IPVDN – Training in virus detection and diagnosis, capacity building, and delivery of IPM packages

1. IPVDN – Training in virus detection and diagnosis, capacity building, and delivery of IPM packages 7th International IPM Symposium Memphis, Tennessee March 27-29, 2012 Capacity building and short term training: Requirements for successful technology transfer for Integrated Pest Management Sue A. Tolin, Virginia Tech stolin@vt.edu

2. The IPM CRSPIPVDN Team – 2009-2014Plant Virus Disease US Participants and Regions c Sue Tolin, Program Leader Judy Brown UAZ Naidu Rayapati WSU MikeDeom UGA Bob Gilbertson UC-D South Asia India Bangladesh Nepal West Africa Mali Senegal Ghana Central Asia Tajikistan Latin America & Caribbean Honduras Guatemala Ecuador Southeast Asia Indonesia Philippines Cambodia East Africa Uganda Kenya Tanzania

3. IPMCRSP - International Plant Virus Disease Network (IPVDN) Objectives: • Define virus disease problems in key crops through surveys and virus diagnosis and detection • Build in-country capacity for research on virus detection, ecology, and epidemiology • Develop and help deliver IPM packages for virus management

4. Virus Diagnosis and Detection are needed for Disease Management • Diagnose virus and identify vector and sources • With in-country capacity building and training • Involve plant pathologists and entomologists • Predict potential management practices • Design experiments to assess success of interventions • Provide models and approaches to be used by IPM scientists and practitioners • Technology transfer to introduce ecologically-based management system practices • Most developing countries lack this capacity !

5. Why do virus diagnosis? • Curious to know the cause of a symptom • One or more viruses? Phytoplasma? Fungi? Bacteria? Insect? • Growers want an answer – What can I spray? • To know what virus or viruses are present to develop a management practice – virus IPM is long term plan • Source? Where did the virus come from? • Can the source be reduced? Weeds? Seeds? • How is the virus transmitted? • Biological – specific vectors transmit certain taxonomic groups of viruses • Mechanical – by human contact during transplant and harvest is important for certain viruses • Seed – Only common for certain taxonomic groups of viruses

6. Why do virus detection? • Monitoreffectiveness of in-field management • Vector control with pesticides • Time of planting, barrier crops to avoid vectors • Host-free periods to reduce virus in vector and weeds • Help breeders develop virus resistant varieties • Provide virus-tested planting material • Citrus tristeza, banana viruses, potato, other vegetatively propagated plants • Seeds free of seed-transmitted viruses • Test for phytosanitary regulations for export • High sensitivity needed

7. Training and Capacity Building • Workshops in host countries • Honduras workshop on virus management • As a part of IPDN workshops • Separate virology workshops • Short term visits by trainees to US laboratories • 1-2 weeks to 6 months • Enhanced capacity • Graduate degree training • Full-time; sandwich programs • Needed for full capacity

8. Comayagua, Honduras workshop for field extension workers from various projects • PURPOSE: To transfer to field extension workers practice-oriented (and some theoretical), up-to-date information on management of virus diseases, focused on their nature/biology, dissemination, transmission, epidemiological aspects, e.g., influence of weather, genotypic differences, cultural practices, and other factors. At the end, participants should be able to understand the rationale of and knowingly apply the recommended management measures to combat viruses. • 1. BASIC PRINCIPLES OF PLANT VIROLOGY. Their importance, nature, and characteristics of the viruses and virus diseases of major relevance to their management. • 2. MANAGEMENT OF VIRUS DISEASES IN CROPS.Strategies and management practices used in the open field and protected environments before, during and after the cultivation cycle.

9. Training Workshops - Lectures • The nature of viruses – what are they! • Types of symptoms caused by viruses • Virus ecology – plants infected, ecological niches in ecosystems • Virus epidemiology – how do viruses get to crop plants • Via vectors? From seed? Other ways? • Virus diagnosis and detection methods • Apply management and monitor results

10. Diagnostic Workshops (IPDN) • Hands-on methods • Immunostrips • Indirect ELISA • Tissue blot immunoassay • Inclusion bodies • PCR for DNA viruses • FTA Cards Univ del Valle de Guatemala – 1 day of Virus Training Optimizing membrane-based technologies for virus identification RT-PCR + sequencing TBIA

11. Virus Diagnosis in Workshops • Immunoassays – reaction of virus-specific antibodies with virus particles • ELISA – DAS, PTA – microtiter plates • Tissue print or blot ELISA on membranes – TBIA • Lateral flow devises on membranes - Immunostrips • Molecular assays – detection of specific nucleic acid sequences of viral genome • Amplification by polymerase chain reaction (PCR) • Nucleic acid recovery methods; Primer Design • Reverse Transcription, Real-time • New sequencing methods and analysis

12. Facts about Virus Diagnosis • Virus can rarely be diagnosed by symptoms • Strains, host and variety, environment, time of infection all cause variations in symptoms • Plants may have multiple viruses • Diagnostic tests are available for only certain known viruses or virus families • There are no rapid diagnostic tests for unknown viruses • New viruses are emerging rapidly • Or, viruses have just now been recognized and characterized

13. Choosing a diagnostic/detection test?Information, Needs, and Capacity • What viruses are known in the crop? • What tests are available for those viruses? • What specificity and sensitivity are needed? • Quarantine and clean planting stock - high • Monitoring for IPM and disease management - lower • What tests can you conduct? • Immunoassays? PCR? Biological? Physical? • What tests can you afford to do?

14. Eggplant Pepper Tomato • Other vectors • Nematode, fungi, mite, grasshopper, beetle Mechanical, contact Major insect vectors Aphid Whitefly Thrips Potyvirus Cucumovirus Begomovirus Tospovirus Potexvirus Tobamovirus Tombusvirus Deformation of plant parts Necrosis Chlorosis Stunting / Distortion Mottle / Mosaic Crop, Dissemination, Vector, Virus genus, Symptom

15. India – Tamil Nadu Agricultural University • First virus workshop July 12-16,2010 • Week-long program, 1.5 day hands-on exercises in virus detection/diagnosis, 1 day field trip. • Tolin and Naidu gave most lectures; some by Indian and Indonesian scientists • All participants spoke at beginning and end of the workshop to introduce their own work, and comment on what they learned.

16. Tamil Nadu Agricultural University, Coimbatore, India Workshops for capacity building in plant virus diseasesIPVDN Global ThemeJuly 2010 Participants: South Asia – India Southeast Asia – Cambodia, Indonesia Central Asia – Uzbekistan

17. Asia Virus Workshop at TNAU:Detection and Diagnosis: Hands-on Activities Biological Detection Symptom Observation Mechanical Transmission See results 4 days later

18. Asia Virus Workshop at TNAU:Detection and Diagnosis: Hands-on Activities Diagnostic Immunoassays Classical ELISA ELISA on Membranes

19. Plant Disease and Insect Pest Diagnostics Workshop – Bogar, Indonesia July 22-23, 2010 • First day, lectures on use of serology and PCR in disease diagnostics, and lecture and lab sessions on diagnosis of virus • Conducted by Drs. Tri Damiyanti and Sri Hendrastuti Hidajat, who had attended the TNAU Workshop the week before • They demonstrated Tissue Blot Immunoassay with nitrocellulose membranes and reagents used by Tolin at TNAU - and first applied in the IPM CRSP in Jamaica in Caribbean Site, Phase II, before the Virus Global Themes

20. Central Asia • IPDN Workshop held June 6-11, 2011 at Tajikistan National University in Tajikistan (Naidu) • A strong need for increased effort in capacity building in diagnosis and management of virus diseases • Candidate from Tajikistan will be sent for short term training at Naidu’s laboratory in potato virus detection and seed certification

21. Planned Activities • IPDN/PVD workshop in May 2012 in Tanzania to train using SOPs for diagnosis – includes 3-4 viruses • Workshop in India – TNAU on Virology • July 13-16, 2012 • Degree training • Washington State University • Univ California – Davis • Honduran MS student to begin - Arizona