7th International IPM Symposium

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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_at_vt.edu

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

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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 !

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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

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Why do virus detection?

• Monitor effectiveness 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

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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

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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.

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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

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Diagnostic Workshops (IPDN)
Univ del Valle de Guatemala 1 day of Virus
Training

• Hands-on methods
• Immunostrips
• Indirect ELISA
• Tissue blot immunoassay

• Inclusion bodies
• PCR for DNA viruses
• FTA Cards

Optimizing membrane-based technologies for virus
identification
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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

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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

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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?

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Crop, Dissemination, Vector, Virus genus, Symptom
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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.

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Workshops for capacity building in plant virus
diseasesIPVDN Global ThemeJuly 2010
Participants South Asia India Southeast Asia
Cambodia, Indonesia Central Asia Uzbekistan
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Asia Virus Workshop at TNAUDetection and
Diagnosis Hands-on Activities
Biological Detection
Symptom Observation
Mechanical Transmission
See results 4 days later
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Asia Virus Workshop at TNAUDetection and
Diagnosis Hands-on Activities
Diagnostic Immunoassays
Classical ELISA
ELISA on Membranes
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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

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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 Naidus laboratory in potato
  virus detection and seed certification

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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