmers,Plants, Fruits and Seeds (Regulation of Import into India) Order,1989 was promulgated, by the Government in suppression of theearlier order. This order is popularly known as PFS Order, 1989.The Order stipulates that:i) no consignment shall be imported into India unlessaccompanied by a valid 'import permit' issued by acompetent authority in India;ii)no consignment shall be imported into India unlessaccompanied by a 'phytosanitary certWcate' issued byan officer authorised by the official plant quarantineservice of the source country;iii) all consignments for consumption, 5.Q:wtng andp~()p~g.aHng or pl9:!lt!1)$ shall be imported only throughentry points notified from time to time. Consignmentsof plants and seeds for sowing or planting shall beimported through lan(l cl,li3to:qt st!'ltjons, s.ea/ C).ir portsat Amritsar, Bombay, Calcutta, Delhi and Madras andsuch other entry points as may be specifically notifiedfrom time to time; .

Plant Quarnatine Regulations and their implementation il1 India 49IV)v)vi)vii)viii)ix)x)xi)the consignments on arrival at any E;:ntry ,point, shallbe inspected by the Plant Protection Adviser to theGovt. of India (PP A) or any other official authorised byhim in accordance with the guidelines issued by himfrom time to time. The,:pPA or the official authorisedby him may, after inspection, fymigation, disinfectionor di~il1:feEjtation as may be considered necessary"accord quarantine clearance or order in public interest,destruction of the consignment or return the same to thesource country;hay, straw or any m.aterial of plant origin except sawdust and wood shavings or peat for horticultural use,shall not be used as packi~g material;import of soil, earth, clay, compost, sand, plant debrisalongwith seed/planting material shall not be permittedexcept for research investigations under a specialperm.it issued by PP A;seeds/plants/planting materials which require postentry-quarantineisolation growing and inspection,shall be grown in facilities approved by the PP AIDeSignated Inspection Authority (DIA) or any other,officer authorised in this regard to conform to theconditions laid down by the PP A;seeds of coarse cereals, pulses and oilseeds shall becleared or rejected after quarantine check within aperiod of 55 days;seeds of vegetables/flowers/ornamentals and bulbs/tubers of flowers shall be cleared /rejected within aperiod of 50 days;import of seed/planting materials of fruits shall bepermitted selectively by the DAC on therecommendations of the concerned Director ofHorticulture/ Ag'riculture subject to quarantineregulations;seeds/planting materials of all species of Allium;Sferculiaceae and Bombaceae; Citrtis;, COCUSi Coffea;Gossypiunt; Arachis; Medicago; Solanum; Hevea; Saccharum;

50 Ram Nathxii)Nicotiana; Trifolium; Helianthus; saplings and budwoodsof flowers and ornamental plants shall be importedsubject to the conditions specified in Schedule II of thePFS Order;the Central Government may, in public interest, relaxany of the conditions of this order relating to importpermit and phytosanitary certificate in relation to theimport of any consignment.Domestic quarantine regulationsSo far eight domestic quarantine regulations have beenpromulgated under the provisions of the DIP Act with a view toprevent spread of certain specific pests and diseases with restricteddistribution in India. These include fluted scale (lcerya purchasi),San Jose scale (Quadraspidiotus perniciosus), banana bunchy top,banana mosaic, potato wart (Synchytrium endLlbioticum), apple scab(Venturia inaequalis), codling moth (Cydia pomonella) and goldennematode (Globodera rostochiensis, G. pallida).Implementation of quarantine regulationsIn the beginning, plant quarantine regulations were beingimplemented by the nontechnical personnel of customs departmentand hence, plant quarantine was being observed in India as a mereformality. The first step to remove this serious lacuna was takenin 1946 1 when the Govt. of India established the Directorate ofPlant Protection, Quarantine and Storage with Plant ProtectionAdviser to Govt· of India as its head and responsibility forenforcing the quarantine regulations was entrusted to thisDirectorate. The first Plant Quarantine and Fumigation Stationunder the directorate was established at Bombay in 1949. Atpresent such stations are in operation at all the international air /sea ports and land custom stations on border check posts withneighbouring countries which are manned by scientific andtechnical staff. Adequate laboratory and isolation growing facilitiesalongwith specialist scientific staff have been created at Bombay,Calcutta l Delhi and Madras to meet the quarantine requirementsof New Policy on Seed Development.However, the directorate does not have the powers of 'searchand seige'. These powers rest only with the customs authorities

Plant Quarnatine Regulations and their implementation in India 51and whenever any seed/planting material is noticed by them inthe accompanied baggage of incoming passengers, it is interceptedand passed onto the quarantine officials. Bulk consignmentsimported as cargo are referred to the quarantine under customsescort. Similarly, consignments imported as post parcels, oftenattract the attention of the postal appraisal staff who forward thesame to plant quarantine authorities for inspection and clearance.In the case of domestic quarantine, while the regulationsregarding inter-state movement/transport of seed/plantingmaterials are framed by the Central Government,these areenforced with the help of the State Governments. Therefore, closecooperation of State Government authorities is essential for thesuccessful implementation of domestic quarantine regulations.Quarantine for germplasmUnder the PFS Order, three types of consignments areimported : (a) bulk consignments of grains/pulses etc. forconsumption, (b) bulk consignment of seed/planting materials forsowing and (c) germplasm of agri-horticultural crops in very smallquantities for research use. The Plant Quarantine and FumigationStations under the Directorate of Plant Protection, Quarantine andStorage undertake quarantine processing and clearance of thefirst 2 categories. The National Bureau of Plant Genetic Resources,being the nodal institution for germplasm exchange has beenauthorised under PFS Order to undertake quarantine clearance ofthe materials under the third category. Director NBPGR, NewDelhi, has been empowered to issue import permit for germplasmmaterials imported for research use in the country both by publicas well as private sectors. For this purpose, NBPGR has a separateDivision of Plant Quarantine having well trained scientific andtechnical staff in the disciplines of Entomology, Nematology andPlant Pathology, very well equipped laboratories and insectproofscreen houses at the headquarters in Delhi and its regional stationat Hyderabad. These facilities are being further strengthened withthe addition of modern equipment and environment controlledgreen houses at Delhi, Hyderabad, Bhowali and Kanpur.Constraints(a)Lot of material may be entering the country asaccompanied baggage and through international mailwithout quarantine check.

52 Ram Nath(b)(c)(d)(e)PFS (amendment) Order, 1991 of January 20,1992 allowsimport of plants, fruits, seeds and any material of plantorigin for consumption without import permit andphytosanitary certificate. Under the pretext of'consumption' small consignments of planting materialmay be imported without quarantine check.Bulk import of materials for planting purposes posesserious risk as thorough examination and treatment isdifficult.The New Policy on Seed Development provides forpost-entry-isolation growing for cuttings and sapJingsof flowers, however, no post-entry isolation growing isrequired for tubers, bulbs, rhizomes etc. which carry farmore risks.Under domestic quarantine, only eight pests/ diseaseshave been covered, while there are number of otherserious pests which are s~ill localised and not coveredunder the regulations. Even in the case of these eightpests, domestic quarantine is not being enforced,properly with the result that these pests continue tospread to new areas/states in the country.SuggestionsSome suggestions for improving the plant quarantinefunctioning in the country are give!l hereunder.(a)(b)A separate 'plant quarantihe declaration card' like theone for customs declaration should be introduced forpassengers visiting India for declaring any seed/planting material brought as accompanied baggage andall such materials should be subjected to quarantinecheck.All incoming international mail (packets/envelop.es)should be passed through some kind of scanningdevice to intercept seed material as is done for securitycheck. .

Plant Quarnatine Regulations and their implementation in India 53(c) Bulk import of planting materials. is required t~ bediscouraged as far as possible and if consideredessential, these should be imported from seed companiesreputed to produce planting materials under strictphytosanitary conditions. .(d) . For effective implementation of domestic quarantine,State Departments of Agriculture/Horticulture shouldhave separate plant quarantine wings/units.(e)Most problems related to plant quarantine functioningin the country are due to lack of proper appreciationof the risk involved with introduction of exotic pestsand the damage they could do to Indian Agriculture-including their adverse effect on the export of agriculturalproducts. There is ,therefore, an urgent need for anaggressive mass education programme through press,electronic media and other publicity channel~. foreducating the farmers, scientists, customs, postalauthorities and all others concerned with theintroduction/ movement of plants and planting materialsabout the need to observe plant quarantine requirementsin the best interest of the country.

•Medium and Long-term Storage of Seed Materialin Gene BankR.K. SaxenaIntroductionThe Indian region is well known for its native wealth of plantgenetic resources with over 800 species of ethnobotanical interest.It has earned a prominent place among the recognised centres oforigin of cultivated plants and the areas of crops diversity of theworld.' India has played a lead role in recent years among thedeveloping nations for scientific and safe conservation as well asaccelerated utilization of PGRs by developing a capable system forthis purpose and accepting responsibility for coordinating suchefforts for the Asia and Pacific Region. Indian initiative has alsosucceeded in evoking keen interest in this subject among SAARCand G15 .developing countries who are now pooling their knowhow as well as other resources for adopting a regional strategy andcoordinated action plan for conservation, inventorisation, evaluationand sustainable use of plant genetic resources.The conservation of plant genetic resources is both safest andcheapest through seeds. It is important ,first to understand thestorage physiology of the seeds to be conserved. The seeds can beclassified into two distinct groups according to their storagephysiology. The first group is described as 'orthodox'. It includesthe majority of arable and horticultural crop species. In these cropsthe seed aging and loss of viability occurs as a function, not onlyof time, but also of temperature and moisture. Thus the storagelife of these seeds can be increased if moisture content andtemperature are reduced.

Medium and Long-term Storage of Seed Material in Gene Bank 55The seeds of many important tropical plantation crops, suchas rubber, cocoa, coconut, many tropical fruits and a number oftimber species of both the tropics and of the temperate latitudesfall into the second group known as the 'recalcitrant' seeds. Theseseeds suffer subcellular damage and cosequent loss of viabilitywhen dried. Moreover, lowering the temperatures to lOoC causeschilling injury and cooling to sub zero temperatures causesfreezing injury in these seeds. In comparison to orthodox seeds,therefore, the storage life of recalcitrant seeds cannot be easilyincreased by manipulating the storage environment.Indian National Gene BankThis national facility for long term ex-situ conservation ofbase collections, set up by NBPGR at New Delhi, has four majorcomponents:a) Seed Repository: Samples of orthodox type seeds aredehydrated at 15°C and 15% RH to around 5% moisture content,sealed in laminated aluminium foils under vacuum and depositedin cold storage at -20°C.b) Tissue Culture Repository: Disease-free tissue cultures ofrecalcitrant seeds (those not capable of withstanding dehydrationand very low temperatures) and vegetatively propagated plantspecies, developed from meristem explants, are maintained usingappropriate growth-limiting media and by subculturing at suitableintervals.c) Cryopreservation facility: Seed samples of selected species'synthetic ~eeds' embryos and gametes are being preserved usingcryopreservation technique, either by immersion in liquid nitrogen(-196°C) or by keeping in its vapour phase (around -150°C)following protocols using computerised freezing-thawing ratesand chemical cryoprotectants.d) Clonal Repository: Germplasm collections of vegetativelypropagated crops like turmeric, ginger, sweet potato find bananaare being maintained under field conditions following suitablecrop rotations and backed by tissue-culture safety duplication.Gene 'Bank Standardsi) Seed sample size: About 3,000 viable seeds in case of selfpollinated crops and twice that number for cross pollinated.

56 R.K. Saxenaspecies. One third of this sample is sealed in a separate packet andstored alongside the original sample under identical conditions(at -20°C) for monitoring germinability at regular intervals.ii) Viability testing: Initial germination tests are carried out ona minimum of 200 seeds drawn randomly from the originalsamples. Acceptable initial germination percentage should behigher than 85% for most species but lower levels are acceptablefor cotton and wild forms. 50-100 randomly drawn seeds are usedfor subsequent viability monitoring tests.iii) Seed drying: Seeds are dried in seed drying cabinetsoperated at 15°C and 15% relative humidity using silica gel asdesiccant based on adsorption system and appropriate air flow.iv) Seed storage conditions: Seeds dried to 5% moisture contentand sealed in laminated aluminium foils. Base collections arestored in modules operating at _20DC. Active collections are storedat 4 a C and 35% relative humidity.v) Regeneration: Undertaken when seed viability falls to 10%of the initial value. 100 plants or mme'-are ·preferred for thispurpose: ~ " .",-,'Seed Storage(a) Short Term Storage: The storage period for such seeds isbetween 1 year and 18 months. In the arid and cool agro-climaticzones, seeds of most cultivated species will maintain a highgermination capacity for as long as 18 months with only the baseshell of a storage. However, in warm and humid environments theviability loss is very rapid in the simple seed warehouse. Also,seeds of soybean, cotton, onion and several flower and tree specieslose germination capacity rapidly under warm humid storage andmay deteriorate in periods as short as 2 or 3 months in commonstorage under these climatic conditions. Therefore, in warm andhumid areas, and for seeds of these species, more substantialstorages are needed. The first requirement is some insulation tokeep the storage as cool as possible. One possibility is a false ceilingwith,ventilation between ceiling.and the roof. Heat inflow will bereduced by thick stone or brick wall or by a layer of insulation inwalls or ceiling. Ventilation fans to bring in cool night air can help,if it is not humid as to raise the seed to moisture contents to a levelthat allows attack by storage fungi.

Medium and Long-term Storage of Seed Material in Gene Bank 57The second requirement is to keep the seed dry. If bags areused, storage on pallets will keep the seed "from direct contact witha damp floor. Sealing the floor against moisture penetration is alsouseful. Storage of seeds in steel bins with tight fitting lids or inmoisture proof bags will solve the problem of moisture penetrationprovided the seeds are already dry enough for sealed storage.(b) Medium Term Storage: The siz~ of the accessions kept inthe medium term are generally larger than those meant for longterm storage. These working collections have higher rate of usageas they are distributed regularly for evaluation and breedingpurposes. The accessions have to be regenerated periodically dueto depletion of stocks, rather than hom loss of viability. Therefore,the time period for storage is about 5 to 10 years and the pressureof extending the longevity as for the base collections is not existing.The active collections are stored at temperatures ranging from O°Cto 10 e e. The seed samples in these collections are stored in varioustypes of containers such as cloth bags, metal cans or glass jars. Ifthe containers are hermetically sealed after drying the samplesthen the relative humidity of the medium term storage is notimportant . If the unsealed containers are used then the relativehumidity should be brought down. This shall, however, increasethe running costs. The method adopted, thus shall be decided bythe frequency of withdrawal of the samples ( A temperature of SOCand 35~40% RH is adequate for maintaining the viability of mostorthodox seeds for 5-10 years).Long Term StorageThe base collections and duplicate base collections aregenerally kept in stores where the strict storage environment aremaintained.The conditions recommended by IBPGR (now IPGRI) forstorage of base collections are probably most suitable andeconomical to maintain by conventional mechanical refrigerationmethods. The preferred standard for base collections is storage at5% ± 1 % seed moisture content in hermetically sealed, moistureproof containers at -20°e. In fact -2oDe is an arbitrary temperature.The 1976 Working Group ofIBPGR suggested that the temperaturestandard could be relaxed to -lOee if the seed bank was restrictedto a few species with good storage characteristics such as the

58 R.K. Saxenacommon cereals. Thus any store which operates at -IOcC is alsoreferred as a long-term store. Present indications suggest thataccessions stored at -IO°C will require monitoring and regenerationtwice as frequently as those at -20°C. Consequently it isreconunended that new facilities should adopt -20°C.Processing of seed for conservation at NBPGRThe NBPGR receives exotic and indigenous germplasm ofagri-horticultural crops (orthodox seeds) from different sources forlong term storage in the National Genebank. Before it is finallystored! conserved, it passes through following channels:1. Registration: Seed received from different sources, arechecked and duplicate samples rejected. Only new arrivals are,registered. The accession number once assigned is not re-assignedeven if seed viability during storage is lost.2. Seed cleaning and purity analysisi) Seeds are inspected and under-sized, shrivelled,immature seeds are screened out, 'ii)iii)iv)Broken and damaged seeds are rejected,Diseased seeds, insect attacked and seeds affected by. nematodes are rejected andGrass and cereal florets, empty glumes, lemmas, sterileflorets of grass and cereals are rejected; weed seeds arealso separated out. Samples of the clean seed lot aresubjected to initial seed viability tests (by germinationtests) following 1ST A guide lines. Seed samples showingviability standard of 85% or more are accepted.3. Seed germinationi) Rolled towel test: Generally medium and large seeds suchas those of c~reals, maize, grain legumes, oilseed crops (sunflower, 'safflower etc.) and vegetables (okra, cucurbits etc.) are tested forge1,"mination by this procedure. Towel papers are spread on topof each other and then soaked in water. A waxy paper is keptbelow the towel paper. On rough side of waxy paper, date ofexperiment and accession number are written. Normally 25 or 50seeds are plc.lll'd on top of the uppermost sheet in a fairly regular

Medium and Long-term Storage of Seed Material in Gene Bank 59pattern so that they are ~pproximately equidistant. Another towelis then placed on top of the seeds. The basal 20-30 mm portionof the longer edge of all towels are turned up to form a lip toprevent the seeds falling out.The towels are then gently and loosely rolled to form a tubeof about 50-60 mm and tied with rubber band and kept ingerminator at a desirable temperature and period. The towels arerolled loosely, (tight rolling would prevent air circulation betweenthem and result in either no germination or abnormal germination)and are placed in the germinator. Where germinator maintainshigh humidity, the rolled towels can be held with rubber band andkept upright in cages within the germinator. If the germinationtest is to be conducted in incubators where relative humidity is notcontrolled, water is sprayed over the towels every 2 to 3 days.ii) Filter paper: Normally 9 cm diameter filter papers are used.In some cases, larger and smaller filter papers and petri dishes areused. The small seeded crops e.g., tomato, brinjal, chillies, Brassicaspecies etc., are subjected to germination test in petri dishes. Thefilter papers are soaked in water and placed in the petri dish.Normally 2 filter papers are required for each dish. Seeds areplaced on top of the filter paper and accession numbe! can bewritten on the petri dish lid by a glass marking pencil. Petri dishesare then kept in the germinator for recording tl:e viabilitypercentage of the accessions.iii) Agar media: It is an alternative to routine germination test.Agar.is a polysaccharide complex, which though insoluble in coldwater, dissolves slowly in hot water to a viscid solution. NewZealand agar or Japan agar is suitable for germination test. Oneper cent agar solution forms a stiff jelly on cooling, on to whichthe seeds can be placed for the germination test. For example, ina germination test at 20°C, the agar remains moist for 2 weeks orso, the initial seed imbibition environment may be more favourablefurther than on in agar paper substrates and potential problemsfrom imbibition injury can thus be reduced.iv) Sand media: The sand consists of uniform particles and havea pH between 6.0 and 7.5. Before use (i) the sand should be sievedwith 0.8 mm diameter holes, the sand particles which pass throughthe sieve are used and (ii) must be sterilized at lS0°C for 2 hours.During heating, the sand must be turned regularly in order to dry

60 R.K. Saxenait uniformly. Sufficient quantity of water is added to aluminiumor plastic dishes (containers) filled with the sterilized sand andseeds are sown in rows on top of the sand and then covered with10-20 mm loose sand. The dish is covered with flat plate, andplaced in the germinator. The dish is labelled with properaccession number. Those accessions having more than 85 per centgermination are processed for drying.v) Seed drying: Each accession is placed either in porous paperbags or muslin cloth bags. Seeds are dried at low relative humidityand at low temperature, i.e., 15 per cent RH. and 15°Ctemperature. Under these conditions, seeds dry within 10-15 days.When seed attains 4-6 per cent moisture, they are packed inaluminium foil and hermetically sealed. These packets are readyfor storage in genebank. All the information pertaining to thegermplasm stored (long term), is properly documented and thecomputerized database is available to the users for their breedingprogramme.Selected ReferencesBass, L.N. (1978). Principles and practices of seed storage.Agriculture Handbook No. 506, U.S. Department of Agriculture.Frankel, D.H. and Soule M.E. (1981). Conservation and evaluation.Cambridge University press, Cambridge.IBPGR (1985). Cost effective long-term seed stores. InternationalBoard for Plant Genetic Resources, Rome.Khanna, P.P. and Singh, N. (1991). Conservation of plant geneticresources. In: Plant genetic resources conservation andmanagement. (Eds. Paroda, RS. and Arora, R.K.) IBPGRpublication.Rana, RS. and Chandel, K.P.S. (1992). Crop genetic resources. In:Sustainable management of natural resources (Eds. Kosho, T.N.and Manju Sharma). Malhotra Publ. House, New Delhi.

IIPromising Introductions and Prioritize Needs ofExotic Germplasm : Cereals and MilletsMansoor KazimIntroductionCereals and millets are basic constituents in our dietaryhabits. The green revolution brought into the country with theintroduction of Mexican wheat and Taiwan paddy is wellrecognized. However, the scope of improvement of our. productionin these crops for home consumption and export is stj.ll restricted.In order to boost up this there is need for continued efforts forimproving the existing germplasm/ cultivars by introducing diversetype of materials as donors which could be profitably utilized inhybridization and crop improvement programmes.In order to provide high yielding, disease resistant, tolerantto both biotic and abiotic stresses, the division of GermplasmExchange at the National Bureau of Plant Genetic Resources havebeen making all possible efforts, since 1976, inception of thisBureau, to import such mate'rials and distribute to researchers inthe country for utilization in thei.r research programmes. Theapproach is two way, one by scanning the world literature andpicking up promising materials for utilization in the country by theBureau and second by meeting the requests made by theresearchers, their specific need oriented. Thus, the decade afterNBPGR's creation (1976-85), 7,29,390 accessions, both germplasmand trial materials have been introduced. This activity has beencontinued since then and each year some 50-56,000 collections havebeen added bringing the total imports to 12,12,757 till end of 1994.

62 Mallsoor KazinzEach year diverse materials, possessing economic traits,disease resistance, wide adaptability and stress tolerance wereintroduced. These promising introductions in each group of cropsare indicated below:I. Promising IntroductionsWheat : Semi-dwarf, oligo culm spring type bread wheat (EC122043), dwarf oligoculm with branching habit (BC 122044 exIsrael), Sabergeg (EC 124298), Ajiba 69 (EC 124299), Mexi-Pak (EC-124300) all from Iril-q. Livorne-WIR 5841 (EC 130595) andRumanico-WIP 5862 with high protein content and cold resistanttypes from USSR, Fortune (EC 133489), hard red grained springwheat with stem solidness and male sterile line (EC 137254) fromUSA. Wialki CN 8242 a variety introduced from Australia derivedfrom a cross involving Madden x Boral is a hard, red grained type,superior in grain quality and possess resistance to Urocystisagropyri and Puccinia graminis race Sr 13. Sukota-N-82 (BC 158114ex Australia) is yet another variety derived from Tenson x IRN 67-451 (North Dakota), a mid season type with good quality andresistance to prevailing field races of Puccinia recondita andUrocystis agropyri.BC 183950-57 from UK having yellow rust resistance, EC258472 from Canada - a rust resistant line, cold tolerant, hardgrain/red grain typed BC 278274.:321 from USSR, BC 328914 and15 with PM II gene for resistance to powdery mildew from Japan,differentials for Helminthosporium leaf blight (BC 336670-73) fromBangladesh, EC 368167-69 male sterile and restorer lines fromFrance, were promising.Among hard wheat (Triticum durum), promising typesintroduced were Coast C-71 (BC 124293), Senator Copella (EC124294), Jwice-6 P (EC-124295) and Sin-AI-Jamel (EC 124296) allfrom Iraq, besides resistant sources to Erysiphe graminis (exCanada), isogenic lines resistant to powdery mildew and yellowrust from USA. Strains resistant to nematode (Heterodera avenae) areEgyptian Cl-12345 (BC 170660), Aus-Po 894 (Ex Denmark),powdery mildew resistant accessions chul/SCC-CCI 41120, CI1121, CI 12632, Transec CI (EC 159057-60) all from USA and springwheat (WWI46-73) from USSR with high number of grains/spike,and EC 198033-37 (T. compactum) also from USSR. Aegilops spp. BC299344-355 from Syria representing 12 different species.

Promising Introductions and Priortize Needs of Exotic Germplasm 63Barley: Stripe rust and powdery mildew resistant line BC 123304'A' from USA, tetraploid variety EC 137248 and diploid EC 37249from UK, dwarf accession with high yielding traits (EC 138742)from Japan, Ee 138784 (ex France) and Ee 141803 (ex USA) are ofpotential value. Oweolbosi Milyang 16 (Be 151529 ex Korea) andH. distichon var. Norhest (Ee 152099 ex Canada) male sterile lineCI 13446 (EC 165103) offer useful genetic variability. Hertizpossessing resistant genes for rust from Denmark. Cytoplasmicmale sterile lines and restorer lines 9EC 322739-41 ex Finland),bacterial leaf spot resistant lines (Ee 328936-39 ex USA). AmongstHordeum spontaneum early maturing, cold tolerant lines Be 339893-944 from Syria and early heading, high yielding and lodgingresistant, suitable for malting industry types Ee 363845-71 fromSyria.Oats: Swan (Ee 102649) high yielder, Irwin (Ee 102650), aXB 164(Mulga x Daler) x Fulghum) x Orient) x Avon (Ee 102653) all fromAustralia provided broad genetic base. Ee 120119, 120744 and Be121448 al ex USA are promising genetic stocks. Species collectionsEe 130444-61 made from Canada, high protein content accessionsEC 163087-94 from USA.Rye : An introduction EC 338685 from USA white seededpossessing aluminium tolerance is promising line.Rice: Cold resistant line (Ee 121003 ex Philippines); blast resistantcollections Be 121807-823, stem borer resistant donors (Ee 121824-25), leaf folder resistant accessions (EC 121828-53) and salinitytolerant types Ee 122934-4{) all from Philippines, Oryza speciescollections BC 127797-801 from Czechoslovakia. Good cookingquality variety Akibare (BC 131474), upland waxy type Hatsukaori(Ee 131484) short stature and high panicle number/plant, Kinfia(Ee 131489) with good grain quality, high adaptability and Roimni(Ee 131510) short statured type from Japan hold promise. Blastdifferential varieties (Be 161320-46), Tungro virus, brown hopper,and stem borer resistant accessions from Philippines, Ee 178772-9100 also from Philippines have tolerance to brown plant hopper,soil stresses, cold tolerance, blast resistance and submergencetolerance. EC 199447-54 BPH resistant lines, 199455-61 bacterialleaf blight resistant lines, EC 199465 resistant to sheath blight, BC199466 resistance to stem borer, Ee 199581-712 aromatic varietiesand BC 201878-975 sheath blight resistant, leaf blight and drought

64 Mansoor Kazimtolerance, all from Philippines. EC 232709-909 BPH resistant linesderived from Oryza officinale crosses, ec 232900-910 salt tolerantlines, EC 232912-15 drought tolerant types, EC-268456-931pofosessing disease resistance togetherwith adaptation to rainfedconditions and submergence tolerance. Salinity tolerant materialEC 286620-965 and 18 different wild species Ee 297481-569 fromPhilippines. EC 315248-63 cytoplasmic male sterile lines andmaintainer lines from Philippines. Ee 334331-43 salinity tolerantlines and Ee 334937-47 lines insensitive to delayed transplantingand photoperiodism, Ee 347769-72 cold tolerant lines, Ee 365154-58 cold tolerance donors and Ee 361662 tolerant line to acidsulphate conditions besides many disease resistant types anddonors. .Maize: Early maturing variety Ee 119433-37 from Mexico and coldresistant types Ee 121736-42 from USSR, Taiwan, white a syntheticvariety Ee 123768 ex Thailand, male sterile lines EC 123862-90 exUSA, maintainer lines Ee 123891 ex USA, Kaneja 470 (EC 155548)from Bulgaria. HI02 (Ee 151581) with yellow grains, dent type,involved line, resistant to races of HeZminthosporium turcicum,Colletotrichum graminicola and Peronosclerospora sorghii and H-l11(EC 151782 ex USA) also resistant to Helminthosporium turcicum(race 122), Collectotrichum graminicola, leaf blight and stalk rot arepromising genetic stocks. EC 191782-83 cold tolE::rant lines fromUSSR, EC 180027 photoperiod insensitive collection from USA,early white semi-dent type Ee 201528 ex Mexico, EC 260679-730white grain types with tropical adaptations, some lines possessingresistance to transport disease (Phyllt;lcora spp.). EC 275542-47possessing resistance to downy mildew and insects from Thailand.Ee 343371 -90 ex Thailand resistant lines for stalk rot. Ee 361476,.78 ex Thailand having resistance to downy mildew and alsoresistance for maize streak virus. EC 369553-68 early yellow - whitegrained hybrids.Sorghum: Species collections (ex USSR) representing S. chinense,S. guinanse, S. cantuorum, S. durra. Sorghum midge resistant line(EC 131437-73), male sterile line Ee 131994, EC 131997 restorer linewith high grain yield all from USA. EC 191789 ex Australia a highlysine content line. Drought hardness type Ee 242786-91 ex Sudan.Acid soil tolerant germplasm EC 315823-52 and EC 331138-48'possessing resistance to army worm and anthracnose all fromUSA. .

Promising Introductions and Priortize Needs of Exotic Germplasm 65ProsomiIlet: An elite accession BC 129 ex USA and several otherpromising collections were introduced from Australia, Italy andUSSR.Fingermillet, Barnyard millet and Foxtail millet : A sizablecollection has been introduced from Syria, USA and Malawipossessing useful traits togetherwith high yields ..II.Priortize need of exotic germplasmThe need for exotic germplasm has already been said earlier.It is much required to incorporate the desirable characters in theexisting germplasm and to pick up those selections for wide useas varieties in crop improvement programmes. These could alsobe utilized as direct introductions as had been in the past Ridleyand New Algerian, and Kent Oat varieties etc. However, thepriorities of particular germplasm is largely need based ofresearchers in the country, recommendations of annual cropworkshops, mini missions on pulses and oilseeds and suggestionsof different crop advisory committees. Some of these, could beindic2!ted below:Wheat:1. Heat tolerant types suitable to warm areas fromICARDA.2. Material suitable for moderate rainfall, low rainfall bothin T. aestivum and T. durum from ICARDA.3. Materials possessing resistance for leaf rust, yellowrust, and stem rust and common bunt (Tilletia foetida)and aphid resistant types from ICARDA and CIMMYT.4. Heat stress types adaptable to warmer areas of countryfrom CIMMYT.5. Drought tolerant types suitable for acid regions ofcountry from CIMMYT.6. Types suitable to high rainfall areas from CIMMYT and7. Spring wheat nurseries for selections of adaptablematerial to Indian conditions from CIMMYT.

66 Mansoor KazimBarley:Maize:1. Early maturing varieties from CIMMYT2. Materials suitable to low rainfall areas and moderaterainfall areas from ICARDA.3. High yielding malting types from ICARDA.4. Naked types suitable for wide cropping areas fromICARDA.1. Introduction of populations for commercial exploitation.2. Introduction of inbreds for production of elite hybridssingleand double crosses.3. Introduction of gene pools/genetic stocks for specifictraits - earliness, leaf blight resistance, downy mildew,corn borer from CIMMYT and other countries - SouthAmerica and Asiatic countries.4. Introduction of vqrieties possessing tolerance towater-logged areas/submerged areas.5. Introduction of sweet corn and popcorn types forcommercial utilization in industry and6. Germplasm possessing resistance to common diseases,and pests to be utilized as donors in crop improvementprogrammes.

Use of Tissue Culture Techniques in Exchange andUtilization of Plant Genetic ResourcesS.R. BhatIntroductionPlants have moved and established themselves far and widefrom their original home aided by both biotic and abiotic agencies.Trade and colonization have greatly helped dissemination ofplants between continents during the past couple of centuries. Forexample, Polynesian voyagers carried and established 27horticultural plants (mostly vegetative propagules) while inhabitingHawaiian islands (Bevacqua, 19.94). Plant introductions havedramatically altered the economy and agrit;::ulture of manycountries. Such historic plant introductions include African oilpalm, rubber, coffee, cinchona, tea etc. (Plucknett et al., 1987).Thanks to free movement of plant species, major world producersof several important crops are now located in places outside thecentres of origin of those crop species.Success of plant breeding based on exotic germplasm duringthe past fifty years has highligl1.ted the importance of plant geneticresources. The dwarfing genes of wheat and rice that ushered ingreen revolution came from obscure exotic introductions, Todayplant genetic resources are regarded as most important forsustaining future food supply. Thus from introduction of new cropspecies as done in the past the focus has now shifted to exchangeof genetic diversity of crops already under cultivation, Tremendouspossibilities of germplasm utilization being made possible bymodern biotechnological innovations have vastly increaSied the

68 S.R. Blzatscope of genetic resources. Thanks to these techniques, geneticmaterial from any organism can now be a potential source for cropimprovement.Germplasm introductions in most countries are regulatedand controlled by government agencies. These regulations aremeant to facilitate safe introduction of germplasm and to avoid/minimise the risks of introduction of unwanted material, weeds,. pests, pathogens etc. Genetic engineering and patenting of genes/crops have, of late, led to conflict about ownership of plant geneticresources. The transfer of genetic resources in future is expectedto be more stringently controlled and proper documentation ofgermplasm exchange/movement will be crucial in settlingownership disputes.Exchange of germplasm involves the following steps:1. Placing an indent with proper documents2. Preparation of material3. Examination and issue of phytosanitary certificate4. Packing and transport5. Receipt of the material and quarantine examination6. Establishment and multiplication7. Release to the indentorGermplasm exchange is needed for research use, cropimprovement and/or for commercial planting. It may be in theform of DNA/RNA, celt tissue, organ or complete plantlets.Depending on the crop and the nature of the materiat supply maytake anytime between three months to three years.Seeds are the main forms of germplasm exchange. Cropspecies with orthodox seed generally pose no problem. In contrastexchange of vegetative propagules (suckers, bulbs, tubers, cuttingsetc.) and recalcitrant seeds require great care and is fraught withserious risks. Tissue culture techniques can help in efficientexchange of germplasm in various ways. These techniques whenproperly employed cart. reduce quarantine risks, save time,facilitate transport and handling, and improve establishment andutilization of the germplasm. Already germplasm of importantcrops such as banana, potato, sweet potato, yam, strawberry,

LIse of Tissue Culture Techniques in Exchange 69blackberry, grapes etc. are routinely exchanged as in vitro cultures.Use of tissue culture techniques at various steps in germplasmexchange and the resulting benefits are briefly discussed below:1. Preparation of material : Availability of material for supply isoften limited and it usually takes a year (for annuals) before theindent can be honoured. In perennial crops it may take evenlonger. In many c10nally propagated crops multiplication rate isvery low (5-10 times per year) and may require special skills andresources. Recourse to tissue culture can alleviate this problem.Rapid multiplication through in vitro culture is already beingpracticed on a commercial scale for many important crops.Further, tissue culture multiplication has been reported for over1000 plant species.Since tissue culture is carried out under controlledconditions, material can be multiplied at any time of the year. Thus,rapid multiplication through in vitro culture can save time andmake material available in adequate quantity to the indentor.2. Assistance in quarantine. Exchange of germplasm alwayscarries the risk of introduction of pests, pathogens, weeds etc.These may be present as mixtures (e.g., weed seeds) or borneexternally or internally. There are numerous instances ofaccidental introduction of unwanted organisms during germplasmexchange. Whereas seed material is more easily accessible forquarantine inspection and sanitisation h'eatments, bulky vegetativepropagules· (suckers, corms, bulbs etc.) and plantlets are difficultto screen effectively. Undesirable organisms have been known toescape through quarantine net especially in materials with soil ,roots, or external coverings (sheaths, husks).Tissue culture involves surface sterilization and cultureunder aseptic conditions. Almost all insects, pests, bacteria andfungi borne externally are eliminated during surface sterilization.Since tissue culture medium contains sugars, vitamins andnutrients, bacteria and fungi, if present, will grow and show upas contaminants in the medium. Similarly, presence of insects andnematodes can be easily detected. Thus when clean. cultures areestablished and used for exchange the risk of introduction ofunwanted organisms is greatly redu.ced. Q~y ()bligate parasitesare giffkult, to detect in, in vitro c.ultures and need speCial attention.

70 S.R. BhatQuarantine procedures for in vitro culture and exchange ofgermplasm are well established for important crops such aspotato, banana, sweet potato, yams and can be adopted for othercrops as well (FAO/IBPGR, 1989).3. . Salvaging diseased material. As stated above, in vitrocultures are usually free from bacterial and fungal contaminations.Even plants infected with viruses and other obligate parasites canbe salvaged through 'meristem culture'. Although White (1934)recorded for the first time the absence of viruses in culturesinitiated from root apices of infected material, it was Morel andMartin's (1952) work with shoot apex culture of dahlia whichdemonstrated the significance of 'meristem culture' for viruselimination. The exact reason for' the absence of viruses inmeristematic zone is unclear. However, meristem culture techniqueis now well established and success has been reported in over 100plant species. Even seed borne virus (pea seed borne mosaic virus)has been eliminated through' meristem culture (Kartha andGamborg, 1978). A modification of this technique called 'in vitro,'_l!P~~9g!~ging~. w"a~ ._g~vi~ed by Navarro .. ~t. al . . (197~) for' theelimination of viruses in citrus .. which is currently in wide use.,_, ...For most of the problem crops ,meristem culture is now usedto establish clean cultures. The plantlets derived from meristemculture need to be tested for ascertaining virus-free status. Thesuccess rate may range from 10-80%. However, once the materialis cleaned it can be rapidly multiplied and can be maintainedindefinitely in disease-free state through tissue culture.4. Assistance in transport : Recalcitrant seeds and vegetativepropagules (cuttings, budwoods) are highly perishable and aredifficult to transport over long distances. Similarly, bulbs,suckers, grafts and seedlings are delicate and need excessivepacking and care during transport. Such bulky materials aredifficult to handle and hence limit exchange by restricting thenumber of samples and also the nU:p1ber of replicate~. By resortingto in vitro culture, recalcitrant seed species and clonal materialscan be easily transported. Over 200 culture tubes (each with3-4 shoots) can be packed in 0.75 cubic feet (approx.O.02 m 3 ) boxand will weigh less than 5 kg. In vitro cultures can withstandrigors of transport and survive at ambient temperatures (IS-30°C)for 3-4 weeks. Even in germplasm collection in vitro techniques

Use of Tissue Culture Techniques in Exchange 71are being employed. For example, efficiency of collection ofcoconut and cocoa germplasm was greatly improved by using invitro techniques (Withers, 1987). Tissue culture techniques canthus reduce the bulk and make handling and transport ofgermplasm more simple and efficient.5. Better product quality through in vitro culture : Seedexchange is often resorted to in perennial tree crops owing todifficulties in conventional clonal multiplication and transport, andalso to minimise quarantine risks. However, in heterozygousplants one is often interested in introducing spedfic, elitegenotypes. (e.g., female plants in monoecious species). By tissueculture multiplication unique genotypic constitution can bemaintained and,made available to the indentor. Similarly, insteadof budwoods or cuttings, rooted plantlets ready for planting in soilcan be supplied thus obviating the need for special skills at thereceiving end. Furthermore, in vitro production of storage organslike tubers, bulbs, rhizomes etc. can help in efficient exchange andestablishment of germplasm.6. Tissue culture at the receiving station/quarantine station: Atthe receiving end also tissue culture techniques can be employedfor salvaging and mass multiplication. Samples of in vitro materialcan also be conserved in tissue culture to meet future demands.This will reduce repeated introduction of the same material andreduce workload on exchange and quarantine: services.7. Rejuvenation and establishment: Poor germinability of seedsamples is a common complaint in germplasm exchange. Lowgerminability may be due to several factors such as dormancy,hard seededness, low viability or vigour etc. So long as embryo isviable, one can hope to recover complete plants through tissueculture. Recently, by embryo culture, Dilday et ai. (1994) were ableto recover a11126 accessions of rice, stored for more than 19 years,that failed to germinate in the field and in standard laboratorygermination tests. Tissue culture can be particularly importantwhen introducing rare germplasm or when sample size is verysmall (e.g., wild species).8. Tissue culture and management of germplasm exchange:In vitro cultures of tropical, subtropical and temperate crops canbe maintained and multiplied under a single roof. Hence overall

72 S.R. Bhatmanagement of germplasm conservation and exchange can beeffectively handled from a single window.Concurrent with developments in genetic engineering andcell genetic manipulation the range of material being exchangedhas also increased. Specialised cell lines giving high yields ofsecondary metabolites, genotypes with unique features are beingdeveloped and used in research and production. Tissue culture isindispensable in the exchange of such germplasm.Thus, the need for in vitro techniques in germplasm exchangeranges from desirable to essential. It is needless to emphasize thatadequate prior research is necessary to develop suitable protocolsfor in vitro culture and plantlet recovery in various crop species.General guidelines for in vitro exchange of germplasm1. Vegetatively propagated plants, as far as possible,should be exchanged through tissue culture.2. Use proliferating shoot cultures or rooted plantlets forexchange. (Avoid callus where stability is important).3. Cultures for exchange should be raised on semisolidmedium (agar or gelrite) and should not containcharcoal or antibiotics.4. In vitro cultures raised in plastic pouches or tubesshould be used for exchange.5. Only disease-free and certified germplasm should beexchanged.6. Information about media for proliferation and rooting,hardening procedures etc. should be provided whileexchanging germplasm in the form of in vitro cultures.7. Proper communication about shipment and expectedarrival time of material be ensured to avoid loss ofgermplasm.8. Proper labelling and directions for handling and storageof package during transit should be provided.

Use of Tissue Culture Techniques in Exchange 73ReferencesBevacqua, RF. (1994). Origin of horticulture in Southeast Asia anddispersal of domesticated plans to the Pacific islands byPolynesian voyagers: The Hawaiian Islands case study. Hart. 'Science 29 : 1226-1229.Dilday, R.H., Yan, W., Lee, F.N., Helms, KS. and Huang, F.H.(1994). Application of embryo rescue in recovering rice (Oryzasativa L.) germplasm. Crop Science 34 : 1636-1638.FAO/ffiPGR (1989). Technical Guidelines for the Safe Movementof Germplasm. FAO /IBPGR, Rome.Kartha, KK and Gamborg, O.L. (1978). Meristem culture techniquesin the production of disease-free plants and freeze-preservationof germplasm of' tuber crops and grain legumes. In: H.Maraite and J.A. Meyer (Eds.). Diseases oJ Tropical Food Crops.Univ. Catholique, Louvian-Ia-Neuve, Belgium. pp. 267-283.Morel, G. and Martin, C. (1952). Guerison de dahlias atteints d'une ,maladie a virus. C. R. Rebd. Seances Acad. Sc. 235 : 1324-1325.Navarro, L., Poistacher, C.N. and Murashige, T. (1975).Improvement of shoot tip grafting in vitro for virus-freeCitrus. J. American Soc. Hort. Sci. 100 : 471-479.Plucknett, D.L., Smith, N.J.H., Williams, J.T. and Mum Annishetty,N. (1987). Gene Banks and World's Food. Princeton UniversityPress, Princeton, New Jersey. pp. 41-71.White, F.R (1934). Multiplication of viruses of tobacco andAucuba mosaics in growing excised tomato root tips.Phytopathology 24 : 1003-1011.Withers, L:A. (1987). In vitro methods for collecting germplasmin the field. FAO/IBPGR Plant Genetic Resources Newsletter69 : 2-6.

Promising Introductions in Horticultural Plants(Fruits, Ornamentals) and Oil SeedsB.P. SinghIntroductionThe Division of Germplasm Exchange of the National Bureauof Plant Genetic Resources (NBPGR) over. the last sixteen years(1976-1994) has introduced 3,81,982 accessions of different agrihorticultureand agri-silviculture crops from over 25 countries ofthe world, strictly under phytosanitary conditions. Some of thesenotable introductions have been highlighted earlier (Paroda et al.,1986, Singh et aI., 1989, Singh and Rana 1993). However, abrief account of some of the significant introductions ofhorticultural plants (fruits), ornamentals and oilseed crops havebeen discussed here.'I. Horticultural Plants (Fruits)In fruit plants over 3125 accessions of different fruit plantshave been introduced during the period 1976-1994. These indud~dtropical! subtropical and temperate fruits.(a)Tropical and subtro'pical £rui~s(i) Banana (Musa paradisica) : The cultivar Lady finger (EC:-160160) reported to possess resistance/tole~ance to bunchy topvirus which is a serious problem in banana, had been introducedas tissue culture raised plants from Australia and is underevaluation at IIHR, Bangalore and TNAU, Coimbatore. The otheruseful introductions include cultivar 'Grand Nain MS (EC-27237)from France and 'Valery' from West Indies. In recent years more

Promising Introductions in Horticultural Plants and Oil Seeds 75cultivars have been introduced from ·.Belgium as tissue culturedmaterial for establishment/ multiplication/ evaluation andutilization.(ii) Citrus spp. : Some of the major problems faced by citrusindustry in India are owing to citrus die back, citrus canker, tristesaand greening viruses, nematodes, Phytophthora, waterlogging and'salinity. Some of the commercial cultivars of sweet orange(c. sinensis), blood drange, acid lime, satsuma mandarins, mandarinhybrids and citrus rootstocks resistant to die-back, nematodes, etc.have been introduced from USA, Japan, Thailand, Peru, Portugaland Belgium. In sweet orange (c. sinensis) cultivars Toroceo Blood(USA), Sunramon (Peru), Vainiglia Snguigno (Portugal); in bloodorange eultivars 'Tha' and 'Thai'; in acid lime (c. aurantifolia)cultivar 'Star! Ruby'; in grape fruit cultivar Moga are prominentintroductions.The prized eultivars of Satsuma mandorm (c. unshiu) Macowitchinclude Mihowase, Oktiwase, Miyagawase, Sogiyama, unshiufrom Japan and in mandarin hybrids like Fortune, Page, Wilking,Pixi and others are useful introductions from USA and Japan.(iii) Datepalm (Phoenix dactylifera> : Cultivars Khalasa, Basshi,Naghal and Khaneji are useful introductions from Sultanate ofOman; cultivars Tayar, Hatemi, Mejnaj, Khalas, Ruziz and Khesabfrom Saudi Arabia; cultivars Zaghlool, Samani, Amri, Saklote fromEgypt are promising introductions. The cultivar Khadrawy hasshown resistance to 'Graphiola' leafspot at Hissar.(iv) Fig (Ficus carica) : 52 cultivars have been introduced fromUSA, Iran, UAE and Egypt. Some of the prominent ones are'Genoa white', 0030 Genoa, 009 Flanden, Candsia, 0032 (Adriatic),Mission and others are some of the useful introductions. These areunder evaluation at llHR, Bangalore.(v) Grapes (Vitis vinifera) : The earlier promising introductionsinclude Thompson seedless, Perlette and Beauty seedless fromUSA and Kishmish beli and Kishrnish chorni from USSR. A goodnumber of commercial cultivars as well as Vitis species have beenintroduced from Australia, Brazil, USA, Russia, Portugal andCanada having desirable characteristics such as earliness, betterberry quality, higher sugar content, disease resistance andsuitability for preparation of wine and raisins and for table

76 B.P. Singhpurposes. The pronusmg cultivars like Ruby seedless, Gordoblanco (L1-9/c/Merbein Riesling from Australia; Totiocha fromBra.zil, Flame seedling 1281, Dogridge (Phylloxera resistant), Pride,Dixie, Wedor and Black Corianth-2 from USA. A number of Vitisspecies viz., V. gigas, V. caribea, V. munsoniana, V. smalliana,V.cinerasia, V. shuttleworthi, V. arizonica and V. monticole are usefulintroductions.(vi) Guava (Psidium guajava) : The cultivars like Beaumont G-35, (attractive pulp colour and aroma), 7-12, 9-35, 11-56 andIndonesia seedless are useful introductions from Australia, besidesVerdie and M 25988 from USA and CHN from China. Species P.sartorianum M. 19701, P. littorale var. longipes M. 6505 and P.longipes M. 25181 have been introduced from USA.(vii) Macademia nut (Macademia tetraphylla) : Macademia nutis a cash fruit in Hawaii and Australia. The cultivar Renown (EC-154155 ex USA) and Nultyglen from Australia and cultivar EC122172 and EC 154155 from USA are interesting introductionswhich are under evaluation at lIHR, Bangalore.(viii) Papaya (Carica papaya) : The useful cultivars like Sunrise,HGAR-29, Maradol and Cawflora from Hawaii (USA) and severalCarica species, viz., C. gondotiana , C. cauliflora, C. sphaerocarpa, C.quaciflia, C. monoica, C. pubescess, and C. stipulata, which are sourcesof resistance to diseases / pests and frost are promising introductions .. (ix) Jiicoticaba (Myrciaria cauliflora) : It is a new fruit introducedfrom Brazil/USA, which bears black berries on the main trunk ofthe plant. The cultivar Sabara blanc M. 8962 (EC 183095) from USAand EC 158116, EC 167627 and Be 167628 from Brazil are recentintroductions .. (x) Pomegranate (Punica granatum) : Cultivars like Wonderfulfrom USA, A. Males, A. Be Hastah, A. alah, A. Agha MohammadAli from Iran are useful introductions.(xi) Mango (Mangifera indica) : Since none of the Indian Il').angocultivars possess good colour, one of the major objectives ofbreeding programme is to develop cultivars with good fruit colour,so G\S to boost export of mangoes. Therefore coloured mangocultivars-Tommy, Zilete, Harden sensa tori, Julie and others fromMiami, Florida (USA) have been introduced.

Promising Introductions in Horticultural Plants and Oil. Seeds 77(b)Temperate fruitsNoteworthy cultivars, genetic stocks/roots stocks and specieshave been introduced in apple, pear, peach, plum, apricot, cherry,walnut, almond, pecan, Chinese gooseberry, feijoa and others.(i) Apple: There has been pressing demand for introduction ofbetter spur type delicious cultivars for early cropping, scabresistant and suitable dwarfing rootstock for high density planting.There is also need to have low chilling cultivars for growing atlower elevations or in plains. Amongst the high chilling types,promising introductions include :Spur type red delicious-II (Ee 43974, USA) - a bud spurt of RedDelicious - it is regular and heavy bearer with medium large fruit,with red splashed skin, ripening in the middle of August. It issuited for high density planting.'Red Baron (Be 115820 USA) - a heavy bearer, with medium size,yellow and bright red fruits maturing in August.Mollies Delicious (USA) - has performed well in HimachalPradesh; fruits are large, very sweet, with good keeping quality.In low chilling types - cultivars Vered (Israel) performed well atlower altitudes in Ranchi hills, Delhi and Udaipur. Fruits smallsized, conical in shape, light yellow, green skin splashed with red,soft flesh, ripens in middle of June.Tropical beauty (USA) is another promising low chilling variety,which gave good performance at Ranchi (Bihar ) and in Orissa.Amongst the rootstocks useful introductions include M7, M9,MM 25, MM 26, MM 101 and M 106.(ii) Pear: Amongst the recent introducti

78 B.P. SinghAmongst low chilling types - promising cultivars include Floradsun,Florida Red, Shan-e-Punjab, and Sunred (nectarine) have becomequite popular in plains of Punjab and u.P.(iv) Plum: Cultivars like Santa Rosa and Starking Delicious fromUSA; Methley and Settler from Kenya and Kanto-5 from Japanhave performed well in Shimla hills.(v) Apricot: Amongst the more recent introductions the cultivarNugget (EC 27791, USA) has been found most promising for hills.The other promising introducti,(;ms include cultivar: Edrassel,Karoda Riland from USA, Krishanesky, Silitronska and Roxanafrom Bulgaria.(vi) Sweet cherry: The cultivars Emperor, Francis and Lambertfrom USA have performed well in Shimla hills. Rootstock MazzardF 12/1 has also given fairly good performance. Among recentintroductions cultivars Early Rivers, Vega, Raimier and Corumfrom Norway and Windsor and Napolean from USA are usefulintroductions.(vii) Walnut: Promising introductions are Lake English, Payne,Tutle-31 and ORTH from USA and EC 31071 from Canada whichperformed well in Shimla hills.(viii) Almond: Promising introductions include Neplus Ultra,NonPareil and Drake, which performed well in H.P. and U.P. hills.The other promising introductions include California Papershell,Marcott and Briggs Hardshell.(ix) Pecan: The cultivar Mahan (BC 2455&, USA) proved mostpromising with regard to yield and quality for cultivation in hills.(x) Chinese gooseberry or Kiwi fruit : The cultivars Allison(Ee 24672) and Hayward from New Zealand have performed wellin Shimla. This is a new introduction which is becoming verypopular in hills.(xi) Feijoa or Pineapple gua~a: Cultivar Nikitskp (EC 24526,USSR) an evergreen hedge cum fruit plant, has given goodperformance in Shimla.II.Ornamental PlantsThe Division of Germplasm Exchange of NBPGR (earlierDivision of Plant Introduction IARI) over the last 22 years

Promising Introductions in Horticultural Plants and Oil Seeds 79(1970-1992) has introduced 1250 accessions of different ornamentalplants from over 23 countries of the world under phytosanitaryconditions. Some of the notable introductions have been highlightedearlier (Rathore, 1981, 1985, 1986; Singh, 1986; Singh and Rana,1990; Singh et al., 1993; Swaroop, 1987; Shane, 1986; Srivastava andRana, 1988). These ornamentals fall under following groups.iiiiiiivvviviiOrnamental treesOrnamental shrubsBulbous ornamentalsFlowering annuals and herbaceous perennialsOrchidsFoliage plantsMiscellaneous plantsSome of the significant introductions of ornam~ntal plants arelisted briefly below.(a)Ornamental trees(i) Chaenomeles sinensis : It is known as flowering qUince. Anintroduction EC 24520 was introduced from USSR in 1963 at Shimlastation. It is a small size tree, which attains a height of 6 lU, bearspink flowers in spring (in the month of April). The bright yellowfruits persist on the plant for a long period; giving an ornamentallook to it.(ii) Ceratonia siliqua: One cultivar Be 36084 has been introducedfrom Spain in 1965 and grown at Shimla Station. It is handsome,slow growing, evergreen tree which may attain a height of about10 m. It bears shining dark green, oval, beautifulleaflets.(iii) Camellia sasanqua : An ornamental tree, native of tropicalAsia, China and Japan. NBPGR has recently introduced rootedplants of three improved cultivars, Polar Ice PI 546367, NA 61950(EC 322327), Snow Flurry PI 546367, NA 61951 (EC 322328) andWinter Rose PI 546368, NA 61952 (EC 3223290) from USA, Whichare under evaluation at Delhi.(iv) Colvillee racemosa : A native of Madagascar. One cultivarEC 164651 had been introduced from USA in 1984. It is a deciduoustree.

80 B.P. Singh(v) Deubentonia puricea : A small leguminous tree, which isnaturalized in South America. One collection Ee 104146 has beenintroduced from USA in 197J and grown at NBPGR RegionalStation, Shimla. The plant bears scarlet flowers.(vi) Paulownia sp. (Empress tree) : One cultivar Be 316474 of P.tomentosa has been introduced from China during 1991 and isunder evaluation at Bangalore. It is fast growing deciduous tree,bears erect panicles of fragrant pale violet panicles during May.(vii) Sophora japonic'a (Japanese Pagoda tree) : One cul.tivar Be155222 has been introduced from USA during i983 and is undertrial at NBPGR Regional Station, Shimla. It is small deciduous tree,pendulous, prostrate stiff branches, forming a dense round head.(viii) Tabebuia chrysotricha : A native of tropical America isevergreen medium size tree, has been introduced from USA in 1977and tried at NBRI, Lucknow. Plants flowered in February-March1983 attaining a height of 2.5 m. It looks pretty in full bloom fromFebruary to April, bears funnel shaped bright yellow flowers,arranged in umbrella characters.(b)Ornamental shrubsNBPGR has int~oduced seeds/ plant propagating materialsof commercial cultivars of flowering shrubs like Bougainvillea,Buddleia, Caliandra, Cotoneaster, Forsythia, Hibiscus, PhiZadeZphus,Rosa spp., Spirea, Vibumum and Weigela from various countries.(i) Bougainvillea: Bougainvilleas are among the most floriferousshrubs of the tropics, producing beautiful colour effects, which canhardly be excelled by any other plant. NBPGR has introduced 38cultivars from New Zealand and Kenya. The prominent ones areHawaiian scarlet, Hawaiian orange, Hawaiian white, Crimsonglory, and Lady Hudson. These are under trial at IARI New Delhi!and IIHR, Bangalore.(ii)BuddleiaB. davidii - Varieties Royal R~d (EC 25938 Hungary) and WhiteBanquet (BC 25939 Hungary) were inh·oduced. These are mediumto tall deciduous shrubs, with thick 20 em long panicles of darkpurple and white flowers respectively, .flowers during rainyseason.

Promising Introductions in Horticultural Plants and Oil Seeds 81B. nivea - One accession EC 25940 has been introduced fromHungary. It is a medium tall shrub, yielding slender 30 cm longpanicles of blue colour.(iii)ChaenomelesC. japonica - Two accessions Ee 27853 and EC 43428 have beenintroduced from Japan and USSR respective.Iy. These are shrubsof about one meter height and produce very attractive big flowersin abundance in early spring varying in shade from salmon toorange, pink to deep scarlet.(iv) Cotoneaster jrigidus : One accession EC 43421 var. Victorywas introduced from USSR. It is evergreen shrub of mediumheight. The round leaves are shiny above and greyish beneath.(v) Rosa spp. : 458 accessions of Rosa sp. and conunercialcultivars have been introduced from UK, Germany, France, USSR,USA, IsraeL Italy, Holland, Australia and Denmark. Someinteresting varieties introduced are Red Cushion (EC 94226) SanDiego (EC 94234), Aquarius (grandiflora) and Cornlenche(grandiflora) (Ee 94242-94243 USA) Mazurka, Fantasia (Be 127598-127599 ex Germany) and Spotless pink (Ee 159476 ex USA).Among the Rosa species are R. ajzeliana, R. beggeriana, R. carolina,R. calif arnica, R. cinnamamea, R. damascena, R. dumalis, R. eglanteria,R. giraldri, R. glutinosa, R. in dora , R. laxa, R. maximowicziana,R. micrantha, R. micrantha, R. moschata, R. pratensis, R. rubiginosa,R. spinasissima, R. stylosa, R. tomentosa, R. virginiana, R. watsonianaand many others.(vi) Spiraea spp.: S. chamaedrifolia is a very hardy shrub ofmedium height. One accession Ee 35562 introduced from Hollandin 1965 has established at NBPGR Station, Shimla . It bears purewhite flowers in large numbers during April.(c)Bulbous ornamentalsThis group of plants includes bulbs, rhizomes, tubers,fascicled roots etc. A large number of exotic species like Amaryllis,Anemone, Begonia, Cyc1aman, Dahlia, Freesia,· Gladiolus,Hycinthus, Iris, Lilium, Narcissus, l'oJianthes and Tulipa havebeen introduced. Some of the prominent one!'> "re listed below.(i) Cyclamen: is a native of Mediterranean regf(lll. These arefinest green house and indoor plants for autumn and winter

82 B.P. Singh. decoration. Flowering period is six months from November toMay. Flowers remain fresh on the plant for 20 days. A newintroduction cv Ideal Osena Pink in Eye ( EC 11822 ex Denmark)bears pink flowers.(ii) Dahlia: A native of Mexico/Central America and northernmost South America. In India it grows in hills as well as plains.NBPGR has introduced 25 cultivars of Dahlia from Holland andUSA. Most prominent ones are Maitre Noyer (EC 106237),Spectacular (Be 606238 Holland), Elegance (BC 106242) . All thesecultivars were made available to Project Coordinator (Floriculture)for their trial! evaluation.(iii) Freesia: A native of South Africa, medium size herbs.Freesias are well known for beds, pots and hanging baskets. Theplant is about 30 cm tall. Inverted bell shaped flowers appear inlarge number from last week of February to first week of April inShimla hills.(iv) Gladiolus: A native of South Africa, has majestic flowerspikes with florets of brilliant colours. NBPGR has introduced 221varieties of Gladiolus from USA, USSR, Egypt and Israel. A newintroduction Mirage (Ee 154209) from USA has performed well atIIHR, Bangalore. Cultivar Margarat Fulton proved tolerant toFusarium wilt.Among the secondary introductions, following varieties ofGladiolus have been evolved through hybridization at IIHR,Bangalore.MeeraNazravaSapnaMan MohanManoharGPIX Friendship (Ee 164736, USA)Black Jack x FriendshipGreen Wood Packer X FriendshipFriendship x G. tristisFriendship x C. tristis(d)Flowering annuals and herbaceous perennialsNotable introductions made by NBPGR include Aster,Antirrhinum, Calendula, Celonia, Columnea, Digitalis, Dianthus,Delphinium, Tagetes (marigold) Zinnia and Chrysanthemum.

Promising Introductions in Horticultural Plants and Oil Seeds 83(e)OrchidsOrchids are most beautiful flowers which.are valued as cutflowers. Most of the commercial hybrids and species generallybelong to the genera Cymbidium, Cattlaya, Dendrobium, Paphiopediumetc. NBPGR has introduced 23 hybrids/ cultivars from Thailand,Australia and passed on to IIHR, Bangalore ..III. . Oil Seed CropsDiverse germplasm of over 27160 accessions of differentoilseed crops from more than 26 countries were introduced during1976-1994 viz., groundnut 3198 accessions; sunflower 3819accessions; sesamum over 3800 accessions; safflower 5321accessions (this includes duplicates); rapeseed & mustard 3069accessions; soybean above 7220 accessions; linseed 542 accessions;castor 120 accessions and selected germplasm collections ofoilpalm (74 accessions). The gennplasm resources were madeavailable to coordinated project in oilseeds/ oilseed researchworkers in the country and public and private sector.(a)Groundnut (Arachis hypogaea)Several promising cultivars like Banting, Petandur, Tapis,Anoa, Typai, Gagas, Rusa (EL.170361-367 ex Indonesia), Taining4,5, Taman Sel. 69 (EC-170433 -455 ex Taiwan) UPLPN-2, ACC 72,CBS 2-25, F 334-33, PI 118200, M 10, CBS 101, CES 102, (BC 168099-106 ex Philippines); Lompong, SK 38, Taiwan 9 (BC 169319-321 exThailand), rust resistant lines (EC 115767-80 USA), droughtresistant early types (BC 115992-993 ex Senegal) were introduced.A large number of Arachis species like A. villosa, A. spinalba,A. batizacoi, A. chacoensis, A. correntina, A. stenosperma, A. duranensis,A. rigonii, A. paraguariensis were. also introduced.(b)Sunflower (Helianthus annuus)The promising introductions include improved cultivars/hybrids, cytoplasmic made sterile lines, maintainers/restorers,source of resistance to Alternaria blight and downy mildew, lineswith high oil content. Some of the useful introductions include birdresistant strains-BRS1, BRS-2, BS5-3 (Be 172473-475 ex USA),CMSHA 207 and HA 207 (ex USA), CM 306, CH 77/ HA 89(sel -2) CH 77/HA 89 (sel-2), RHA 265, CMS 400, CH 77/HA 89

84 B.P. Singh(Sel-3), hybrid strains HX 113, Hy 280, Cargill 205, Kargill 26 fromAustralia, lodging resistant line Ee 327993 (ex USA), compositerestorer germplasm population BC 328126 (ex USA) and differentHelianthus species like H. argophyllus, H. petiotaris var. petiolaris,H. praccox ssp. ranyonii, H. praecox ssp. hirtus, H. anomalous,H. bolanderi, H. paradoxus, H. niveus, H. ~eglastus, H. tuberOlLSUS,H. muttallii, H. pumilis, and others. In Sesamum dwarf, lessdehiscent types namely PI 189082, PI 189229, PI 210687, PI 231033,PI 231034, PI 250099, PI 250100, PI 231096, PI 481875 are importantintroductions. The cultivars like Giza 159, 24, 25 (EC 10696-698),Suweon 5,9 (BC 11403-404 ex Korea), Aceitera 76-5, Glauca 76-125,Venezuela 52-76-1340 (BC 121370-372 ex Venezuela), Has ava,Panta (ex Nigeria) Be 327476-52 highly drought tolerant & highyi;elding from Sudan. A few related species like S. angolense, S.alatum, S. capense, S. triphythum from Italy.(c) Safflower (Carthamus tinctorius L.) .Germplasm collection from diverse sources like USA, IBPGR,USSR, Australia, Turkey, Hungary, Germany etc. was introduced.These include commercial high yielding varieties, thinhulled types, namely Gila 4510, Oleic lead, Reduced hull 1,2,3,4,T-410 (62), Partal hill 62,63,61,43, Peoy (53), Peoy (62) 14.5 USB(86), USB-45, BC 303268, EC 276881, BC 301333 and leaf 75 andsaffire and related species C. tenus, C. palaestinus, C. lanatus,C. lanatus sub. sp. turkestanicum, C. oXljcantha, C glancus.(d) Rapeseed mustard (Brassica spp.)In Raya or mustard (Brassica juncea) high yielding germplasmcollections (BC 302158-302173 ex USA), germplasm lines BC302465-302469 (ex Sweden) disease resistant/ tolerant, lines withhigh oil content (EC 302884-302885 ex Canada BC 303460-313463ex UK) are useful introductions.(e)Rapeseed (B. campestris)Commercial cultivars Torch, Echo 1984, Tobin 1986 (Ec-302886-302888 ex Canada) germplasm collections EC 305574-305578 ex Bangladesh, and high yielding breeding lines BC 312379-312386 ex Canada were introduced. In Toria (B. campestris var. toria)promising introductions include BC 204233, BC 204234 BC 223406which are early and fairly high yielding.

Promising Introductions in Horticultural Plants and Oil Seeds 85(f)Gobhi sarson (8. napus)Cultivars Be 302149-302157 ex USA and Wester 1987(EC 302839 ex Canada) are useful introductions.(g)Soybean (Glycine max)Improved cultivars!lines AGS 190, AGS 292 (EC 301882-.301885 ex- Taiwan), disease resistant lines Cordon S-L 028 AC(EC 302092) high yielding, disease resistant collection EC 303542-303579, EC 310338-310340 ex USA are useful introductions. Theother high yielding germplasm lines include EC 172611, EC 93741,BC 251477, G 230, G 309.Amongst the Glycine species collections introduced areG .• canescens, G. cladestina, G. latifolia, G. arneria, G. latrobaena, G.tabacina, G. tomenletta and G. glancus.(h) Linseed (Linum usitatissimum)Promising introductions included cultivars Bison (ex USA)Sibrisk, Avangard (EC USSR) Bionda (EC Nigeria), Mesten~BRD,Hosszutroti, Farmosa, Rocket, Coneorent, Rostlen, and a largenumber of other useful materials. The accessions EC 109490 andBC 110289 showed resistance to powdery mildew at Akola.Amongst the Linum species introduced were L. perenne, 1.altaricum, L. flavum, L. bienne, 1. africanum and L. angustifolium.(i)Castor (Ricinus communis)Some useful introductions include CNESI, NSL 88837 (Ee370307 ex USA), BGRC 25402, RGRC 25420, ZM! A 8033, ZM/A5057 ZM/ A 5236, 2M/ A 5335 all from Zambia, and accession EC168959-989 ex USA). The collection BC 168483 has given excellentperformance at Akola. .(j)Oilpalm (Elaeis guineensis)Earlier useful introductions were from Malaysia and Indonesia(Be 159876~877). Recently high yielding germplasm lines havebeen introduced from Africa, Belgium and UK. Efforts have alsobeen made for introduction of germplasm Perilla (Perilla jrutescens),niger (Guizotia abyssinica) and oil yielding halophytes like Salicorniabiglovii from USA.

IIPromising Introductions and Prioritize Needs forExotic Germplasm of Grain Legumes, Fibre Cropsand Under Utilized Crop PlantsR.V. SinghIntroductionExchange of economic pl~.nts is' a prerequisite for embarkingon any crop improvement programme and also for introducingnew crops from one part of the world to another part. Coffee, anative of south east Africa reached India through Saudi Arabia,Solanum tuberosum, Nicotiana tabacum an~ Lycopersicon esculentumall native to South America were introduced in India during early10th century and Trifolium alexandrium used as a green legumefodder in India was introduced in the early years of this centuryfrom Egypt. More recently, new crops like soybean (Glycine max)rich in protein and oil, introduced from USA; sunflower (Helianthusannuus) rich in oil introduced from USSR and USA; jojoba(Simmondsia chinensis) from USA, Chinese gooseberry (Actinidiachinensis) from New Zealand and many others. Therefore, lookingat the importance of exotic introductions, a brief account of recentpromising introductions of different legumes, fibre and underutilized crops with their futUre needs and priorities is presentedin this paper for information and further utilization in the country.Grain LegumesThe grain legum~s constitute a group of major crops such aschickpea, pigeonpea, cowpea, black gram, green gram, lentil,soybean, french bean, horse g!am, faba bean, and others like rice

Promising Introductions and Prioritize Needs for Exotic Germplasm 87bean, mucuna, and adzuki bean. These occupy an important placein the di~tary systems of Indian people meeting the proteinrequirements of the predominantly vegetarian population. Someof the pulses serve as excellent forage and grain concentrates tofeed the large cattle population and some of them are excellentgreen manure crops adding much needed humus and major plantnutrients to the soil.In order to augment the germplasm of grain legumes geneticresources, a fairly good number of introductions were made fromexotic sources by Germplasm Exchange Division of NBPGR, NewDelhi.Promising introductions'Chickpea : Germplasm collections with desirable attributes, viz.,bold seeded (Ee 267486-501), leaf minor resistant (EC 267425-436),Ascochyta blight resistant (EC 267483-487, BC 367319-320) and coldtolerant (EC 267559-522), J:righ yielding, and disease resistant (EC286030-033) all from ICARDA (Syria); improved cultivars fromUSSR, basal branching kabuli type (EC 223125, EC 223513), earlymaturing (EC 280187, 280226, 280227 and 280228); Cicer speciescollections, viz., C. canariensis, C. yamashitae and C. echinospermumpossessing source of resistance/tolerance to biotic and abioticstresses.Pigeonpea: Very early type collections/cultivars from Australia(BC 158219-158248), Ethiopia (EC 244842-244871), Kenya (EC21~614-215643) particularly Que~t (EC 284065 ex Australia),profusely branched, bold seeded types (EC 16855, 168489), bean flyand bruchid resistant lines (BC 245976-77) from A VRDC (Taiwan);YMV and leaf crinkle resistant line (EC 27268) and wild relatedspecies, viz., C. einarens and C. karstingi. .Mungbean: Germplasm collections possessing resistance toCercospora leaf spot, mungbean mottle virus, powdery mildew, podwithering, drought and lodging with early maturity, large yellow, seeded and photoperiod insensitive (EC 232498-512 ex Canada);bruchid resistant (EC 245970-80 ex Taiwan); early maturing,resistant to lodging and powdery mildew (EC WOO, EC 182498 exTaiwan); widely adaptable, photoperiod insensitive and resistantto TMV (VZ 984, EC 19817i ex Taiwan); the cultivars BC 35537 andEC 33883 have performed well at Jodhp.1;1;r. Accessions introduced-_ ': j

88 R.V. Singhfrom Taiwan such as EC 318985, 19059 and other differentgermplasm lines resistant to powdery mildew, charcoal rot, leafcrinkle virus,drought and flood tolerant, high yielding, uniform inmaturity, photoinsensitive and also found stable in production.Cowpea: BC 130163 from USSR a bushy type, early maturing (68-70 days) gave an average yield of 9 q/ha at Jodhpur. EC 5000 fromRhodesia, a bushy and photoinsensitive was noted for higher grainyield (14-15 q/ha): Be 107155 ex USA is also a high yieldingintroduction, aphid resistant (Be 216845-46 ex USA); extra earlymaturity coupled with higher grain yield and bruchid resistant (Ee169416-17, Be 169720-22 ex Nigeria). The cultivars Rituraj andAssem have been developed by the Bureau using Ee 26410 exMexico and BC 21622 ex Philippines as one of the parents.Accession Ee 336589-90 (Nigeria), were observed as aphid,bruchids, thrips and Striga resistant while Ee 305728 has long podsand Be 243988 and Ee 244980 are resistant to foliar diseases.Fieldpea : Cultivar Harbhajan (Ee 33866 ex Portugal) an earlymaturing, short statured, yellow seeded with dual purpose type,is an excellent introduction (Be 292160 ex USSR) and is reportedto be resistant to Ascochyta blight and fungal rot. The collectionsBC 15184 and Be 1741 are resistant to pea seed borne mosaic virus.Ee 271572-75 ( ex France) possessed resistance to pea seed bornemosaic virus. BC 109235, -15272 and EC 322745 have been observedto be free from powdery mildew at Delhi.Lentil: High yielding, bold seeded ,collections (BC 284207-237 exICARDA,. Syria); the early type collection Be 280267 and boldseeded Be 280286 gave good performance at Bhowali station.Ee33214, -23315, -233239 were identified as early promising types andEe 223193, -267585 and EC 267662 as late promising types,French bean: Diverse germplasm collections have been introducedfrom CIAT Colombia, USA, USSR, Nigeria and other countries.Amongst the notable introductions for high grain yield are: Ee57080, -43036, -94171, -9939 (UK) and BC 97829 (USA), respectively.The exotic introductions (PI 302538, PI 168980, 0 PI 281849 andPI 810871 ex USA) have been observed to be free from powderymildew (Erysiphe polygont) at Bhowali. The accession BC 62161-10,Be 22607, PI 170639 and PI 289441 possessed resistance/toleranceto Cercospora and angular leaf spot. Ee 290517, -271559 are good

Promising Introductions and Prioritize Needs for Exotic Germplaslll 89in grain yield while Ee 271522 possess bold seeds, Ee 290516 anearly maturing (51 days at Shimla), Ee 1711528 have longest pod(19 cm) and Ee 29051 has the h.ighest pod numbers (22/plant) Ee127384 good for fodder; Ee 8938, PI 339491 high in grain yield andIP 281849 and PI 168980 were found free. from powdery mildewat Bhowali.Khesari: Be 106886 and EC 106892 (USSR) have given goodperformance in respect of grain yield at Delhi. Ee 322620-31 (Syria)have low neurotoxin contents. The related wild species of Lathyrus,viz., L. tingitanus, L. 5zowitsci, L. cinera, L. clayme1'lum and L. ochrushave also been introduced for further utilizatio? in the improvementprogramme.Mothbean: Diverse germplasm collections for various desirabletraits (Ee 251876-253424) have been introduced from USA.Fababean: High yielding germplasm collections EC 284356-375(Syria); improved varieties/cultivars Giza-3, Giza-402 (Ee 183567-68 ex Syria); evergreen and long podded accessions (EC 191573-574 .ex Newzealand) are some of the notable introductions. Ee303650-71 are resistant to chocolate leaf spot disease while Be303672-91 (ex Syria) was observed as resistant to Ascochyta blight.Ricebean: Collection possessing resistance/tolerance to Cercosporaand Uromyces (Be 108080) and high grain yielder (Ee 1243'6), earlymaturing (Ee 90453, -182228) have been introduced. Accession BC32244 have high protein content.Soybean: Germplasm collections, viz. Ee 39703, Be 76726· andBe 95809 are rich in oil content (23.6%) and Ee 127512 (24%);collections with high level of field resistance against bacterialpustules Xanthomo1'las campestris (Ee 34094, 95806,106991,992 andBe 241920 ex USA); germplasm lines with high seed longevity (Ee241949, Ee 2i±1920); T 280 and T 307 lines are low in linolenic acid;L 861436 lacks lip oxygenase; L 81-~871 and L 83-4387 lack trypsininhibitor; PI 317334 B and PI 297550 are photoperiod insensitive.EC 250574, EC 287481, 172668, 251418 and EC 274673 were highin grain yield at Akola. Accessions Be 172575, 172630, 172648,172657, 232044, 232048, 251411, 251418, 251457, 251457, 274707were found to be resistant to pod shattering at Akola. Accessionswhich have high seed germinability (80-94%) are BC 19729,39495,172587,172638,172654,251369,251418,251459,251771,244673 and

90 R.V. SinghEe 241472. EC 308316-17 possessed characters like salt tolerant andhigh protein contl;'nt EC 308322-23 rust resistant; Ee 308325resistant to foliar feeders introduced from USA. Ee 333855-60,resistant to soybean rust; Be 333864 soybean cyst nematoderesistant; BC 333897-906, high oil and protein rich lines; Be 333907-92 resistant to Phytophthora megasperma, non shattering and withhigh protein content were also introduced from USA.Winged bean: The germplasm collections which gave higher yieldat Akola included Be 38955A (1058 kg/hal and Ee 114273B (971kg/ha). A selection AKWB-l from EC 114273 B (Indonesia) hasbeen recommended for release during VIIth AICRP on underutilized and under-exploited plants workshop held at Bangalore.Accession BC 114273-B out yielded (106 q/ha) green pod yield andfound significantly superior over check (Akola local). It is also ahigh grain yielder (10 q/ha).Promising Introductions in Fibre CropsIn India, the major fibre crops are cotton and jute and theyhave an important place in the economy of the country. Some ofthe promising exotic introductions are given hereunder:Cotton! Accessions Be 314280-82 ex Syria (Aleppo-40 and Aleppo-33/1) are tolerant to Verticillium wilt; Be 317455-63 ex Turkey ishigh yielding, early maturing, high ginning outturn and fibre,length (28.5 rom and above). BC 323682-686 and Be 326150-166(U~A) are high yielding while BC 323682-686 (USA) are boll weevilresistant; Be 34398, BC 343987 are dwarf types; Ee 34417 early in .flowering; EC 344544, has long fibrE!; Ee 344895, a bushy type;EC 345771 is resistant to Heliothis spp. EC 341979, high yieldingupland cotton; Be 340239, 334244, 334245 high yielding;Ee 340239, okra leaf type; EC 340240 a male restorer line,EC 340242 is glandless, all introduced from USA. BC 335928 andEC 342984 (Turkey) are high yielding. Be 346025 is resistant toFusarium wilt. BC 346026 is resistant to Lygus; Ee 346027, resistantto Heliothis spp., EC 346031-51 has long fibre and Ee 350370 abushy type, ail from USA. Be 361387~405 high yielding hybridcotton from Israel. PI 506414, PI 506415 and PI 506416 combine okraleaf traits and have high yield potentials and also resistant to pinkboll worm (Pectinophora gossypiella). EC 344571":733 (USA) are

Promising Introductions and Prioritize Needs for Exotic Germplasm 91resistant to boll worm; BC 343926 (USA), long staple variety withhigh lint recovery; EC 338693 (Turkey) has high ginning percentagetogether with resistance to Verticillium wilt.Jute: Most accessions of jute were introduced from Bangladesh,Thailand and China having different desirable traits which includeCorchorus species, viz., C. olitorius, C. capsularis, C. trilocularis, C.facicularis, c. tridesns, C. aestuana, C. distonius etc.Different Hibiscus species were also introduced such as H.acetosella, H. acutangula, H. calyphyllus, H. cannbinus, H. subdariffaall from Bangladesh and various accessions of Crotalaria. jUl1ceafrom Italy, and its related species from Hungary. Other allied fibrecrops like Agave americana and Agave sislana have shown goodadaptations in arid zones and studies are in progress to evaluatefibre production potential under varying habitats of arid zones ofIndia.Promising Introductions of Under Utilized and Under ExploitedCropsA few interesting and successful introductions of fodderutility for arid habitats are Tamarugo (Pl'osopis tamarugo), kumbat(Acacia senegal), and A. albida. Brachychiton populneum, a droughttolerant shade tree of Australian origin, provides finest fodder andis also useful as wind breaker when planted in multiple rows.Prosopis tomarugo, ftom tropical America, is salt tolerant, droughtresistant and supplies pods/leaves as fodder for goats throughoutthe year. Kumut (Acacia senegal Wild), provides best grade of gumarabic, which has wide uses as an emulsifying agent, demulcentand protecting agent in the manufacture of adhesives, textiles,leather, water paints, polishes, inks, ceramics, food products andpharmaceutical preparations. Cassia sturtii, an Australian shrub isanot!ler drought resistant fodder type for recent introduction.Amongst the Acacias, A. albida is of great promise of arid and semiaridhabitats. Its pods and foliage provide good fodder. Fresh podyield is reported to be 100 tons/ha/year.Among the· Artiplex species, a few promising ones areA. halimus (EC 129767 ex Tunisia), A. canescens (Ee' 129768 exTunisia) and A. nummularia (EC 129766 ex Tunisia) which areperennial, evergreen and salt tolerant, have performed well atJodhpur and hold promise for arid region.

92 R.V. SinghAmong the hydrocarbons and industrial plants is Jojoba(Simmondsia chinensis). A hardy shrub, native of north MexicoCanada, South-North USA, and a source of liquid wax. More than35 accessions of this have been introduced from USA, USSR andTanzania and have been under trial in Jodhpur, Gujarat and otherstates. At NBPGR Regional Station, Jodhpur one female plant ofEC 33198 ex USA gave very good seed yield. In guayule­(Parthenium argentatum), a shrub of tropical Mexican deserts, 65accessions have been introduced from USA and Mexico. All partsof the plant contain good quality of latex which when purifiedgives rubber. The variety Arizona No.2 (ex USA) has given goodperformance. The rubber content of two year old plantations wasfound to be 6 to 8%.Among the species of genus Euphorbia, E. lathyrus and E.tirucalli hold promise as hydrocarbon plants. E.lathyrus also knownas Gopher plant is annual, biennial or perennial of Mediterraneanorigin, an important source of crude oil which can .be effectivelyexploited for gasoline agriculture.E. tirucalli also known as milk bush, a shrub or small treenative of East Africa has become naturalised in India since too long.Its yielding capacity of latex of the plant has been reported to be15 to 20 barrels of oil/ acre/ year. A few collections have beenprocured from USA and are under trial at CAZRI, Jodhpur andother places. The most encouraging results have been obtainedwith Euphorbia antisyphilitica, a desert shrub of Mexican origin,which contains a high quality wax commercially known ascandeli1la wax. Wax content of 2.5% on dry weight basis has beenextracted from plants raised at the CAZRl, Jodhpur. However,recently some of the collections at the institute farm Jodhpur'havebeen reported to yield 4.5% of wax. This wax has been approvedby cordite factory at Nilgiris as per their specification and. requirements. Presently, India's entire quantity of candelilla waxrequirement is imported. Plants can be propagated easily fromstem cuttings. There is considerable fufure for raising E. antisyphiliticain the arid climate of the Thar desert.In recent years, much emphasis has been placed on Ipil-Ipil(Leucaena leucocephala), a successful multipurpose plant. TheBureau has introduced diverse germplasm numbering 335 fromAustralia, Philippines, Colombia, France, Malawi, Sierra Leone,-- ,

Promising Introductions and Prioritize Needs for Exotic Germplasm 93UK .and USA. This fast growing tree, provides fodder, greenmanure, fuel and can withstand drought and is promising for semiarid regions. An introduction K8 (EC 124343 ex Philippines)performed well at CAZRI for fodder purpose. Another varietySalvador (EC 12270 ex Australia) has performed well in Gujaratand Kerala for fodder and fuel.Germplasm material of Casuarina equisetifolia, C. stricta andC. glauca procured from Australia, Philippines and USA is undertrial at CAZRI, Jodhpur / IGFRI, JhansilMettuplayam, (TN) andmany other places in the country. Some newly introduced varietiesare reported to possess much better fuel yielding capabilities overthe existing types grown in Tamilnadu, particularly in coastal areaswith sandy wastelands or even on alkaline soils.In grain amaranths, EC 289378 (59 g/plant) was observedhigh in grain yield at Shimla station while accession EC 345800 exUSA possessed long inflorescence. Different related species ofAmaranthus were also introduced for their further utilization in thebreeding programmes to transfer desirable traits. Likewisebuckwheat, chenopods and Cuphea and their related species werealso introduced from various countries. In buckwheat BC 218772and EC 218753 were identified as best grain yielder at Sh.imlastation .. In chenopods (Chenopodium quinoa, BC 180010), with 6.5 q/ha was also found a promising introduction. In C1lphea, EC 1.33508gave highest seed yield (4.8 q/ha) at Shimla.A new improved strain of Cynara cardunculus (BC 151169 exArgentina) has been introduced which yields oil, fibre and fodder.Priority needs for exotic germplasm introductionsThere is need to introduce· exotic germplasm resourcespossessing resistance to yellow mosaic leaf virus and nematodes·in mung bean, urid bean and in mothbean resistant to Anthracnoseand bacterial blight. .IIi. French bean, resistant to Ascochyta, Fusarium and podborer in chickpea and cowpea; wilt and pod fly in pigeonpea; highyielding, medium maturing, bold seeded types' in lentil; highyielding, medium maturing lines in faba bean. Adequate germplasmlines in crops like Vigna subterranea need to be introduced into th'earid/semi drier plains.

94 R.V. SinghDonor Parents required for immediate useA large number of pests and diseases thrive on pulsesdevouring 20 to 80 per cent of the produce in one crop or another.Donor parents are required to develop multiple disease resistantvarieties in different pulses. Lines resistant against Phytophthora inpigeon pea; Ascochyta and BotrYtis in chickpea; rust in pea; wilt inlentil and powdery mildew in mung bean and mid bean. Linesresistant to insect pests like: pod borer complex in pigeonpea andchickpea; resistance to"abiotic stresses, drought in all pulses waterlogging and frost in pigeonpea and salt tolerance in chickpea.Plant type and quality: Germplasm with better partition and highharvest index are to be introduced because most of the present daycultivars are poor in partitioning the photosynthates betweenvegetative parts and grains which results in poor harvest index.Transfer for desired genes from wild to cultivated species cancreate noble characters or extreme forms, because genetic potentialof wild species is qUite high. One of the methods to decrease theduration of the crop would be to evolve lines which are early andinsensitive to photoperiod. Thus these are the two importantcharacters for which wild relatives of pigeonpea could beexploited. These spe.cies include Atylosia platycarpa, A. sericea andRhynchosia rothii. Besides these traits, wild species of Atylosia andRhynchosia also possess inbuilt resistance for one or the otherimportant diseases of pigeonpea. It is suggested that, crossesbetween Cajanus cajan and other wild species should be made.Characters worth incorporation are resistance to spotted miteTetranychus urticae in A. acutiolia; hairy pods (repel pod eatinginsects) in A. pluriflora and A'. reticulata and drought! frost toleranceof Atylosia species in general. The variation for methionine andcystosine (the limiting amino acids of Cajanus cajan) contents is notlarge enough to be utilized in breeding programmes.The harvest ind.ex of pigeon pea is generally higher with therabi sown than with the kharif sown crop. Besides, there is areduced attack of Alternaria blight on the rabi sown pigeonpea.Therefore, there is a need to introduce such well adaptedgermplasm lines which can be grown well in rabi season and canbe well set with different crop rotations. Introduction of newsource of genetic male sterility is important to develop hybrids inpigeon pea for increasing the production. Germplasm having high, , ,

Promising Introductions and Prioritize Needs for Exotic Germplasm 95heterosis with respect to early and high yields should also beintroduced to increase the yield of hybrids.In soybean, germplasm resources having high protein and oilcontent, resistant/ tolerant to acidic and saline soils and cytoplasmicmale sterile lines for exploitation of hybrid vigour need to beintroduced on priority.Fibre cropsCotton: In the last four decades, Gossypium hirsutum cottons havereplaced desi cottons in 30 lakh ha under rainfed conditions. Byand large, the desi cotton cultivation in the rainfed conditions is'now confined primarily to dry and unproductive areas, which isa major cause for low productivity. Three major possibilities tobring about a rapid breakthrough in rainfed cotton productivitymust include: (i) evolution of new plant ideo types in desi andAmerican cottons, (ii) development of superior hybrids withmedium staple and short duration and (iii) incorporation ofretPstance factors from various sources.(a) Ideal new plant types : Breeding for a new plant type forrainfed conditions is just gaining momentum, but it is yet to takeproper shape. The appropriate genotype would be the one that hasa reduced duration and near determinate type with high bollbearing potential so as to minimize the risks and losses arisingfrom environmental adversities and lapses in management.Diminutive plant type, both in structure and stature with compactzone of fruiting having even a semic1uster fruiting habit, reasonablylarge bolls, fruiting at a lower node on main stem and endowedwith other components of yield, earliness and resistance are likelyto be more useful. In desi cottons, evolution of these kinds of planttypes would need developmental breeding for establishment ofsuch basic forms for plant stature, bigger boll size than at presentand early maturity before taking up recombination breeding. Butin G. hirsutum the required components have already beenidentified.(b) Hybrid cottons: The cultivation of hybrid cottons commencedin the seventies in India and now widely practiced in 10-12% ofthe total cotton area. Till today hand emasculation and handpollination have proved to be the only way of producing hybddcotton seeds for commercial cultiyation and the cost per kg of seedto the farmer is exhorbitant.

96 R.V. Singh(c) Resistance approach: Incorporation of chemical changes inplant composition as terpenoids, phenols and those which help tomake cotton plant less vulnerable to pests aJ;ld even diseases isnecessary. The development of MAR and 'MDR lines through thesetechniques is a possible solution to cut costs of plant protection,reduce risks to the grower and environmental pollution. Lines forbollworm resistance with dense glands flower buds and redpigment trait in combir).ation with high terpenoid level specificallyfor Heliothis resistance are being established as a possible solutionto control bollworm menace. Diploid wild genome introgressioninto· the cultivated background is known to contribute forresistance to bacterial blight, rust, fusarium wilt, drought andinsect pests.Germplasm represents the most important biological tool tobring about a real technological breakthrough in rainfed cotton.Atleast 15000 germplasm stocks are known to be available in majorgenetic resources centres of the world ih cotton of which nearly6000 lines have been assembled in India. Recently two more newspecies G. Jryxellii and C. binatum have been identified. This hasopened up the scope for creating a large number of new nuclearcytoplasmic combinations of the wild species genomes.A to G oncultivated A and AD genomic backgrounds. Our gene pool alsoneeds enlargement through the acquisition of exotic types fromUSA, USSR, China and Australia besides undertaking newexpeditions in the old and new worlds. Therefore, emphasisshould be given for the introduction of exotic germplasm resourcesin co~ton as discussed' above for the various traits. Germplasm isalso required in jute and other allied fibres for future needs.Under utilized and under exploited cropsThere is an urgent need to introduce new exotic germplasmin different under utilized crops particularly in Amaranthus, Jojoba,Ipil-Ipil, Euphorbia, chenopods, A trip lex and other new potentialcrops. .Plant resources that require introduction: A new oil yielding cropworth trying for the arid lands is Cuphea. The seeds of Cupheaspecies belonging to family Lythraceae, contain 16-42% oil and thatthese oils have high levels of lauric acid and their medium chainfatty acids. Efforts have been made in USA to collect wild 'CupJ1eagermplasm from Mexico and Brazil which are recognised as

Promising Introductions and Prioritize Needs for Exotic Germplas1ll 97c~ntres of diversity for this genus. This genus containsapproximately 260 species of which only 5 grow wild in USA. Thespecies C. wrightii Gray appears to have agronomic potential inarid areas. Similarly other new crops for arid areas are Lesquerella,Grindelia and Baccharis. Lesquerella a so:urce of hydroxylatedvegetable oil of industrial importance as chemical feed stocks forthe production of lubricants, plasticizers, surfactants andpharmaceuticals. Seed oils of various species of Lesquerella havebeen discovered to contain sizable quantities of 3 hydroxy fattyacids. The oil percentage reported ranges from 11-39%. Some 20species of Lesquerella have been field tested in Arizona. L. jendleriappears to have the most promise for domestication and use as anew crop.Research at the Bioresources Research Faculty of the Universityof Arizona, Office of Arid Lands Sludes. (OALS) in Tucson, hasidentified species of Grindelia, as having potential for the productionof feed stocks for biocrude production in arid lands. Grindeliaappears to be one of the most promising of the resin producinggenera of the compositae. Grindelia camporum Grene is an aridadapted plant, used in industries. Baccharis is a genus of familycompositae, consists of over .300 species of dioecion, sometimesevergreen shrubs, native to North and South America.The desert broom and rosin bush (B. sarothrodes Gray) is anerect, small leaved evergreen shrub, that is well adapted to thedrought, heat, cold and high salinity conditions. In the naturalhabitat of Sono'ran desert and surrounding arid areas.The buffalo gourd (Cucul'bita joetidissima HBK) is a semixerophytic cucurbit native to arid lands, ranges of south westernUnited States and northern Mexico. As a: perennial, the buffalogourd develops large storage roots containing substantial quantityof starch. The fruit of wild buffalo gourd are 5-7 cm. in diameterand contain 200-300 seeds, with 30-40% edible oil and ·30-35%protein.Much of these plants would need to be grown in marginalland, unused areas, not occupied by agricultural crops. Amongother under-utilised plants of such known potentialities areBrosimum alicastrum, Pittosporum hylliraeoides and Calliganumcomosum used as fodder types.

98 R.V. SinghSuggested ReadingsAsthana, A.N. (1988). Collection and Utilization of GeneticDiversity in Pulse Crops. In: Plant Genetic Resources­Indian Prespective (Edited: Paroda, R.S., Arora, R.K. andChandel, KP.S.), NBPGR, New Delhi, pp 243-254Bhale, N.L. and Narayanan, S. S. (1989). Recent Breeding Approachesin Rainfed Cotton. Proc. National Seminar on" IntegratedManagement Approach for Maximising crop Production inRainfed and Problem Areas, held at lARI, New Delhi, Feb.26-28, 1986 pp 305-312Narsinghani, V. G. and Tiwari, Anamika (1991). Inheritance of, Fusarium wilt Resistance in Peas. Froc. Golden JubileeCelebr-ations, Symposium on Grain Legumes~ held at IARI,New'Delhi, Feb. 9-11, 1991 pp. 279-282Singh, B. P. and Singh, R.V. (1989). Introduction of Crop Plants forRainfed and Problem areas. Proc. National Seminar onIntegrated Management Approach for Maximising cropProduction in, Rainfed and Problem Areas, held at lARI,New Delhi, p. 313-319Singh, B. P.~ Singh, R.V. and Koppar, M. N. (1991). GermplasmResources in Pulse crops-Introduction, Collection,Evaluation and their Conservation. Proc. Golden JubileeCelebrations, Symposium on Grain Legumes, held at IARINew Delhi, p.199-203.Singh, R.M., Raina Rajni, Nandan, R. and Singh, u.P. (l991).Pigeonpea improvement-current status and futurestrategies. Proc. Golden Jubilee Celebrations, Symposium onGrain Legumes, held at JARI, New Delhi, p. 212-225NBPGR (1990 to 1994). Annual Reports, National Bureau of PlantGenetic Rsources, New Delhi

Priorities for Promising Introductions of Medicinal,Aromatic, Spices and Condiments GermplasmDeep ChandIntroductionThe importance of medicinal and aromatic plants, spices andcondiments is well recognized since ages. From time immemorial,these were being utilized in Indian systems of medicine. However,concerted efforts were not made for their improvement so far.Many of plants belonging to this group are of Indian origin or havebeen introduced from other parts of the world, mainly from Indo­ClUna, IndoMalaysian, Central Asian, Mediterranean and European­Siberian regions of diversity. Zeven and Zhukovsky (1975)identified twelve centres of diversity of the economic plants. Outof these, Indo-China IndoMalaysian, Hindustani centre, centralAsian, Mediterranean and European-Siberian centres are importantfor more common plants of this group (Table-I).Centres of Diversity in Medicinal and Aromatic PlantsAbout 1100 species are utilized in Indian medicines forproduction of different formulation and most of these still comefrom the wild growth in forests and several are introduced in thepast such as opium poppy, psyllium, senna, cinchona, celery andperiwinkle (Gupta et aI., 1988). For meeting the large demand ofAyurvedic, Unani and Homeo medicines manufacturing industries,it is necessary to introduce more exotic collections from theircentres of diversity. As a result of efforts made throughcorrespondence to the foreign sources, the Division of GermplasmExchange of the Bureau successfully introduc;:ed 1175 {including'

100 Deep Chandcondiments). new introductions during preceeding years i.e.1990-94 (Table-2).Indian pharmacopoeia recognizes 85 . drug plants whoseingrediE;!nts are used in various pharmaceutical preparations. Theorigin/diversity of some of the important commercially grownplants under cultivation and deserve priority in our nationaleconomy are given below :Cinchona (Cinchona ledgerian.a): 65 species distributed in theAndes at elevations of 800 to 2800 m which occur mainly in Peru,Bolivia, Columbia and Ecuador; Psyllium (Plantago ovata) isindigenous to Mediterranean region; sarpgandha (Rauvolfiaserpentina) widely distributed in the sub Himalayan tract fromPunjab to Nepal, Sikkim, Bhutan, Assam and western Ghats andthe Andamans; Periwinkle (Catharanthus roseus) is supposed to bethe native of Madagaskar which escaped in tropics and sub tropicsof the old and new worlds (Uphot 1968).Indian senna (Cassia angustifolia) is native to Sudan, SaudiArabia but now nature.lized in Tamil Nadu, which is mainproducer in the world; Foxglove (Digitalis purpurea) is native towestern Europe and now cultivated in hilly regions in northernIndia. Pyrethrum (Chrysanthemum cinereriifolium) is a native ofDalmatia (Yugoslavia); an important insecticidal plant undercultivation (Kenya contributing 70% production of the world)(Gnadinger, 1945).Opium poppy (Papaver somniferum) native of Asia minor andindigenous to Mediterranean region which was introduced intoIndia in the early sixteenth century (India produces about 70% ofthe world's production and 90% is exported. Henbane (Hyoscyamusniger) is native of Europe, central Asia, India and Tropical Africa(Good, 1953) as well as of USA and USSR (Hocking, 1947),Egyptian henbane (H. nzuticus) grows wild in Egyptian deserts(arid zones of the Middle east), asvagandha (Withania somnifera) isreported in Mediterranean region and several countries of southeast and west Asia, viz., Baluchistan (Pakistan), Malaysia, Indonesia,Libya, Nigeria and Ghana from where genetic stocks can beprocured. .Some of the major aromatic herbs which are entering intointernational trade, viz., Peppermint (Mentha piperita) is widely

Priorities for Promising Plant Introductions 101used in medicines and in preparation of mint tea; spearmint(M. spicata) used for culinary purposes are popular throughout theworld specially in Egypt and several European countries. Ocimumsp., main centres of diversity are Africa, South America (Brazil) andAsia which are potential source of oil rich in camphor, geraniol,linalool, linalyl acetate, methyl chavicol, eugenol and thymol forsuccessful utilization in industry. Egypt, France, Italy, USA andWest Germany are major sources for assembling diverse germplasm.Sweet majoran (Majorana hortensis) is cultivated throughout Europeand mediterranean region as well as north and south America.France, Egypt and Chili are the major exporters in the world. Ammimajus is indigenous to Egypt and widely distributed in. Europe,mediterranean region, Abyssinica and West Africa; Rosemary(Rosmarium officinalis) grows wild throughout the mediterraneanarea whereas France, Spain, Portugal are the major source ofgermplasm; Sage (Salvia officinalis) is native to mediterraneanregion but most common in Yugoslavia and Albania; Celery(Apium graveolens) native to Europe extends from Sweden toAlgeria, Egypt, Abyssinia and in Asia from the Caucasus toBaluchistan and mountains of India. European dill (Anethumgraveolens) culinary herb extensively grows in India and Europe ,foruse as aromatic stimulant in Ayurvedic and Unani medicines.Centres of Diversity in Spices and CondimentsWild related taxa of major spices reported from the Indiansub continent and from the Western Ghats forest area in southIndia (Arora et aI., 1984). Sl,lb mountaneous tracts of Western Ghatsin India is believed to be the centre of diversity of black pepper.From this centre, it is believed to have spread tQ the other peppergrowing countries in south east Asia including Thailand, Malaysia,Indonesia, Philippines and Sri Lanka.Cardamon (Elettaria cardamomum) : It occurs in the evergreenforests of Western Ghats and' also in Sri Lanka. Guatemala andTanzania are other important cardamon growing countries fromwhere germplasm can be obtained for utilization in breedingprograrrune. Clove (Eugenia caryophyllus) indigenous to MoluccasIslands in Indonesia (Purseglove, 1977). Nutmeg (Myristica Jragrans)is indigenous to the eastern Islands of Moluccas (Indonesia ) andits nearest relative M. argentea is cultivated in some part of NewGuinea (Sinclair, 1958); Cinnamon (Cinnamomum verum) is native

102 Deep Chandof Sri Lanka and South West Indies. Ginger (Zingiber officinale),the country of origin is not known but certainty presumed to bein the region of India or China (Purseglove, 1977). Other importantginger growing countries are Nigeria, Sierra Leone, Jamaica,Taiwan and Australia; Turmeric (Curcuma Zanga syn. C. domestica),place of origin is presumed to be South Asia and cultivated specieshave naturalized in some areas of north eastern India and Islandsof Java:Condiments : Grain spices or minor spices constitute a group ofcondiments which have considerable opportunity for cropimprovement by introducing high yielding, better qualitygermplasm (genotypes) from mediterranean region.Utilization of Exotic GermplasmThe main recepients of germplasm introductions made by theBureau were the net work of All India Coordinated Project onMedicinal and Aromatic Plants, National Research Centre forSpices, various coordinating centres and scientists handlinggermplasm in agricultural universities and other centres. Besides,the Bureau too evaluated and conducted performance trials of suchmaterials at its headquarters and some of its regional stations.These efforts culminated in the identification of several promisinggenotypes, some of which are released as primary introductions.Promising IntroductionsIntroduction activities resulted in selection of promisingmaterials in crop plants. In this context mention may be made ofsome of the recent promising introductions in the following :Opium poppy: Accession (EC 179777 ex Hungry) is tall, late inflowering and bear small sized capsules rich in alkaloid content,line (Ee 196429 ex Finland) yields morphine (0.40-0.82%) and subsp. satigerum (Ee 232605 ex West Germany) presently being usedin specific crossing programmes.Licorice: A selection of the accession (Ee 114303 ex USSR) nowplaced under cultivation in Western and Central parts of thecountry produces 20-30 cm long thick stolens containing 5.5-8.0%of total glycyrrhizic acid acceptable to user ,industry in the country(Gupta et al., 1988). Another introduction (EC 120170) provedrei a tively resistant to rhizctonia root rot and showed response tohigher yield at closer planting.

Priorities for Promising Plant Introductions 103Psyllium: Accession (Be 321635) established well at Anand centre,mutants from accession (Be 42706) developed (Ee 42706-1, 42706-11 A, 42706-2/P2) were found promising in multilocation testingat some of the centres.Henbane : In the genus Hyoscyamus (H. aureus var. rhodes)accession (Be 251945) performed better with higher foliage yieldcontaining 0.207% total alkaloids. H. muticus (Be 251936, BC251938) produced higher herbage yield with average of 0.511% oftotal tropane alkaloids tested at Indore centre and H. aibus (BC146198) observed superior genotype providing dry herbage yield(24.67 g/plant) and total alkaloids (0.089%). H. niger (Ee 251943) __performed well prOViding higher fresh and dry herb yield of 408.33glplant at Indore centre. Ee 115996 of foxglove (Digitalis lanata.)from Poland was selected for higher content of glycoside in tRefoliage at Solan centre. Twenty accessions of different medicinaland aromatic plants established well at different coordinatedcentres at GAU, Anand (Gujarat), Yercaud (TN), Solan (HP),Indore (MP) and NBPGR Regional Station, Bhowali. These areDigitalis purpurea (Ee 202761 & Ee 303213), D. ambigua (Ee 333801-02 ex Japan), Aconitum nepallus (Ee 303220), Solanum laciniatum(303294), Salvia sclarea (Be 182901-02, Ee 314221, Ee 314226 andEe 314327), S. officinalis (Ee 314321). Seed setting started underSolan conditions in Silybum marinum (Ee 314324 Ee 308204 exHungary), Anacyclus pyrethrum (Ee 281897 ex France), A. depress us(Ee 340085), Melissa officinalis (EC 273873), Valeriana officinalis (Ee314322), Hypericum perforatum (BC 303214), Pimpinella saxifraga (EC314319), Papaver bracteatum (Ee 179994) Asparagus officinalis (EC280562-63 ex Russia), Satureja hortensis (Ee 328517 ex Iran),Hyssopus officinalis (Be 174790). "Periwinkle (Catharanthus roseus syn. Vinca rosea) : AICP on M& AP has identified "an accession (BC 120837 ex Poland) havingtotalleaf alkaloid yield (25.215 kg/ha), total root alkaloid yield(2.49%) and alkaloids Ajmalicine + serpentine (0.33%) whichoutyielded in different agrodimatic conditions (Singh et al., 1992).Ocimum : Its oil is used in perfumery and food flavouringindustry. Ocimilm basilicum (Be 176934 from France) with thehighest percentage of'oil (0.43%) and linalool (76.86%) and 43 exoticgermplasm lines were identified under Delhi condition for majorchemical components. EC 338781, Be 338775 and Ee 338778 are

104 Deep Chandmethyl chavicol types (0.960 to 91.24%), accessions (EC 338795,338779, 338784,338776) are linalool types (0.266- 69.19%) whereasgenotype (EC 312264) is methyl cinnamate type (16.56%) andaccessions (Be 282721) is eugenol type.In spices and condiments, some of the accessions established andperformed better under GAU, Main Spices Research Stati~m,Jegudan viz., cumin (EC 279053-54, 279081, 232684 and 243373)coriander (Be 279047-48, 232665-66, -69). Another accession highyielding and very early type (EC 357849), mid maturity types (EC363989, 363477-78), late maturity type accessions (EC 361712,363962 & 68), bold seeded (EC 363965-66 & 363980) and smallseeded genotype (BC 363981) were also observed.In fenugreek accessions like Be 257566, -206024, -243376, -241499),in fennel (Ee 257565 and Ee 257575), dill seed (Be 196641-42,232673-79), narrow and broad leaf variety of Piper nigru11l (Ee241341) introduced from Indonesia and now at NRC for spices forfurther evaluation were found promising.Priority Needs for IntroductionAlthough, NBPGR has introduced sufficient number of exoticgermplasm in medicinal, aromatic plants and spices and condimentswhich are representing quite large amount of crop variability withrespect to various desirable traits but now our emphasis is on forintroduction of germplasm resources with the following attributes.In opium poppy, annual temperate type collection from Finland,Hungary, Tasmania and Austria with high morphine content(more than 1%) for morphine prQduction and germplasm whichhas high range of variability in yield of latex, Senna germplasm .from Egypt, Sudan and Red Sea area having rich sinoside content(upto 4.5%A'f late flowering or more leafy biomass type are requiredto be introM:uced.'In Psyllium, high swelling mucilagenous seed germplasm, nonshattering which response to inputs with more g~netic variabilityin respect of spike length, tillering and resistance to downy mildewgenotypes. To widen the research work in Psyllium more materialshould be procured from Iran and Iraq. Further, Plantagomacrocarpa found over slightly alkaline wet lands in West Coastarea of North America provide useful genes for indehiseritcapsules which are required to be introduced. Germplasm of

Priorities for Promising Plant Introductions 105Periwinkle with high total alkaloid content (2.5%), more branching,thick root and leafy biomass should be procured from Brazil,Canada and Madagascar.Solanum laciniatum germplasm rich in sola so dine content which isconsidered to be an important raw material for commercialproduction of steroidal drugs should be introduced from Vietnam.Egyptian henbane germplasm with short duration, large biomassand Andrographis paniculata germplasm rich in andrographaloidcontent upto 2% and wild races from Sri Lanka are reqUired to beintroduced. In V£;tiver germplasm with high oil yield and superioraroma need to .be introduced from Indonesia, Reunion Island,Haiti, Guatemala, Mexico and Brazil.Oilpalm: Priority should be to introduce proven DXP combinationsand 30-103 P. pisifera tissue cultured plantlets from Nigeria anddurra parental lines from Malaysia, Ivory Coast and CentralAmerican regions.Black pepper : Disease resistant germplasm from growingcountries in south east Asia including Thailand, Malaysia,Indonesia and Sri Lanka need to be introduced.In Clove various genetic stocks from Tanzania and Mauritius onpriority basis, and in Cardamom from Guatemala, Tanzania andSri Lanka; in Cinnamon from Sri Lanka and the Seychelles Islands;in Nut meg (Myristlca argentea) nearest relative of nut meg shouldbe introduced from New Guinea for breeding programme.rt is necessary to introduce better quality, diseases resistant,high yielding germplasm material in all the seed spices such ascumin, coriander, fennel, fenugreek, celery, dill seed, caraway frommediterranean region for satisfying the requirements of thebreeders. This would help the country in augmenting the capacityfor production of essential oils and alkaloids which are muchrequired in drug pharmaceutical and essential oil industries.Selected ReferencesArora, RK. and E.R. Nayar (1984). Wild relatives of Crop Plants inIndia. NBPGR Scientific Monograph No.7 pp. 90Gnadinger r C.B. (1945), Pyrethrum Flowers Supplement. Me LaughlinGormley King CO' r Minneapolis r Minnesota, USA.

106 Deep ChandGood, Ronald (1953). The Geography of Flowering Plants. 2nd edn.,Longmans, Green & Co, London.Hocking, G.M. (1947). Economic Botany 1 : 306-316.Pursegove, J.W. (1977). Tropical Crops. Vol. I & II. Longmans GroupLimited, London.Sinclair, J. (1958). Gardeners Bull. Singapore, 16 : 205-466.Singh, B.M., Pareek, S.K., MandaI 5., Maheshwari, M.L. and Gupta,R. (1992). Variability in periwinkle (Catharanthus roseus).Indian Journal of Agricultural Sciences 62 : 47-50.Uphof, J.e. Th (1968). Dictionanj oj Economic Plants. 2nd ed StechertHafner Service Agency Inc., New York.Zeven, A.e. and Zhukovskyn, P. (1975). Dictionary of CultivatedPlants and their centres of diversity for Agriculture, Wageningen,pp.219.Table 1. Centres of diversity of the common medicinal andaromatic plants (Mega centres of cultivated plants ofZhukovsky, 1970)Centre of originPlan t species1. Chinese-Japan centre Panax ginseng, P. quinquifolius, Ginkgobi/oba, Plantago major~."'"U2. Indo-China Indonesian Cyrnhopogon citratus, C. nardus, Vetiveriacentrezizanioides (syn = V. odorata) -r:Gliilsaromaticus, Ocimum basilicum O.gratissimum, Pogosternon cablin3. Australian centre Duboisia leichhardtii, D. myoporides,Solanum laciniatum4. Hindustani centre Rauvolfia serpentina, Cymbopogon martinii,Vet£veria zizanoides, Jasminumgrandiflorum, J. samback, Plantago ovata,Atropa balladonna, Datura metal5. Central Asian centre Jasminum officinale, Papaver somniferum6. Near Eastern centre Pimpinella anisum, Papaver somniferum.

Priorities for Promising Plant Introductions 107Contd ..... (Table 1).Centre of origin7. Mediterranean centre8. African centrePlant speciesSilybum marianum, Lavandula officinalis(1. angustifolia) many cultivated varietiesare hybrids with wild relatives and 1.latifloia, Majorana hortensis, Melissaoffinali'l, Mentha aquatica, M. longifolia,M. puleqium (Eruopeto Iran), Rosmarinusofficina lis, Salvia officinale cultivated inmany gardens in temperate and tropicalcountries. S. sclarea Iran andTranscaucasia, Satureja hortensis CentralEurope and Siberia, Aloe barbadensisSpread to other regions i.e. Saudi Arabia,EastAfrica,N.W. India, S. Chian andcultivated in W. Indies, Urginea maritima,Algeria, Digitalis purpurea, Hyoscyamusniger, Papaver somniferumCommiphora ssp., Pelargonium graveolenscultivated in Algeria and Reunion Island,P. odoratiimum9. European-Siberian centre Cannabis sativa the wild form (c. ruderatis)is found in CentralAsia (centre-6);Humulus lupulus especially cultivated inEurope and N. America, Anthmis nobilis,Artemisia absinthium, A. maritima Europeto Mongolia, Carum carvi cultivated inNorth India and Sudan, Mentha'cordiaca,M. arvensis, M. longifolia, M. spicatea, M.viridis, Origanum vulgare, Glycyrrhizaglabra, Aconitum vulgare, Aconitum.napellus, Digitalis Zanata, D. pUrpurea,Levisticum officinale & Valeriana officinalis10. South American centre Cinchona ledgeriana and C. officinal is,Andes mountains of S. Peru, Bolvia and,S. Ecuador. There are many Cinchonaspecies found and great diverSity ofbotanical varieties is caused by naturalhybridization between the species andvarieties.

108Deep ChandContd ..... (Table 1).Centre of origin11. Central American andMexian centre12. North American centrePlant speciesTagetes erecta the area of the greatestdiversity is in South central Mexico;Vanilla fragrans (syn= V. plantifolio) aperennial vine cultivated in the tropicesand S. mexico for its aromatic fruitsAgriculture must have been introducedfrom centre-11 (central American andMexico centre). Panax quinquefolia(American ginseng), Mentha canadesllsis,(American wild mint) and DaturastramoniumTable 2.Germplasm introduction of medicinal and aromaticplants, spices and condimentsCrop species Accessions Source countryL Aconitum napellus 4 France, Germany2. Aloe sp. 49 France,J apan,Spain3. Artemisia annua 6 France, ltaly,Japan4. Atropa belladonna 4 Japan, Netherlands, Poland5. Anacyclus sp. 1 France6. Cassia angustifulia 3 Egypt, Japan7. Chrysanthemum cinera- 1 Germanyriifolium8. Datura sp. 20 Hungary, Netherlands9. Digitalis ptlrpurea 35 Germany, Hungary, USA10. Duboisia leichhardtii 2 Australia11. Geranium rotundifolium 2 Denmark, Egypt12. Gentiana Iutea 2 France, Japan13. Glaucium flavum 6 Denmark, France,Germany,Japan, Russia14. Hyoscyamus albus 4 Netherlands15. H. niger 4 Japan

Priorities for Promising Plant Introductions 109Contd ..... (Table 2.)Crop species Accessions Source country16. Hyssopus officinalis 8 Denmark, Hungary, J~pan,Netherlands17, Hypericum perforatum 2 France18 Jasminum sp. 21 Egypt, UK19 Lavandula angllstifolus 2 France, Italy, Netherlands,USA20 Levisticum officinale 5 France, Hungary, Italy,Netherlands21. Metricaria chamomilla 6 Hungary, Italy, Japan,USA22. Melissa officinalis 3 Denmark, France,Netherlands23. Mentha sp. 76 Australia, Denmark,France, Germany Italy,Netherlands, USA24. Ocimum basilicum 85 Australia, Egypt, France,Hungary, Italy, USA25 Origanum vulgare 3 France, Poland, USA26 Papaver somniferum 292 Australia, Bulgaria,Czechoslovakia, DenmarkGermany,Hungary, Japan,Netherlands,USA27 Panax sp. 4 Japan, USA28 Pelargonium sp. 1 USA29 Plantago psyllium 4 Japan30. Rauvolfia serpentina 2 Japan31. Rosmarinus officinalis 4 Egypt, Italy, USA32. Solanum varum 8 Egypt, Nepal, Netherlands33. Salvia officinalis 6 Egypt, Italy, Japan34. S. clarea 4 Bulgaria,France, Japan,Netherlands35. Satureja hortensis 10 Bulgaria, France, Italy36. Silybum marianum 5 France37 Valeriana officinalis 4 Germany38 Vinca rosea 10 France, Japan, USA39 Withania somnifera 5 France, Japan,Netherlands, USA

IIPromising Introductions and Prioritize Needs ofExotic Germplasm in Different Groups ofVegetable CropsPratibha BrahmiIntroductionThe Indian sub-continent had been assigned as one of thecentres of origin of different crop plants by Vavilov, and a goodnumber of vegetables like brinjal, cucumber, ridge and spongegourd, and root crops have been identified as native to thiscountry. Zevan and Dewet have also identified Hindustani centre,as an important centre of diversity for some of the vegetable cropsand their related species. This centre also possesses rich diversityand is a secondary centre for crops like Vigna unguiculata,Abelmoschus esculentus, Capsicum annuum and Citrullus lanatus.India has a past histroy of vegetable cultivation and somevegetable crops were introduced well before the Christian era.The domestication and diversification in some of theintroduced vegetable crops has not been properly studied, andalso duly less appreciated. European temperate type vegetableshave acclimatised to warm and humid conditions of north India,such that these have been transformed into a cosmopolitan cropadapted to wide range of tropical and sub-tropical conditions.Wide range of variability now available in chillies introduced byPortuguese in 15th centuary is another example of diversification.Several other examples can be quoted as in tomato, a comparitivelylate introduction, into India. As a result of rich diversificationalready there in native and introduced kind, prioritization of future

Promising Introductions and Prioritize Needs of Exotic Germplasm 111introductions and also conservation of existing types need to becodified, for proper utilization of genetic resources towards greaterenhancement of vegetable crops' yield and nutritive value. TheInternational Board of Plant Genetic Resources (now IPGRI­International Plant Genetic Research Institute) has indicated thefollowing criteria for priority such as :(1) Economic importance(2) Loss of genetic diversity(3) Nutritional value(4) Present level of researchIn India, genetic erosion is a serious problem in some of thevegetable crops. The classical example is of okra variety Pusasawani released in 1959-60, that swept off native varieties of okra,out of cultivation. This should not be allowed to occur in otherhorticultural crops like garden pea, cauliflower, watermelon etc.This highlights the need for (1) expeditions to other countriesespecially to Africa, Latin America and South East Asia(2) explorations within the country, to collect indigenous germplasm,their documentation evaluation and finally conservation.During 1986 to 1994, 14390 samples of exotic germplasm ofdifferent temperate, tropical and sub-tropical vegetable crops havebeen introduced by NBPGR from over 40 countries. Specificexpeditions have been organised with IPGRI in vegetable cropslike eggplant, okra, and curcurbits, cucumber, melons andpumpkin. In future intensive efforts should be directed to collectonion, chillies and cauliflower (specially tropical types).Promising introductions of the pastIn vegetable crops there are some outstanding exampleswhere introductions have been straight away taken by farmers andthere are also cases where some introductions have been utilizedsuccessfully in breeding new varieties. A few note worthyexamples are cited below :TomatoGarden peaSioux and La'Bonita (from USA), Balkan(from Russia), Roma (from Italy)Bonneville (from USA), Arkel (from France),Early Badger (froni USA)

112Pratibha BralzmiWater melonCabbageCapsicumAsahi Yamato (from Japan), Sugar Baby(from USA)Golden Acre (from Denmark)California Wonder and Yolo Wonder (fromUSA)CowpeaPusa Barsati (a selection from Philippinescollection).Franchbean Contender, Giant Stringless, KentuckyWonder (all from USA), Watex (from WestGermany)Ca uliflower Improved Japanese (from USA)CarrotNantes (from France)LettuceGreat Lakes (from USA)Cucumber Poinsette (from USA)BrinjalBlack Beauty (from USA)During recent years 1986-94, NBPGR has made significantcontributions towards introduction of vegetable germplasmpossessing tolerance to diseases and stress conditions includingdrought, salinity and heat, tolerance. Introductions of differentcrops from AVRDC, Taiwan, viz., cabbage, cauliflower, tomato,chinese cabbage have all heat tolerance and varying degree ofresistance to diseases specific to these crops. In tomato germplasmresistant to viruses and nematodes have proved useful. Besidesdifferent species of Lycopel'sicon which are being used in breedingprogrammes, viz., L. pimpinellifolium, L. hirsutum, L. cheesmani etc.,have also been introduced.Varieties resistant to bacterial wilt,fusarium wilt and nematodes have been received from USA andCanada.Carrot variety Beta III is a high carotene variety from USA.This has been repeatedly introduced and have also been suppliedto many projects on human nutrition. Also breeding lines of carrotwith male sterile and restorer lines with stumpy as well as taperingroot types have been received from Nertherlands.In cucumber, gynocious lines Gy4 and Gy5 have also provedvery useful in cucumber breeding prllgrammes for evolving highyielding all female plants with multiple disease resistance. In

Promising introductions and Prioritize Needs of Exotic Germplasm 113muskmelon also breeding line AC 70-54 has resistance to gummystem blight, powdery and downey mildew. In onion, Egyptianvarieties Giza 6 and Giza 20 have good potential with large darkred bulbs. Different Allium species have also been introduced forutilization in breeding programmes. Large cloves and colouredbulb types of garlic have been received from France and Sultanateof Oman. In cauliflower unusual green curded variety withtolerance to frost and good texture and flavour have been receivedfrom UK . In watermelon, efforts were made to introduce Fusariumwilt resistant varieties from USA.Future emphasisA brief oultline is given below in respect of some importantvegetable crops, about the requirements of germplasm and theirevaluation in relation to breeding investigations.Tomato: The collections in tomato should be augmented to meetthe requirement of producing varieties of various ecological zonesof wet tropics, drought prone areas, and high temperature zones,Production of good fruit set in hot weather with major resistanceto virus diseases, leaf blight, bacterial wilt, fruit borer andnematodes would have to be taken into consideration whilemaking introductions.Chillies : Hot pepper cultivars especially perennial varieties willhave to be collected, to screen for virus resistance. Sweet peppervarieties suitable for tropical conditions will have to be introducedfrom South America. Besides, new varieties with export potentialwill have to be evaluated under Indian condition.Watermelon: There is a gradual erosion of native germplasm dueto large scale cultivation of variety SugarBaby. Some of the nativevarieties with good storage potential under room temperatureconditions should be saved and also allied species of Citrullus willhave to be introduced from African countries.Muskmelon: Although diversity in muskmelon is being collected.Some native non dessert and cooking forms mistakingly identifiedas cucumber should be collected and also primitive cultivars fromnorth eastern India need to be collected. Resistance to virusdiseases and downy mildew and also, resistance to fruit fly, issought from exotic germplasm particularly from. central Asia,Middle East, and Africa.

114 Pratiblta BrahmiCucumber: Even though India is credited as centre of origin forthis crop, breeding and improvement have been very meagreespecially in native varieties suitable for hot and wet weatherconditions. There is also tremendous scope with gynoecious linesin breeding programmes.Pumpkin: A good amount of variability in Cucurbita moschata willhave to be screened for fruit set under long day and short dayconditions and resistance to virus and powdery mildew in exoticcollections from Central and South America. Similarly, in bottlegourd, germplasm needs to be enriched from African sources.Bitter gourd: Resistance to virus and fruitfly will be of importancein evaluation while new collections from South East Asia may beintroducedOnion: Primitive onion cultivars from Asian, African and CentralAmerican countries with photoinsensitivity must be augmented.Allium sp. with resistance to thrips, purple blotch, blight andstorage diseases, also need to be introduced and evaluated.Garlic : There is strong need to introduce garlic cultivars fromsubtropical countries and Europe to test their export potential.French bean: French bean cultivars tolerant to high temperatureconditions especially for the plains of north India and high levelof pest and disease resistance from central and south Americancountries are required.Cabbage: Apart from high temperature, black rot, soft rot anddiamond back moth are some of the important problems limitingproductivity in cabbage. Tropical cabbages developed in Taiwanand Philippines will have to be introduced and evaluated forIndian conditions so that new varieties can be developed.Okra: New material possessing resistance to yellow vein mosaicfrom Africa, should be introduced for intensive resistance breedingprogramme. Varieties with export potential should also be tested.These are only brief outlines for future programmes. Breedersshould emphasize the need in vegetable workshops and other foraso that NBPGR can be guided for future programmes.

IIGenebank Management System (GMS) SoftwareR.L.SapraIntroductionThe Genebank Management System (GMS) Software User'sGuide, the GMS software and the Guidebook for Genetic ResourcesDocumentation are three recent outputs of a project funded jointlyby IBPGR and the International Development Research Centre(IDRC), Ottawa, Canada. The Guidebook as well as personalcomputer software would help genebanks in developing their owntailor-made documentation system. The GMS has been specificallydesigned for the efficient management of information associatedwith the procedures commonly performed by gene banks. Thesoftware is designed for small to medium-sized genebanks thatare not currently using computer based documentation system forthe management of their data. However, the specifications of thesoftware have provided basis for even larger genebanks to use itand evaluation of the GMS software should be made bydocumentation and genebank personnel to determine whether itis suitable for use at the genebank (Perry et al., 1993)Software/HardwareThe GMS has been written in Paradox ApplicationLanguage (PAL) (Borland International) and works in PCenvironment (Microprocessors with 80286 and onward). However,GMS has been optimized for 80386 with 1 MB RAM. It is veryeasy to instill! the software. The installation of either the siteor world database requires 2.5 MB of space, but it has beenrecommended that at least 10 MB of free space be available forworking with the system.

116 R.L. SapraLogical Software OrganisationGMS basically consists of three major programs : DescriptorDefinition, Site Database, World Database. Descriptor definitionprogram (gbdefine) allows the user to define descriptors forvarious tables for individual crop and import them into thesoftware. It is worth mentioning here tha.t the descriptors list mayvary from crop to crop. However, the descriptors required for theregistration of accessions are same for all the crops. Thus in theentire system an accession is uniquely defined on the basis of asingle registration record and this accession number further helpsin linking the other tables.SitelWorld DatabaseThe software is installed in two separate directories: one forthe site database and the other for the world database. The basicdifference in the two is that the former is concerned with theinformation specific to an institute/genebank whereas the latterhelps in compiling the information originated from many institutesor genebanks. The world database does not store management data.DocumentlManagement ModeWithin the site database, the software has been organised intwo modes : Document and Management. These two modes arelinked together through registration table which contains uniqueaccession identifiers (accession numbers) as shown in the logical'data model. Each mode contains four tables. Document modestores and monitors information concerning registration of newaccessions, collecting iniJrmation, regeneration/multiplicationand characterization/ preliminary evaluation of the germplasm.The four tables comprising the Document mode are: Registration,Passport, Regeneration and Characterization. There is a singlerecord for registration and passport and one or more records forregeneration and characterization for each accession as shown inthe logical data model.The Management mode stores and monitors informationconcerning the registration of accessions, accessions maintenanceprocedures, inventory of accessions and their distribution etc. Thetables comprising management are : Registration, Accessionmanagement, Inventory and Movement. There is a single recordfor registration and accession management and one or morerecords for inventory and movement for each accession.

Genebank Management System (GMS) software 117Logical Data Model of GMS------------------------------r----------------T-------------------------------1Document Mode I Management Mode IPassportRegenerationCharacterizationI RegistrationIIIIIIII III IIII IIII1IIIIIIIIIII IAccessionI ManagementIII IIIIIIIIIII, IInventoryMovementI ______________________________ I L ________________! _______________________________ ~I !Registration table is a common table for both the documents.If a registration record is added in Document mode, another doesnot have to be added in Management mode.GMS Functions And UtilitiesThe GMS is highly user friendly and menu driven software.Through menus one can create masks (views) which are subsetsof descriptors for use in reports and data entry forms. In addition,it is possible to add new records, with automatic or manualassignment of numbers, search capabilities (create, modify, save,and delete) and reports in various formats. Utilities submenuallows for importing and exporting of data, deleting andcompressing (packing) of records in order to save the disk space.Importing of data through ASCII files requires a proper format.The report option can be used to create fixed field ASCII fileswhich can be used in other database management systems.III!I !II!III

118 R.L. SapraDescriptor Data TypesThe descriptor definition program makes a provision for 13data types : Absent/Present, Alplia, Choice, Country, Date,Latitude, Longitude, Low /High, No/Yes, Number, Percent, Poor /Good and Upppercase. Once these descriptors have been definedand have been used by the site or world database to store data,their data type cannot be changed. The country descriptor alsoprovides an access to pop-up menu of country names in data entry.Searching capabilitiesThe Search option lacks many features that are commonlyfound in some database management system software. Null ormissing values cannot be searched. The system also does not havethe facilities to work out the statistical parameters (Mean or. Standard Deviation etc.) or to perform various transformations orcalculations. Also, searching is somewhat slower than in otherdatabase management system software (Perry et al., 1993).ReferencesPerry, M.e, K.A. Painting and W.G. Ayad. 1993. GenebankManagement System Software User's Guide.International Board for Plant Genetic Resources, 1993.pp : 1-102.

IINew Policy on Seed Development and Role ofDesignated Inspection AuthoritiesRadhey ShyamIntroductionThe New Policy on Seed Development (NPSD) was introducedby the Government of India in October, 1988 with an objective tomake available to the Indian farmers the best genetic materialsavailable anywhere in the world to increase productivity, farmincomes and export earnings. Simultaneously, while importingseeds/planting materials, care has to be taken to ensure that thereis absolutely no compromise on the Plant Quarantine requirements!procedures in order to preclude the entry of exotic pests, diseasesand weeds detrimental to Indian Agriculture.Emphasis of the policyThe said policy has two basic aspects. Firstly, liberalisationof imports alongwith streamlining of Plant Quarantine proceduresand secondly, certain incentives to encourage domestic seedindustry. The policy has laid special emphasis on the followingitems:a) Import of high quality seeds/planting materials;b) Strengthening/ modernisation of Plant Quarantine facilities;c) Effective observance of Plant Quarantine requirements/procedures;d) Incentives to the domestic seed industry.

120 Radltey ShyamEligibility under policyThis policy for the import of seeds and planting materialscovers the following crQPs :1) Seeds of wheat and paddy;2) Seeds of coarse cereals; oilseeds and pulses;3) Seed,s of vegetables, flowers and ornamental plants;4) Tubers and bulbs of flowers;.5) Cuttings/saplings etc. of flowers;6) Seeds/planting materials of fruits.The policy does not encourage the bulk import of seeds ofwheat and paddy as leAR has already good collaboration withInternational Research Institutes, namely, IRRI, Manila andCIMMYT, Mexico. Further, the policy envisages that farmers needhigh yielding seeds from abroad, but at the same time repetitivebulk imports are not encouraged. As regards seeds/plantingmaterials of vegetables, flowers and ornamental plants, the policyhas much liberalisation and seeds of these crops are permitted tobe imported under OGL basis. As far as seeds/planting materialsof fruits are concerned, the policy envisages the requirement of thelicence from CCI&E.Import of seeds of coarse cereals, oil seeds and pulsesImport of such seeds for sowing are allowed for a period notexceeding 2 years by companies which have technical/financialcollaboration agreements for production of seeds with companiesabroad, proviaed the foreign supplier agrees to supply parent lineseeds / nucleus or breeder seeds / technology to the Indian companywithin a period of two years from the date of the import of the firstcommercial comignment after its import has been rec,-Imended byMinistry of Agriculture (Deptt. of Agri. & Cooperation).A certain quantity of seeds of coarse cereals / pulses andoilseeds for sowing. sought to be imported by eligible importersis given to ICAR 01 such farms which are accredited by ICAR fortrial/evaluation. After trial/evaluation for one crop season, therCAR intimates the results, agro-dimatic zone-wise within threemonths of the season to DAC. After the receipt of the results ofthe lCAR trial/evaluation, an eligible importer is entitled to apply

New Policy on Seed Development and Role of DIAs 121 ,for the import of such seeds on commercial basis to the DAC. DACmay within 30 days of the receipt of the application, reject theapplication or recommend it to the CCI&E for grant of an ImportLicence to the importer.Agencies eligible to importThe agencies as mentioned below are eligible to import theseeds/planting material:a) Department of Agriculture/Horticulture of the StateGovernments, State Agricultural Universities and reARb) Seed producing Indian companies/firms, afterregistration with the National Seeds Corporationc) National Seeds Corporation and State Seeds Corporationd) Food processing industrial unitse) Growers of vegetables/flow~rs registered with theDirector of Agriculture/Horticulture of the State, Governments.Samples to NBPGR Gene BankAll the importers are required to make available certainquantity of specified seeds / germplasm/ parental lines as per listenclosed (Annexure-' A') to NBPGR for testing / accessioning to theGene B::mk.Strengthening of Plant Quarantine facilitiesIn the past :various exotic pests and pathogens have enteredinto the country through the seeds/planting materials. Many ofthese pests have since become widespread, therefore, it isimperative that while importing seeds/planting materials, PlantQuarantine regulations are adhered strictly.Import of seeds/planting materialsThere are 26 Plant Quarantine and Fumigation Stations.(PQ & FS) located at 10 International Airports, 9 Seaports and 7Land Frontiers. But for sake of this policy, the im:ports of seeds/planting materials are allowed at present only through PlantQuarantine and Fumigation' Stations, at Amritsar, Bombay,Calcutta, Madras and New Delhi. Consequently, the PlantQuarantine facilities at thes~ 5 places are being strengthened/modernised.

122 Radhey ShyamThe import of seeds/planting materials is regulated as perprovisions of The Plants, Fruits and Seeds (Regulation of importinto India) Order, 1989 (PFS Order), issued under DIP Act, 1914.The pre-entry requirements for flowers and ornamental plants,flowers, vegetables seeds (FVS), bulbs and tubers (BT) and cuttings&. saplings (CS) are as under:Requirementi) Import permit (E)ii) Phytosanitary Certificate(PSC:E)iii) Additional Declarationa) Freedom from soil (E)b) Freedom from weedsc) Plant Quarantineobjectsvi) Approval of PEQfacilitiesFVS(E =existing; R ==required; NR =Not required)Plant Quarantine screening at the port of entryAll the imported consignments of seeds/planting materialsare subjected to Plant Quarantine screening at the port of entry.The screening may include visual inspection, laboratoryinvestigations, fumigation/chemical treatment and grow-out tests.Post-entry Quarantine (PEQ) and role of DIAs. Under Plants, Fruits and Seeds (Regulation of import intoIndia), Order, 1989, Cacoa and all species of the family Sterculiaceaeand Bombaceae, all species of Citrus (lemon, lime, orange, grapefruit etc.), Coconut seeds and plants (all species of Cocoa), Potato(all species of Solanum), Sugarcane (all species of Saccharum),Sunflower (all species of Helianthus), Tobacco (all species ofNicotiana), Wheat (all species of Triticum), cuttings/saplings/budwoods of flowers and ornamental plants and planting materials/seeds of fruits are subjected to post-entry quarantine inspection.In fact, no exclusive post-entry quarantine station is available withthe National Plant Quarantine Service at present.The post-entry quarantine facilities in which the importedplants/planting materials required to be grown are to be createdRRRRRNRBTRRRNRRNRcsRRRNRRR

New Policy on Seed Development and Role of DIAs 123 .by the importers vide guidelines given at Annexure 'B' and thepost-entry quarantine inspections are undertaken by the DesignatedInspection Agencies (DIAs) (Head of Plant Pathology, StateAgricultural Universities and certain Institutes under reAR). Aprocedure has been prescribed for preliminary inspection of theimported material at the port of entry by the concerned PlantQuarantine and Fumigation Station and forwardal of the samefurther to the concerned DIA through importer for post-entryquarantine purpose. The guidelines for DIAs are furnished atAnnexure 'C.Accordingly, the role played by the notified DIAs has becomevery responsible and significant for undertaking PEQ inspectiOns/monitoring from plant quarantine angle of the above mentionedseeds/planting materials. Hence, it is imperative that all DIAs arewell acquainted with the details of the plant quarantine procedures/requirements. ,The imported consignments are released by the concernedPlant Quarantine and Fumigation Station at the port of entry onlyon production of approval of DIA by the importers. Such importedmaterials are released at the port of entry after initial tests/laboratory investigations under intimation to the DIA concernedfor further monitoring for presence/absence of the exotic pests/diseases of Plant Quarantine significance for the specified periodto be grown in PEQ facifities created by the importer. In order tofacilitate smooth working of PEQ inspections, 41 DIAs have beennotified in different State/UTs, as per list annexed at Annexure 'D'.Sampling of the materialsThe sampling procedures in respect of various importedagricultural items are as under :-a) FVS - The sampling techniques shall be as perInternational Seed Testing Association (ISTA) Rulesand guidelines.b) BT & CS - The minimum sample size is consideredto be 10 numbers. However, in case of bulk imports,the representative sample for V.l. and 1.1. would not beless than 0.1%.

124 Radhey ShyamInspection requirementsThe inspection in respect of the categories mentioned abovecomprises of the following specific investigations/parameters :-InspectionFVSBT' CSi),Samplingii) V.Liii) L.r.a) X-Radiographyb) Washing Testc) Nematodedetectiond) Incubation fordiseases / pestsiv) G.T.v) PEQvi) Total duration(days)As perISTARulesRRRRRRNR45Min.or 0.1%RNRRRRRNR5010 Nos(R = Required; NR := Not required)Release of the consignmentsIn case the imported consignment exhibits the presence ormanifestation of any exotic pests, diseases or nematodes, the entireconsignment 'is rejected and is destroyed/deported. If thematerials show the presence of pests, diseases and nematodesalready reported in India, these are released after suitabletreatment to bring down the incidence to the prescribed levels,otherwise such consignments are also disposed off as prescribedby PPA.After Plant Quarantine screening/investigations, if theconsignment is found free from exotic pests and diseases, the sameis recommended for the release.Simplification by DACRNRRR"RNRR by DIAMin. 25Max. 45The Plant Quarantine procedures/requirements are reviewedfrom time to time by Deptt. of Agriculture and Co-operation in the

New Policy on Seed Development and Role of DIAs 125Ministry of Agriculture in order to streamline/simplify with aview to mitigate the problems of the imports/exports. Theproced1,lres have been streamlined so far as per details givenbelow:-Importi)ii)iii)iv)v)vi)vii)viii)ix)Issuance of Import permits (IPs) has been decentralised.Now, IPs are be:ing issued from Headquarters Office,Faridabad and Plant Quarantine Stations at Amritsar,Bombay, Calcutta, Madras and New Delhi. Earlier,these permits were issued only from HeadquartersOffice, Faridabad.The clearance period in case of seeds has been reducedfrom 45 days to 30-45 days and in case of bulbs/tubersfrom 50 days to, 35-45 days.The plants/plant materials requiring post-entryquarantine (PEQ) are released . within 1-3 days and 7days in case the laboratory tests are required at the portof entry.Tissue culture materials are released within six hoursat the port of entry.The requirement of Import Permit (IP) in case of plantsand plant materials meant for consumption and IP andPhytosanltary Certificates (PSCs) in case of the materialsimported through foreign post channels and asaccompanied baggage by air has been done away.The requirement" of the permit for the import ofMushroom Spawn Culture in respect of 100% ExportOriented Units has been dispensed with.The condition of permit for the import of Tissue Culturematerials of plant origin and flowers has been relaxed.The import of true potato seeds (TPS) and potato microtubers(in vitro) in tube!? has been allowed to generalpublic.The bulbs and tubers are subjected to PEQ, instead ofgrow-out test at port of entry, at the specific request ofthe importer.

126 Radlzcy ShyamExportThe list of the Officers in the Agriculture Department ofStates/Union Territories authorised to inspect/treat the plants/plant materials and to issue Phytosanitary Certificates in lieuthereof has been revised and enlarged based on 'therecommendations received from the States/UTs in order to extendmore facilities for export certification work.Import of GermplasmNational Bureau of Plant Genetic Resources (NBPGR) is thenodal agency for the exchange of germplasm material of all agrihorticulturaland silvi-agricultural crops for research purposes bythe institutions under the leAR system and private organisations.NBPGR has been authorised to issue import permit £01; gennplasmmaterials to authorised reAR 'institutes and private agenciesengaged in research work in accordance with the provisions of PFSOrder, 1989.ANNEXURE _ J AIMinimum weight of sample to be submitted to Gene Bank atNBPGR for safe custodyS1. ~ame ofNo. the Crop1. 2.Oil seed crops1. Rape Seed & Mustard2. Groundnut (Pod)3. Soyabean4. Sunflower5. Safflower6. Castor7. Linseed8. Niger9. SesamePulses1.2.3.4.BlackgramCowpeaMothbeanGramSize of thesample (gms.)3.10100050020090500151577004002001000

New Policy on Seed Development and Role of DIAs 127Contd ..... (Annexure A)I. 2. 3.5. Horsegram 5006. Greengram 1207. Lentil 608. Pea 9009. Khesari 7010. Redgram 300II. Frenchbean 700Fodder Crops1. Barseem 62. Lucerne 53. Oat 1204. Forage sorghum 905. Guar 5006. Sudangrass 257. Sweetclover 58. Guinea grass 29. Buffal grass 610. Setaria grass 2Millets11. Maize 90012. Sorghum (grain) 9013. Bajra 25Small Millets14. Common millet 1515. Finger millet 1016. Italian millet 917. Japanese millet (Barnyard millet) 818. Kodo millet 1019. Little millet 3ANNEXURE IB;Guidelines for importers for establishing PEQ facilities1. The importers of cuttings and saplings etc. of flowerswill be required to establish post entry quarantinefacilities, as prescribed by the Plant Protection Adviser.These facilities may include the establishment of a glasshouse/plastic house/any other specified facility, in adefined area, in isolation.

128 Radhey Shyam2. Imported cuttings, saplings, budwoods etc. of flowersshall be grown in the prescribed PEQ facility for aperiod specified in the import permit granted by thePPA.3. The importer shall facilitate the Designated InspectionAgency (DIA) to undertake inspection at specifiedintervals of time.4. The importer shall inform the DIA about the plantingdates well in advance. If there is any occurrence of pestor disease symptoms, he shall immediately inform theDIA. The importer shall abide by the decision of theOrA.5. No plant or parts thereof whether IhriPg or dead shallbe removed during the pendency of post entryquarantine without the approval of the DIA.6. A qualified Plant Pathologist/Entomologist/PlantVirologist/Horticulturist should be engaged asconsultant at the cost of the importer for looking afterthe plants under post entry quarantine.7. On the detection of any exotic pest or disease, the entireimported material shall be destroyed forthwith in aprescribed manner under intimation to PP A.8. During inspection, the multidisciplinary team of theDIA shall take special notice of symptoms of any plantdiseases specially systemic and viral diseases. Thedisease affected plants shall be destroyed in thepresence of DIA to avoid possible contamination of thesurrounding plants and environment by pathogens andthe soil shall be disinfected with formalin.9. The importer shall pay the cost of post entry quarantineinspection, as prescribed by PP A. He shall make thedeposit, as may be laid down, on the quarantineclearance of the consignment by the PP A. Theconsignment shall be released only after the importermakes the payment in full of the OIA inspection feesto the PPA.

New Policy on Seed Development and Role of DIAs 12910. The importer shall maintain basic inspection tools likehand lenses, microscopes, chemicals, dissection box,glass vials and petriplates.11. A log book shall be maintained for proper record ofincoming and outgoing material as well as the dates ofinspection and detailed Teport by the DIA.12. A register containing details of imported material,planting date, manifestation of diseases and appearanceof pests and control measures taken shall be maintained.13. The PEQ areas shall have the provision of lockingarrangements for security purposes.ANNEXURE 'e'Guidelines for Designated Inspection Agencies under the PlantsFruits and Seeds (Regulation of import in~o India) Order, 19891. Post entry quarantine (PEQ) is essential for importedcuttings, saplings, budwoods, etc. of flowers.2. Eligible categories of importers permitted to importcuttings, saplings, budwoods, etc. of flowers mustestablish post entry quarantine facilities, as may beprescribed by the PP A.3. A certificate shall be issued by the PP A or theDesignated Inspection Agencies in the States authorisedby the PP A under the Plants, Fruits and Seeds(Regulation of import into India) Order, 1989 forconducting PEQ inspections, that the importer hasestablished the prescribed PEQ facilities, after verificationand evaluation ofPEQ facilities created by the importer.4. Imported cuttings, saplings, budwoods etc. of flowersshall be grown in the prescribed PEQ facility for aperiod specified in the import permit granted by thePPA.5. The DIA shall be located in the State AgriculturalUniversity.

130Radhey Shymn6.7.A multidisciplinary team of the DIA consisting of aPlant Pathologist/Virologist/Entomologist shall inspectthe imported planting material at the time of plantingin the specified PEQ facility,Post planting observations shall be taken in thefollowing manner:Cuttings: The first observation would depend upon thegrowth of the plant but normally it should be undertakenwithin 20-25 days followed by second observationwithin 40-45 days of planting,Saplings : The first observation after planting shalldepend upon the growth of the plant, which maynormally be within 2-3 weeks.During inspection, special notice should be taken ofsymptoms of any plant disease, especially systemic andviral diseases,Disease affected plants should be destroyed in thepresence of the DIA, as prescribed by PP A, to avoid anypossible contamination by pathogens of the surroundingenvironment and plants,In the event of observance of any exotic pest or disease,the material shall be forthwith destroyed in theprescribed manner under intimation to PP A.8. The final report of PEQ inspection shall be sent to PP Afor information and record within 15 days of the 'finalobservation.ANNEXURE- '0'List of Designated Inspection Authorities (DIAs) notified underThe Plants, Fruits and Seeds (Regulation of import into India)Order 1989,51. State/Union JurisdictionNo, Territory1 2 31. Andhra Pradesh Entire StateDesignated InspectionAuthoritiesHead, Division of PlantPathology, Andhl'LI PradeshAgricultul'al Univel'sity,Hydcmbad,

New Policy on $eed Development and Role of DIAs 131Contd ..... (Annexure 'D')12342.3.4.5.6.7.8.9.AssamArunachal PradeshAndaman andNicobar Islands'BiharBiharDaman and DiuChandigarhDelhi10. Goa11. Gujarat12. HaryanaEntire StateEntire StateEntire Union. TerritoryExcept North~mcj. SouthChhota NagpurRegion.North and SouthChhota Nagpur,Santhal Region.Entire UnionTerritoryEntire UnionTerritoryEntire UnionTerritoryEntire StateEntire StateEntire StateHead, Division of PlantPathology, Assam AgriculturalUniversity, Jorhat.Joint Director, Indian Councilof Agricultural Research,Research Complex for NorthFastern Hill Region, ArunachalI'radesh Centre, Basal',Arunachal Pradesh.Officer-in-Charge,Indian Council of AgriculturalResearch, Research Complex,Port Blair.Head, Division of PlantPathology, Rajendra .Agricultural SanthalUniversity, Pus a, BiharHead, Division of PlantPathology, Bisra AgriculturalUniversity, Ranchi, Bihar.Head, Division of PlantPathology, Gujarat AgriculturalUniversity, Banaskantha.Head, Division of PlantPathology, Punjab AgriculturalUniversity, Ludhiana.Head, Division of PlantPathology and Mycology,Indian Agricultural ResearchInstitute, New Delhi.Officer-in-Charge, IndianCouncil of Agricultul'ulResearch, Reseal'ch Complex forGoa, Ele. Farm, Ele, Old Goa-403402.Head, Division of PlantPathology, Gujarat AgriculturalUniversity, Dantiwada.Bead, Division of PlantPathology, c.C.S.Agricultural University, Bissar.

132Radhey ShyamContd ..... (Annexure 'D')1 23413. HimachalPradesh14: Himachal Pradesh15. Jammu andKashmir16. Kamataka17. Karnataka18. Karnataka19. Kerala20. Lakshadweep21. Madhya PradeshEntire State(Agriculture)Entire State. (Horticultureand Forestry)Entire StateShimoga,Chhitterdurga,South Kanada,Chikmagloor,Kolar, Bangalore,Hassan, Kurg,Mandya, MysoreBelgaon, Bellary,Bidar, Bijapur,Dharwar, Gulbarga,Raichur, andUttar KannadaEntire StateEntire StateEntire UnionTerritoryExcept RaipurDurg, RajnandgaonBilaspur, Raigarh,Surguja andBastar.Head, Division of PlantPathology, Himachal PradeshKrishi Vishva Vidyalaya,Palampur.Head, Division of PlantPathology, Dr. Y.S. ParmarUniversity of Horticulture andForestry, Solan.Head, Division of PlantPathology, Shere-KashmirAgricultural UniVersity ofScience and Technology,Srinagar /Jammu.Head, DiviElion of PlantPathology, University ofAgricultural Sciences,Bangalore. .Head. Division of PlantPathology, University ofAgricultural Sciences,Dharwar ..Director, Indian Institute ofHorticultural Research,Bangalore (Kamataka).Head, Division of PlantPathology, Kerala AgriculturalUniversity, Trichur.Head Division of PlantPathology, Kerala AgriculturalUniversity, Trichur.Head, Division of PlantPathology, Jawahar Lal NehruKrishi Vishwa Vidhalaya,Jabalpur.

New Policy on Seed Development and Role of DIAs 133Contd..... (Annexure 'D')1 23422, Madhya Pradesh23. Maharashtra24. Maharashtra25. Maharashtra26. Maharashtra27. Manipur28. Meghalaya29. Mizoram .30. Nagaland31. OrissaRaipur, Durg,Rajnandgaon,Bilaspur, Raigarh,Surguja and Bastar.Konkan andRevenue Divisionof BombayRevenue Divisionof Pune and NasikRevenue Divisionof Aurangabad(7 Districts)Revenue Divisionof Nagpur andAmravatiEntire StateEntire StateEntire StateEntire StateEntire StateHead, Division of PlantPathology, Indira GandhiKrishi VishvaVidyalaya, RaipurHead, Division of· PlantPathology, Konkan KrishiVidyapeeth, Dapoli.Head, Division of PlantPathology, Mahatma PhuleAgricultural University, Rahuri.Head, Division of PlantPathology, MarathwadaAgricultural University,Parbhani.Head, Division of PlantPathology, Punjab Rao KrishiVidyapeeth, Akola.Joint Director, Indian Councilof Agricultural Research,Research Complex for NorthEastern Hill Region, ManipurCentre, Manipur.Officer-in-Charge, IndianCouncil of AgriculturalResearch, Research ComplexMaghalaya.Joint Director, Indian Councilof Agricultural Research,Research Complex for NorthEast Hill Region, MizoramCentre, Kelasib, Mizoram.Joint Director, Indian Councilfor Agricultural Research,Research Complex for NorthEastern Hill Region, NagalandCentre, Jharnapani, Nagaland.Head, Division of PlantPathology, Orissa University ofAgriculture and Technology,Bhubaneswar.

134Radlzey SltyamContd ..... (Annexure '0')23432. Pondicherry33. Punj

IISymptoms of Fungal and Bacterial DiseasesP.e. AgarwalIntroductionDisease in plants may be defined as any variation from thenormal as expressed either by the interruption of physiologicalactivities or by structural changes which are sufficiently permanentto check development, cause abnormal formations, or lead to thepremature death of a part of a plant or of the entire plant. Thefact that a plant is diseased comes to our attention mainly throughthe symptoms, i.e., how the plant looks after the pathogen hasestablished itself in the host. In other words symptoms are theexpressions of any reaction of a host to a disease causing agent.In many cases the symptoms become apparent only after thedamage to the host is done. In most cases the symptoms becomeapparent over a long period of time, and in increasing intensities.In a few cases the pathogen, though present in the host, is eithera virulent or too mildly virulent to cause any damage to the hostand hence no apparent disease symptom is produced. Such a hostis known as a symptomless carrier.Among common type of symptoms produced by fungi andbacteria are: leaf spots and leaf blights, anthracnose, damping off,root rot, wilt, foot, stem and stalk rot, powdery mildew, downymildew, rust, smut, bunt, scab, and canker.Sometimes only one type of symptom is produced, e.g., "leafspots, but increased severity will frequently lead to othersymptoms. Leaf spots may coalesce and defoliation occurs .. This

136 P.c. Agarwalmay further result in stunting of the whole plant. Some pathogensproduce a wide range of symptoms. Xanthomonas campestris pv.·malvacearum causes angular leaf spot, veinal necrosis, boll rot, andstem dieback (black arm). The type and severity of symptomsproduced depends on sevet'i'l factors including climatic conditions,resistance of the host, site and method of infection and possiblythe amount of in'oculum.The mechanism involved in symptom expression is complex.Toxins may be produced to cause necrosis or chlorosis, as withFusarium oxysporum, Pseudomonas syringae pv. phaseolicola; enzymesmay be involved, as with soft rotting Erwinia spp. that producepectinases; plasmids may be involved, as with Agrobacteriumtumefaciens to produce a tumor, etc.Symptoms on seedFungal pathogens produce various types of symptoms onseed which may be specific or non-specific. Purple discolourationof a part of whole soybean seed indicates infection of Cercosporakikuchii. In case of downy mildew of soybean (Peronosporamanshurica) dull white crusts are present on the seed. Distinct 'eyespot' with grey centre and brown to dark brown margins on wheatseed shows infection of Drechslera sorokiniana (seedling blight/rootrot). Shrivelled, whitish/pinkish light weight seeds of wheat andmaize indicate infection by Fusarium spp. (foot rot, ear scab). Blackcoloured, dotted structures on the surface of groundnut, sesame,maize seed reveal infection of charcoal rot pathogen (Macrophominaphaseolina). In Karnal bunt of wheat (Neovossia indica), seed is. converted into dark brown/black powdery mass to varyingdegrees along the furrow. In case of paddy bunt (Neovossia horrida),minute pustules or streaks of black colour brush through theglumes, the seed may be partly or wholly converted into blacksp·ore mass. In the case of coriander gall disease (Protomycesmacrosporus) seed shows hypertrophy of a part of entire seed. Inthe case of wheat bunts, covered smut of barley and oat, grain smutof sorghum, pearlmillet etc. entire seed except seed coat isconverted into black powdery mass of spores, while in ergotdiseases of cereals and grasses individual grains are converted intolong, black hard sclerotia.Xanthomonas campestris pv. phaseoli in bean is detectable bythe presence of the va~nished masses of bacteria in the hilum area.

Symptoms of Fungal and Bacterial Diseases 137Xanthomonas c011lpestl'is pv. ph as eo Ii and Pseudomonas syringae pv.phaseolicola produce a bluish-white fluorescence on bean seeds.Symptoms on host-plantLeaf spotSpots are the most common symptoms observed on a widevariety of plant species. Spots start as minute, chlorotic flecks,which later increase in size and become necrotic. Spots may appearin different shapes, sizes and colours like round, elliptic, elongated,oval or irregular and pale, brown, dark brown, reddish to black.Examples: Spindle shaped (Pyricularia onjzae); long stripes(Drechslera graminea), oval, brown with grey centre {Drechsleraoryzae)j circular to slightly angular with grey centre and reddishborder (Cercospora spp.); spots with concentric rings (Alternariaspp.); and water soaked and raised on lower surface and depressedon upper surface (Xanthomonas vesicatoria).Leaf blightBlight implies sudden and extensive damage to leaves.Under favourable conditions, the spot increase in size, coalescewith each other and occupy larger area of the infected leaves. Atthis stage the leaves appear blighted (scorched). Beside leaves,blight appears on stem. Ultimately whole plant may die. Examples:. Early blight of potato (Alternaria solani); late blight of potato(Phytophthora infestans); maize leaf blight (Drechslera maydis);Phyllosticta leaf blight of maize (Phyllosticta maydis); Halo blightof beans (Pseudomonas phaseolicola); Common blight of beans(Xanthomonas phaseoli); Phomopsis blight of egg plant (Phomopsisvexans); Bacterial blight of pea (Pseudomonas pisi); and Ascochytablights of peas (Ascochyta spp.).AnthracnoseThe anthracnose has been used for two distinct types ofdiseases, one producing typical necrotic spot like lesion of deadtissues and the other like raised border around a more or lessdepressed centre. It develops most commonly on the youngsucculent parts of stems, petioles leaves, fruits and rarely on roots.In case of beans, usually the lesions are dark brown, in moistweather, pinkish spore masses can often be seen. Elongated spots.appear on the veins on the lower side of the leaf and later spread

138 P.e. Agarwalto upper surface. Petioles, stems, and hypocotyle are oftenaffected, causing death of the plant. On the immature bean pod,small brown spot appear which enlarge rapidly upto a diameterof 1 cm. They become dark brown to black and are depressed inthe center. Sometimes the developing bean seeds becomesinfected. Small, black, oval to elongated fruiting structures(acervuli) develop on stems and leaf sheaths as the plants approachmaturity, which serve as the most reliable diagnostic feature.These acervuli have raised appearance due to the presence ofnumerous black setae (spines) that protrude above the plant tissue.Other hosts are maize, sorghum, cucumber etc. Examples: Beananthracnose (Colletotrichum lindemuthianum); anthracnose ofcucurbits (C. lagenarium) and anthracnose of chillies (c. capsici andC. piperatum).Damping offA disease of germinating seeds and seedlings is collectivelyknown as damping off and seedling blights. It is of two types:Pre-emergence : Emergence is poor even with seeds of highgerminative capacity; there are patches (in the nursery) with noseedling at ail. Post-emergence: Very serious d~sease particularly, in crops raised through nurseries. Seedlings that have emergedoften show water soaking, browning or shrivelling of the stemtissues at soil level and subsequent death of the seedlings. Whenthe plants are pulled up they are found to have extensive browningand rotting of the smaller roots or stem lesions at soil level. Sometimes, seedlings may show drooping symptoms as if due to waterstress, although there may be enough water in the nursery bed.Fungi involved are: Pythium, Phytophthora, Fusarium, Rhizoctoniaand Sclerotinia.Foot, Stem and Stalk rotInfection start at or above soil level Le. the basal portion ofstem (foot rot) in the form of small brownish discolouration whichgradually increases in size and finally girdles the whole stem. This.gives rise to a number of symptoms in the Sho0ti growth is checkedand the plant becomes stunted, as the plant can not obtain thewater and nutrients for its growth. Dropping and wilting of theleaves takes place, which later on tum yellow, some drop andeventually the plant collapses and die. Sometimes the rotting

Symptoms of Fungal and Bacterial Diseases 139extend toward the stem and downwards in the roots producingsame symptoms of discolouration and softening of. tissues.Otherwise in general, roots remain healthy. Examples: Stem andfoot rot of papaya (Pythium aphanidermatum)i red rot of sugarcane(Colletotrichum falcatum); foot rot of linseed (Phoma exigua var.linicola); stem rot of jute (Macrophomina phaseolina).Powdery mildewInitial symptom starts as minute, pale coloured spot on theupper surface of the leaves. From these spots whitish, powderygrowth of the fungus develo~~". Later on, the powdery growthspread an.d gradually cover the whole leaf, and then to all aerialparts including stem and fruits. Minute pin head sized, darkcoloured bodies (c1eistothecia) may develop and scattered in thepowdery growth. Ultimately, the leaves turn greyish or lightbrown and finally wither away. Examples: Powdery mildew of pea(£rysiphe polygoni); powdery mildew of wheat (E. grnminis); grapepowdery mildew (Uncinula necator). . .Downy mildewOn Monocots : The plants are infected systematically from earlygrowth stage. Leaves appear pale yellow, severe stunting of plantand excessive tillering. The leaves show chlorotic stripes ofvarying size on the upper surface. On the underside or on bothside of the leaves, there is a whitish downy growth of the fungus.Many severely diseased plants die during the tillering to earlyelongation growth stage. Plants may remain sterile. In case ofpearl millet, the diseased plants may develop thickened andtwisted leaves and usually fail to head. The floral parts aretransformed into green leafy structures (green ear phase). Theseplants rarely produce viable grain. Examples: green ear of bajra(Sclerospora graminicola); downy mildew of sorghum(Peronosclerospora sorghi).On Dicots : Upper surface of the leaves (infected plants) turnyellowish, grey, light yellow with fine downy growth of whitegrey, purplish or bluish colour. Symptoms also appear on stem,twigs, petioles and fruits in the form of downy growth. In caseof severe infection, inflorescence may become deformed due tohypertrophy. Examples: Downy miidew of crucifers (Peronosporaparasitica); blue mold of tobacco (P. tabacina)j downy mildew of

140 P. C. Agarwalsoybean (P. manshurica); downy mildew of grapes (Plasmoparaviticola)i downy mildew of sunflower (P. halstedii).RustRust fungi are host specific, symptoms appear on leaf- stem,leaf sheath, ear, and even pod. Symptoms may appear initially assmall flecks, round or elongated; scattered all over the leaf surfaceor in the form of stripes on stem or leaf sheath. Colour of thesepustules may be either pale yellow, orange, brown, red or black.Examples: Wheat stem rust (Puccinia graminis f. sp. tritici); wheatleaf rust (Puccinia reco1tdifa); brown rust of barley (Puccinia hordei);linseed rust (Melampsora lini); sugarbeet rust (Uromyces betae);maize rust (Puccinia polysora); safflower rust (P. carthami); onion,leek and garlic rust (P. allii).SmutThere are three basic types of smuts, which are distinguishedby the way in which they infect a host plant. (a) Corn smut: spores(Ustilago maydis) can penetrate the plant and cause disease at anygrowing point (local ,infection); including the tasset leaves andalso developing corn (maize) kernels; (b) Covered smut: infectsyoung barley, oat or wheat seedlings (seedling infection); (c) Loosesmut (like loose smut of wheat and barley-Ustilago nuda) infects theovary of cereal plants when plant is flowering. (floral infection).In case of pearl millet smut (Tolyposporium penicillariae) only fewgrains are replaced by spore mass. The smut sari are projected,green at initial stage then turns brown in colour. Examples: Headsmut of sorghum (Sorosporium reillianum); loose smut of oat(Ustilago avenae); covered smut of barley (U. hordei); loose smutof wheat (U. tritid).In case of leaf smuts symptoms are restricted to leaf and leafsheath, in the form of greyish black, slightly swollen bands, stripesor elongated, linear spots. Leaves may shred along the infectedportions which contain spore mass of the fungus, ultimately thespore masses are released in the air and soil. Examples : Smut(En tyloma oryzae); flag smut of wheat (Urocystis tritici); Onion smut(U. cepulae).BuntLike ear smuts, in case of bunts, the symptoms are onlyvisible at the time of heading. In hill bunt of wheat (Tilletia caries

Symptoms of Fungal and Bacterial Diseases 141and T. foetida) ears are greener than the normal ears, later onglumes open apart due to bunted grains. Bunted grains are smallerand darker than normal ones. In case of Kamal bunt of wheat(Neovossia indica) and paddy bunt (N. horrida) only few grains ina ear show infection. Grains are usually partially infected. At thetime of ear head maturity, outer glumes spread and inner glumesexpand, thereby exposing the bunted grains. Spore masses remaincovered with the pericarp. The bunts can be recognised by adistinctive smell of decaying fish.Gall and WartThese symptoms may appear on any plant part both aerialand underground. These out growths are produced due touncontrolled multiplication of cells, (i.e. hypertrophy = increase insize of plant cells, or hyperplasia = increase in number of cells).Examples: potato wart (Synchytrium endobioticum); coriander gall(Protontyces mac/'ospom) and crown gall of apple (Agrobacteriumtumefaciens) .ScabThe term scab refers to a roughened or crust like lesion onthe surface of the host. Examples: Common scab of potato(s treptomyces scabies), scab of cucurbi ts (Cladosporium cucumerinum).CankersCankers or warty outgrowths are formed on leaves, twigs,and fruits. They result from the necrosis of tissues and reactionof undamaged tissues to produce cork cells. Such diseases aremostly localized. Examples: citrus canker caused by X. campestrispv. citri and bacterial canker of tomato (Corynebacteriummichiganense).

IIDetection Procedures for Fungal Plant Pathogensand Salvaging of Infected GermplasmA. Majumdq.rIntroductionAmong the pathogens of plant quarantine significance, fungiform an important group and their association and spread with themovement of germplasm is well documented in the literature.Detection of fungi in seeds or planting material requires verycareful examination and use of specialised techniques. Thetolerance level is zero for any exotic pathogen intercepted fromimported germplasm. Hence, the detection techniques must besensitive enough to detect the lowest level of infection. Techniquesused for detection of fungal pathogens.in quarantine laboratory aredescribed below.Detection techniquesDetection techniques can be grouped into two broadcategories:(a)(b)Generalised tests: which help in detecting a wide rangeof pests/pathogens.Specialized tests: used for the detection of specificpests and pathogens.A. Generalised Tests1. Visual examination: This is carried out by inspection of dryseed with unaided eye, hand lens or under low power ofstereoscopic microscope. Each seed sample should be first'

-Detection Procedures for Fungal Plant Patl10gens and Salvaging 143examined visually and under low power of stereobinocularmicroscope before subjecting to incubation or any other specializedtests. By this method it is possible to detect sclerotia, smut balls,discolouration, malformations, hypertrophies, fungal spores andother fructifications such as pycnidia, perithecia etc. Also we candetect inert matter which includes broken pieces of plant parts,sand or soil particles, stones and seeds of other crops includingweed seeds.During quarantine processing of germplasm samples ofvarious agri-horticultural crops with this method we couldintercept spore crusts of Peronospora manshurica (downy mildewof soybean); uredio and teleutospores of Puccinia carthamii (rust ofsafflower); bunted grains due to Neovossia indica (Kamal bunt);Tilletia caries and T. foetida (wheat bunts); ergot sclerotia (Clavicepspurpurea) in wheat, rye, barley, oats and a number of grasses etc.2. Microscopic examination of suspensions obtained by seedwashing: The method is used only for surface-borne, contaminatingfungi. The seed is shaken along with water (having a few dropsof detergent) for a fixed period with the help of a shaker ormanually. The resultant suspension may be examined directlyunder a stereobinocular microscope or the suspension may beconcentrated by centrifugation. The concentrate (sediment) can bediluted in small quantities of water and observed under thecompound microscope. The method has been used for detectionof smuts, bunts, downy mildews, powdery mildews, rusts andalso spores of Alternaria, Cercospora, Drechslera, Fusarium,Pyricularia etc.3. . Incubation method: Incubation tests can be used successfullyagainst surface borne as well as internal infections. After plantingof seed, the incubation period gives an opportunity to the dormantmycelium of fungal spores to grow alongwith the host. Mostcommonly used incubation tests are dealt below:(a) The blotter method: Usually 400 seeds of each sample tobe tested are placed on 3 layers of moist blotters in plasticpetritplates (10 to 25 seeds per plate, depending upon the size ofthe seed) and incubated under near ultraviolet light/whitefluorescent tubes in alternating cycles of 12 hours light/ darknessfor 7 days at 20°C±2°C. Identification is mainly based upon thestereobinocular observation. However, slides are made and

144 A. Majumdarobserved under a compound microscope, whenever required. Incase of stored seeds, saprophytes such as species of Alternaria,Aspergillus, Mucor, Penicillium and Rhizopus may create hindrancesin observations. Using this method in our laboratory we coulddetect a large number of seed-borne pathogens including thosewhich are not reported from India.Sometimes the growth of seedlings in the blotter testinterferes during recording of observations under stereobinocularmicroscope. To overcome this problem the germination of seedsto be tested by the blotter method may be retarded or stopped byusing a 0.1-0.2% water solution of the sodium salt of2A-dichlorophenoxyacetic acid (the herbicide 2A-D) in place ofpure water for dipping the blotters (Neergaard, 1969). Thisprocedure is widely used in testing the seeds of crucifers for Plzomalinganz, the black rot pathogen.Another way of retarding the germination of seed is by usingdeep freezing method (Limonard, 1966, 1968). In this case seedsafter plating are kept at 20°C for 24 hours, then shifted to -20"Cfor the nest 24 hours and finally incubated at 20"C for theremaining 5 days. Exposure to -20°C kills the germinating seeds,thus, making available enough food material for better growth ofcertain fungi. The method has been used for the detection ofDrechslera, Fusarium and Septaria infection in cereals and grassesbut can also be used for many other seed borne fungal pathogens.Rolled paper towel method can be applied for fungi whichproduce visible symptoms on seedlings.(b) Agar plate method: The principle of recording in the agarplate test is macroscopic examination of fungal colonies. Themethod is qUite reliable and quick but one has to be wellacquainted with colony characters of different fungi on agar media.Seeds are generally pre-treated with mild disinfectant suchas sodium hypochlorite, before plating on agar to prevent overgrowth on the seeds by saprophytic fungi. A soak for 10 min. ina 1% solution of chlorine in water is the usual pretr~atment butin certain seeds it has been observed that the pretreatment from30 seconds to 40 min. has little effect on the test results (Hewett,1979). Other pretreatment chemicals used are Ca (OCl)2' H 20z andHgCl2_' Except with the mercury pretreatment washing of seedsis not necessary. .

Detection Procedures for Fungal Plant Pathogens and Salvaging 145Potato dextrose agar (PDA) is the most commonly used agarmedium; other media used are: malt extract agar (MEA), oat mealagar (OMA) etc. Antibiotics are often added to limit the growthof bacteria. Usually 5 to 10 seeds per plate are placed andincubated for 7 days at 20 to 25°C. The method is used for thedetection of Drechslera teres and D. graminea in barley; D. avena inoats, Colletotrichll7n lini in linseed; Ascochyta spp. in legumes, etc.However, the method is unsuitable for slow growing pathogensbecause they are overgrown by other pathogens or saprophytes,e.g. Pyricularia oYljzae is readily overgrown by Trichaniella padwickii.4. Examination of symptoms developed on seedlings grown insoil or sand : Sowing the seeds in sterilized soil, sand, or gravel,etc. provides more natural conditions and under such conditionsinfected seeds and seedlings may develop symptoms comparableto those developed under field conditions.(a) Sand method: The seeds are sown at the depth of 3-4 em incoarse grained sand. For small seeds, the depth js of 1 em only.The method is used for cereals in Germany. It is also applied fordetection of Drechslera, Septaria and Fusarium infections in wheat,oats and barley. In Israel, a combined routine method is used fordetection of Colletotrichum gossypii (Neergaard, 1969). The methodis time consuming but provides information comparable to fieldperformance.(b) Soil method : A uniform soil mixture composed of 4 partsof clay and 6 parts of peat with fertilizers is used for sowing. Themethod is used in Sweden where the soil is sold in plastic bags.The medium for the test is prepared by mixing 3 parts of abovesoil mixture, 1 part of sterilized sand and 0.6 parts of water for5 minutes to get uniformity and filled in plastic multipot trays.Plates after sowing of seeds are covered with plastic bags to avoidwater loss throughout the incubation. Cereals are kept at 10 D C forSeptaria and Drechslera. The symptoms are then recorded and thepercentage of attacked seedlings is calculated.5. Grow out test: Certain seed-borne diseases need' longerperiods for their expression than provided in the normal incubl1 t It mtests; specially some bacterial and viral diseases. In plantquarantine system, growing on tests have gre?-t importance, asexemplified from Portuguese quarantine import inspection of seed

146 A. Majumdarsamples. Three devastating pathogens which were not known tooccur in Portugal at the time of inspection were intercepted.For certain seed-borne pathogens, routine laboratory testingof seed samples is regard~d unsatisfactory. In such casesinspections of representative samples against Drechslera graminea(leaf stripe) and Ustilago nuda (loose smut), is undertaken. Also,similar procedures are adopted for rice seed certification schemesin Portugal.B. Specialized TestsSpecific seed health testing methods have been developed fromtime to time in seed pathology for a variety of fungal and bacterialpathogens. Such methods are specific in the sense, that they havebeen developed for a particular pathogen in a crop and mostlyrestricted for that purpose only. In certain cases such methodshave been used for some additional pathogens or crops (Table-i).Table 1.Specialised methods developed for detection of fungalpathogens .Test Method :r'athogen CropSeedling symptoms test Septoria nodorum WheatDrechslera graminea BarleyD. teres BarleyD. sorokiniana BarleyOxgall agar method! Septoria nodorum WheatFluorescent methodHoldfast method! Phoma betae SugarbeetMangan's water Agarm.ethodPeptone-PeNB agar Fusarium spp. CerealsGuaiacol-Agar Pyricularia oryzae RiceDrechslera oryzae RiceEmbryo count method Ustilago nuda Wheat, BarleyNaOH soak method Neovossia indica WheatTilletia barclay ana Rice

Detection Procedures for Fungal Plant Pathogens and Salvaging 147Salvaging of infected germplasm1. Mechanical separationThe seed lots are cleaned mechanically by separating the soilclods, plant debris, weeds, discoloured, deformed and shrivelledseeds showing visual symptoms of the disease by hand pickingunder the stereobinocular microscope. Whereas, the vegetativepropagules are cleaned by excising the infected portion.2. Spirit washFor eradicating the seed-borne rust spores of Pucciniacarthamii from safflower, a modified technique of seed washing isgenerally adopted. The contaminated seeds are taken in a testtube. Ethyl alcohol and a pinch of river sand is added to it. Thecontents are stirred on the mechanical stirrer for 30 seconds. Thespores adhering to the seed surface get agitated and separate outin the alcohol.3. Acid washConcentrated sulphuric acid is used for destroying the rustspores of Uromyces betae in' sugarbeet seeds. The contaminatedseeds are stirred with a glass rod in the acid forI min. The t.:-:eatedseeds are immediately washed under the running tap water toremove the traces of acid and then sun dried.4. Hot water treatment (HWT)HWT technique is used for controlli 19 various seed-bornepathogens like fungi, bacteria and nematodes. Some of the fungiwhich can be successfully controlled by HWT are:Fungi Host TemperaturelTimeAlternaria brassicae Brassica spp. 50 a C for 30 minA. brassicicolaFusarium solani Cucumis spp. 55 a C for 15 minPhoma betae Sugarbeet 50°C for 30 minPhoma lingam Brassica spp. 50 a C for 30 minPhomopsis vexans Brinjal 50°C for 30 min

148 A. Majumdar5. Chemical treatmentChemicals are generally used for eradicating seed-bornepathogens. Various fungicides are available which can be used asseed dressing or dips for the vegetative propagules. The chemicalsalso give protection against soil-borne fungi. Systemic fungicides. are widely used at present. The metalaxyl seed treatment has beeneffective against the downy mildew of pearl millet, maize,sorghum, poppy, sugarcane, sunflower and brassicas. Oxathins,(vitavax and plantvax) are used for controlling seed-borneinfection of smuts. Benomyl compounds are broad spectrum andare effective against many seed borne fungi.6. Post entry quarantine growingInfected seeds can be sown in the green house / glass houseor in isolation nurseries in the field. The harvest from only thedisease-free plants is collected. This method is used for thechemically treated germplasm of wheat, barley and triticale againstloose smut; and sunflower against downy mildew infection.The treatments used in plant quarantine are mostly eradicativeand sometimes protective also. Plant materials infected withdisease organisms of economic significance that are not present inthe country and for which no satisfactory treatment is available,are not released.ReferencesHewett, P.o. (1979). Pre-treatment in seed health testing. Durationof hypochlorite pre-treatment in the agar plate test forAscochyta spp. Seed Sci. & Technol. 7 : 83-85.Limonard, T. (1966). A modified blotter test for seed health. Neth.J. PL. Path. 72 : 319-321.Limonard, T. (1968). Ecological aspects of seed health testing.Proc. Int. Seed Test. Assc. 33 : 167 p.Neergaard, P. (1969). Plenodomus lingam, black-leg of crucifers.Occurrence in Danish seed lots for export, and control byGermisan hot water treatment. Friesia 9 : 167-179,_

Detection of Bacterial Pathogens and Salvaging ofInfected GermplasmBaleshwar SinghIntroductionInfected/ contaminated seed and other propagative materialsare a major source of primary inoculum as well as an importantmeans for local and long distance dissemination for most of theimportant phytobacterial diseases of major agri~horticulturalcrops. Therefore, during exchange of germplasm material, thereis always a risk of introducing exotic pests and pathogens intoareas where they were not known to occur earlier. This threatrelates not only to the possible introduction of a pathogen but alsoof new, more virulent strains or races of pathogenic bacteria.Hence, it is essential to test incoming planting materials for thepresence of pathogenic bacteria to preclude their introduction intothe country. The successful detection and control of bacteria inseed and other planting materials depend upon the availability ofquick and accurate methods for detection and isolation of the targetpathogen. The following methods are employed for the detectionof plant pathogenic bacteria in seeds and other propagativematerials and for salvaging the infected/ cont~minated materials.Technique for Detection of Bacteria1. Visual examination of seedsXanthomonas campestris pv. phaseoli causing bacteria,1 blight inbeans, and Curtobacterium flaccumjaciens pv. flaccumjaciens causingvascular wilt in bean are detectable by the presence of yellow

150 . Baleshwar Singhdiscolouration around the hilum. Pseudomonas syringae pv.phaseolicola, which causes halo blight of bean, produce a bluishwhitefluorescence in white-seeded beans when observed underultra-violet light (Wharton, 1967). Seeds showing such symptomsshould be removed from the samples. Similarly, discoloured,deformed and shrivelled seeds may carry the pathogen whichshould be sorted out by visual examination.2. Incubation testsThese methods are generally used for detection of seed bornefungal pathogens but can also be used for the detection of seedborne bacteria.(a) Blotter test: This method is used for detection of manybacterial pathogens in seeds like Xanthomonas campestris pv.campestris which causes black rot in crucifers, X. campestris pv.phaseoIi causing bacterial' blight in beans, X. campestris pv.vesicatoria causing bacterial leaf spot in chilli and tomato. To detectthe presence of X. campestris pv. campestris the crucifer seeds areplaced on moist blotter papers in plastic Petriplates and incubatedat 22±1°C. The typical black rot symptoms may be seen on theseedlings after 8 days of incubation. The bacterial association withthe symptoms may be observed by bacterial ooze test undermicroscope. Then the associated bacterium is isolated from theinfected parts on agar medium, purified and identified. Forbacterial ooze test, a piece is cut through infected tissue with sharpblade or razor and the piece is placed in a drop of water on slideunder cover glass. The slide is examined under microscope. If theinfection is due to bacteria, cloudy mass of bacterial cells is seenoozing out from the cut ends of the tissue piece. In vascularinfection bacteria ooze out forcefully at distinct pointscorresponding to the vascular strands as seen in bacterial blight ofbean caused by X. campestris pv. phaseoli whereas inparenchymatous infection the oozing of bacteria is slow, diffusedand throughout the cut ends e.g. in bacterial leaf spot of chilli andtomato caused by X. campestris pv. vesicatoria.(b) Paper towel method: The seeds are sown on moist paper'towel, then rolll7d and incubated. The symptoms of the disease canbe observed after 8 days of incubation. Xanthomonas oryzae pv.oryzae causing bacterial leaf blight disease in rice, X. campestris pv.

Detection of Bacterial Pathogens and Salvaging of Infected Germplasm 151campestris causing black rot in crucifers are detected by thismethod.(c) Agar plating method : The first agar plating method fordetection of X. campestris pv. campestris in Brassica seeds wasdeveloped by Lundsgaard (1973). The method has been modifiedand used for the detection of many seed borne bacteria in differentcrops. The bacterium in seeds may be detected as given below:(i) Direct plating: X. campestris pv. campestris in crucifer seeds;X. campestris pv. phaseoli, P. syringae pv. phaseolicola bean;Clavibacter michiganensis subsp. michiganensis in tomato can bedetected by direct plating the seeds on the agar medium.Randhawa and Schaad (1984) have developed semi-selective agarmedia (BSCAA) and successfully detected X. campestris pv.campestris in crucifer seeds on this medium.Plating with seed washing: X. campestris pv. campestris(ii)causes black rot in crucifers, X. campestris pv. carotae causesbacterial blight in carrot and other pathogens can be detected bythis method. For detection of X. campestrispv. campestris in cruciferseeds, 10000 seeds (about 40 g) are soaked in 0.85% NaCI solution(3-5°C) containing 0.02% Tween-20,S mg benlate 50% WP and 0.5m1 of a 1:10 dilution of Bravo 500. The contents are shaken on arotary shaker for 1.5 hr at 3-5°C, then filtered through sterile cheesecloth and rinsed with 25 ml sterile water. Seeds are discarded andthe liquid is centrifuged at 12,000 rpm for 10 min. at 2°C to pelletthe bacteria. The supernatant is discarded and each pellet is.suspended in 3 ml of sterile 0.85% NaCl (saline). The suspensionis diluted serially to 1:10, 1:100, 1:1000 and 0.1 m1 of each dill,ltionis plated individually in plates of semi-selective media (Randhawaand Schaad, 1984). The plates are incubated at 30°C and observedfor suspected bacterial colonies after 3 and 5 days.(iii) Plating after seed maceration: Bacterial pathogens can bedetected by seed maceration technique. The surface sterilizedseeds are soaked in sterile buffer water or distilled water at room.temperature for 3-4 hr and then crushed with pestle and mortar.The paste is then transferred to sterile water columns (quantity ofwater depends on the quantity and size of the seed), stirred wellon shaker or in centrifuge at 12,000 rpm and allowed to settledown. The supe:rnatant is discarded. The pellet is re-suspended

152 BalesJnvtly Si/lghin 10 ml of sterile buffer and is serially diluted from 10- 1 to 10- 5 or10- 6 dilutions. Suspension from each dilution is then streaked onthe nutrient agar medium or semi-selective agar media in plates.Alternatively 1 .ml of suspension from each dilution is pouredseparately first in Petri plates, 25 ml of cooled medium (45 11 C) thenis poured and the plates are gently rotated to mix the suspensionin the medium and the plates are incubated at 27±1 (Ie for 4-5 days.The colonies of the pathogen will appear after 48 hr which can bepicked up, purified and identified.3. Phage plaque method or phage sensitivity testMost of the bacteria are sensitive to bacteriophages and asa result they are lysed. The phage sensitivity test has been usedsuccessfully to detect X. cantpestris pv. plzaseoli in bean seeds; X.oryzae pv. oryzae in rice and Clnvibacter michiganensis subsp.michiganel1sis in tomato seeds,The seed leachate/extract is mixed in the nutrient agarmedium and poured in the sterilized Pet.riplates. The phagesuspension is spotted on to the surface of the agar medium witha pipette and the plates are dried in controlled conditions to drythe drop into the surface of the medium! the seed extract andphage suspension are mixed in cooled agar medium (4S°C) andpoured in sterile Petriplates. The lytic zone can be observed afterincubating the plates at 25°C for 24 hr. Formation of the lytic zonesaround the phage spot confirms the identity! presence of thebacterium.4. Serological techniquesAmong the possible ways to detect bacteria in seed or in'propagative materials, serological techniques are more suited forroutine application because of their specificity, rapidity, quickdetection or identification of bacteria and easy application to largeamounts of (seed) samples. Some of the serological tes~s, used fordetecting and identifying seed~borne bacteria are as follows.Agglutination test: Pseudomonas syringae pv. phaseolicola, the(a)halo blight pathogen, in bean (Taylor, 1970) und Pseudomonasjuscavaginae, the bacterial sheath brown rot pathogen in rice(Duveiller et al" 1988) have been detected by this method. In thistest the bacterium reacts with antiserum and forms a fine

Detection of Bacterial Pathogens and Salvaging of Infected Gcrmplasm 153precipitate. Agglutination test can be performed in culture tubesas well as on glass slides.(b) Immunofluorescence (IF) test: Immunofluorescence is asimple and rapid test which is more commonly used in clinicaldiagnosis of the agents of human diseases. Nowadays IF testsare also used for the detection of plant pathogenic bacteria in seedsand other plant parts. P. syringae pv. phaseolico/a and X. campestrispv. phaseoli, the halo blight and bacterial blight pathogens,respectively in bean seeds can be detected by direct IF and indirectIF test. In direct IF, the slide prepared for IF test is treated withantibacterium conjugate and after mounting, the slide is examinedwith a super pressure mercury lamp and a 1-2 filter system forincident illumination with blue light for FITe excitation whichshowed clear IF positive cells. For indirect IF test, the slide is firstreacted with bacterial antiserum then with antibacterium conjugateand the mounted slides are examined for IF positive number ofbacterial cells.The IF test can also be performed on agar medium by IFcolony staining. After the appearance of the colony on agarmedium, the agar medium is treated with antibacterial conjugateantiserum and the agar can be checked with a UV microscope atlow magnification for fluorescing colonies.' The halo blightpathogen in bean and bacterial canker in tomato can be detectedby this method.(c) Immunodiffusion: The basic principle of this test is that theantigen and antiserum are placed opposite to each other at certaindistance in soft gel (0.75 to 1%) of agar. The soluble antigen andantibodies diffuse through the gel and at the place of meeting fromantigen-antibody complex, a precipitin line appears.(i) Direct double diffusion: The halo blight pathogen in beanis detected by this method. The seed extract/leachate is inoculatedin agar plates and the plates are incubated. The bacterial coloniesare observed on agar under binocular after 24-48 hr of incubation.Then wells are made in agar medium at a distance from thebacterial colonies to be tested and diluted antiserum is added tothe wells. The precipitation line may be seen after 24-48 hr.(ii) Ouchterlony double diffusion (ODD) : Clavibactermichiganensis subsp. sepedonicus which causes ring rot in potato is

154 . Baleshwar Singhdetected by ODD test. The iilgar mediu_!ll (composition: agar 8.0g; NaCI I 8.5 g; NaN0 31200 mg; distilled water 1 lit) is prepared,melted and poured in Petriplates. Wells are cut in the agar in apattern of 6 pheripheral wells surrounding a center well. Undilutedserum is placed in the central well. A known positive sample isplaced in two opposite pheripheral wells and unknown samplesin the remaining four pheripheral wells. Agar plates are incubatedovernight at room temperature. In a positive test, the precipitinband can be seen between the sample and antibody well whichmay fuse with the control precipitin band.(d) Enzyme-linked immunosorbent assay (ELISA) : ELISA.. procedure is a simple,. rapid, very sensitive technique and has beenused to detect several plant pathogens including bacteria in seeds.In ELISA the antigen: antibody reaction is visualized by enzymaticconversion of a chromogenic substrate. The sandwich techniquehas the greatest potential for detection of bacteria and is mostcommonly used. In this case the first step involves the trappingof antiserum in the wells of ELISA microtitre plate. Then theground seed/plant extract suspected to harbour the bacteria i.e.antigen is put in the same well. If bacterium is present in theextract, it is trapped by the antiserum. In the third step, the sameantiserum (bound to the enzyme) is put in the wells. This allowsto form a "Sandwich'l of antiserum-bacteria-antiserum (bound toenzyme). If this complex is formed, the enzyme molecules areliberated. The presence of bacteria is indirectly estimated by'reacting the liberated enzyme with the suitable substrate to forma coloured product. Many bacteria have been detected by thismethod.G~owing on test(a) Growing on seedlings by water agar method: This methodwas developed by Srinivasan et al. (1973) to detect X. campestris pv.campestris in crucifer seed. The seeds, after surface sterilization withaureofungin solution, are plated on 1.5% water agar in Petriplatesand incubated at 20°C in darkness. The germinated seeds andseedlings when examined on 8th day under stereo-binocularmicroscope, show the black rot symptom. The bacterial ooze testconfirms the bacterial association with the symptoms. On isolationsfrom the infected parts, smooth, butyrous, glistening, yellowcolonies of X. campestris pv. qampestris will appear after 72 hr.

Detection of Bacterial Pathogens and Salvaging oj Infected Germplasm 155(b) Growing on seedling in green house' : Many bacterialpathogens such as X. campestris pv. phaseoli in be anSi X. campepstrispv. malvacearum in cotton; X. campestris pv. carotae in carrot .andPseudomonas syrin'gae pv. phaseolicola in beans have been detectedby growing-on test. The symptoms develop on grown seedlings /plants. The association of the bacterium with the disease may beobserved by bacterial ooze test and the bacterium is isolated on theagar medium, purified and then identified.6. Host plant inoculationThyr (1969) and Saettler '(1971) used this method for thedetection of Clavibacter michiganensis ssp. michiganensis andXanthomonas campestris pv. phaseoli in tomato and bean seedsrespectively. The seed exu:act is inoculated in seedlings/plants byinjection or spray inoculation technique and the inoculated plantsare kept in hlimicl chamber for the development of the symptoms.Venette et a( (1987) inoculqted the seed extract in 'the seeds byvacuum infiltration and the seeds were sown in humid chamberfor growing the seedling~ to detect seed-borne bacterial pathogensin beans.Techniques for salvaging infected germplasmKeeping in view the importance of the germplasm mat~rial, effortsshould be made to salvage it so as to save the germplasm. Variousmethods used in plant quarantine to salvage seeds and plantingmaterials infected with bacterial pathogens are as fo116ws.1. Roguing-out infected plants in post-entry quarantine nursery(PEQN)The seedlings/plants raised in net house/field and showingsymptoms of bade rial diseases are uprooted and burnt immediately.This method is feasible where specific symptoms are produced bythe pathogenic bacteria like bacterial blight of beans, carrot, andcowpea. The, seeds are collected from healthy plants only andreleased to the indentor.2. Laboratory treatmentSOther salvaging methods used in the plant quarantine laboratoryfor eradicating pathogenic bacteria from infected seeds and

156 Baleshwar Singhplanting material are hot water treatment (HWT), dry heattreatment and use of chemicals. Some of the recommendedtreatments for different crops/pathogens are given in Table-I.Table 1.Laboratory methods for salvaging seeds and plantingmaterials infected with pathogenic bacteriaCrop Pathogen TreatmentHot water TreatmentCabbage Xanthomonas campestris HWT of seed at S2"C for 30 minpv. cllmpestrisCarrot X. campestris pv. HWT of seed at 50-S2"C for 10 min.carotaeor 128"F for 11 to 12 minCauliflower Erwinia carotol1ora HWT of seed at SO"C for 18 minsub sp. carotovoraXanthO/1l011aS campestrispv. cCl1npestrisHWT at 122"F for 25 minChilli E. caratovora sub sp. HWT of seed at 50"C forcarotovora20.min., 4 days before sowingX. campestris pv. HWT of seed at SOnC £01' 10 min.vesicatoriaCotton X. caml'estris pv, Immersion of seed in hot watermalvacearumat 60°C for 10 min.Cowpea X. campestris pv. HWT qf seed at SO"C for 30vignicolaminGuar X. campestris pv. HWT of seed at 50"C for 10cyamapsidismin.Larkspur E. carotovora sub sp. Seed treatment at 50"C andatrosepticaS5"C for 10 min.Rice X. oryzae pv. oryzae Seed soaking in normal water for 12hr then at 53"C for 30 min. 01'soaking for 12 h in ceresan(500ppm)+ Agrimycin -100 (250 ppm) andlater HWT at 52"C for 30 min.Pseudomonasfuscovagil1aeHWT of seed at 55"C for 20 minSesame X. campestris pv. HWT at 51-52°C for 10 min.sesami

Detection of Bacterial Pathogens and Salvaging of Infected Germplas11l 157Contd ... (Table 1)CropPathogenTreatmentTomatoDry heat treatmentLettuceX. campestris pv.vesicatoriaPseudomonas cic1lOriiSeed immersion in hot water at50"C for 1 hri at 55°C for 30 min orat 60"C for 15 min.Seed treatment at 70"C for 1-4 daysPea Pseudomonas syringae pv. Dry heat treatment at 65"Cpisifor 72 hr.Rice Pseudomonas avenae Dry heat treatment of seed at 65"Cfor 6 daysChemical treatmentCarrot Xcampestris pv.carotaLoCauliflower P. syringae pv.mactelicolaCrucifers X campestris pv.campestrisCottonX. campestl'is pv.malvacearumGuarSoybeanTomatoTurnipWheatPseudomonas fitscovaginae Heat treatment at 65"C for 6 daysPseudomonas glumae Heat treatment at 65"C for 6 daysX. campestris pv.cyamopsidisX. campestris pv.glycz'llesX. campestris pv.vesicatoria. E. carotovora subsp.carotovoraP. syringae pv.atrojaciel1sSeed treatment with 0.1%HgCl z for 10 min.Seed steeping in 1% HgCl,solution for 1'0 min. •Seed disinfection by 3 min.immersion in 1:1000 HgC1 2solution.Seed treatment with 1%ceres an or treatment for delintingwith cone. H 2S0 4(1:17 acid: seed)for 50 or 10 minSeed treatment with ceres an WP@ 0.2%; thiram 0.2%; streptocycline0.01% or Agrimycin-100 @ 0.01%.Seed treatment with fundazol@ 3 kg/toSeed treatment with 1:1000HgC1 2•Seed treatment with 80% TMTD@ 8 g/kg.Seed treatment with 80% TMTD@ 2 kg/t.

158 Baleshwar SinghReferencesDuveiller, E., Myajima, K, Snacken, F., Autrique, A. and Maraite,H. (1988).' Characteristics of Pseudomonas fuscovaginae anddifferentiation from other fluorescent pseudomonads occurringon rice in Burundi. J. Phytopathol. 122: 97-107.Lundrgaard, T. (1973). A method for detection of Xanthomonascampestris (Pammel) Dawson in Brassica seeds. StatensPla.ntetilsyn 21 : 34-38.Randhawa, P. and Schaad, N.W. (1984). Selective isolation ofXanthomonas campestris pv. campestris from crucifer seeds.Phytopathology 74: 268-272.,Saettler, A.W. (1971). Seedling infection as an aid in identifyingbean blight bacteria. Plant Dis. Reporter 55: 703-706.Srinivasan, M.C., Neergaard, P. and Mathur, S.B. (1971). Atechniqe for detection of Xanthomonas campestris in seedhealth testing of crucifers. Seed Sci. & Tech. 1 : 853-859.Taylor, J.D. (1970). Bacteriophage and serological methods for theidentification of Pseudomonas phaseolicoZa (Burkh.) Dowson.Annals Applied BioI. 66 : 387-395. .Thyr, BD: (1969). Assaying tomato seed for Corynebacteriummichiganense. Plant Dis. Reporter 53: 858-860.Venette, J.R., Lampa, R.S., Albaugh, D.A. and Nayes, J.B. (1987).Presumptive procedure (dome test) for detection of seedbornebacterial pathogens in dry beans. Plant Disease 71:984-990.Wharton, A.L. (1967). D~tection of infection by Pseudomonasphaseolicola (Burkh:) Dowson in white seeded dwarf seedstocks. Annals ~pplied BioI. 60 : 305-312.

Symptoms Cause,d by Plant VirusesD.B. ParakhIntroductionViruses constitute the most serious pathological problems ofthe living world. In agriculture, they were' recognised as ' growthreducingfactors' for a long time. They are, therefore, the subjectof intensive research. Present knowledge about viruses is that theyare infectious agents (often cause disease), invisible with lightmicroscope (sub-microscopic), small enough to pass through abacteria-proof filter, lacking a metabolism of their own anddependent on living host cells for multiplication~ Virusesconstitute small packages of genetic information of either nucleicacid (ribonucleic acid, RNA or de-oxy ribonucleic acid! DNA)encapsidated together or separately and enclosed in a coat of oneor more than one types of protein. Few viruses have a lipoprotein,besides its nucleic acid and protein.Plant-virus interactions'1There is an intimate relationship between a virus and its hostplant. The process of multiplication of virus in its host cellsdisorganizes the host's physiology and results in 'disease'.. Thevirus is then a pathogen causing disease. Once the plant isdiseased, the virus multiplication leads. to visible deviations orsymptoms. A symptom is defined as any perceptible change inthe plant or its functions, indicating a disturbance in the normalcourse of the physiological process. The science of the stuc4f ofsymptoms is called symptomatology. Different kinds of symptoms

160 D.B. Parakhare produced due to virus infection. A group of symptoms is calledsyndrome and may be characteristic of causal virus(s). Symptomsmay be grouped according to their effect on host such as growth,colour, tissue life-span, shape of plant part or organ, plant anatomyor physiology and water content. Diagnosis and correct descriptionof plant virus symptoms are important because viruses aregenerally named after their main symptoms on a particular host,e.g., a mosaic causing virus in tobacco is named as 'tobacco mosaicvirus' (TMV).Symptoms of virus diseases1. Reduction of growth: Many plant viruses cause a growthreduction similar in proportion in all organs, so that plants remainmorphologically almost normal. The term dwarfing and stuntingare used in describing growth reductions such as peanut stunt andbarley yellow dwarf. .2. Colour deviation: Most obvious and better known symptomsare colour changes, especially in foliage. Most of the colourchanges are in the chlorophyll and, therefore, the disorders areessentialiy the same for leaves, young stems and fruits. Someimportant colour deviations are as follows:(i) Mosaic: When the various shades of green and yellowpigmentations are usually irregularly angular but distinctivelysharply demarcated, the symptom is described as mosaic, such astobacco mosaic or abutHon mosaic.(ii) Mottle: In contrast to mosaic, when various shades ofpigments (green and yellow) are diffusely bordered with no clearcut demarcation, the symptom is described as mottling. But thereare various intermediates.(iii) Spots: In this case, the discoloured parts are sharplybordered but circular, systemic infection may also form light ringsar01..md a normally green centre or even concentric rings of darkand light tissue called ring spotting. Such spots are frequentlycause~ by ring spot viruses such as tobacco ringspot, tomatoringspot, etc.(iv) Line pattern : Systemically infected leaves, for instance ofwoody plants, may show beautiful patterns known as line patternas in rose leaves or oak-line pattern.

Symptoms Caused by Plant Viruses 161(v) Striping: Mosaic in leaves with parallel veins look likechlorotic or yellow striping or streaking as is apparent from barleystripe mosaic, maize streak, wheat streak mosaic, etc.(vi) Chlorosis : Colour changes may be evenly distributedthroughout organs or entire plants as in chlorosis in which lesschlorophyll is produced.(vii) Yellowing: Yellow pigments when predominant causeyellowing. Examples of such diseases caused by phloem limitedluteoviruses (luteus == yellow) are barley yellow dwarf. Suchsymptoms may largely result from degeneration of the host'stransport system. With certain other viruses and hosts, chlorosisor yellowing may be restricted to tissues adjoining the veins,resulting in bright vein-yellowing or yellow-vein-banding as withseveral phloem limited whitefly-transmitted virus diseases, suchas okra yellow vein mosaic, mung and urd bean yellow mosaic,soybean yellow vein mosaic, etc.(viii) Reddening: Other pigments may increase due to certainvirus infections causing reddening or purpling, as with carrot redleafand cotton anthocyanosis (purple colour in plants is due toanthocyanin pigment).(ix) Browning and bronzing: Browning occurs alongwith deathof tissue as in case of pea early-browning virus. Bronzing mayresult from superficial death (necrosis) of epidermis over normaltissue.3. Necrosis: It is rapid death of local tissue and is usuallyaccompanied by blackening or browning. It is often characteristicof local lesions on inoculated leaves of test plants. It may alsoresult from degeneration of internal vascular tissue such as phloemcausing phloem necrosis as in case of potato leaf roll virus orsometimes developing into visible vein necrosis or necrotic. streaking of petioles and stems. Stem tips or buds may rapidlyreact with such necrosis causing tip necrosis or bud necrosis. Suchnecrosis may proceed further and the whole plant may die ratherrapidly, as in case of bean infected with virulent strain of beancommon mosaic virus. Necrotic rings are formed in tomatoesinfected by a necrotic strain of tomato spotted wilt virus. Necroticline patterns are sometimes observed on pep seeds due to pea seedborne mosaic virus infected plants.

162 D.B. Parakh4. Malformations: Virus infections som.etimes may also lead todeformed plants or organs called malformations. Malformation isa pathological defor.mation that is caused by disturbance ofharmonious tissue development. It differs from stunting anddwarfing which are due to general growth reduction. Malform.ationsare of many types. When growth correlation is more directlydisturbed by virus infection, the malformations are primary.Secondary malformations become evident only after the plant hasdeveloped other symptoms that lead to the malformations.5. Anatomical and Cytological deviations : As virus activitystarts in: the host's cells, its structure (anatomy) and function(physiology) are altered. Cells decrease in size (hypotrophy) orriumber reduced (hypoplasia) or may increase in size (hypertrophy)and number (hyperplasia). Hyperplasia is also called as proliferationwhen cells multiply excessively.Inclusion bodies are the product of host-virus interaction.here, a large number of virus particles get arranged in a particularsequence and shape, which is now known to be characteristic foreach protein group of plant viruses in presence of protein.Inclusion bodies may occur either in cytoplasm or nucleus. Theseare easily stained and detected even in light microscope thusleading to detection of viruses in host tissue.6. Water deficiency symptoms: These are caused because ofshortage of water, due to excessive transpiration or obstruction ofwater supply through vascular tissue causing wilting or moreseriously, withering. This is irreversible desiccation of tissuewhich causes its death. Etching symptoms are caused due todesiccation of superficial tissue as in tobacco etch disease.Constraints in identifying viral disease based on symptoms1. Symptomless carriers : In some diseases, the wrus infects itshost and multiplies within, but causes no vi~ible externalsymptoms. Such hosts are described as symptomless virus carriersand the virus infecting it as latent virus, e.g., potato infected withlatent mosaic virus.2. Masked symptoms : There are instances when the plants areinfected with virus but do not show distingUishable symptoms dueto absence of favourable environmental co~ditions. Such maskedsymptoms are seen in potato infected by potato mild mosaic virus

Symptoms Caused by Plant Vinlses 163which expresses symptom only when the temperature is around16°C.3. Deviation~ resembling symptoms of virus disea&e: SymptolnSof certain virus diseases closely resemble deviations due to geneticaberrations, physiological non-parasitic disturbances, insecttoxemias, micro-organisms such as mycoplasma-like-organisms(MLOs), spirdplasma, rickettsia-like-organisms (RLOs), rickettsialike-bacteria(RLB) and viroids. Many of the transmissible diseasesymptoms thought to be caused by viruses were infact later provedto be caused by MLO, RLO, RLB and viroids. Sometimes. differentagents lead to identical deviations, especially with malformationssuch as enations and tumours may result from attacks by bacteria,mite, insects, nematodes or a virus. Symptoms produced by soilnutritional disorders such as mineral deficiencies and toxicities areoften confused with virus or virus-like symptoms.Despite problems encountered in identifying symptoms ofviral diseases, it should be noted that vIrus symptoms often havea characteristic distribution or the sequence of symptoms in virusinfectedplants because of the typical way in which the virusinfects, and associates with specific tissue, spreads and multipliesin the plant.Laboratory detection of viral symptoms in germplasmA variety of viral symptoms on seed such a~ discolouration,deformation, necrosis etc. on seed coat have been identified to beassociated with particular virus infection, e.g., mottling on soybeanseeds due to soybean mosaic virus (SMV) , shrivelled seeds ofmungbean and urdbean due to urdbean leaf crinkle virus (ULCV),necrotic line pattern or I and split seed coat of pea due to pea seed. borne mosaic virus (PSbMV), necrotic spots of broad bean due tobroadbean stain virus (BBSV), colouration of part of seed coat ofcowpea due to cowpea aphid-borne mosaic virus (CApMV), andpigmentation of seed coat of soybean cv. iwate-wasekurome dueto soybean stunt virus.Similarly, germplasm cuttings or tissue culture material mayalso show paleness, yellowing or virus-like symptom, but theseneed actual virus-indexing procedures to detect any association ofpathogen. Such sYlflptoms, infact, can be also caused byenvironmental factors or nutritional disorders during transit.

164 D.B. ParakhReferencesBos,L. (1978). Symptoms of virus diseases in plants. CAPD,Wageningen, 250 pp.CIMMYT (1983). Common diseases of small grain cereals: A guide toidentification. (Ed. Zillinsky), 141 pp.Singh, S.R. and Allen, D.J. (1979). Cowpea pests a.nd disea.ses.IITA, Manual Series. No.2. Nigeria, 113 pp.Nemeth, M. (1986). Virus, Mycoplasma and Rickettsia Diseases ofFruit Trees. Forestry Science. Akademiai Kiado, Budapest,Hungary, 841 pp.The American Phytopathological SOciety. The CompendiumSeries, APS press: and Insects, 1986; Corn Diseases (2nd Ed.)1980; Cotton Diseases, 1981; Elm Diseases, 1981; Onion andGarlic diseases, 1994; Ornamental Foliage Plant Diseases,1987; Pea Diseases, 1984; Peanut Diseases, 1984; PotatoDiseases, 1981; Rhododendron and Azalea Diseases, 1986;Rose Diseases, 1983; Sorghum Diseases, 1986; SoybeanDiseases (2nd Ed.), 1982; Strawberry Diseases, 1983; TropicalFruit Diseases 1993; Turfgrass Diseases (2nd Ed.), 1993;Wheat Diseases (2nd Ed.), 1987.

Techniques for Detection of Plant VirusesShamsher Singh and R.K. KhetarpalIntroductionThe germplasm material is exchanged and conserved mainlyin the form of true seeds. However, germplasm lines of certainvegetatively propagated plant species such as sweet potato, yams,garlic, banana, citrus, ginger, etc. are nowadays exchanged andconserved in in vitro cultures. Seeds are efficient carriers of plantviruses. More than a hundred viruses are known to be transmittedin plant species belonging to 26 families. Also the plantingmaterials of in vitro cultures, as mentioned above can be a potentialsource of many threatening virus diseases unless they aremultiplied by meristem tip culture and subjected to thermotherapyand virus indexing to ensure their freedom from viruses. Thedetection and identificati9n of viruses become imperative to restrictor minimise their movement in areas previously considered asdisease-free and also in conserving germplasm in a state free frominfection.A strategy comprising of a combination of various virusdetection techniques is employed to ensure that germplasmmaterials being introduced are free from exotic viruses or th.eirvirulent strains. The various techniques employed for detectingplant viruses are given below ;

166 Shamsher Singh and RK. KhetarpalA. General methods for detectiona) Symptomatology : The symptoms of virus diseases aregenerally observed on plants/planting material during' visualinspection. They may be grouped according to their ~ffect on hostsuch as growth reduction, colour deviation, necrosis andmalformations. Also during visual inspection of seeds, a varietyof symptoms, viz; mottling on soybean seed coat due to soybeanmosaic virus, split or stained seed coat on pea seeds due to peaseed borne mosaic virus, necrotic spots on broad bean due to broadbean stain virus, shrivelled seeds of mungbean and uidbeancaused by prdbean leaf crinkle virus etc. have been found to beassociated. But, it has not been possible to demonstrate a strictcorrelation between the presence of virus symptom on seed andac'tual seed to plant transmission of virus disease. The onlyconclusion which of virus drawn is that the seed samples havebeen harvested from infected mother plants. Therefore, seed freefrom the seed symptoms should be handled for exchang~ andconservation.b) Infectivity test: Viruses are by definition transmissible andthis prop.erty of viruses gives us ways and means by which theycan be transmi,tted experimentally. The simplest test by which aplant virus can be transmitted is the infectivity test. This istypically done to determine whether an observed host plantsymptom is caused by a viral infection or a symptomless plant(with latent infection). In this test virus containing extract ofcrushed infected leaves is taken in distilled water or a buffer andrubbed (inoculated) on leaves of certain 'young herbaceous plantswith a finger or cotton swab. Since viruses are passive parasites,they require a wound to enter cells to cause an infection. Therefore,carborundum powder or celite is added to infected extract.Some plants can be infected by sap inoculations more readilythan others. In general herbaceous plants are infected more easilythan woody oneSj dicotyledons than monocotyledons and youngseedlings than older plants. Moreover, some species are particularly'easy to infect and are susceptible to many viruses. Some plantfamilies such as Solanaceae and Leguniinoseae, seem to containmore of these species, than other families. Certain useful speciesfor detecting, culturing or assaying a w~de range of plant virusesare:

Techniques for Detection of Plant Viruses 167ChenopodiaceaeCucurbitaceaeGraminaeLeguminoseaeSolanaceaeChenopodium amaranticolor, C. quinoaCucumis sativtlsTriticum aestivum, Hordeum vulgarePhaseolus vulgaris, Vicia Jaba, Vigna unguiculataNicotiana tabacum, N. clevelandii, N. glutinosa,Petunia hybrida, Datura stramonium.Apart from this mechanical inoculation,' grafting scions,especially in case of fruit trees on healthy root stocks or on suitablewoody indicator plants is an accurate and reliable screeningmethod for viruses. . .c) Staining of inclusion bodies: Inclusion bodies are aggregatesof virus particles or virus induced proteins or consist of specialstructures characteristic of infections by viruses -either-·in thecytoplasm or in the nucleus. These are essentially a product ofinteraction between the virus and the host cell which can readilybe detected with a light microscope. Inclusion bodies are presentin epidermal tissues, mesoderm (underlying tissue of epidermalstrip) and in phloem. The detection and diagnosis of plant virusinfection is achieved by cutting free-hand sections with a razorblade and after brief treatment with certain stains such as AzureA and O-G combination. The tissue is observed under lightmicroscope (best with oil immersion) and inclusion bodies arelocated and characterised.d) Electron microscopy: To detect virus particles in plant tissue,electron microscopy (EM) is an indispensable tool. It reveals thesize and shape of virus particles. Samples from seedlings of growout test or inoculated indicator host plant can be subjected toelection microscopic examination. Based on EM· results, thepresence of virus can be established, but which virus is associatedwith the disease cannot be known. For further characterization ofvirus, serological and other tests are done.B. Advance methods for detectionSerological diagnosis of plant viruses has undergone arevolutionary change with the development of Enzyme LinkedImmunosorbent Assay i.e~ ELISA. Apart from this recentdevelopment of cDNA probes and polymerase chain reaction(peR) permits to detect a very low concentration of virus in the

168 Shamsher Singh and R.K. Klletarpalhost tissue with enhanced sensitivity.described below.These techniques area) Enzyme linked immunosorbent assay (ELISA) ; ELISA isrelatively simple, rapid, very sensitive and requires small amountof antiserum. Of the various forms of ELISA tests, the doubleantibody sandwich form (DA-ELISA) is most commonly used. Inthis case virus specific C\ntibodies are adsorbed to a solid surface(microtiter plates) and to this is added the sample suspected tocontain the viral antigen which is followed by the addition ofenzyme labelled specific antibody. The labelled antibodies bind tothe antigen which is already round to the coating antibodies.Finally the substrate is hydrolysed and the development of colourin the end product is measured as absorbance value in a multiscanspectrophotometer. The colour change in the substrate isproportional to the amount of enzyme present which in turn isproportional to antigen concentration.The repetitive nature of most of the operations involved inELISA make this technique well suited to automation and permitssimultaneous testing of a large number of samples.b) Dot-immunobinding assay (DIBA) : It is a variant of ELISAtest wherein, instead of using microtitre plates as solidsupport,nitrocellulose membranes are used. A few microlitres ofextract of infected samples are blotted on this membrane which isthen submerged in primary antibody (crude antiserum). Theprecipitated specific antibody is then detected with enzymelabelled second antibody or protein A.DIBA has an edge over conventional ELISA as it does notrequire any special equipment, it is relatively inexpensive, itrequires only a crude specific antiserum to each of the viruses anda single universal enzyme conjugate. Above all the blottedmembranes can be mailed to long dis~ances for further processingin a centralized laboratory.c) Immunosorbent electron microscopy (lSEM) : It is a verysensitive technique coupling serology with electron microscopy.In this case the electron microscope grids are pretreated withspecific antiserum which facilitates the adherence of virus particleson the grid by several folds. The virus particles are then

Techniques for Detection of Plant Viruses 169specifically trapped and decorated with the antiserum and thecontaminants are washed away.d) Complimentary DNA (eDNA) probes: This method is basedon the fact that complimentary strands of nucleic acid hybridizewith one another. It has a potential of detecting extremely lowlevel of inoculum or latent infections in plant/planting materials.Basically the method involves the immobilization of a spotor dot of sap extract from the plant under test on a solid matrixand the detection of viral nucleic acid sequences in that spot byuse of a hybQ"dization probe. For preparing a DNA probe, DNA,complimentary to RNA is synthesized from the RNA templateusing.the enzyme reverse transcriptase. If this DNA is then madedouble stranded it can be cloned in a bacterial plasmid and thusimmortalised. As much qf the DNA copy of the viral sequences,as required, can be prepared and it can also be manipulated usingrecombinant DNA techniques. The probe nucleic acid usuallycarries reporter molecules which can be detected when it ishybridized to the immobilIzed viral sequences. 'e) Polymerase chain reaction (PCR) : This involves rapid andhighly specific in-vitro amplification of selected DNA sequences,for which specific primers are synthesized. The p.roduct obtainedfrom this cycle amplification process is separated by gelelectrophoresis and identified by specific cDNA probes. With itsrelative simplicity and high sensitivity (detecting picogramquantities of viral nucleic acids in infected tissues) PCR method hashigh potential in virus indexing programmes. However, the prerequisiteof having known viral sequences to allow synthesis ofsuitable primers limits its application to well characterised viruses.C) Virus indexing _For poorly characterized or unknown viruses detectionmethods either are not available or are not sensitive. In such casesbioassays using sap and graft inoculation to a range of sensitiveindicators are still the only methods available, . Conclusions basedon negative results in these methods are reliable only when testingis repetitive and includes alternative metho9.s using different virusproperties for detection.

170 Shamsher Singh and R.K. KhetarpalDouble stranded RNA (ds RNA) analysis is a rapid tool thatcan supplement the information obtained from. bio-assays. Analysisof dsRNA is based on the isolation of disease specific dsRNAs fromvirus infected tissues and their electrophoretic separation on a gel,which is then stained and viewed. However, the presence of nonviraldsRNAs in healthy plants and the apparent absence of dsRNA profile in some viruses, may result in false negatives or falsepositives. Negative dsRNA tests should be confirmed by othermethods before plant material is indexed as virus free.

IISymptoms of Insect Damage in CropsShashi BhallaIntroductionThe struggle between wan and insects began long before thedawn of civilization. I.nse_S~s are most numerous m~mbers ofanimal kingdom, which is attributed to their s;naU size, widea£laptab.ility and s!lperior body structure. About lQI.QQO of the640,000 species of insects are injurious to plants and of these about-?Q9pm be considered as 1!);ajor pests in India. Annual losses dueto insect pests have been estimated to about p per cent. No otherclass of animals is so intimately involved in the intric~cies andcomplexities of the biological world as are insec;ts.Extent of insect injuryInsects have been found associated with all the plant partsi.e. leaves, flowers, fruits, buds, seeds, bark, wood and roots. Infact,~;o pla;},t part is free from their damage. Nearly all the injurydone by insects results directly or indirectly from their attempt tosecure food. ¥any insE;!cts are physically observed on the _Rlantsbut q_the!s are I10ctumal in their :v.£lbit and they .J:rlde under ~oilclods, fallen leaves or bark crevices during the day time. Thoughthey may not be visible during the day time, they causeconsiderable damage to the crops.An insect or a group of insects produces specific symptomsof damage by ~egtng or o_vipositing activities. Therefore, it isessential for a pJa.p...! l:>r~~der or ,plant explorer to be acquainted with

172 Shashi Bhallathese symptoms of insect damage to collect healthy ger:rJ.1.plasmmaterial. As pest risk is involved in the import of both vegetativelypropagated material and ~~eds, the symptoms of insect damage arevery important for the quarantine worker to detect infestation inthe germplasm under exchange to prevent the introduction offoreign pests into the country. The damage due to insect pests maybe direct or indirect. .Direct damage is caused by biting and chewing off the plant parts,piercing of plant tissues and sucking of plant juices. Much of thedamage from seed infesting insects is done directly to the kernels.Grain is reduced in weight due to consumption by adults andimmature stages.Indirect damage is due to the secondary invasion of host plantsthrough the wounds caused by feeding ~nd other activities of theinsect. Piercing and sucking of the plant juices results in fungusand virus develop,ment followed by plant decay, wilting, systemicyellowing, blotching or mosaicing etc. Insects and mites serve asvectors of a number of virus diseases. In storage, they produc~heat and change the humidity. Germination is impaired bydamage either to germ or endosperm.On the ~asis of insect feeding and its effect on external andinternal plant tissues or parts, these have been categorized asExternal feeders and Internal feeders.'External feeders: Either they feed on the plant tissues from outsideand produce eaten out areas of different extents or they roll or webtogether the plant parts as leaves, flowers and flower buds theyinfest and feed within.Internal feeders: They cause damage by remaining within theplant tissues during a part or all of their destructive stages. Thisis accomplished by adult thrusting their eggs into the tissues eitherby ovipositor or the larvae gain entry by eating/boring their wayin. These include leaf-miners/.borers (stem-borers, fruit-borer), gallmakers, gallery makers, and seed feeders.On the basis of the feeding habits of the insects, they can becategorized as biting and chewing type which bite and chew offthe plant parts and sap feeders (piercing and sucking type) whichpierce the plant tissue and suck the sap.

Symptoms of Insect Damage in Crops 173Biting and chewing insectsThese insects chew off the plant parts and grind them up andswallow them because they possess such mouth parts. This groupincludes adults and nymphs/larvae of crickets, grasshoppers,beetles, weevils, wasps, flies (maggots), butterflies and moths(caterpillars) .Sap feedersThese insects feed on the grovdng. pl..eyl1.~s by :g!.~XS!ng andsJlSkip.g out the cells from the plants within. Only li_guid PQrtim:tsof the plant are s~9_J£~d, although the insect itself rema~sext~:cr.g_liY on the plant. This is accomplished by an extremelyslgnder, shm:p, pointed portion of the beak which is thrust into theplant and. through which the sap is sucked. This group illc1udesleaf-hQP.J?~s, ~_Qs, V{hH~_tU.es, SSales and thrp"s. They generallyprefer tende!.fug~. As a result of 8~E..c~i~g, the leaves develop:ygUowJ;:rrQ,wn patches or even c;!ld up or sl;1rivel, 'thus devitalizingthe plants. Besides they also ~_b,Qney: ..d£.w which attracts ~and also favours the development of ~s.apr.op.hyl~s in theform of blacy sooty moulds on leaves, stem and flowers. Manyof these pests infect their salivar:. secretion at the time of feedingTwhic1! due to presence of certam toxins damage the plant tissuefurther. Sap-feeders are important as they are generally found withthe v~etativ:elY_J?r.opag.ete.d !llaterials. Important insect groupscause the following symptoms of damage.Aphids : They feed on ~S, flQY.'L~rs, b.tanclJ.es and cause injuryby extracting the plant sap. some of them feed on cluster of r.Q.Q!:s('1VJ/.flJ.p1Iis .raseus). Some species transmit number of vU:ll§dj_~.eases.Thrips : Lacerate the epidermis of tender leaves mostly near thetips resulting in silvering or whitening of leaf surface due toremoval of cell contents. 'Affected leaves roll up and turn pale.They also suck the s~ from ~s.Psyllids ; They produce white wogllyJ,!ryal, secretions on leav~s, .~o.is and ll}flQ!§~gI1ce~" Heavy sooty mould can develop afterhoneydew deposit in case of severe infestation. Nymphs producegalls on leaves and adults transmit di~~Mles.

174 Shashi BlzallaJassids : Commonly called ~~~LhgpE~X~' They suck the juices fromveins and veinlets mainly from th,e undersurfaces "Of leaves whichbecome brown, tum upwards and dry out in cases of severeinfestatio;t:-thls feeding causes c;hlQro.ti~.lines parallel to the veinsand plants become ~u.JJ.ted..Scales : ~(}YE:!r the tender k_a;yes and shoots and $ck th~ _sap. Sootymould development is commonly noticed, e.g., G:re,en scale Coccus1!iridis on coffee.Pest darnage is probably most conveniently consideredaccording to the part of the plant body attacked as damaged leaves,f1OW~,rs and..buds, {nuts, stems, r.Qots, tubers and s.eed~.Foliage feedersThe insects feed on the plant tissues from outside. This groupincludes grasshoppers, beetles, weevils, their' grubs and manycaterpillars, e.g., armyworms, hairy caterpillars, cutworms etc.The leaves ~ay be eaten to different extents thus causing reductionin the assimilative area of the leaf.The leaves could be eaten from edges (i) with regular notches(Curculionidae), (ii) with semicircular pieces of lamina missing(adults of leaf cutting ants) and (iii) with irregularly eaten out areas(grasshoppers, locust). The leaf lamina is damaged to differentextents ranging from windowed lamina (Diamond back moth),holed lamina (adult flea beetles, Chrysomellidae) to skeletonizedlamina (Epilachna, citrus flea beetles etc.). The rice hispa,DichZadispa armigera scrapes the upper surface of leaf blade andleaves intact the lower epidermis. Leaf curled under or generallydistorted, sometimes completely distorted into a bunchy lump oftissue is the characteristic damage by the sucking pests. Adults andnymphs of some thrips make tiny epidermal feeding lesions whichgive the lamina a silvery, bronzed or scarified appearance. Leavesare also damaged by the galls either on upper or lower surface,sometimes arranged randomly, distributed peripherally or alongthe side veins. These are made by the feeding larvae of gall midges,gall wasps and some psyllids.Leaf webbersMany ~Sl ~tles and ~es form webs on 1~.They roll up the leaves by binding the edges with strands of silk

Symptoms of Insect Damage in Crops 175and leaf lamina is eaten within the protective rolt e.g., Bananaskipper (Erionota spp.) and cotton leaf roller, Sylepta derogata. Theyalso bind together the leaves to make nest- like structures as Redant, Oecophylla smaragdina. Some caterpillars (Pyralidae,Lasiocampidae etc.) web the leaves and shoots with silk and makea tent or web in which they live and eat the leaves. Often the webtraps a large number of faecal pellets.Leaf and stem-minersTiny larvae of many Qg.fi!ties and lUQtQS live in between thetwo epidermal layers of leaves and tS;!1g~r_ green st~m and feed onthe food material inside, they are referred to as ~:r.s. Minersare polyphagous ins.~cts. Sometimes they are serious pests andinhibit plant growth by interferring with the photosyntheticactivity of the plant. The mines may be either ~~1 mines or121Qtches or sometimes starting as a tunnel and ending as a blotchmine. In serious cases, the attacked leaves wither away altogether.The'se mines also serve as poUlts where ~.ase and c!~say maystart. C_ea l~af miner is a very serious pest and inhibits plantgrowth. The

176 Sltashi Bhallacoffee stem borer, Xylotrechus quadripes, bore into the main stem,make tunnels into the lower part of stem and into the main root.The tunnels made in the bark appear as ridges and plants may wilt.m. case of shot hole borer of coffee, Xylosandrus compactus, infestedbranches gradually wilt and turn yellow and fall off. The grubs ofcotton stem weevil, Pemphesulus offinis Fst., tunnel round the stemfeeding on the soft tissues and result in the formation of gall likeswelling at the'site of injury. In pink boll worm affected crops,flowers fail to open, squares fall and bolls are eaten out. The mangostem borer, Batocera rufomaculata grubs feed by tunnelling throughthe bark of branches and cause wilting. There are abol,lt 20 borers(caterpillars) that attack sugarcane in India. A few beetles havealso been found boring into growing canes and decaying stems.In some cases one larva bores inside the plant and kills thegrowing point resulting in dead heart and then moves to adjacentshoots causing several dead hearts in a clump.Gall makersThere are ~ertain insects which in their immature or adultstage live in the plant tissues and their activities lead to the plantdeformities. These may be r.2!:!!!£l, ~..heJ:,ical, and e.!mJ.g~.ted· ors..l?in~uitr,"!ls~~res and are ~own as gall~", which are :R-est spes;ific.These galls provide shelt~r and food to the insects. The gallforming insects include many '!:.l2!:!ids, ~les, lJlic;!g"es, ~igslt@lps, ~ps and some w.,g!'l:~d1s. Due to the formation of galls thegrowth of the' plant may be impaired and setting of fruits, grainsand seeds may be adversely affected. The galls may be simple a:scurling of leaves or simple enlargements of affected portions or ofcomplex structures as some galls produced by rollid _Q-ggs.Mostly some species belonging to families Cecidomyiidae,Cynipidae, Aphididae, Psyllidae and Aleyrodidae and orderThrysanoptera (Thrips) cause galls on the differen~ parts of theplant. The rice gall midge causes damage by producing tubulargalls popularly: known as Silver shoots which are the leaf sheathsand tillers produced into galls. 1;'hese do not bear any panicles.Early infestation results in P!ofuse tillering of plants and new tillersget infested. Infestation at later ,stage results in branching athigher nodes, just below the silver shoots. On bitter gourd, stemgalls are formed by Lasioptera falcata. Flower galls are producedby midges, Contarinia sorghicola on sorghum and,by'blossom midgeon mango.

Symptoms of Insect Damage in Crops 177Flower and bud feedersAdult blister be.e.tles (Meloidae) chew the petals of manyplants. Adult flower h.eg.t~~s (Scarabaeidae) m~~.§_m.ajL~21~~ inthe .£.~.~ls. l:hriz;?s, both ad.!!lts and nymp}1s, scarify the bases of~ls. Tiny black beetles make feeding scars at the base of thepetals. Flower deformation is caused by ~DlQggQ.ts as gall midgelarvae. Caterpillars of some Tortricidae bore into the large buds.Large caterpillars as semi-loopers feed on the flowers by makinglarge holes. Maruca testulalis, webs together the flowers and budsand feeds on the opening buds thus causing reduction in seedproduction. Gall mites of family Eriophyidae infest buds of someplants and cause a disorder called 'big bud'.Fruits feedersThe fruit flies, fruit worms and wepvils cause considerabledamage to different fruits. The fruit flies of family Tephritidaeattack almost all larger types of fruits. They puncture the fruitsurface for oviposition. The larvae develop internally as the fruitripens by feeding on the pulp resulting in fruit rot with visiblebrown patches on the exterior. The infested fruits fall prematurely.The color of rotting patch is brownish in citrus, mango, apple,peach, apricot and black in penr and banana. The green fruit wormcauses large, irregular cavities in immature apples which at the. time of feeding look deformed, with brown disks on healed overcavities. The grubs of walnut weevil Alcidodes porrectirostris, boreinside the fruit and feed on the kernel turning it to a black mass.The pomegranate fruit borer, Virachola isocrates, caterpillars boreinto the fruit and feed on its internal contents. Characteristicallythe anal segment of the caterpillar is often seen plugging the borehole. Some heteropteran bugs of families Miridae, Coreidae andPentatomidae have toxic saliva which causes necrosis at thefeeding site on fruits. This usually becomes infected with fungiand bacteria resulting in rotting, death and premature fruit fall.Bark feedersA considerable number of insect species damage the bark ofthe host and make different forms of galleries on the trunk andtwigs. The branch or whole tree may die by feeding of larvae insidE"and girdling of the bark. Termites also cause damage to the barksof plantation crops and forest trees. Q!u~s of Xyloborus perforans

178 Shashi Bhallatunnel into the dead wood and construct irregular and extensivegalleries inside the stem of arecanut. The caterpillars, Atherastiscirculata and Ptochoryctes rosaria, build their galleries on the trunkand branches of rubber plant filling them with feacal matter andsilk. They feed on the dead bark inside and create irregular,depre:?sed channels into stems. This interferes with latexcollection. The bark beetles, Scolytus rugulosus and Phloeotribusliminaries attack the trunk and branches of the peach tree. Galleriesof borers are found in the inner bark radiating from the main orparent gallery and terminating in sman, round exit holes throughthe bark.Root feedersThere are a number of insects which are found in the §.oil andlive by feeding on the r.99ts of plants and lrees by ch~wing, ~9ring,sy_(;}gng or fQrming galls, thus causing loss of water and nutrientabsorbing tissues. These prevent proper establishment andgrowth. These insects may include cti9s,ets, t&nnites, ~bi~~ grt,lbs,several b~es and vx~ey:ils. Though the damage caused to rootmay vary depending on the portion of root infested. Generallythe attacked plants show Shm!iPg, discoiouration, withering andin severe infe.s!£pon, death of the J?lant. The larvae may feedexternally on roots as in wireworms, weevils, grubs and whitegrubs. Grubs bore or tunnel into the root as Longitarsusbelgaumensis on sunhernp plants. Root. aphid, Tetraneuranigriabdominalis suck the sap from the roots. The woolly aphid,Eriosoma lanigera spends its life cycle in the soil and both adultsand nymphs suck the sap from the roots of apple causing theformation of root galls. The rhizome weevil, Cosmopolites sordidus,grubs bore into banana rhizome and cause death of the plant.Tuber feedersTubers are also attacked by insects. v.y:ireYY,Qrms bore thetubers by making ~els. Potato tuber moth larvae boredown the stem into the tubers by making the tunnels where rottingoccurs due to secondary infection. The tubers of sweet potato areriddled with holes by the larvae of weevil Cylas formicarius. Bothlarvae and adults of yarn beetle bore the yam tubers with widetunnels. These weevils remove the stored food from tubers andcorms and affect next season's growth.

Symptoms of Insect Damage in Crops 179In addition to insects, mites (Acarina) is another importantgroup which accompanies the vegetative propagules. Mites havebecome major pests of economic significance in recent years dueto adoption of improved agronomic practices and use of moderninsecticides. They are either ~g. or ..1?i.~.J:~il)g and &,J.lcking type.They damage the crop plants in number of ways:(a) They suck thecellular material resulting in the formation of characteristic whiteblotches on leaves and devitalizati on of plantsi (b) Eriophyid mitescause severe deformities in plant parts; and (iii) a few mitestransmit virus diseases.:,

180 Shashi Bhallainternal feeders, (ii) secondary feeders feed on the broken grainsor the grains damaged by the other E.~~t$.General symptoms of seed damage are:(i) the whole seed is consumed or reduced to dust,(ii) the internal contents of grain are eaten leaving onlyoutershell,(iii) the germinal portion is scraped/ devoured,(iv) scarving of grain surface or burrowing holes in them(v) tum: lling of the seeds(vi) webbing together of the grains.The t;tm.pr~ peetle, '[rogoderma granarium Everts, larvae feedfrom thl" germpoint and consume the entire kernel leaving onlythe shen. Infested material is full wi~h £rass, cast skins of larvaeand fragments of insects etc. Larval feeding of cigarette beetle,Lasioderma serricorne (P.) results in tunnelling of seeds. The dryfruit moth, Cadra cautella feeds on the germpoint of seeds. In heavyinfestation, larval:! cover all the a, ~i1able surface with webbing.The rice moth, Corcyra cephalonica larvae spin dense silken tubesand web the grain kernels into the walls of the tubes. They producedense webbing as they become full Qrown. Red flour beetle,Tribolium castaneum causes serious damage to the grain productsand processed food. Insect causes damage to broken seeds but notthe whole grain. This insect also imparts a nauseous smell andtaste to the infested material.The internal feeders spend a part or entire larval and pupalstages within the seed only to emerge as adults. Among them isa group of pests whi~h multiplies under favourable conditions anda number of generations are passed (Callosobruchus spp. Sitophilusspp., Rhizopertha domi'nica) but in others, after development. iscomplete, the insect remains quiescent till the next favourableseason (Bruchus pisorum). In case of the lesser grain borer, bothlarvae and adult stages are equally injurious to the grain. Larvaeenter the grail .. generally near the soft embryo and develop there.As a result of larval feeding whole grain is reduced to hollowbroken shells. The damaged kernels are always surrounded bypowder from the chewed up grain. The drug store beetle, Stegobiumpaniceum (Linn.) grub tunnels into stored products like turmeric,

Symptoms of Insect Damage in Crops 181ginger, coriander and dry vegetables. The Angoumois grain moth,Sitotrogra cerealella , caterpillars feed on the endosperm or germuntil fully grown. It then eats out a channel to -the outside of theseed and makes a weakly fastened flap at the exit by cutting theshell, through which the adult emerges. A large number ofbruchids infest the seeds of leguminous and non-leguminousplants both in field as well as in, stores. The beetle lays numberof small oval, yellowish eggs on the seeds/pods. Hatched larvaeea t their way into the seed and feed within. Full groyvn larva beforeentering into pupation, cuts a disc in the seed coat, through whichthe adult beetle emerges by pushing open the disc called the'operculum'.The seed lots infested with internal feeders, therefore, don'tbear any external symptoms of their presence within unless theyemerge as ad1,llt or leave the seed for pupation. Seeds .of 320 plantgenera are known to carry hidden infestation, especially bybruchids and chalcids. These seeds don't show any externalsymptoms of damage by which the infestation could be detected.These infested seeds can easily be detected by )$-ray"!,~9t.qgt:aE~y,Insects as disseminators of plant diseasesA serious phase of insect injury is the destruction, caused bytheir direct feeding Le. the connection of insects with the ravagesof plant diseases. There are number of insects as Vi.hlt~f1iel', ~t\!4~,l~~( h.QE12.~rs and tbtiFs which damage the plant by sucking thesap and are responsible for transmitting the plant diseases indifferent ways.1. Mechanical transmission(i)(ii)(iii)Feed and oviposition marks predispose plants to viral,fungal and bacterial infection.They may actually disseminate the pathogens from oneplant to the other.Pathogen is carded inside their body and injectedhypodermically into the plant as they feed.2. Biological transmission: Most viruses depend upon the activityof an insect vector for transmission. The vector is usually also anintermediate host, as is the case with most aphid and whiteflyhosts.

182 . Sltashi BhallaAs there is an increasing need to accelerate the collection,preservation and utilization of Plant Genetic Resources tlu'oughintroduction programmes, quarantine measures will be moreeffective with the increased knowledge of insect distribution,biology and mainly with the symptoms of insect damage. This willfacilitate introductions through transfer to increase the germplasmin genebank without expanding the current stocks of pests.ReferencesAnon. (1983). Agricultural Entomology, vol. II. All India ScientificWriters Society, New Delhi, 436 pp.Comstock, J.H. (1948). An Introduction to Entomology. ComstockPublishing co. Inc. Itachca, 1064 pp.Hill, Dennis, S. (1993). Agricultural Insect Pests of the Tropics andtheir Control (Fourth edition) Publ. Manas Saiki a, Delhi, pp.746.Metcalf, C.L. and Flint, W.P. (1979). Destructive and useful insects,their habit and control. (Fourth ed.) 1087 pp. TMH Publ. Co.Pradhan, S. (1969). Insect pests of crops Pub!. National Book Trustof India, 208 pp.Verma, B.R. and Kapur, M. L. (1990). Eriophyoid Mites (Acari;Eriophyoidea) of Arid ·zone Fruit Crops and Quarantine.Indian J. Ent., 52(2) : 249-252.

IIMethods for Detection of Insects and Mites inImported GermplasmB.R. VermaIntroductionPests (insects and mites) attack and damage a large numberof agri-horticultural plants. Some of the pests get associated withseeds and planting materials (internally or externally) and movefrom one place to another through men's activities. While the.movement of planting material is desirable, the movement of pestsin this manner is objectionable specially from quarantine point ofview. Examination and detection of pests, therefore, is an essentialprocedure for phytosanitory certification purposes Oones, 1972) inplant quarantine. The inspection and detection are required (i) atthe point of origin, (ii) during the transit and (iii) at destination.Pest spectrumAgricultural imports include materials for consumption,processing, commercial planting and research purposes. The firstthree groups of materials are usually imported in bulk comprisingof relatively fewer varieties, while for research purposes, number. of accessions (samples) is very large but quantity per sample maybe very small. However, germplasm material for research has highquarantine risk. Pests accompanying germplasm material couldbe broadly divided into two groups :(a) Pests of vegetative propagules: The risk involved in importof vegetatively propagated plant material (bulbs, budwood,cuttings, tubers etc.) is higher than the seed. The number of insects

184 B.R. Vermaand mites involved is extremely large in plant propagules ascompared to seeds. The vegetatively propagated plant materialharbours eggs, larvae and pupae deep inside the plant_ tissue anddo not bear any external symptoms of their presence till theemergence. Insects and mites belonging to some families arefound hidden inside the plant tissues and need careful examination.(b) Seed 'pests : The seeds may get infested while developing andmaturing on plant, on threshing grounds, during transport or instores. The pests involved can broadly be grouped into twocategories viz; (i) internal feeders including chalcidoids, majorityof bruchids and some other weevils and beetles and (ii) externalfeeders which continue to feed externally and multiply even in thestored seed and cause more serious losses. .The former category includes seed infesting chalcidoids,which except for the adult stage pass their life cycle in the seed.Eggs are deposited through the seed pod into the developing milkystage seeds. The hatching la:r;va feeds, grows and pupates in theseed itself only to emerge as an adult, since such insects can becarried with apparently healthy seed shipments~ they pose aquarantine problem.In the international exchange of seed which can be infestedwith phytophagous chal~idoids, bruchids and some weevils andbeetles, it becomes necessary to either use especial detectionprocedures or SUitably disinfest or reject the imported seeds. Thedetection procedure can be time consuming or seed consuming,whereas the rejection of the planting material would defeat thevery purpose of import.Methods for detection of insects and mitesDetection techniques may be divided hi.to two· broadcategories : active and passive. Active techniques use externalinputs, either physical (flotation or radiograph) or chemical(staining or transparency m.ethod) to derive information in the'form of an output such as film image (X-ray) or colour change (byusing acid fuchsin, genetion violet or berberin sulphate) in anindicator material, or an electronically measurable change inresistance, cOIl).plex ~mpedance, protein spin or the like. Passivetechniques do not use external inputs and depend on naturallyoccurring outputs such as infrared energy radiated by the insect,

Methods for Detection of Insects and Mites in Imported Germplasm 185sounds produced by insect feeding, or an aromatic vapour or gasproduced by insect metabolism or respiration.Active techniques are reliable, have some potential forgenerating personal hazards or environmental pollution, or foraltering or'destroying the sample (hardness) under test. Passivetedmiques, on the other hand, simply monitor an insect producedoutput, sound and CO 2determipation come in this category.However, these have inherent disadvantages s~nce not all lifestages of insect produce detectable sound or urine or CO 2bec

186 B.R.. Vermakernels that constitute to the amount of dockage, webbing, excreta,scraped germ, or even any other tell' tale signs like the egg andoperculain stage of the seed coat. On slightest suspicion ofinfestation the seed/plant parts is cut open and examined todetermine if it is infested and if yes, with what ?(a) Visual method without previous seed treatment: Certainpests like the Sitophilus spp., Sitotroga cerealella and Rhizoperthadominica, the infested seeds bear a circular hole through which theadult emerges. Though the infestation due to these pests is saidto be of hidden type, it no longer remains so, once the adultemerges. The visual examination of the seeds infested with thesepests would give away the presence of holes on the seed surface.It is generally believed that every seed containing an emergencehole indicates approximately 5 times as many seeds havinginfestation still within the seed.This method is not only tedious and time consuming but isalso approximate. It gives an idea about the pest population inthe sample.(b) Visual method with previous treatment(i) Fluorescence 1?1ethod : The seeds are partially embedded ina layer of drying cement. After the cement hardens, the seeds aresanded off to almost half and the remainder exposed to UV light,the infested seeds fluoresce because of the insect waste productsand can be counted.This method though accurate, is useful on sample basis onlyand can be used to estimate the population in a quantity of seed.This method is laborious, time consuming and inaccurate becausethe insect waste loses its property to fluoresce after certain periodof time.(ii) Semi-transpare~t method: In this method a strong alkalisol~tion (like NaOH) is· used to partially digest the seed so thatthey become sufficiently translucent to show the internal infestation.This method is laborious and not practicable because very oftenthe coloured seeds do not become translucent enough despiteprotracted digestion.2. Transparency methodThe seeds or plant parts are soaked in a solution containing2 parts of distilled water, 2 parts of phenol crystals, 2 parts of lacti12

Methods for Detection of Insects and Mites in Imported Germplasm 187acid and 1 part of glycerine (by weight). This renders themsufficiently transparent to reveal internal infestation of larvae,pupae or adults and holes and tunnels made by hatching larvae.The solution is prepared by heating the distilled water and thenadding the rest of the ingredients Kaura (1959).This technique is hazardous to workers due to exposure tofumes of phenol ( a nerve poison) which is a major drawback, ifa large number of seed samples are to be handled. The viabilityis lost and hence .the valuable imported material is unfit forbreeding purposes,- This is a limitation when sample size is verysmall and especially in quarantine where 100 per cent sample isto be examined and cleared for indenter.3. Staining methodStaining methods make use of the capacity of certain dyes tostain the gelatinous egg plugs deposited by the insect on seed orany part of the plant. Some of more successful staining methodsare:a) Adq fuchsin technique for phytophagous chalcidoids : It iswell known that in freshly oviposited green pods, little quantityof watery substances oozes out of the area around the ovipositionpunctures which gradually develops a rust coloured lesion - aminute spot which can be seen with the help of a binocular on theseed coat of infested seeds. In course of time the surrounding seedcoat develops rust coloured lesion which increases in size, as thelarva is full grown the seed attains brownish colour and becomessome what shriveled.The eggs could be detected by making the green pods andseeds transparent in lacto-phenol medium (Koura, 1959) and 3 percent acid fuchsin stain (BDH) added to it. After 24 hrs. eggs couldbe recovered by dissecting the seeds, placed beneath the seed-coatin developing cotyledons or between the two cotyledons near oraround the germpoint (Verma, 1986). Before examinanon underbinocular, the fresh lactophenol medium is to be changed to avoidthe tinting of eggs with chlorophyll.b) Detection oj eggs ,in vegetative propagules : pirectdetermination of the site of egg deposition is difficult in thoseinsects that lay their eggs within plant tissue. The eggs of

188 B.R. Vermaaleurodids, aphids, delphacids, psyllids, capsids, cicadellids(Homoptera), thrips (Thysanoptera), Cecidomyiids (Diptera),eurytomids (Hymenoptera) and eriophyoids (Acarina) could be'detected in situ by using a simple staining technique.The plant parts conlaining insect eggs are bleached in boilingwater for 5-7 minutes, kept in 95% ethyl alcohol for 3 days, rinsedin water and then immersed in 1% aqueous acid fuchsin solutionfor 2 days. Plant parts then are washed under running water untilstained eggs are differentiated from destained plant tis~:ue. Eggscould be counted using a 20x binocular microscope (Khan &Saxena, 1986, Pablo, 1977). The plant parts could be stored inglycerine for an indefinite period without affecting the colour ofthe stained eggs. Alcohol could be .saved by decolorizing carbon.This method also has certain limitations. Prolonged storageof the plant tissue in alcohol caused tinting of eggs withchlorophyll, making their differentiation from the surroundingplant tissue difficult (Gifford and Trahan, 1967). This techniqueobviates the use of harmful and costly chemicals such as lactic acidand phenol and fume hood facilities are needed. This method canalso be used for detecting the eggs of crop pests that deposit theireggs inside soft plant tissues.4. Sound detectionBrain (1924) was one of the early scientists to publish ondetecting the chewing and moving sounds of insect larvae inagricultural commodities with electromechanical devices. Hereported that he could detect fruit borers in stems and weevilsinside grain. The limiting factors are sensor sensitivity, sensornoise and ambient noise. If sufficient sensitivity is developed thissystem could detect accurately early instar larvae in fruits, nuts,grain and stem of vegetatively propagated material. It is mucheasier and instant way than the X-ray radiography wherefrequency range of the insect could also be calculated. Acousticscan be defined as the generation, transmission and reception ofenergy in the form of vibrational wa,ves in matter (Kinsler et aI.,1982). These vibrational waves can be transmitted through liquids,solids and gas~s. When these vibrational waves travel throughdifferent media, special sensor configurations are required todetect the acoustic energy, depending on the media, it is travellingthrough. The accelerometer is normally used to detect the energy

Methods for Detection of Insects and Mites in Imported Gennplasm 189transmitted through solids, the hydrophone through liquids andthe microphone through gases or air. The vibrations can beconverted into electric signals and could be displayed on a meter /software/printer and recorded (Vick et aI., 1988).5. X~Ray radiographyX-rays are in everyday usage for medical purpose andsomewhat less so in industry. Their use for the analysis of seedquality dates back to the fifties when Simak and Gustafson (1953)and Simak (1953) demonstrated that soft X-ray after passingthrough seeds produce a pattern on X-ray plate. Thus, it is possiblein X-ray picture to recognise the structure, viability and extent ofdevelopment of embryo and endosperm (Swaminathan andKamra, 1961) and also to detect mechanical injuries as well asdamage caused by insects, pathogens and bacteria (Yuasa, 1926;Milner et al., 1950; Wadhi et al., 1967). The insect detection workthrough radiography is fairly common in many countries. Thistechnique has been noted as the lUost accurate method of studyingthe extent of damage, the stage of insect development inside theseed/ stem and the identity of insect could also be ascertainedwithout damaging the seed. Detection of pathogens iscomparatively more difficult since creating contrast between hosttissue and pathogen is so far unsuccessful. Only when the fungalmycelia are in a mass, e.g., in some tea seeds, they become visible.However, the pathological changes accompanied by the pathogenicattack become clea~ at times due to shrivelling.Seeds infested with phytophagous chalcidoids, bruchids andcertain other insects do not present any external evidence for thepresence of infestation on seed. A list of 322 plant genera has beendrawn up, the seeds of which have been known to carry suchinfestation. Whenever seeds of any of the listed plant genera arereceived, these are subjected to X-ray radiogrCij'hy at 22 kv, 3 rnafor 15 seconds at the distance of 30 cm. However, dose-ratedepends on the seed hardness. The infested seeds, if present, arehand picked and only healthy seeds are released to the indentor.The initial cost of X-ray machine and the ongoing cost· of X­ray films and chemicals to develop the X-ray film are high andexamination of each of the individual grain on the X-ray film forinsects is labour intensive. Detection of infestation in very smallseeds (e.g., tobacco and Trifolium spp.) is difficult and cumbersome.

190 B.R. VermaThe technique such as X-ray micrography or projection microradiographyneed to be developed so that radiograph of smallseeds could be enlarged and accurately analysed.6. Simple flotation methodIt has been reported by Kurdjumove (1913) that dry seeds offorest trees infested with phytophagous chalcidoids can beseparated from uninfested dry seeds by simple flotation method.This method was standardised (Verma, 1986) and modified forSesbania green seeds infested with immature stages of seed chalcid,Bruchophagus mellipes Gahan. Infestation can be detected bysplitting open the seed, removing the submerged ~eeds from waterand dissecting them carefully under binocular. The seeds removedfrom the bottom of container contain mostly eggs and insubsefluent removing, seeds may contain larvae, pupae and deadadults. (Verma, 1986).7. Post entry quarantine detection methodsInsects like-capsids (mirids), psyllids, cicadellids, cecidol1)yids,eurytomids, eriophyoids, borers etc. may not be detected at thetime of laboratory examination because (i) eggs/larvae may befeeding deep inside the plant tissue, (ii) perishable nature ofpropagating material makes it difficult for a detailed specializedexamination in a short time and (iii) this type of material representsmost serious quarantine risk Therefore, such type of material maybe detained to allow the eggs to hatch. Suspected plant materialis grown isolated in glass houses/net houses/screen houses andinspected regularly. Plant material may also be grown in thirdcountry (Intermediate Quarantine) where particular host is notavailable for cultivation. Regular survey in the post-entry quarantine(PEQ) nursery is required. As and when the symptoms developand identification of the emerged adult stages is completed, theplanting material may suitably be treated, destroyed or releasedto the indentor. In some cases material is grown for one seasonand healthy produce of the crop is released to the indento~.8. Processing of interceptionsFor adequate detention, control and regular work rapid identificationcan not be relegated to be handled as some time later. Once theinfestation is detected, the identity of the pest needs to beestablished speedily to determine the quarantine risk and the

Methods for Detection of Insects and Mites in Imported Germplasm 191suitable quarantine treatment. The identification of pests isdifficult because a few institutions have an insect reference. collection and information on faunistic studies is lacking. However,NBPGR has developed an insect reference collection for over 250species intercepted from plant material under exchange. Udaigiriand Wadhi (1982), Hinton and Corbet (1972), La! (1987) andJeppson et al. (1975) have compiled keys to the identification ofbruchids, stored grain beetles, seed chalcids and mites. Thecollected specimens of insects and mites may also be sent foridentification to IAR!, New Delhi, FRI, Dehradun, ZSI, Calcutta,TNAU, Coimbatore and BM (NH), London. A logic requisite isto have well documented records of pests, identification keys, theirhost range, nature of damage, biology, distribution and controlmeasures. Knowledge on exotic species is very useful to evaluatea particular line of battle during and after detection of pests.ReferencesBrain, c.K. (1924). Preliminary note on the adaptation of certainradio principles to insect investigation work. Arm. Univ.Stellenbosch. 2: 45-47.Gifford, J.R. and G.B. Trahan (1967). Staining teclmique for eggsof rice water weevils oviposited intracellularly in the tissueof the leaf sheaths of rice. J. Econ. Entomol. 62: 740-741.Harein, P.K. (19 ). Methods for detecting hidden infestation ingrains. Pest Control 28 : 35-36, 38.Hinton, H.E. and Corbet, A.S. (1972). Common Insect pests ofstored food products. A guide to their identification. BritishMuseum (Natural History), London, Economic Series No. 15:1-62.Jeppson, L.R., Keifer, H.H. and Baker E.W. (1975). Mites injuriousto economic plants. Dniv. Calif. Press, California XXII + 614,74 plates.Jones, H.L. (1972). A critique of the status of plant regulatory andquarantine activities in the United States. Ann. Rev. Entomol.17 : 453-460.

B.R. VermaKhan, Z.R. and Saxena, RC. (1986). Technique for locating planthopper (Homoptera : Delphacidae) and leaf hopper(Homoptera : Cicadellidae) eggs in rice plants. J. Eean.Entamal. 79 : 271-273.Kinsler, L.E., Frey, A.R.; Coppers, A.B., Sanders, J.V. (1982).Fundamentals of acoustics,3rd Edition, John Wiley and Sons,Inc. New York.Koura, A. (1959). A new transparency method for detectinginternal infestation of grain. Grain Storage News Letter, 1(1):12.Lal, B. (1987). Taxonomic studies on seed inhabiting chalcidoidsfrom India Ph. D. Thesis, IARI, New Delhi 149 pp.Milner, M; Lee, M.R. and Katz, R. (1950). Application of X-raytechniques to the detection of internal insect infestation ofgrain. J. Eean. Entamal. 43 : 933~935.Simak, M. (1953). The X-ray contrast method for seed testing. Scotspine Pinus silvestris; Medd. F. Stat. Skagsforsk Inst. 47 :1-22.Simak, M. and Gustafsson, A. (1953). X-ray photography andsensitivity in forest tree species. Hereditas Lund. 39 : 458-68.Swarninathan, M.S. and Kamra, S.K. (1961). X-ray analysis of theanatomy and viability of seeds of some economic plants.Indian J. Gen. and pl. Breeding 21 : 129-135.Udaigiri, S. and Wadhi, S.R. (1982). A key to world bruchid genera.NBPGR Sci. Manogr. No.5: 1-16.Verma, B.R. (1986). Detection technique for phytophagouschalcidoids in green seeds. Indian J. Ent. 48 : 225-227.VickI KW., Webb, J.e., Weaver, B.A. and Litzkow, C. (1988).Sound detection of stored product insects that feed insidekernels of grain. J. Eean. Entomal. 81 : 1489-1493.Wadhl, S.R., Vf7rma, B.R.; Thomas, Sares and Rattan Lal (1967).Detection of phytophagous chalcidoids in seeds for quarantinepurposes. Indian J. Ent. 29 : 197-199.Yuasa, H. (1926). On the advantages of the X-ray examination Ofcertain classes of material and insects subject to the plantquarantine regulations. Proc. 3rd Pan Pacific. Congo Tokyo.1 : 1141.

Plant Quarantine Treatments for SalvagingGermplasm infested with Insects and MitesManju Lata KapurIntroductionThe undeniable need for improved plant varieties to cope upwith the demand of ever increasing human population, calls forthe conservation and exchange of plant genetic material on a globalscale. This involves risk of widespread distribution of plant pestsand pathogens alongwith the movement of germplasm, whichmay harm the agriculture and the environment of new areas, andthus, vitiate the benefits .of any plant _introduction or breedingprogramme. The basic tenets of Plant Quarantine include,regulations to allow germplasm exchange with no or minimumpest risk, inspection of material to detect the presence of pests andtreatment of infested material. Last one is the most instrumentalin promoting the positive image of plant quarantine as it ensuressalvaging of material rendering its introduction permissible.Rejection of a sample on the basis of mere presence of aninsect pest, may not be a wise preposition. Therefore, severalmethods are employed to eradicate the insects from infested seedor planting material and pest free material is made available to theuser scientists. Some of the common salvaging methods used inquarantine entomology laboratory are discussed hereunder.Traits of a Quarantine TreatmentPlant Quarantine Treatment is a method which can be usedto lower or liquidate the pest-risk by destroying or eradicating a

194 Manju Lata Kapurpest, thus preventing its dispersal to an area where it is not knownto occur or is 'of limited distribution. Following traits are of majorconsideration while developing a quarantine treatment.a) Pest tolerance: Theoretically, quarantine pest exclusionimplies pest tolerance limit after a quarantine treatment as zero,i.e., ,it must give 100% kill of the pest. But practically it is notpossi~le. Therefore, the concept of 'probit 9' has been acceptedin quarantine circles (Baker 1939). Statistically, it is survival rateupto three individuals for a total population of 100,000 individuals,determined by a minimum of three separate tests at 95%confidence level. 'Probit 9' is not valid for pests which can developfrom a single organism, or when the pest/its hardiest stage is notavailable for testing. However, it may be feasible to use data fromsmall scale tests and literature on the efficacy of a test along witha 'safety factor' i.e. a slightly higher dosage or longer exposureperiod than the routine control treatments., .b) Commodity tolerance: There is very fine margin between thelethal dose (Probit 9) for insect mortality and phytotoxicity ofplants (Benschoter 1963). A quarantine treatment should cause noor minimum acceptable changes in the commodity, which aregenerally in terms of modifications in appearance, odour, taste,viability and vitality. While using a Quarantine Treatment thatincludes 'safety factor' as mentioned above, commodity tolerancemust be considered, although in the interest of pest security,indentor has to strike a compromise to some extent.c) Residue tolerance : It implies that a quarantine treatmentshould leave no or minimum residue in commodity so that it doesnot harm man, animals or environment. The safe limit of residuetolerance is established by authorised environmental agencies fora Quarantine Treatment before it is put to use, or else tolerancelimit is considered zero. This is disregarded in the interest of pestsecurity only when commodity is not required to be treatedrepeatedly. .d) Time factor: A quarantine treatment must not be very timeconsuming and it should cause only a minimum delay in thequarantine clearance' of the material.

Plant Quarnatine Treatments jor Salvaging Gennp/asm Infested with Insects and Mites 195Treatment vs. InfestationThe kind of treatment to be employed depends upon the typeof infestation to be taken care of. Germplasm to be quarantinedin National Bureau of Plant Genetic Resources comprises mainiyof seeds of various agri-horticultural crops while 2-3% of it is alsoin the form of vegetative propagules like bulbs, corms, cuttings,root stocks, tubers and seedlings, which harbour a variety ofinsects and mites of diverse habits. It is essential for a quarantinepersonnel to establish the systematic identity .of the interceptedpests to know their habits, so as to be able to employ a suitable/prescribed quarantine treatment schedule as the latter are specificto the pest as well as the material to be salvaged.Quarantine Treatments and their schedulesTreatments commonly used for the quarantine of germplasminclude X-ray radiography, fUmigation, pesticidal dip/spraytreatment, mechanical cleaning, tissue culture, cultural proceduresetc. Apart from these, quarantine treatments commonly used forfresh commodities are cold, heat, controlled atmosphere andcombination treatments. Although, limited for germplasm, theiruse is gaining popularity as environment friendly and alternatetreatments, specially to the use of chemicals, viz., methyl bromide,which has been designated as an ozone layer depletor and is likelyto be phased out in near future. Each of the above treatment isdiscussed below. .X-ray radiographyThis technique ~.hL.?Lal." 196'l~ is used against insectswhich live inside the seeds with no apparent external symptomsof their presence within. Over 300 plant genera have been listedfrom literature, seeds of which are reported to harbour such insectsand these are, therefore, essentially subjected to X-ray radiography.The seeds are spread in a single layer and are exposed to X-rays .at 22 kv, 3 rna for 10 seconds from 30 em distance. Infested seedsare clearly distinguishable from healthy ones on the X-ray plateafter development. Corresponding (infested) seeds are then handpicked from the original sample which is kept undisturbed. Thisis nondestructive and fool proof method of excluding insects' butis very cumbersome and is feasible only in small samples ofpreferably bold seeds.

196 Manju Lata KapurFumigationThis is one of the most effective methods used as quarantinetreatment. Due to technicalities involved specifically trainedpersonnel are required to undertake the operations.Fumigation is a process in which chemicals effective astoxicant gas (fumigants) are used in an enclosed space tokill the pest organisms.A fumigant may be a gas (methyl bromide), liquid'(ethylene dichIoride~carbon tetrachloride mixture), solid(aluminium phosphide) or a crystalline powder(naphthalene) at room temperature, but for effective plantquarantine usage it must be highly volatile with goodpenetrability ., Specially designed dose dispensers are required to applygaseous fumigants, whereas heating may be required forvolatilization of liquids and solids.A fumigation schedule generally comprises dosage/concentration of fumigant (C), exposure period (T),temperature and pressure. 'Following factors need to be considered while undertakinga fumigation.a) Concentration Time Product (CTP) is constant for a schedule.After a minimum concentration build up, exposure period can beadjusted or vice-versa to have required CTP.b) Sorption is uptake of gas by solids in a system. This resultsin the loss of some fumigant and necessitates monitoring ofconcentration during the fumigation.c) Penetrability is capacity of a gas to diffuse through air andthe commodity and this can be improved by using fans/blowersor vacuum i.e. reducing air (pressure).d) Temperature affects physical absorption of a fumigant as wellas the insect respiration, thus determines the dosage requirement.e) Permeability of the containers holding the commodity andairtightness of the enclosed space are also equally important indetermining the efficacy of a fumigation.

Plant Quarnatine Treatments for Salvaging Gem/plasm Infested with Insects and Mites 197f) Equipments required to conduct a fumigation include anairtight enclosed space/chamber with or without a pump, fan orblower, a heating mantle, dose dispenser, concentration mOnitoringunit, leak detector, pressure gauge and temperature recorder.On the basis of pressure at which a fumigation is tonductedit is termed as atmospheric or vacuum fumigation. .a) Atmospheric fumigation: This is a fumigation conducted atno!mal air pressure (NAP) in any airtight enclosure i.e. apolyethylene bag, under a tarpaulin or tent, a laboratory autoclaveor a steel drum, which can retain a fUmigant during the exposureperiod without appreciable loss through leakage. Airtightness canbe tested by an open arm manometer filled with kerosene.Ahnospheric fumigation takes care of most of the insects/mites/stages thereof that are external! surface feeders. For examplefumigation with ethylene dichloride- carbon tetrachloride ceOCT)@ 320 mg/1 for 48 h at NAP and 30°C is effective for most of thecommon stored product insects. .b) Vacuum fumigation : In this process most of the air in thechamber is removed. For this purpose, a specially designed /constructed chamber is required which is usually made up of steeland is capable of withstanding external pressure upto oneatmosphere. The installation includes a pump to evacuate the· .chamber, and valves and ducts for introduction and exhaustion offumigant. The primary opject is to hasten the penetratiqn of thefumigant through tightly packed material or internal .infestation.The operation generally gets completed in 1-4&1/2 h as comparedto atmospheric fumigation which takes a minimum of 24 h.Fumigants which can be ~afely used in vacuum include ethyleneoxide imd carbondioxide mixture, hydrogen cyanide gas andmethyl broJl!ide. Specific quarantine fumigation schedules havebeen summed up in Plant Quarantine Treatment Manual, USDA(Anonymous,1985). Fumigation with MB @ 64 mg/l or HCN@ 47.1 mg/l for 2 h at reduced pressure equivalent to 200 mm ofHg at 27°C gives quarantine' control of internal infestation ofphytophagous chacidoids in the seeds of Sesbania, Coriander andfennel (Verma, 1991).Aeration ,is ventilation of fumigant after treatment so as toremove the fumigant from the space and the material.Complete diffusion back to the atmosphere/desorption

198 Manju Lata Kapurmay take upto 24 hours or even longer at temperaturesbelow lOC)C, but can be hastened by forced aeration throughthe use of fans/blowers or repeated air-washings byproducing and releasing vacuum.Safety p.recautions are a must, while undertaking anyfumigation as all fumigants are highly toxic to all livingforms. A fumigator must know the threshold limits of the. fumigants under use and also the maximum period ofexposure above which it is hazardous. Gas masks mustalways be used. Gas leak detectors, concentration monitoringunits, first aid and antidotes should always be kept readilyavailable.Pesticidal dip/spray treatmentAlthough fumigation is the most practical method in plantquarantine for eliminating insect infestation, some plant materialsare not tolerant to dosage required to care for a pest in interest.Under such circumstances approved pesticidal dip treatment maybe given. Plants should be entirely submerged in solution for aperiod of 30 seconds to 2 minutes. Agitating the plants while.immersed in solution will eliminate air pockets and dislodgeresidues which could prevent complete pesticide coverage. Similarlya spray should completely drench the plant. Only freshly preparedsolutions should be used. Despite all J use of chemical dips/spraysfor control of pests as plant quarantine measure is limited as theseare effective only against surface feeders. A complete control isnot always feasible because of lack of penetration due to thepresence of wax or fine hair on insect bodies.M;echanical cleaningPhysical cleaning or excision of infested plant parts is alsoused as quarantine safeguard, especially when pest incidence islow and consignment is comparatively smal~. By this method pestorganisms are physically removed along with the infested plantparts, e.g., insects and mites with galls, larvae with fruit pulp,borers with branches and miners with leaves etc.Tissue cultureIn heavily infested planting material, salvaging is alsopossible by excising the healthy portion of the plant and using it

Plant Quarnatine Treatments for Salvaging Gerrnplasm Infested with Insects and Mites 199for tissue culture under aseptic conditions to produce healthyplants.AerosolsInsecticide particles dispensed through gas are mainly used. as space sprays in plant quarantine to disinfest, cargoes andinspection sites. These are effective against flying/ exposed insects.Hot water treatment (HWT)This is primarily used against pathogens, but various fruitflies have also been controlled by this method. For examplemediterranean fruit fly -15 minutes immession at 50°C (Armstrong,1989).Vapour heat treatment (VHT)Air, heated and saturated with moisture is used to raise thetemperature of commodity to the reqUired point. Specificallydesigned airtight, insulated chambers with a false floor, ducts,thermocouples and temperature'recording system are required toundertake VHT against various fruit flies. The schedule prescribesan approach period during which a commodity is heated upto therequired temperature and held at that during treatment period.Commodity is then cooled immediately and air dried. Forexample, VHT for oriental fruit-fly llh approach period from23.3°C to 44.4°C to be held 'for 8&3/4 h (Sec et al., 1974).Microwave HeatingShort exposures of 100-130 seconds to raise the temperatureupto 53°C-57°C have been found effective against tobacco leafinsects (Herose et al., 1975).Controlled Atmosphere TreatmentsThe use of manipulated natural atmosphere in terms of CO 2,02 and N z content to treat a commodity is called controlled ormodified atmosphere treatment. This involves removing of lifesupporting O 2by adding CO/N 2• These treatments are quitesimilar to fumigation in respect of their requirements, i.e., airtightchambers and gas dispensing units etc. In fact these can be anappropriate substitute for fumigation because gases involved donot leave any harmful residue and prov~de superior conditions to

200 Manju Lata Kapurnormal storage. The treatments have been successfully usedagainst insects ill fruits, vegetables, cut frowers and also in storedgrain, e.g., inN2~ 02 atmospheres, O 2levels below 1.2% for 1 weekat 350C and above, and for 24 weeks at 15°C give complete controlof stored grain insects. (Banks and Anis, 1977). In CO 2atmospheres, 60% CO 2gives 95% kill of most of stored grain pestsafter 4 days exposure at 27°C Gay, 1971) whereas, initial 70% CO , 2subsequently maintained at 35% for 10 days at 20°C also gavecomplete kill (Banks, 1979).Combination treatmentsCombination or multiple treatments have been found moreeffective in the sense that the use of cold, hot or controlledatmosphere treatments in conjunction with each other or fumigation,gives an advantage of lower effective dose, milder temperatureextremes or shorter exposure periods which alone would not beeffective or result in unacceptable injury to the commodity. Forexample unrooted cuttings of chrysanthemum stored at 0-2"C for4 days followed by fUnUgation with MB @ CTP 54 for 4 h at 15°Cagainst Spodop,tera littorallis (Powell, 1979).With the present knowledge of the treatments/salvaging. techniques to get rid of plant pests, healthy genetic units can besalvaged/ developed for safe exchange of virtually any germplasm.Hence Plant Quarantine can be projected with pride as a filter andnot a barrier against global exchange of plant genetic resources.ReferencesAnonymous (1985). Plant Quarantine Treatment Manual, USDA.Armstrong, J.W. (1989). Development of hot water immersionquarantine treatment for Hawaiian grown 'Brazilian bananas./. Econ. Ent. 75 : 787. .Baker, A. C. (1939). The basis for treatm~nt procedure wherefruit-flies are involved as a condition of entry into the U.S.Circ. 551, USDA.Banks, H. J. and Anis, P.c. (1977). Suggested procedures forcontrolled atmosphere storage of dry grain. Canberra,CSIRO Tech. Paper 13 : 23 p.

Plant Quarnatine Treatments for Salvaging Germplasm Infested with Insects and Mites 201Banks, H.J. (1979). Recent advances in the use of modifiedatmospheres for stored product pest control. Proc. II Jnt.Working Can! Stored Prod. Entomol. Ibadan Nigeria, 1978 :198.Benschoter, C. A. (1963). Evaluation of EDB as a fumigant forcitrus and mangoes infested by mexican fruitfly. J. econ. Ent.g:m. .Herose, T.; Abe, I.; Kohno, M.; Suzuki, T.; Oshima, K. and Okakara,l.T. (1975). The use of microwave heating to control insectsin cigarette manufacturing. J. Microwave Power 10: 181.Jay,E. G. (1971). Suggested conditions and procedures for usingcarbon dioxide to control insects in grain storage facilities.ARS Bulletin : 51.Powell, D. F. (1979). Combining methyl bromide fumigation withcold storage to eradicate Spodoptera littoralis on chrysanthemumcuttings. Plant Pathol. 28 : 178.Seo,S.T.; Hu, B.K.S.; Komura, H.i Lee, G.Y.L. and Harris, E.J. (1974).Dacus dorsalis, vapour heat treatment in papayas. J. Econ. Ent.67 : 240.Verma, B.R. (1991). Vacuum fumigation schedule for seedinhabiting chalcidoids. J. Entomol. Res. 15: 229.Wadhi, S.R.; Verma, B.R.; Thomas, Soares and Rattan Lal (1967).Detection of phytophagus chalcidoids in seeds for quarantinepurposes. Indian J. Ent. 29 : 197.

,IISymptoms of Nematode DamageV.K. 'Mathur and Nandini GokteIntroductionDuring the last two decades, there has been tremendousincrease in collection of plant germplasm and its exchange acrossthe national and international boundaries for use in cropimprovement programmes. However, it has aiso led to increasedrisk of introduction and spread at exotic as well as virulentpopulations of indigenous pests and pathogens (includingnematodes) to new areas.' It is therefore, essential for plantexplorers and breeders to become acquainted with symptoms ofnematode infection to avoid collection and exchange of nematodeinfected plant propagules.Plant parasitic nematodes are obligate parasites and exceptfor a few species, which attack buds, leaves or stem, rp.ost speciesattack subterranean plant parts. Nematode feeding on plant tissuesresult in mechanical or biochemical injuries which get expressionin manifestation of symptoms. The symptoms of injuries causedby plant parasitic nematodes can be divided into twocategories: 1) Nonspecific symptoms 2) Specific symptoms.Non-specific symptoms'Symptoms of nematode attack can be either due to damagedroot system or direct damage to plant parts. ,Root systemsdamaged by nematodes usually get manifested in form ofnonspecific symptoms as chlorosis of leaves, reduced vigour, slowdie back, smalling of petioles and leaves, redu~ed ability to tolerate

Symptoms of Nematode Damage 203adverse conditions (as lack of moisture, temperature extremes,nutrient deficiency), reduced tillering and general weakening 'ofplant. There may also be reduction in quality as well as quantityof flowers, seeds and fruits. In standing crop, patches of unthriftyplants are usually indicative of nematode problem. These symptomsresult from inadequate supply of water and nutrients to aerial partsdue to improper functioning of root systems.Specific symptoms of nematode damageSpecific symptbms of nematode damage may be evident onabove ground parts and! or on subterranean parts. The expressionof symptoms varies .with nematode species and host plantinvolved. Most often the infested plants exhibit a combination ofsymptoms. The follO"~ing description can serve to familiarisegermplasm collectors of symptoms of nematode attack so thatcollection and exchange of nematode infected germplasm materi~lcan be avoided.Symptoms on above ground partsDead or devitalised buds : Growing point or buds of plants mayget killed due to infestation by nematodes as Aphelenchoidesparietinus, Abesseyi and A fragariae. The term 'blind' denotes thiscondition. Abesseyi on strawberry causes dropping off of flower'buds.Seed galls : Seed galls formed by Anguina trifici offers the mostoutstanding example. Nematode larvae invade flower primordiawhich instead of developing into seed gets transformed intorounded nematode gall. Nematode galls are green initially butwith crop maturity turn brown black. Gall formed on Festuca grassby A. agrostis are fatal to animals due to association of bacteria withgalls.Distorted, shrunken, discolored seeds: Seed borne nematodes asAphelenchoides besseyi and Ditylenchus dipsaci attack inflorescence ofplants and result in formation of withered, shrivelled andmalformed seeds. Seeds showing such symptoms may harbourheavy population of nematodes as usually no symptoms areexpressed at low levels of nematode infection.Crinkling of leaves : Crinkled, folded leaves usually result fromattack by Anguina spp. and Aphelenchoides spp. A.besseyi on

204 V.K. Mathur and Nandini Goktestrawberry causes a condition known as' Crimp, Summer Crimpor Summer Dwarf. Affected plants show crinkled, folded leavesdark brown in colour which are also quite brittle. A. tritici infectionon wheat also results:··l similar crinkled twisted leaves. A. fragariaecauses a condition in strawberry called as' Spring Crimp'. Crinkledleaves are also associated with infestation of Nothotylenchus acrison alfalfa.Leaf galls and spikkles : Galls on leaves are produced by somespecies of Anguina. Small, slightly raised, yellow pimple like areascalled as 'Spikkles' on Narcissus leaves are associated withDitylenchus dipsaci infection.Leaf spots, lesions and discolorations: Spots or lesions on leavesmay be indicative of infection by species of Aphelenchoides. Foliarnematodes enter through stomata causing destruction of leafparenchyma which gets apparent as leaf spots or lesions.A. ritzemabosi and A. fragariae cause leaf spots on Chrysanthemumand Begonia respectively. Spots first appear on undersurface assmall yellow areas whiC;h turn to brown and later black. The spotsspread and coalasce leading to destruction of l~af.A. ritzemabosi causes interveinal discolouration in Strawberryand Chrysanthemum. Discolouration is also evident in Aster leavesinfected with D. dipsaci. White tip disease of rice caused by A.besseyi, is characte:rized by upper 2-5 cm of leaves becoming yellowthen white and ultimately brown and necrotic.Stem galls: Stem galls are produced by many species of Anguina.A. tumajaciens produces greenish or reddish galls on Cyanodontransvaalensis. Similarly, A. pastulicola infection results in pustulelike stem galls on grasses.Necrosis and discolouration of stem: Necrosis and discolourationof stem 4ue to nematode attack may range from light or paleyellowishness to different shades of reddishness or even blackcolour. Rhadinaphelenchus cocophilus causes formation of brown redring in transverse secti-on of stem in coconut. A band of necrotictissue is formed followed by appear~nce of reddish colour due tomortality of cells. Stem discolouration is also evident in pine treesinfected .with Bursaphelenchus xylophilus.Distortion of stem : Stem distortion and twisting are evident ininfestation of Anguina tritici on wheat.

Symptoms of Nematode Damage 205Abnormal growth and swellings: Nematode infection may causeabnormal growth of stem and leaves as a result of hypertrophy andhyperplasia. Such swellings can often be of diagnostic importance.A. tritici produces swellings near basal part in about 3 week oldseedlings. Swellings in infected tissue are also caused by D. dipsacion narcissus and A. ritzemabosi on strawberry.Below ground symptomsRoot galls : PreSt'net.' of galls on roots is characteristic symptomproduced b\ r\ lIlt-knot nematodes (Meloidogyne spp.), Falserootknot I1L'matode (Nacobbus spp.), and Ditylenchus radicicola.Small root galls are also produced by Hemicycliophcra p.renaria onCitrus and Xiphinema diversicaudatum en roses. The size and formof galls formed varies with the species involved, infection leveLhost species and age of plant. In papaya, root-knot nematodeproduces galls as large as golf balls while in grapE-vine galls arEquite small. Nacobbus sp. produces galls in strands or bead likefashion along the root.Root lesions: Small or large dark necrotic lesiom are producedby migratory endoparasites as. species of lesion nematodePratylenchus, burrowing nematode Radopholus similis and rice roctnematode, Hirschmanniella sp. The lesions IDay ultimately coalasceleading to browning of roots. On large roots, lesions can formcallused margin. Root lef'ions may somtimes be caused l::yectoparasites as Cricone11;.oides sp.Excessive root branching : Nematodes as Mtloidogyne sPF.,Heterodera spp. and Nacobbus spp. can stimulate development ofbranch rootlets near the pOint of invasion leading to excessive rQ(ltbranching variously called as' witche's broom, hairy root andbearding'.Discolouration, necrosis and decortication of re.ots : Nematodefeeding specially by ecto and semiendo parasites result in smallsuperficiG'.1 areas of necrosis which may spread resulting incomplete browning of roots. These sympt.:>ms < re indicative ofinfection by species of Xiphinema, Tylenchullls, Htlicotylenchu5 alldAphelenchoides parietinus. In citrus, feeder ['Dots infected with T.semipenetrans are slightly thicker than healthy on('s and have dirtyappearance due to adhering.of soil particle.::; to gelatinous eggsacsof nE:ma tode. Heavy nematode infection results in cC'rtical ~ ~oughing.

206 V.K. Mathur and Nandini GokteDevitalised root tips : These symptoms are caused due topenetration of nematodes just behind root tip resulting in ceasationof root tip growth. Stubby root caused by Trichodorus sp. and insome plants by Pratylenchus spp. is characterised by numerousshort stubby roots arranged in cluster. Coarse root is caused byMeloidogyne sp. and Belonolaimwt sp. Abrupt halt in growth oflateral roots results in appearance of main root devoid of smallrootlets. Feeding by species as Xiphinema and Meloidogyne near theroot tip results in unequal growth rate on two sides of a root givinga curl to tip area referred to as curly tip.Pearly root: Small pearly creamy white ,pin head sized, sphericalbodies attached to roots may be females of cyst forming genera asHeterodera spp. and Globodera spp. These can be seen with nakedeye or with a hand lens after gently washing the root system toremove the adhering soil and debris.Rots: Nematode infection in many cases incite and aggravateactivity of other microorganisms. Many secondary microbes alsoenter through wounds created by nematode resulting ultimatelyin rotting of tissues. Yam nematode Scutellonema bradys and dryrot nematode Ditylenchus destructor offer classic examples of thistype of nematode damage. Infestation of potato by D. destructor isfirst characterized by small discoloured superficial spots followedby drying and cracking of skin. Microorganisms enter throughcracks and ultimately cause rotting.Disease complexes: Another facet of symptomatology of nematodeinfection is interaction of nematodes with other microorganisms indisease complexes. Yellow ear rot of wheat is one such typicalexample and is caused by interaction between nematode A. triticiand bacterium Corynebacterium michiganense pv. tritici. The diseaseis recognised by appearance of yellowish slimy bacterial mass onear head and curling of leaves. Similarly cauliflower disease ofstrawberry is caused by involvement of both nematode A. fragariaeand bacterium Corynebacterium fascians. Symptoms produced indisease complexes are usually at variance with those produced bythe component organisms occurring singly.ConclusionsCollection/ exchange of germplasm and quarantine activitiesare equally necessary to strengthen and develop agriculture of the

Symptoms of Nematode Damage 207, country. Plant breeders and explorers can contribute towardsprevention of introduction and spread of pests and pathogensincluding nematodes by following guidelines such as 1) plantmaterial showing one or more symptoms should not be collectedor exchanged, 2) plant propagules should be free of soil as soilserves as vehicle of dissemination of plant parasitic nematodes, 3)all the seed/plant material imported from abroad should berouted through plant quarantine channels, and finally 4) domesticplant quarantine regulations be not contravened.ReferencesDropkin, V.H.(1980). Introduction to Plant Nematologtj. John Wiley& Sons, 293 pp.Swarup, G., Dasgupta, D.R. and Koshy, P.K. (Eds.) (1989). PlantDiseases. Anmol Publications, 395pp.

IIDetection Techniques for Plant Parasitic Nematodesin Imported Germplasm Material for ResearchArjun LalIntrol,iudionAmong the multicellular animals, nematodes are one of theoldest existing forms of life. Nematodes show considerablevariation in their habitat. They are found in all oceans, from thepolar region to the equator, from the literal zone to the abyssaldepths; they colonize freshwater lakes, riyers and marshes and alltypes of soil from the antarctic to the tropics; they parasitize inostgroups of animals (including other nematodes), and a wide varietyof algae, fungi, and higher plants. Of the total nematode species,about 25% are free-living (soil and fresh water), 10% are plantparasites, 15% are animal parasites and remaining 50% are marine.In agriculture, we are mainly concerned with those nematodeswhich attack crops and reduce yields directly or indirectly 'inassociation with other plant pathogens like fungi, bacteria andviruses.Nematodes attacking plantsA plant growing in soil is often attacked by one or morespecies of plant parasitic nematodes. Plant parasitic nematodes arefound in and about roots of host plant, or in the leaves, stems, buds,flowers and seeds of plantjs. There a~129.E:!.2.!~q9 ~p~cies of pl~ntpa~~s )).ematodes . attacking various species of plants. Theirgreatest diversity of form occurs, amongst parasites of roots. Agrain of galled wh~at lnight contain 30,000 nematodes. One gramof coconut root could yield about 400 Radopholus similis. By causing

Detection Techniques for Plant Parasitic Nematodes 209mechanical injury, they predispose the plant to other pathogenslike fungi and bacteria. Some plant parasitic nematodes are knownto transmit plant viruses of economic importance.Length of survival in nematodesMost plant parasitic nematodes are subjected to unfavourableconditions sometime duril1g their life cycle and the persistence ofa species in a habitat confirms that the survival mechanisms areoperative. The periods of survival for some economically importantnematode species are given in Table-l.Table 1.Length of survival in some nematodes of quarantineimportanceNematode species Length of Survival conditionssurvivalAnguina tritici 32 years In galls at SoCAnguina agrostis 4 years In galls at room temp.Bursaphelenc1lUs xylophilus 1 year In wood logsDitylel1chus dipsaci 23 years Plant tissue at room temp.Ditylenchus destructor 5 months Peanut hulls at 10 DCGlobodera rostochiensis 2.5 years In cysts at room temp.Heterodera avenae 5.5 years In cysts at SoC & 75% RHHeterodera glycines 6 years Cysts in dry soil at °-20 °CHeterodera schachtii 3 months Encysted eggs in soil at 62°CPratylenchus penetrans 11 months In dry soil in greenhouseRadopholus similis 6 months Without host in field soilIt is clear from Table 1 that certain nematode species havegreater or lesser survival capacity than others. In other wordslength of survival depends on factors like temperature, aeration,moisture, rate of water loss, stage of life cycle and amount of fatsand carbohydrates in nematode body. If the nematodes are notkilled in plant material or seed, activity usually resumes rapidlyupon return of favourable environment.Means of nematode spreadPlant parasitic nematodes move relatively short distancesunder their own power. In a sandy soil under optimum moistureand temperature conditions, a nematode will travel about 30 cm/year. Being very minute in size (average body length 0.5 mmi body

210 Arju1l Lalwidth 0.02 rom), nematodes often go undetected in germplasm asthey contaminate soil or infest/infect plants, roots, tubers, bulbs,seeds etc. When infected plants and plant materials are collectedand moved, the nematodes travel with them. One of the worstparasitic nematodes (Ditylenchus dipsaci) have moved around theworld in this manner. Studies of the nematode movement andspread reveal that man is often the culprit, unintentionallycontributing to the distribution of these pests.N ow we shall discuss some of the commonly used techniquesto detect and diagnose the nematode infested plant material orseed and simple laboratory methods used to recover nematodesfrom infested soil or plant material.Nematode detection techniquesI. Detection of nematodes in the laboratory. Germplasm material received in the quarantine laboratory issubjected to detailed examination by employing various nematodedetection techniques depending upon the kind of plant material,size of the sample and availability of time. Some commonly usedtechniques are described below :a) Examination of the plant material: Examination of the plant·tissues under the microscope is the most conclusive and directevidence of nematode association with the infested plant material.This method is most suitable for nematodes producing distinctsymptoms on plants as their females become swollen.b) Observation of symptoms: Many species of plant parasiticnematodes cause stunting, yellowing, or wilting of their hostplants in the field. Nematode-plant interactions also result indevelopment of some specific signs of abnormal plant tissuegrowth (Table-2). It may be possible to find nematodes by directexamination when careful inspection of the plant/plant materialreveals-****Galls or swellings on roots, tubers, or rhizomesWhite, yellow, or brown pinhead size bodies adheringto rootsSwollen or malformed leaf, stem, or other tissueRoot lesions, or unusual root proliferation

Detection Techniques jar. Plant Parasitic Nematodes 211TalJle 2.Detection of nematodes on the basis of symptoms inseed or planting materialCropNematodeSymptoms of damageSeedsWheatFaba beanOnion'RicePeanutAnguina tritieiDitylenclttts dipsaciDitylenchus dipsaciApl1elencllOides besseyiAphelenchoides arachidisDitylenchus destructorBulbs, Corms, Rhizomes & TubersBlack, misshapen seed galls.Shrivelled and discolouredseeds, sometimes withnematode wool attached.No distinct symptoms on seed,nematodes emerging fromsoaked seeds examinedmicroscopically:No observable symptoms butthe infested seeds are lighterand nematodes from soakedseeds can be recovered byremoving the seed coatmicroscopically.Shrivelled seeds with darkbrown testas.Seed blemished or unsound,pods and hulls dark coloured.Onion,garlicGingerTurmericBanana, Abacaplantain(Mllsa spp.)Ditylenclms dipsaciMeloidogyne spp.Radopholus similisRadopholus similisRadopholus similisPratylenchus coffeaePratylenchus goodeyiHelicotylenchusmuiticinetu5Internal necrosis or rot of(Allium spp.) sets and cloves,sometimes with nematode wq,6lattached.'Light brown, wa terv areas onsurface, often in Q'onstrictionswith internal necrosis.Small, sunken, watery lesionson surface.Shallow, water-soakedbrownish les10ns and rotting.Purple to c;iark brown necroticlesions thrbughout cortex ofattached roots, and similarlesions in/buter tissues of corm.Purple fu dark brown necroticlesions in periphery of cortexof attached roots.

212Arjun LalContd ..... (Table 2)CropNematodeSymptoms of damageTaroYam(Dioscoreaspp.) .PotatoHirschmanniella miticausaScutellonema bradysPratylenchus coffeaeRadopholus similisMeloidogyne .spp.Pratylenchus spp.Meloidogyne spp.Globodera spp.Ditylenchus destructorSeedling transplants (symptoms after uprooting)Vegetables,rice, coffee etc.RootstocksGrapevine,peach, etc.CitrusMeloidogyne spp.NacobbllS spp.Pratylenchus spp.Hirschmanniella spp.Radopholus spp.Heterodera spp.Meloidogyne spp.Pratylenchus spp.Tylenchulus semipenetransInternal red necrotic streaks.Internal dark brown dry rotin periphery of tuberobserved when cut or whenepidermis is scraped away.Whole tubers spongy to touchwith surface cracks.Uneven, knobbly tubers;necrotic spots internally.Uneven tubers pitted with darkdry necrotic lesions.Watery tubers, swellings andlesions on surface.Cysts on tuber or in adheringsoil (difficult to detect).Surface cracking and internaldark brown rot and peripheralrot.Swellings of galls on roots.Discrete swellings or galls onroots.Yellow or brown lesions onroot.surface necrosis seen aftercleaning.Spidery or distorted root growthsometimes with white specks(females) on the surface in olderseedlings. .Galls on roots, sometimesaccompanied by rotting.Brown lesions on root surface.Irregular, dirty roots with soiladhering to surface.

Detection Techniques for Plant Parasitic Nematodes 213If any of these symptoms are observed on the plants Iplanting materials, dissection can then be undertaken to confirmthe nematode presence.c) Dissection techniques: For examination of nematode infectedplant tissue, following things are required: stereoscopic dissectingmicroscope, petri dishes or Syracuse dishes, fluorescent illuminator,dissecting needles, fine-bladed knife, scalpel or razor blade. Cutaffected roots, b~bs, leaves or buds into small pieces. Place a fewpieces in a petri dish or Syracuse dish containing water and notethe presence of nematodes under microscope by cutting open thetissues with dissecting needles. If any nematodes are found, pickthem up and process for identification.True seeds infected with nematodes do not show any clearcut symptoms and, therefore, difficult to separate from healthyones. However, in some cases (paddy, onion, peanut, foxtail millet,fababean, alfalfa) infected seeds are discoloured, shriveled andlighter in weight. Nematodes from such seeds are recovered bysoaking the seeds in water at 25 DC for about 24 hrs. and observingunder stereoscopic microscope after crushing the seeds or removingthe seed coat. It is always better to examine as many seeds aspossible. Important nematode species reported as seed borne orcarried on seed are listed in Table-3.d) Staining techniques: It is often difficult to detect nematodesin plant tissues as they are colourless or translucent. Staining issimple and helps in differentiating nematodes from plant tissues(specially roots). The most common staining technique used forroot samples is to colour the tissue with acid, fuchsin dissolved inlactophenol or lactoglycerol (0.05%), then destain it with clearlactophenolllactoglycerol. The nematodes retain red stain morestrongly than does the root tissue.e) Soil examination: Some plant parasitic nematodes (cystforming and ectoparasitic ones) can spread with small amounts ofsoil mixed with seeds or adhered to roots. To detect the presenceof cysts in soil, 5-10 gm of soil (removed from seeds by sieving orhand picking the clods) is mixed with 20 m1 water and directlyobserved under stereoscopic microscope at 15 to 50 X magnification.Brown to dark brown cysts can be seen if the soil is infested. Collectall cysts by a forecep or brush and send for identification afterproper labeling. ,

214 Arjun LalTable 3.Some important seed-borne nematodes of quarantineimportanceNematode speciesAnguina tritidAnguina agrostis*Aphelenchoides besseyiAphelencltoides arachidisDitylenchus destructor'Ditylenchus . dipsaci*Pratylenchus brachyurHsRhadinaphelenchus .cocophilus*Common nameSeed gall nematodeBent grass nematodeWhite tip nematodePeanut testa nematodePeanut pod rot nematodeStem & Bulb nematodeLesion nematodeRed ring nematodeHost plantsWheat, barleyBentgrassPaddy, Setaria,Panicum, StylosanthesPeanutPeanutAlfa alfa, onion,. garlic, faba bean,tulip, GladiolusPeanutCoconut, oil palm* Nematode species not yet reported from India.f) Molecular aids to nematode diagnosisAs stated in the beginning, only a few nematode-plantinteractions produce distinctive, readily observable symptoms onroots, stems, leaves, bulbs and seeds. As a result, the diagnosis ofnematode- induced plant damage is dependent upon isolating andidentifying plant parasitic nematodes from infested plant tissuesor soil samples. Over the last two decades, efforts have been madeto diagnose and identify nematodes by analysing proteins, lipids,carbohydrates and most recently DNA sequences. In fact, significantprogress has been made in developing these molecular aids fornematode diagnosis, for example, the use of esterase or malatedehydrogenase phenotypes to separate Meloidogyne arenaria, M.javanica, M. incognita, and M. hapIa; use of monoclonal antibodiesand DNA probes to separate Globodera rostochiensis from G. pallidaland Bursaphelenchus xylophilus from B. mucronatus.Techniques to detect DNA sequences differences betweenorganisms can be divided into three basic approaches: detection ofRestriction Fragment Length. Polymorphisms (RFLPs) 'betweennematode DNA samples; use of DNA probes in dot blot assays andPolymer chain Reactions (peR). In general, most DNA studies forspecies, and race{pathotype separation have made comparisons

Detection Techniques for Plant Parasitic Nematodes 215between RFLPs in total DNA (genomic and mitochondrial DNA(mt DNA) of unknown nematode populations against ~pecies andraces standards.It is very clear that use of biotechnology based methods ofnematode diagnosis (early detection, identification andquantification of populations in soil and plant tissues) Will increasespecially in quarantine services and in certification of -nematodefree seed and root stocks. These methods have been used to detectviruses and bacteria in propagated plant material for quarantineand certification purposes as they have proved to be efficient}accurate and user friendly. However} further research is requiredfor quantification and direct use of DNA probes or monoclonalantibodies on samples extracted from soil or plant tissues asopposed to isolated nematodes.II.Detection of nematodes in Post-Entry QuarantineThe field that is to be used for planting introducedgermplasm material must be completely free from species ofnematodes for which material is to examined under PEQ as wellas other indigenous nematode species. Detection of a nematodespecies will depend upon the average density and number ofsamples taken per unit area. Conversely, the probability that aninfestation of given density will be detected depends on thequantity of soil examined and on the number of points from whichthe sample is derived. There are about 2 million Htres of top soilper hectare upto 20 cm depth and thus it is practically not possibleto detect all the nematodes present in soil. However, percentagechances of detection increases with increase in number of samplesper unit area.For detection of nematodes on a crop grown in PEQ,individual plant should be carefully observed for development ofsymptoms. Soil and plant (induding roots, stems} leaves) samplesshould be collected from suspected plants and analysed in thelaboratory for presence of nematodes of quarantine importance. If. any exotic nematode species is detected, the plants must bedestroyed and PEQ area must be cordoned off, treated with heavydosages of nematicides and not allowed to grow any crop till it iscertified to be free from nematodes.-Above discussed techniques are simple and efficient to detectthe presence or association of nematodes in soil or plant material

216 Arjun Lalcollected or introduced, if they are used carefully. For recovery ofnematodes from infested soil or plant material in large numbersfor identification and further studies, several nematode extractiontechniques are used by the nematologists which require specialequipments.a) To extract nematodes from soil : If the sample size is verysmall (100 g or less), the soil may be placed directly in a funneland processed by the mist extraction or Baermann Funneltechnique. Larger soil samples are first processed by the combinedscreening-funnel technique or the gravity-screening techniquefollowed by mist extraction of the residue.To extract cysts-forming nematodes from soil: The centrifugalflotation tecruuque is most efficient in recovery of cysts from soil.To recover sluggish nematodes from soil: Nematodes havingvery little locomotion power such as Criconemoides, Hemicycliophoraare recovered by centrifugal-flotation methods.b) To extract nematodes from roots: Infected foots are cut intosmall pieces, and processed by mist extraction or Baermann funnel,or gently washed and subjected to jar or plastic bag incubation.Mist extraction is effective in recovering both endoparasitic and~ctoparasitic nematodes. Jar incubation of washed roots recoversendoparasites and migratory endoparasites only.e) To extract nematodes frolP other plant parts: To extractnematodes from tubers, bulbs, rhizomes, leaves etc. the mistextraction method is most effective but Baermann funnel methodcan also be used.Each technique has its own advantages and disadv~ntagesand selection of a method may vary with the purpose of thestudies.Suggested ReadingsCurran, J. and Robinson, M.P. (1993). Molecular aids to nematodediagnosis. In: Plant parasitic Nematodes in Temperate Agriculture(ed. Evans et al.) : 545-564, CAB!, UK.Lal, A. and Mathur, V.K. (1988). Seed health testing for plantparasitic nematodes. In: Proc. Seed Sci. & Tech.( Yadava, T.P.& Chandgi Ram, eds.), pp. 318-320, HAD Press, Hisar.Southey, J.F. (1986). Laboratory methods for work with soil and plantnematodes.· MAFF, HM:SO London: 1-202.

Treatment Schedules for Eradication ofNematodes from GermplasmRajanIntroductionAvailability of pest (nematode) free germplasm material fromdiverse origins is an important requirement in breeding a cropvariety of desired characteristics. With the movement of germplasmthe nematodes and nematode problems are known to spread fromone region/ country to another. Through men's activities, nematodesare disseminated either due to unconscious efforts or throughmovement of numerous types of plant materials. The importationand exportation of seeds and more so the vegetatively propagatedmaterial such as rooted plants, bulbs, rhizomes and tubers with orwithout soil must always be regarded as a major breach in anyphytosanitary barrier and should be allowed only when the stockis guaranteed to have been in 'soil or growing media free fromquarantine parasites. Most of the times this requirement is not metand a question is posed, what to do with the material which hasbeen imported at a high cost, when examination reveals it to beinfested. It is not always possible to advise for destruction of theinfested material. The other, most practical solution to the problemis to salvage such material.A large number of treatment schedules are available forbringing down the nematode populations to certain levels. Thenematodes which escape/survive through such treatments maymultiply under new environmental conditions and establishthemselves. So the whole objective of keeping an'area virgin from

218 Ra'ancertain species of nematodes fails. The tolerance limit in quarantineis zero ftherefore, special treatments, with 100% efficiency are amust to cope with problem of nematode dispersal alongwithgermplasm. Such disinfestation procedures mainly comprise of hotwater treatment, nematicidal dip (chemotherapy), mechanicalcleaning and I or fumigation.Salvaging methods for nematodesHeat therapy; An excellent review of physicC!-1 methods of controlof plant parasitic nematodes is given in Southey (1978). The mostimportant physical method of nematode control is application ofenergy as heat to produce a temperature exceeding the limit oftolerance of the nematodes. Hot water treatments are commonlyused to control nematodes in planting material moving in nationaland international trade. Fortunately, many nematodes are lesstolerant to heat than their host plants, especially when the plantsare at a dormant stage, as with bulbs, tubers, perennial root stocksetc. Such hot water treatments to planting material, therefore,generally meet quarantine requirements in salvaging the material.(a)Treatment for the bulbsHot water treatment has been used for more than 80 yearsto control Ditylenchus dipsaci in flowering bulbs, includingnarcissus, hyacinth and some cultivars of tulip. As infested bulbsare symptomless it is good practice to treat all bulbs. This isparticularly importanfwhen bulbs are being exported to a differentcountry /region, as D. dipsaci can easily be introduced with infestedbulbs.In the Netherlands D. dipsaci in flower bulbs ranks high inimportance among quarantine pests. The Dutch plant healthregulations require inspection of all bulb crops and infested stocksmust be given a prescribed hot water treatment. Importednarcissus bulbs are invariably treated whether infested or not. Thetreatment is a severe one (four hours at not less than 45"C) beinggeared more to destruction 6f D. dipsaci than to ensuringsatisfactory growth of the bulbs. In the same country, investigationsrevealed that hot water treatment for control of D. dipsaci in bulbsof narcissi at 43.5°C for three to four hours is sufficient and showedthat similar treatment also controls the nematodes in hyacinths,

Treatment Schedules for Eradication of Nematodes from Germplasm 219violets, muscari, gladiolus and potatoes. The chief aim of much ofthis work was to test the tolerance of the nost plant to hot water.Several refinements have improved the efficiency of hotwater treatment for control of D. dipsaci in narcissus bulbs andlessened the risk of damage to the bulbs. These include early lifting,storage at 25-35°C followed by presoaking, immediately before hotwater treatment, use of a wetter and formaldehyde in the treatmenttank, use of anti foaming agents and increasing the temperatures.Bulbs may require drying after treatment to avoid hazardsin subsequent storage. Further the treatment has limited usefulnessbecause of certain drawbacks. The margin between killing ofendoparasitic nematodes and phytotoxicity is usually very narrow.HWT can be applied only when bulbs are dormant.(b)Treatment for the rootsHWT is also used to control nematodes on grapevinerootlings, Meloidogyne javanica in South Africa (Smith, 1982) and M.javanica and the citrus nematode in Australia. Dormant rootlingsare treated for either 15 min. at 50°C or 5 min. at 52°C in SouthAfrica and for 5 min. at 51°C in Australia. Immediately after HWTthe rootlings are plunged into cold water to minimise damage tothe plants.Control of Pratylenchus penetrans in raspberry root cuttingswas attempted by a chemical bare root dip in several chemicals andby a hot water treatment. Only the hot water treatment waseffective in killing the nematodes without injuring the plants. Fivecultivars were tested with hot water. The nematodes wereeliminated without injury to the cultivars (McElroy, 1973). Thetreatment schedules developed in the Plant Quarantine Lab atNBPGR., for salvaging germplasm are mentioned in Table l.(c)Treatments for the tubers, corms, stools and suckersHot water treatment can be used as a preventive measure tocontrol endoparasitic nematodes that are disseminated on or incorms, tubers, rhizomes and other planting materials. An applicationof hot water treatment for control of Aphelenchoides ritze11labosi inchrysanthemums is by treating the dormant stools. Here theproblem is simpler than with bulbs, since one is dealing withnematodes that are either external contaminants or occur within

220 RajanTable 1.Treatment schedules developed in P.Q. Lab at NBPGRfor salvaging germplasmPlant material Nematode Treatmet1. Vetiver Heterodera a) HWT of roots at 50°C forzeae15 min. ora) Dipping roots in Demacur-1500 ppm for 15 min. oriC)Dipping roots in Nemagon-1000 ppm for 15 min.2. Mentha spicata Pratylenchus a) HWT of roots at 48 0 C forhamatus, and30 min. orMeloidogyne b) Dipping roots in DBCPincognita,0.1 % for 15 min.3. Peach plants P. pen.etrans a) HWT of roots at 470C forfor 15 min. followed byHWT at 'SooC for 20 minand finally treating at 52"Cfor 15 min.4. Rosa indica P. zeae arid a) HWT of roots at 48°C forHelt'cotylenchus10 min. followed bygaleatustreatment at SO"C for 5 min.5. Apple rooted P. vttlnus a) HWT of roots at SO"C forcuttings15 min.6. Proso millet, or Aphelenchoides a) Presoak seeds in 1% HP2Foxtail millet or besseyi for 3 hours followed byForage millet seedsHWT at 48°C for 15 min. orb) Without presoaking HWTof seeds at 50°C for 10 min.7. Banana Radopholus a) HWT of suckers at 55"C forsimilis5 min. orb) Dipping suckers in 0.05%Formalin at 50°C for 10min.8. Chinese potato M. incognita a) HWT of tubers at 53"C for10 min. orb) Dipping suckers in 0.05%formalin at 50 n e for 10min.

Treatment Schedules for Eradication of Nematodes from Germplasm 221Contd ..... (Table 1)Plant9. Grapevine10. PomegranatematerialM. incognita andM. javanicaM. incognitaNematode Treatmeta) HWT of roots at 4snC for20 min. followed bytreatment at SO°C for 10min.a) HWT of roots at SO°C for10 min. followed bytreatment at S2°C for 10min.shoot remains and leaf fragments etc. attached to the stools - theheat does not have to penetrate to worms deep seated wi,thin planttissue. A perfect control could be achieved by treating stools at 46°C for 5 min with less damage to the plants than from the previouslyrecommended treatment of 43.3°C for 20-30 min. Although theshort treatment is usually safer and more convenient. Someworkers found that a few varieties were less harmed by the oldertreatment of 43.3°C for 20-30 min. and they found it possible toclassify chrysanthemum varieties according to their tolerance ofboth systems of treatmentFollowing success with 46°C treatment for chrysanthemums,scientists treated strawberry runners at that temperature for 7 min.and obtained good control of D. dipsaci without damage to the. plants. Slightly longer treatment appeared to be necessary to kill"Aphelenchoides spp. in strawberries, and for these it was suggested10 min. It was found that a more severe treatment 49°C for 20 min.was necessary to produce nematode free runners for. a strawberrycertification scheme, potted plants being treated upside down andonly the crown and leaves immersed.The nematodes M. incognita and Rotylenchulus reniformiswere completely eliminated from heavily infested sweet potatoplants by immersing the pl~nts in water at SO°C for 3-5 min. Insome cases immersion for longer periods up to 20 min. killed orseverely damaged plants (~artin, 1970).Hot water treatment of rhizomes of bananas to' controlburrowing nematode is improved by peeling all necrotic tissuefrom the corms before treatment. HWT of pared banana sets is

222 Ra'anpracticed in Australia and other countries to control the nematodes.Sets are treated for 25 min. at 55°C.With hot water treatment, problem is to kill the quiescent preadultjuveniles or wool on ,the outsides or between dry scales.Efficient HWT of planting material is obtained only if the materialbeing treate~ is dormant and free of soil.(d)Treatment for the s,eedsWhite tip a seed borne disease of rice caused by A. besseyi iseasily controlled by HWT of seed for 15 min, at 52-54"C For thesame nematode on Setaria and other minor millets, time andtemperature requirements are slightly different ..Heat tolerance and heat hardeningThe chances of successful hot water treatment increase as thedifference between thermal sensitivity of host and nematodesincreases, with the latter being more sensitive. The smaller thisdifference, more accurately the temperature and time of thetreatment must be controlled, and the more important it becomesto treat only the most heat resistant plant propagules available.As a precautionary measure material should be plunged into coolwater to terminate the treatment.ft should be realised that both the host and the nematode aremore tolerant to heat in a dehydrated and dormant stage than ina stage of active metabolism. In many cases the plant can be grownor pretreated so as to produce tolerant stock. Rhizomes of bananagrown in relatively dry soil were more tolerant to hot watertreatment to control R. similis than were mOTe succulent ones ofthe same size grown in relatively moist peaty soil in Surinam.In Britain and The Netherlands, early lifting and subsequentstorage at 25-35°C made it possible to treat narcissus bulbs forcontrol of D. dipsaci at temperatures up to 47.7"C instead of 43.3"Cfor three hours, without damage to the bulbs. However, thisadvantage has to be balanced against the fact that warm storageincreases the resistance of the nematodes as well. Good results ofhot water treatment of strawberry runners to control leaf and stemnematodes were obtained in Ireland, when runners were taken inSeptember, preheated in warm water and immersed in cold waterafter hot water treatment at 45 L1 C for 15 minutes. Thermal injmywas reduced and uniformity of growth increased after hot water

Treatment Schedules for Eradication of Nematodes from GermpZasm 223treatment of rose plants infected with Pratylenchus vulnus by (i) firstinducing cold hardiness by storing at 2.2°C for 3 weeks then(ii) holding at 37.8°C for 24 h to induce heat hardening, followedby (iii) hot water treatment of the root system at 48,3°C for 35minutes (Towson and Lear, 1982).Vapour heat and hot air treatmentsVapour heat treatment, Le., exposure to heated, moistureladen air, have been used to control various pests. Air is passedthrough a mixing chamber with steam and cold water sprays,balanced to give required temperature, and mixture is circulatedthrough an insulated treatment chamber containing the bulbs orother plants. The method has some possible advantages over hotwater treatment in reducing risk of fungus infection facilitatingsubsequent handling and drying. Some expei'iments indicated thatsome of the nematodes survived plain vapour heat treatments justas they did hot water treatment without formalin, unless thetreatment was severe enough to damage the bulbs,Chitwood and Blanton suggested less severe vapour heattreatments preceded by a formalin pre-soak. Results show that hotwater treatment for 30 min. was effective at 48°C and above,whereas with vapour heat 30 min. at Sl°C was necessary forcomplete control. However, in this instance the plants were saidto withstand all treatments without injury. Dry heat is known tobe less effective than moist heat for treating nematode infestedplants. All the nematodes are heat sensitive when active in waterand most resistant when in desiccated state.Microwaves'There is not much experience yet on the application of heatby microwaves to disinfect living plant material of nematodes.Hendrick et al. (1982) completely eradicated root knot nematodeswithout significantly decreasing the vigour of rose plants bytreatment with microwaves for 30 seconds at 600 W. Results wereas good as with hot water treatment at 48°C for 30 min.Chemotherapy(a)Bulbs and setsIn 1917, in a lecture Ramsbottm described his experiments oncontrol of D. dipsaci in nards sus bulbs by immersion in water and

224 Rajanvarious chemical solutions at different temperatures. Hastings(1933) quoted Weiss of the U.S. Department of Agriculture whohad shown that active stages of D. dipsaci escaping into the waterof the bath could survive even after four hours at 43.3°C and hadproposed the addition of one or two per cent formalin to the water.Several workers have found that Formaldehyde is not verynematicidal at low temperatuIes but becomes much more so astemperature rises. Hot-water formalin treatments at 43.3°C havealso been used in U.S.A. against D. destructor in iris bulbs andAphelenchoides fragariae in lily bulbs.D. ,dipsaci, in tulips and narcissus can be controlled bydipping the bulbs in the organophosphate thionazin, the treatmentbeing 2500 ppm for ~;5-3 h. Dips of banana sets are a regularpractice, best control being reported with fenal1).iphos at 100 ppmfor 5 min. (Decker et aI., 1971). .(b)SeedsThe seed transmitted nematodes can be controlled byfumigation with methyl bromide in an air tight chamber, but thedosage (concentration * time product) required will depend on theoil and moisture content of the seed. In onion seed, dosage in, excess of 800 mg h/L needed to control the cryptobiotic stage ofthe stem nematode, are phytotoxic, but lucerene seed will toleratedosage upto 2500 mg h/L which is in'excess of the effective dosage.Methyl bromide has been shown to be effective against Anguinaagrostis in bent grass. Generally seeds should be at about 10%moisture content to avoid phytotoxic damage by methyl bromide.(c)Bare root dipsDipping seedlings and transplants in non volatile chemicalsis an effective method of controlling nematodes. M. hapla Chitwoodin roses was controlled by a dip in phenarruphos or ethoprophos.at 100 PPll'l: for 30 min; and the immersion of seed potato piecesin fenamiphos at 1800 ppm was very effective against severalnematodes. Dipping ,tobacco transplants in oxamyl is a good wayof checking introduction of several nematodes.Total control of M. incognitl1; was obtained with fensulfothionand thlonazin (1000 F\Pm) on peach plants (Ponchillia,1973). Boththe treatments were, not phytotoxic. Thionazin at 2000 ppmapparently eradicated the n~matode, M. hapia on peony roots

Treatment Schedules for Eradication of Nematodes from Germplasm 225without phytotoxic effects (Malek, 1974). M. ~apla were completelyeliminated from rose plants when the washed roots were dippedfor 39 min. in 0.1% solution of prophos (Mocap) and Nemacur at16°C (Dale, 1973).Uptake of nematicides by dormant plant parts may beinsufficient to eliminate nematodes occurring in the centre of thematerial (e.g., leaf nematodes in lily bulbs). Therefore, someworkers tried control of root kilot nematode'by immersion of plantmate:rial in nematicidal solutions at different temperatures. Thebioassay showed that immersion in phenamiphos for 60 min. at16°C or 30 min. at ~4°C eradicated M. javanica (Suatmadji, 1982).Zem et ill. (1980) reported that nematicides applied as a hot solution. were efficient in controlling nematodes but had adverse effects onplant growth.Other methodsMechanical seed cleaning is considered the major reason forthe disappearance of the wheat seed gall nematode, formerly acommon and serious pest in north - west Europe. In generalnematode infested seeds are generally shrunken, small andsometimes have'discolouration on the surface. The easiest way ofsalvaging is to extract all such seeds for further examination andtreat them, if needed. As symptoms are not clear in case of certainseed borne nematodes, such as A. agrostis in grass seed, A. besseyiin rice seed and D. dipsaci occurring in small particles of debrisassociated with onion, clover and lucerene seed, more refined seedcleaning methods might be expected to assist in the control of theirspread alongwith healthy seeds. Treatment for the whole lot ofsample is recommended.Ectoparasitic nematodes on root systems or other undergroundplant parts may also be effectively removed by light brushing orrubbing of the plant parts in a tank or a stream of water.Fumigation .with methyl bromide rarely penetrates plant tissuesufficiently to kill endoparasitic nematodes.Precautionary measuresWhenever scientists have a real choice between differentmeans of propagation for a particular crop, the one which preventstransfer of nematodes in planting material or enables thenematologists to examine material for nematode symptoms before

226 Ra'anplanting should be recommended. A number of such alternativesare available. For example stem cuttings of potato, cassava andsweet potato do not carry nematodes whereas tubers or roots oftencontain Meloidogyne and other nematodes. Seeds of pyrethrum arefree of M. hapla. Stolons taken from the base of vines of blackpepper (Piper nigrum L.) could carry R. simi/is and MeloidogJ-Jne sp.but the movement of stem cuttings will ensure that gennplasmexchanged is free from nematodes. In yam (Dioscorea spp.) the useof tuber pieces rather than mature whole tubers for exchange willenable the examination for symptoms of dry rot caused byScutellonema bradys or P. coffeae and other nematodes.The leaf bearing tips of mature corms of taro (c. esclilenta)are generally free from H. miticallsa and should be used inpreference to cormels or suckers which can be infected. In additiontransport of nematodes on planting material can be prevented byacquring seeds rather than seedlings, e.g., with perennial cropssuch as coffee which can be infested with the root parasitesMeloidogyne, PratylencJllls and other genera. Use of tissue culturefor exchange of germplasm can prohibit entry of several pests andpathogens. Techniques have been standardised in case of severalcrops, e.g., garlic, banana, etc.ConclusionsGermplasm procured for import or export to some countryneeds examination and presence of nematodes which is very likelyin underground parts and vegetatively propagative materialsrequire some treatment to make it healthy for exchange.Treatment schedules are carefully planned recommendationsfor salvaging plant or plant parts with 100 1 )10 efficiency ofelimination of nematodes without any or very less damage to thehost plant. For such recommendations thorough investigations aremade with regard to temperature and time interval- tolerance incase of heat treatment and concentrations time as well astemperature in case of nematicidal dip treatments. Specialprecautions are needed as most of the times the treatment whichis lethal to the nematodes is also lethal to the host. To meet suchconditions either dormant stages of the host or uninfestedpropagative parts of the plant or heat hardening of such parts bypre treatment is suggested. Treatments vary from host to host andspecies to species of nematode infesting it. Development and

Treatment Schedules jar Eradication oj Nematodes from Germplasm 227application of proper treatment schedule not only makes healthyexchange of germplasm possible but also avoids economic lossesincurred on the material for its repeated procurement.ReferencesAdesiyan, S.O. (1977) Studies on the effect of gamma radiation(from Cobalt 60 source) on storage life of white yam(Dioscorea rotundata val' efon) infected with Scutellonemabradys. Ann. app!. BioI. 86: 213-218.Dale, P.S. (1973) Elimination of root knot nematodes from roses bychemical bare root dips. N.ZI Exp. Agric. 1 : 121-122.Hastings, R.J. (1933). Treatment of narcissus bulbs with hot water.Gdl1rs C/zron. 94 : 313-314.Malek, R.B. (1974). Control of MeloidogtJne hapla on peony bychemical bare root dip. Pl. Dis. Reptr. 58 : 997-999.Martin, W.J. (1970). Elimination of root knot and reniformnema todes and scurf infections from rootlets of sweet potatoplants by hot water treatment. Pl. Dis. Reptr. 54 : 1056-1058.McElroy, P.D. (1973). Control of PratylenclUls penetrans in raspberryroot cuttings. Pl. Dis. Repfr. 57 : 492-495.Ponchillia, P.E. (1973). Control of Meloidogync incognita on peach:chemical bare root dips. PI. Dis. Reptr. 57 : 489-492.Southey, J.F. (1978). Physical Methods of Control in PlantNematology, Technical Bulletin No.7. Third Edition, HMSO,London.Suatmadji, RW. (1982). Control ofroot knot nematodes, MeloidogynejavanicCl, in rooted stocks of grapevine, Vitis vinifern byimmersion in nematicide solutions at different temperaturesand in hot water. Nematol. Medit. 10 : 119-125Towson, A.J. and Lear, B. (1982). Control of nematodes in roseplants by hot water treatment preceded by heat hardening.NefllCltoiogicCl 28 : 339-353.Zem, A.c., Alves, E.J. and Santos-Rodrigues, J.A. (1980). Efficiencyof different treatments for the control of nematodes in bananaplanting material. Soc. Brasileira de Nematologia ; 65-83.

IIPlant Genetic Resources activities at IPGRIOffice for South AsiaR.K. AroraIntroduction. The International Plant Genetic Resources Institute(IPGRI) isthe legal successor to the International Board for Plant GeneticResources (ffiPGR), and became operational in 1993. IPGRI is aninstitute of the Consultative Group on International AgriculturalResearch (CGIAR). The CGIAR was established in 1971 andsupports a network of 18 international agricultural research centres .induding IPGRI. .IPGRI has four main elements / objectives in its work on plantgenetic resources: .1. Strengthening national programmes2. Contributing to international collaboration3. Improving strategies and technologies for conservation4. Providing an international information serviceThe mandate of IPGRI is to advance the conservation and useof plant genetic resources for the benefit of present and futuregenerations. IPGRI's mission is to encourage, support and engagein activities to strengthen the conservation and use of plant geneticresources worldwide with special emphasis on the needs ofdeveloping countries. IPGRI considers national programmes to bethe basic elements of any global effort in pl~nt genetic resources.,IPGRI will undertake research and training and provide scientific

Plant Genetic Resources activities at IPGRI office for South Asia 229and technical advice and information. The strategy of the IPGRI"Diversity for Development" elaborately deals with IPGRI'sprojections. Its programme is built on the basis of multidisciplinaryprojects, comprising a series of activities. .To facilitate overall collaboration in its activities, the institutehas adopted and implemented a structure of eight programme,groups; five regional and three thematic. The regional strUctu~~·comprises five Regional Groups which are responsible for theinstitute's work in Sub-Saharan Africa, West Asia and NorthAfrica, Asia, the Pacific and Oceania, the' Americas and Europe.The Asia, Pacific and Oceania (APO) Regional Office is located atSingapore and has two more offices under it - the Office for EastAsia at Beijing and the Office for South Asia at New Delhi.Fig. 1 gives the location of regional offices. These Regional Groupsare responsible for developing and reviewing regional strategiesin PGR activities. In addition to the five Regional Groups, threeThematic Groups have been established at Headquarters in Rome,Italy. These Groups are responsible for developing and coordinatingresearch and information work of inter-regional or global relevancein their respective subject areas. The Groups are also responsiblefor providing scientific and technical support to the regions. Thethree Groups comprise; Genetic Diversity, GermplasmMainten.anceand Use and Documentation, Ipformation and Training.IPGRI office for South AsiaThe IPGRI office for South Asia is located in the premises ofthe National Bureau of Plant Genetic Resources, New Delhi. Theestablishment of this Office (initially Regional Office for South &Southeast Asia) was formalized with GOI under a Memorandumof Understanding with Indian Council of Agricultural Research(leAR) in November, 1987 and the Office became operational inJuly, 1988. The South Asia region covered by this Office inlcudesBangladesh, Bhutan, India, Maldives, Nepal and Sri Lanka. It isvery diverse agro-climatically, topographically, culturally and aseat of diversification for several crop plants; rice, grain legumes,minor millets, spices, oilseeds, several tropical fruits & vegetablesetc. With India, its.programme was jointly developed under thebiennial work plans with ICAR/NBPGR, for plant geneticresources activities as per national priorities.

230 RK. AroraObjectivesThe major mandate is to promote and strengthen plantgenetic resources activities in the region with emphasis on theirconservation and use. Emphasis is on the folowing activities:a) . Assist the national plant genetic resources programmesin the region in strengthening their PGR activitiesthro~g~ advice, small projects, etc., as per r~quest.b) 'Coordinate and support PGR activities on a regionallevel wherever desired by the countries and whenevermeaningful (South Asian PGR national coordinatorsmeeting~ workshops on crop networks, training coursese1c,r'-c) Create awareness on national and regional level andpromote the conservation and utilization of PGRd) Coordinate/undertake information synthesis anddocumentation of data on plant genetic resources heldby national programmes.e) Liaise with lARes and other regional, globalorganizations/FAD and the NGOs; andf) Disseminate 1?GR information/literature etc.Activities undertakenExplorationThe primary focus on exploration activity has been to :i) Facilitate joint explorations in the region and to provideguidelines/logistics. Crops/national programmesinvolved in this activity are : egg plant & okra­Bangladesh (NBPGR-BARI), Nepal (NBPGR-NARC),Sri Lanka (NBPGR-PGRC); Sesame-Bangladesh (NBPGR­BARI); Buckwheat-Bhutan (proposed).ii)iii)iv)Support within-country explorations on priority i.e.,okra, eggplant, maize, sesame undertaken by NBPGRAssess gaps in collecting, including wild relatives,genetic erosion, national priorities.Assist in preparing project-proposals, country-reportsto prioritize germplasm collecting activities.

Plant Genetic Resources activities at IPGRI office for South Asia 231Characterization, Evaluation and DocumentationSome of the regional activities in which IPGR1: is involvedinclude:i) Preparation of descriptors such as of buckwheatii)iii)ConservationChoice of crops of national/regional importanceAssess status of evaluation efforts on germplasm heldby national programmes and promote this activity,including preparation of crop-catalogues.IPGRI advocates complementary conservation strategiesinvolving in-situ, ex~sitlt and other approaches. More emphasis. ison ex-situ conservation technologies vis-a-vis functioning ofgenebanks. It emphasizes on seed conservation and the use ofcomplementary conservation strategies. These include :(i)(ii)Trainingex-situ conservation of assessed material in genebank:base col1e~tions/ active collections; field-genebanks forvegetatively propagated species, and in-vitroconservation and cryopreservation.provide guidelines for seed conservation aspects inoverall perspective, and provide technical/scientificadvisory role and assistance.To develop more trained personnel in the field of plantgenetic resources, IPGRI organizes specific and broad~based shortPGR training courses. Since its inception, it has supported severaltraining programmes organized/ conducted by NBPGR, such as :i) F AO Regional Training course on collection andconservation of crops of local importance.ii)On-job training for South Asia region on characterization,evalu~tion and conserv'}tion of orthodox seeds.Further, it has supported sponsoring M.Sc. training courseat Birmingham University, (candidates from Sri Lanka, India),regionally / globally on case to case basis, to promote know~howin national programmes and develop local expertise.

232 RK. AroraWorkshopslMeetingsTo develop better dialogue among national programmes inthe region, the office organized the South Asia NationalCoordinators' meetings in 1990 at NBPGR, New Delhi, and in 1992at PGRC, Peradeniya (Sri Lanka). The Proceedings of the firstmeeting contain country reports of all national programmes andare very informative to draw upon gUidelines on future work.Recommendations through NPs provide action plan in collaboration, with countries of the region.As this region is a centre of diversity for several crops anda strong work-base exists at national progralllmes, IPGRI has usedtheir expertise and organized international workshop such as onokra genetic resources at NBPGR in 1990, and 011 sesamum'Evaluation and im~rovement' in 1993.Other programmeslBPGR has also facilitated visits of scientists from the regionto IPGRI sponsored programmes (regionally, globally) whichinclude:·1. Trainings2. Workshops3. 'Symposia .4. Working Groups/meetingsUse of r~gional expertise is also encouraged and made useof in different activities in the region. The office also helps inprOViding other services such as genetic resources information,sources etc.Information dissemination(i)Two way approach is followed :provide IPGRI publications to promote concern on PGRactivities and advance research thereof, IBPGR has produced>200 publications which include:F AO-IBPGR NewsletterAnnual RepQrtGene FlowWorkshop IWorking Group Proceedings

Plant Genetic Resources activities at [PGRI office for South Asia 233(ii)Crop NetworksDescriptors for CropsPGR special publicationsNational programmes are regularly supplied with abovepublications. Also selected publica~ions go to senior scientistsconcerned with PGR activities upon request and as permailing list.The office also undertakes prGmotion of such activitiesthrough publication of :.l\~gional Newsletter for Asia, the Pacific and OceaniaBooks and Regional workshop/network proceedingsR~ports on Project evaluationIn overall perspective, the office also provides advisory /info!mation service for PGR infor~ation on all aspects.IPGRI supported projectsTo carry out PGR programmes in India with NBPGR/ICAR,activities have been funded by IPGRI and operated through jointworkplans now under completion (1990-92/93). These activitiesinitiated since 1989, include the following projects:1. Collection of okra and eggplant germplasm' in SouthAsia2. Characterization, evaluation and documentation ofokra & eggplant germplasm3. Collection of sesame germplasm4. Characterization, evaluation and documentation ofsesame germplasm5. Collectionof maize germplasm from Himalayan region6. Characterization, evaluation and documentation ofIndian maize germplasm7. Biological mechanisms determining recalcitrance inseeds of tea, cocoa and jack£ruitAlso supported a project with the National AgriculturalResearch Centre, Hill Crops Improvement Programme, Kabre,Nepal on evaluation of barley, buckwheat, amaranth, finger millet,foxtail millet, proso millet etc. which has been completed.

234 . R.K. AroraNew'programme structureWith IPGRI coming into operation, and better emphasis onPGR conservation and development, the Regional Office for Asia,the Pacific and Oceania (APO), headed by the Regional DirectorDr. Ken Riley, based at Singapore; with its two offices-South Asia(New Delhi) and East Asia (Beijing) which are already operational,prioritized activities to assist national pI:ogrammes in the regionto improve their plant genetic resources activities and capabilities;lay more emphasis on conservation and use of underutilized cropsimportant to the region, strengthen crop networks and to layincreased emphasis on PGR training to develop resource persomlelin the region.Considering the heterogenity in the PGRnational programmesin the region, emphasis will be laid to develop national programmes·in Nepal, Bhutan and Bangladesh. IPGRl consultation and advice,and also assistance in preparing project proposals will be available.Emphasis on underutilized crops would include tropical fruits,buckwheat, sesame, Lathyrus and taros and yams. Capturing oflocal knowledge towards enhanced use of resources woulddemand emphasis on ethnobotanical studies. In strengtheningactivities on crop networks, okra, sweet potato, minor millets,apart from bamboo and rattan, banana and coconut will assumeimportance. In this context, new projects supported includeassesment of diversity in okra using molecular markers (NBPGR,India) and ethnobotanial studies on taros and yams in Bangladesh(BARI, Bangladesh).To conclude, IPGRI is conpnitted to collaborate/coordinatePGR activities with national programmes and in the region, inidentifying their needs and priorities, and devising plans for actionaccordingly.

IIIntellectual Property Rights: Protection ofPlant VarietiesR.S. RanaIntroductionRights of inventors to obtain legal protection for exclusive useand marketing of their creative irinovations are widely recognizedand granted in the form of patents in many developed countries.These are intended to reward the inventors by enabling them toearn exclusive profits from the commercial application of theirinventions over a reasonable period of time and also to ensure thatthe new inventions and discoveries are fully utilized on acommercial scale for public good, particularly in situations whereinve:t;'ltors do not have adequate reso~rces to do so on their own.Licensing is the primary mechanism by which intellectual propertyrights are ttansferred from inventors to investors in a way thatbenefits may reach the users.Breeders' RightsIntellectual property protection of plant varieties is somewhatdifferent from that of the other forms of technology because a newplant variety does not normally arise out of a single innovativestep. It is rather developedirCa' cumulative manner in discernablestages "dth addition of a new advantage or genetic improvementat each level to a basic genulype which often happens to be apo.Vdar locally adapted variety. Fven though a new plant varietymay not involve an inventive step in the strict sense, its economicusefulness to society is unquestionable. It is, hence, important toencourage development of new varieties by providing incentives

236 R.S. Ranasuch as Breeder's Rights over exclusive production and marketingof seed of an improved variety.General Agreement on Tariffs and Trade (GATT with its newform of World Trade Organization) was finally concluded . andsigned by over 120 nations in December, 1993. India is among the64 countries who had ratified this multilateral agreement by June1994 and the agreement is legally binding. It includesTrade Related Intellectual Property Rights (TRIPS). GATT hasbeen replaced by World Trade Organisation (WTO) since 1stJanuary, 1995.All the contracting parties are bound to provide certainminimum levels of protection to new plant varieties through aneffective Sui generis system or patenting or a suitable combinationof both.Criteria for Granting Protection to a New Plant Varietyi) Novelty: The criterion of 'novelty' replaces that of 'nonobvious'which is the requirement for protection of a newinvention. It is used in the sense that the new variety should nothave been ,commercially exploited for more than a year beforegranting protection.ii) Distinctiveness: The new variety must possess a trait thatmake it distinguishable from other known varieties by one or more~dentifiable morphological, physiological, or other characteristics.iii) UnHormity: The new variety must be uniform in appearanceunder specified environment of its adaptation ..iv) Stability: The new variety must be stable in appearance andits claimed characteristics over successive generations underspecified environment.Union for the Protection of (New) Plant Varieties (UPOV)This International Union, founded in 1961 in Geneva(Switzerland) has 24 member countries and has held severalconventions. India is not yet a member of UPOV. Provisions ofthe 1991 Convention are far more strict in protection regime ascompared to ,those of the 1978 Convention. Although the basicinternational treaty prOVisiOns, as amended by the 1991 Convention,have not been accepted by most national governments so far, yet

Intellectual J:roperty Rights,' Protection of Plant Varieties 237the proposed amendments provide an indication of what is tocome. Any country can choose to opt for provisions of the 1978Convention provided it becomes a member of the DPaV by31st December, 1995.Positive points of the 1978 Convention included exemptionsfor use of seeds of protected varieties by farmers (including rightto sell seeds though not on commercial scale) and researchers.These exemptions are proposed to be modiiied by the 1991Convention which has sought to extend Breeder's rights to coveruse of harvested plant part. Whereas, the period of varietyprotection was 15 years (18 years in case of grapevines and trees)under 1978 Convention it is sought to be extended to 20 years (25years in case of grapevines and trees) by the 1991 Convention.Burden of proof of 'not~guilty' will also be on the accused. Anothernoteworthy deviation from the 1978 Convention is a new conceptof an 1/ essentially derived variety". This provision is likely to haveimplication in the context of biotechnology. Consider a case wherea biotechnologist inserts a new gene into a variety protected underPlant Breeder's Rights. Under this new concept, a holder of PlantBreeder's Rights protection can claim corresponding rights over avariety that differs only slightly from the protected variety.Another noteworthy difference between provisions of thetwo conventions pertains to the number of species required to beprotected by member countrieS'. Whereas, the 1978 Conventionrequires protection to cover 24 plant species over a period of eightyears, the 1991 Convention demands protection to cover the entireplant kingdom.The USA sceneBreeder's rights were first granted in USA with the passageof the Plant Patent Act (PPA) of 1930 that covers asexuallypropagated new varieties. Prior to this time, common notion wasthat plants and other living organisms were not eligible for patentprotection because all living organisms were believed to beproducts of nature. The new Act provided that 'Whoever inventsor discovers and asexually reproduces any distinct and new varietyof plant, including cultivated species, mutants, hybrids, and newlyfound seedlings, other than a tuber propagated plant or a plantfound in an unCUltivated state, may obtain a patent thereof, subjectto the conditions and requirements of this title'. The rationale for

238 R.S. Ranaexclusion of tuber propagated plants was that their propagatingand edible portions of the plant were the same (stems). Commonunderstanding at that time was that only asexually reproducedplants were capable of reproducing true-to-type. Hence, sexuallyreproducing plants were excluded from protection.Protection of plant varieties that are reproduced sexually wasgranted by the Plant Variety Protection Act (PVP A) of 1970. Plantbreeding had reached a .stage by that time to develop generalopinion that new sexually reproduced varieties also could bereplicated true-to-type when self-pollinated. Noteworthy provisionsof PVP A are mandatory license, exemption for planting of savedseed, exemptions for sales by persons whose primary occupationis farming, and also an exemption for research use. Protectionunder PVP A does not extended to fungi, bacteria, or firstgeneration hybrid plants.PVP A does not provide protection to commercial hybrids insexually propagated crops (like maize) because hybrids do notreproduce true-to-type in such crops. Protection of hybrid' seedscan be achieved by protecting the inbred parent lines that are usedto produce hybrid seeds and this can be done as trade secrets.Thus, the option of protecting inbred parent seed as trade secrethas been used as a strategy by breeders for protecting theirproprietary inbred lines and also for retaining reasonable controlof their exclusive genetic stocks.A more restrictive regime of patenting, called, Utility Patents,is also being used now by breeders. US Supreme Court's decision(1980) in "Diamond vs. Chakrabarty" opened up this option. Priorto this decision, it was commonly believed that utility patents werenot obtainable for plants. Utility patents were, however, beinggranted for methods of hybridizing and breeding plants.It is important to \assess the type and scope of protectionrequired by a breeder dr other researcher while deciding whichform of intellectual prop'erty protection is appropriate for a given'invention' in plant biotechnology. Protection under PVPA coversthe whole plant only and extends to the plants of the variety asdesr:ribed in the document. In contrast, the utility patent systemprovides for claims of varying scope that can be granted dependingon the nature of the 'invention'. It is noteworthy that, only utilitypatent allows for protv('tion of genes and other parts of pl~nts.

Intellectual Property Rights : Protection of Plant Varieties 239Farmers' RightsIt will be desirable to mention Farmer's Rights while we arediscussing Breeder's Rights, According to FAG resolution no. 5189, Farmers' Rights arise from the past, present and futurecontributions of farmer&- in conserving, improving and makingavailable plant genetic resources, particularly those in the centresof origin I diversity. Attempt is made sometimes to treat theserights as privileges. This is not acceptable because privileges areconsidered optional and can be withdrawn.Farmers' Rights·are basic rights and can not be compromised.These rights should essentially include those arising from nonformalinnovations linked to accural of monetary benefits wheredue, the right to save, use and sell their own seed, the right to theirharvested plant materials and also the right to quality seeds.Burden of proof of innocence or otherwise should be on theaccusers and not on the farmers. Whereas Farmers' Rights arewidely..acknowledged, mechanisms of their implementation arestill being debated.It is worth recalling that agriculture began around 10,000years ago. If this time span is condensed into an hour,conservation of plant germplasm in botanic gardens and otherliving collections began within the last minute while gene bankswith long term storage capabilities appeared just a few secondsback. Plant genetic resources have been selected, developed usedand safeguarded by farmer families and farming communitiesthrough successive generations for most of the time. The keyquestion is whom to reward and how? Tribal people, ruralcommunities and farmer families deserve attention in this context.Proposed Plant Variety Protect~.onLegislation in IndiaAgricultural plants alongwith life-saving drugs were excludedfrom the purview of Indian Patent Act of 1970 that permittedprocess patenting only and not the end-product patenting. Thisact was amended by the Government of India through apresidential ordinance issued on 31 Decembe.r, 1994 that allowedpatenting of produce and extended it to all pharmaceuticals andalso to agriculture. An effort is now underway to develop aneffective Sui generis system for plant variety protection and providea suitable legal framework as reguired under GATT provisions.

240 R.S. RanaEssential features of the 1978 upav Convention are beingconsidered for adoption retaining exemptions for Farmers' Rightsand also for Researcher's use. It is proposed to entitle farmers toclaim suitable compensation from the breeder of the protectedvariety / dealer in the event of failure in the stated performance., Period of protection is being proposed to be 15 years forannual crop plants and 18 years for fruit trees and vines. It is beingmade compulsory to deposit a reference seed sample in theNational Gene Bank and also to do cataloguing in the NationalRegister. Certification :may also be made compulsory. There willbe a provision for compulsory licensing also. .It is being proposed to set up a National Authority for plantvariety protection, i.e., protection of Breeder's, Farmers' andResearch Use Rights. There will also be an Appellate Board toresolve arising disputes. NBPGR is expected to playa vital rolein implementation of the proposed legislation and efforts havealready been undertaken to develop the anticipated capabilitiesand scientific competence.The key point under debate is how to ensure' unrestrictedflow of plant genetic resources for research purpose and alsosharing of research information while providing adequate legalprotection to plant variety across countries.International Convention on Biological Diversity and FAOInternational understanding of Plant Genetic ResourcesThe International Convention on Biological Diversity (CBD),signed by 174 countries, became legally binding with effect from29 December 1993. India ha$ signed and also ratified this historicaltreaty which recognised'sovereign rights of nations over theirgenetic resources and also for determining access to them basedon prior informed consent and linking to transfer to relevant. technologies and sharing of benefits.Prior to the CBD, India was sigiltatory to the InternationalUndertaking on Plant Genetic Resources that was developed byFAO of the United Nations in 1983 and adopted by 112 countries.This International Undertaking was based on the concept thatplant genetic resources were the common heritage of manki,nd.Text of the International Undertaking is being revised now through

Intellectual Property Rights : Protection of Plant Varieties 241step by step process to bring it in harmony with the provisions ofthe CBD. The revised International Undertaking is expected tobecome a protocol to the CBD providing a mechanism for itsimplementation.The CBD recognised two issues to be outstanding thatrequired further consideration to resolve them. These included themechanisms for implementing Farmers' Rights and the status ofex-situ germplasm collections that were not acquired according toprovisions of the CBD, i.e., ass~mbled prior to December 1993.These matters have now been taken up by the FAO Councilfollowing a resolution (3/93) adopted by the conference ofContracting Parties.International Agricultural Research Centres under the CGIARsystem have placed their global germplasm collections underauspices of the FAO upon joining the International Network of exsitucollections through an agreement signed 011 26 Odober 1994.An International Teclmical Conference on genetic resources forfood and agriculture, sponsored by the FAO, has been planned tobe held in June 1996 at Leipzig (Germany) to overview presentstatus of genetic resources in the world and to develop acomprehensive global plan of action in this context.

IIThe Indian N atjonal Gene BankR.S. RanaIntroductionIt is noteworthy that countries that are strikingly rich in. biological diversity happen to be in tropical and sub-tropical partsof globe in marked contrast to those in temperate regions. It is alsoremarkable that most of these nations are als,? technologically poorand have not been able to make full use of their biological resourcesfor human welfare. Among the biodiversity rich countries, IndIahas taken a 'lead in recent years in ex situ conservation andutilization of plant genetic resources (PGR) through the vigorousefforts of National Bureau of Plant Genetic Resources (NBPGR)located at New Delhi. The mission of NBPGR is collection,introduction, characterisation, evaluation, documentation,maintenance, conservation and distribution of germplastn of cropplants and their wild relatives.The Indian National Gene Bank at New Delhi, operated andmanaged by NBPGR, has developed excellent facilities for holdingaround 1.5 million accessions of base collections of germplasmunder long term storage conditions in seed repository, eryo-tanksand in vitro repository. Base collections are linked to activegermplasm collections for safety duplication anp also to provideeasy access for users. The Indian Gene Bank co~pares well withsimilar facilities developed by most advanced nations in terms ofboth international standards as well as size of germpl,asmholdings, Ideally, every apparently unique accession with'appropriate passport data and documentation should be in· theNational Gene Bank. Limitations of resources such as staff, time,

The Indian National Gene Bank 243space and funds, however, impose constraints that need to beovercome to reach this ideal.Plant wealth of IndiaOver 15,000 species of flwering plants have been documentedin the Indian region. The Indian gene centre, one of the original8 Vavilovian Centres of origin of cultivated .plants and theirdiversity, is known to be particularly rich in genetic resources ofcultivated plants and their wild relatives. In addition to more than20 important agriMhorticultural crops that originated here, over1000 species are recognised to be of known ethno-botanicalinterest. Seven biosphere reserves, 67 national parks and over 400sancturies and other reserve areas have already been establishedby the Department of Environment & Forests to promote in situconservation of naturally growing biological diversity in differentparts of the country. Besides native species, a large number of cropplants have also been brought here by traders, travellers andmissionaries from foreign lands. They got naturalised, diversifiedand selected upon in due course in different parts of the countryunder intensive care of local farmers and tribal communitiesGermplasm collectionsCrop improvement programmes began in India during thebeginning of this century with the establishment of severalagricultural research institutes. Landraces and locally adaptedcultivars grown by farmers in different agro-climatic situations arebeing collected since then. A large number of such collectionsestimated to be around four million (including possible duplicates)still lie dispersed at various research centres. Systematic collectionand safe conservation of germplasm to ensure their continuedavailability to plant breeders have been assigned top priority inIndia in view of the feared replacement of primitive cultivars byhigh yielding i!Uproved varieties.National Plant Genetic Resources SystemThe Indian National PGR System comprises a network ofbas,e collections of germplasm of different crops kept under longterm storage and linked to numerous crop-specific active collectionsthat are maintained. at appropriate locations. NBPGR is the nodalorganisation for developing, operating and coordinating this

244 RS. Ranasystem. It is also the national facility in India for ex situconservation of genetic resources of agri-horticultural crops andother economic plants including their wild relatives. Establishedin 1976, this Bureau has a staff of over 560 persons working at NewDelhi and 12 regional stations Ibase centres located in representativeagro-ecological zones. It has developed strong linkages with alarge number of crop-based institutes, national research centres,All India Coordinated Crops Improvement Projects c;tnd stateagricultural universities. It interacts actively with the FADprogrammes in the region and also collaborates with severalInternational Agricultural Research Centres of CGIAR. South AsiaOffice of IPGRI is also located in the NBPGR campus. Mandateof NBPGR includes developing policy guidelines and extendingconsultancy on bi9diversity and PGRs.Base collections of germplasm are kept by NBPGR in theNational Gene Bank. The Bureau is also developing computeriseddatabase on PGR. Crop-wise components of base collections arealso maintained at over 30 National Active Germplasm Sites anddistributed to users from there. Bureau's scientists add. around10,000 new accessions of germplasm each year through plannedexplorations and also introduce around 60,000 exotic seed samplesannually from abroad. All these materials are characterised,evaluated, documented and conserved. Inventories and cataloguesare brought out and Field Days organised to promote their use.Germplasm advisory committees help the Bureau byreviewing the progress of activities, identifying shortcomings andsuggesting ways and means of overcoming them. NBPGR has theauthority on behalf of Govt. of India,. under the New SeedDevelopment Policy (1988), to issue import permits for introductionof plant germplasm for research purpose. It has also beenauthorised to conduct plant quarantine inspection and to issuephytosanitary certificates. The Union Department of Agricultureproposes to enact a Sui generis system of Plant Variety Protectionfor recognition of breeders' and farmers' rights. Likewise, theUnion Department of Environment & Forests is likely to issue aPublic Notice soon under the Export-Import Policy of the ForeignTrade (D&R) Act 1992 for regulation of transfer of genetic resourcesout of India. NBPGR will be assigned key roles in implementationof both these proposed legislations.

The Indian National Gene Bank 245Indian National Gene BankThis national facility for long term ex situ conservation of basecollections, set up by NBPGR at New Delhi, has four majorcomponents:(i) Seed Repository : Samples of orthodox type seeds aredehydrated at 15°C and 15% RH to around 5% moisture content,sealed in laminated aluminium foils under vaccum and depositedin cold storage at -20°C.(ii) Tissue Culture Repository : Disease-free tissue cultures ofrecalcitrant seeds (those not capable of withstanding dehydrationand very low temprature) and vegetatively propagated plantspecies, developed from meristem explants r are maintained usingappropriate growth-limiting media and by subculturing at suitableintervals.(iii) Cryopreservation Facility: Seed samples of selected speciesI synthetic seeds' embroys and gametes are being preserved usingcryopreservation technique, either by immersion in liqUid nitrogen(-196 D C) or by keeping in its vapour phase (around -150°C)following protocols using computerised freezing-thawing ratesand chemical cryoprotectants.(iv) Clonal Repository: Germplasm collections of vegetativelypropagated crops like turmeric, ginger, sweet potato and bananaare being maintained under field conditions following suitablecrop rotations and backed by tissue-culture s~fety duplication.Priorities of the,Gene BankThe Indian Gene Bank serves as the national facility for longterm conservation of genetic resources of cultivated plants, andtheir wild relatives and also species of economic importance.Priorities include landraces, locally adapted traditional cultivars offarmers, primitive and old varieties, prized genetic stocks and wildand weedy relatives of cultivated plants. Seed samples of allimproved crop varieties released at the national level for commercialproduction as well as of the varieties granted protection are alsokept as reference materials. The Indian Gene Bank has alsoaccepted global responsibility for providing safety duplication ofgermplasm holdings of several crops on behalf of lARes underterms set out in memoranda of understanding.

Z46R.S. RanaGene Bank Standards(i) Seed sample size: About 3,000 viable seeds in case of selfpollinated crops and twice that number for cross-pollinatedspecies. One third of this samples is sealed in a separate packetand stored alongside the original sample under identicalconditions(at _20DC) for monitoring germinability at regularintervals.(ii) Viability testing: Initial germination tests are carried out ona minimum of 200 seeds drawn randomly from the originalsamples.Acceptable initial germination percentage should be higherthan 90% for most species but lower level$ are acceptable for somevegetables an.d wild forms. .50-100 randomly drawn seeds are used for subsequentviability monitoring tests.(iii) Seed drying : Seeds are dried in seed drying cabinetsoperated at 15 D C and 15% relative humidity uSing, silica gel asdesiccant based on adsorption system and appropriate air flow.(iv) Seed storage conditions : Seeds are dried to 5% moisturecontent and sealed jn laminated aluminium foils. Base collectionsare stored in modules operating at -20°C. Active collections arestored at 40C and 35% relative hUmidity.(v) Regeneration: Undertaken when seed viability falls to 85%of the initial value. '100 plants or more are preferred for thispurpose.Capacity of the Indian N ationa! Gene BankA built-in cold storage vault of 1003m size was obtained fromUK in 1983 to gain experience in medium and long term seedstorage. It comprised two compartments, on.e operating at 4°C andthe other at-20°C. Four more modules were subsequently, ..procured, assembled; installed and made operative maintaining-20°C. Two of them had ,a capacity of lO03m each while the othertwo were larger with a size of 176 3 m each. A long term storagecapacity for OVer 2,00,000 seed samples was thus created, backedfully by diesel generator sets. A sizeable capacity for in vitro. repository and cryopreservation has been developed recently to

The Indian National Gene Bank' 247complement the seed repository and this capacity is being furtherexpanded.Infrastructure for installing twelve more modules for longterm storage of around 800,000 seed samples has also beencompleted. When fully developed by 1996 1 the Indian NationalGene Bank will have a capacity for long term storage of over onemillion accessions. -Germplasm holdings in the Indian National Gene Bank andPGR system are listed in Table 1. Information provided inTable 2 presents a global scenario showing known numbers ofgermplasm accessions held by gene banks around the wo:r1d.Indian National Gene Bank ranks third after gene banks of USAand Russia in terms of size of collections. In fact, its capacity forlong term storage is next to that of USA at pres~nt and it Can beeasily expanded ~hen required.Table 1.Germplasm Holdings in Indian National Gene Bankand PGR System as on 31.12.1994ComponentsNo. of accessionsSeed samples dehydrated to 5% moisture 139,576content, sealed in laminated aluminiumJoils and stored at ~20°C .Seed samples, gametes, meristems cryopreserved 875in liquid nitrogen (~196°C)Accessions maintained in tissue/cell cultures 765Reference sample~ stored at 4°C, RH 35% 27,350Accessions under multiplication and 120,226char~cterisation in N13PGR networkAdditional accessions under multiplication 188,263and upgrading in the Indian PGR System

248 R.S. RanaTable 2.Categ_oryEstimated Gobal Germplasm CollectionsNo. of accessionsIARCs 510,000Other international and regional gene banks 40,000Dev'eloped countries2,230,000Developing countries1,550,000Others30,000Total4,360,000Research NeedsReal strength of Gene Bank lies not only in the size of itsholdings and safety of its storage conditions but also in the geneticdiversity among its collections and the extent of informationavailable on each of its accessions. Duplicates among its holdings,burden the system and contribute to wastage of space and energy.They need to be identified and removed so as to minimiseredundancy. NBPGR is gradually developing a strong base in.collaboration with several leading research centres for more precisecharacterisation of its accessions of proven value at biochemicaland molecular levels employing techniques like isozyme profiling,RFLP, RAPD-PCR and MSV. Time has now come to develop andemploy multi-locus DNA probes for DNA finger printing of mostprized native accessions as also single-locus probes for screeningavailable collections for detecting genotypes possessing desirabletraits or co-adapted gene complexes.

LIST OF PARTICIPANTS OF TRAINING COURSEON EXCHANGE AND QUARANTINE OFPLANT GENETIC RESOURCES(January 10 ~ 24 ,1995)1. Shri D.K. Agg

25012. Dr. S.K. KaushikScientist (Plant Breeding),Central Potato ResearchInstitute, Shimla,Himachal Pradesh-l71 001.13. Dr. Angrez Singh KhattraVegetable Botanist,Department of VegetableCrops, Punjab AgriculturalUniversity, Ludhiana,Punjab.14. Shri B. Kt'islmamoorthyScientist (SG), Plant Breeding,National Research Centre forSpices, Post Bag 1701,Marikunnu P.O.,Calicut-673012, Kerala.15. Shri K. MohanasundaramAssoc. Professor, Breeding(AICRIP), School of Genetics,Tamil Nadu AgriculturalUniversity, Coimbatore-641003,Tamil Nadu.16. Dr. N.D. RautScientist, Seed TechnologyResearch, Dept. of PlantBreeding & Genetics, JNKVV,Jab~tlpur-482004, MadhyaPradesh17. Dr. A.K. RoyScientist, Indian Grassland &Fodder Research Institute,Jhansi-284003, Uttar Pradesh.18. Dr. Rajiv Kumar SharmaDivision of Vegetable Crops,Indian Agricultural ResearchInstitute, Delhi- 110012.List of Participants19. Dr. Suneel SharmaAsstt. Horticulturist, CCSHaryana Agricultural University,Hisar, Haryana.20. Dr. S.K. SaxenaSr. Scientist,Division of Fruits &Horticultural Technology,Indian Agricultural Rese