Pest Diagnostics in Phytosanitary Systems

1. To create awareness on pest diagnostics and its application in Phytosanitary systems
2. To enhance participants knowledge and skills on pest diagnostics
3. To enhance knowledge and skills on the application of pest diagnostic protocols
4. To create awareness and enhance knowledge on principles of pest management

1. Principles in pest diagnosis
2. Approaches to pest diagnosis
3. Diagnostic protocols in phytosanitary systems
4. Detection and identification of specific groups of pests
5. Reference collection and documentation
6. Principles of pest management

1.1.1 Principles in pest diagnosis (1 hr)

1. Definitions, importance and application of diagnosis,
2. Concepts of regulated and non-regulated quarantine pests;
3. Laboratory techniques applied at quarantine stations;
4. Factors to consider in diagnosis: cropping history, sampling including sampling for soil-borne pathogens, transportation, sample handling, storage and disposal. Refer to ISPM 5, 27

1.1.2 Approaches to pest diagnosis (2Hours)

1. Visual examination: symptoms, signs and damage;
2. morphological and cultural characteristics:: microscopy, isolation and purification,
3. biochemical and physiological: biochemical tests, fatty analysis, immunoassay/serological methods, molecular based -PCR based methods, indicator plants and indexing, pathogenicity
4. Modern techniques: digital and remote based diagnosis, rapid diagnostic kits,.

1.1.3 Diagnostic protocols in phytosanitary systems (3 Hours)

1. Importance of protocols, general requirements for diagnostic protocols
2. Format of diagnostic protocols: pest information - (identity to species level, hosts range, biology of the pest life cycle, distribution, transmission and dissemination pathways; taxonomic information; pest detection and identification methods – signs and symptoms, type of damage, methods of detection, description of morphology; comparison with similar or related species, use of reference specimens
3. use of diagnostic protocol, publication of diagnostic protocols. Refer to ISPM 27

1.1.4 Detection and identification of specific groups of pests (4 Hours)

Pest specific identification procedures including visual examination (symptoms, signs and damage), rapid diagnostic kits, indicator plants (virus/bacteria detection), pest sampling, barcoding, confirmation of results for new pests or unconfirmed cases.

Arthropods (Insecta, Arachnida)

1. Sampling: trapping, collection, preservation.
2. identification: morphology, microscopy, X-ray, identification keys, pictures, reference specimens, molecular, identification keys, software.

Molluscs

1. Sampling, collection and preservation
2. visual examination (morphology), molecular techniques, pictures.

Nematodes

1. Sampling, extraction and preparation of specimens,
2. selection of identification tool: software, manuals, identification by morphology (microscopy), molecular methods and bioassay.

Fungi

1. Sampling and sample collection, sample packaging and storage,
2. Use of symptoms and signs,
3. Isolation and purification: extraction, baiting, isolation, culturing and purification, use of selective media,
4. Identification: morphology (staining, microscopy), pathogenicity tests, molecular methods, chemotaxonomy (secondary metabolites), rapid diagnostic kits, immunoassay.

Bacteria and phytoplasmas

1. Sampling and sample collection, packaging and storage,
2. Use of symptoms and signs: bacterial streaming, bacterial ooze,
3. isolation and purification: selective media, cultural characteristics (immunoflorescence, pigmentation and colouration)
4. identification: staining and microscopy, pathogenicity tests, molecular methods, biochemical tests, chemotaxonomy, rapid diagnostic kits, immunoassay, fatty acids.

Viruses

1. Sampling, packaging and specimen storage, symptoms,
2. Extraction and purification,
3. Identification: characterization, morphology (electron microscopy), use of indicator plants and grafting, immunoassay methods, molecular methods, rapid diagnostic kits.

Weeds

1. Sampling and sample collection, preservation and preparation of herbarium
2. Identification: visual, identification keys, molecular techniques,.

1.1.5 Reference collection and documentation (2 Hours)

1. Definitions, types of reference collections,
2. Preservation of specific groups of pests: preservation of bacteria, fungi, viruses, nematodes, insects, insect boxes, herbarium
3. Creation and management of reference collections:site, staff, collection updates,
4. Reference laboratories, reference samples, herbaria and live collections
5. Documentation: diagnostic reports, list of pest and pest list and pest databases.

1.1.6 Principles of pest management (3 Hours)

1. Pest management methods: scouting, monitoring, identification, good agricultural practices, pheromone and semio-chemical methods, chemical, biological, physical, mechanical, host plant resistance, GMO, bio-pesticides and IPM
2. Field and green house sanitation, greenhouse growth media preparation, water source management
3. Postharvest management: precooling, sorting, grading;, postharvest treatment - fumigation, irradiation, waxing, hot water treatment, heat treatment

1.2 Practical exercises (16 Hours)

Arthropod (insecta and arachnida)

Visual observation (display materials showing different pest damage), trapping/collection, preservation and morphological identification.

Nematodes

Visual observation (display materials showing nematode damage), extraction, identification.

Fungi

Visual examination (display of material showing fungi damage), demonstration of packaging and storage, selective media, isolation, morphology (staining, microscopy), rapid diagnostic kits.

Bacteria and phytoplasma

Visual examination (display of material showing bacterial damage), bacterial streaming, bacterial oozes, isolation, selective media, cultural characteristics (pigmentation and colouration), pathogenicity tests, molecular, biochemical, rapid diagnostic kits, immunoassay.

Viruses

Visual examination (display of material showing viral damage), indicator plants and grafting, immunoassay, molecular, rapid diagnostic kits.

Weeds

Visual examination (display weeds), weed seeds especially in grains and legumes, identification and herbarium.

1.3 Case studies (3 Hours)

1. Emerging and new quarantine pests important to participating countries:
2. Virus such as those causing Maize lethal necrotic disease (MLND), Banana bunchytop virus
3. Bacteria such as ralstonia, Banana xanthomonas wilt and Dickeya
4. Weeds such as Oxalis latifolia, striga,
5. Nematodes such Potato cyst nematode,
6. Arthropods such as Fall armyworm, False codling moth, fruitflies, khapra beetle
7. Fungi such as fusarium oxysporum tropical race 4 (TR4)
8. CABI Plantwise initiative on identification of pests and plant health clinics; and other relevant case studies.

1. Crop Protection Compendium. http://www.cabi.org/cpc/
2. ISPM 04: 1995 Requirements for the establishment of pest free areas
3. ISPM 06: 1997 Guidelines for surveillance
4. ISPM 08: 2017 Determination of pest status in an area
5. ISPM 16: 2017 Regulated non-quarantine pests: Concept and application
6. ISPM 17: 2017 Pest reporting
7. ISPM 19: 2016 Guidelines on lists of regulated pests
8. ISPM 26: 2015 Establishment of pest free areas for fruit flies (Tephritidae)
9. ISPM 27: 2016 Diagnostic protocols for regulated pests
10. ISPM 22: 2016 Requirements for the establishment of areas of low pest prevalence
11. ISPM 29: 2017 Recognition of pest free areas and areas of low pest prevalence
12. ISPM 30: 2017 Establishment of areas of low pest prevalence for fruit flies (Tephritidae)
13. ArcGIS information software, version 10: www.esri.com
14. Open data kit (ODK): https://opendatakit.org/
15. Quantum GIS (QGIS): https://www.qgis.org/en/site/
16. Trapping guidelines for areawide fruitfly programmes: by International Atomic Energy Agency of UN: https://www.iaea.org/publications/6916/trapping-guidelines-for-area-wide-fruit-fly-programmes