Diagnostics, Phytosanitation, Phytosanitary Regulations ...gcp21.org/Tanzania/LavaKumar.pdf ·...
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Diagnostics PhytosanitationDiagnostics, Phytosanitation, Phytosanitary Regulations and Germplasm Transfer
An overview
Lava KumarIITA, PMB 5320, Ibadan, Nigeria
L k @ [email protected]
Some terms
Pest = Pathogens, insects, mites, nematodes, etc.Pest Pathogens, insects, mites, nematodes, etc.Any species, strain, biotype injurious to plants and plant products
Virus negative: plants tested negative to a particular virusCBSD free: Free of all virus species presently known to cause CBSDCBSD‐free: Free of all virus species presently known to cause CBSDVirus clean or clean germplasm: Seldom used in official documents
Diagnosis: Identification of the cause of the problemDiagnosis: Identification of the cause of the problem
Diagnostic tools: To ascertain presence, absence or quantify pests
Germplasm(Gene Bank / Breeders)
Collection missions Distribution missions
C ti / d ti Utilization/marketingConservation/production Utilization/marketing
Indexing for Germplasm HealthGlobalization of Pathogens
Germplasm exchange: risks
•Movement of germplasm involves a risk of accidentally introducing quarantine pests.
•Germplasm sometimes are contaminated ith h f l t
quarantine pests.
with harmful pests.
•Pathogens harmful to the same species (e.g. cassava to cassava)
•Can also carry pathogens that do not harm the carrier, but harmful to other species
(e.g. cassava carrying spores of maize pathogen)(spores, insect eggs, etc)p gg
Clean germplasm is prerequisite for safety of conserved germplasm andClean germplasm is prerequisite for safety of conserved germplasm and for the safe distribution of germplasm
I i f i di USInterception of non‐indigenous pests at US ports
• 725,000 pest (non‐indigenous insects, mites, molluscs, nematodes, plant pathogens and weeds) interceptions recorded in US Port‐interception network from 1984 to 2000.
• 62% of intercepted pests were associated with baggage, p p gg g
• 30% were associated with cargo and
• 7% were associated with plant propagative material• 7% were associated with plant propagative material.
• 50,750 in 17 years (ca. 3,000 interceptions per year)
Source: McCullough et al., 2006, Biological invasions 8: 611‐630.
•Measures to remove pests infested/infected plants •Elimination of pests from infected/infested tissues (therapy)
Phytosanitatoin
Elimination of pests from infected/infested tissues (therapy)
Phytosanitaory Germplasm y yRegulations
pTransfer
•Policy framework and exing
international standards for risk assessment, risk reduction, phytosanitation,
Inde
Diagnostics, p y ,
safe exchange of germplasm and otherwww iipc int •Tools for detection and identification
of pests in the germplasm
www.iipc.int
International regulations and technical guidelinestechnical guidelines
International Standards for Phytosanitary Measures (ISPMs) of FAO IPPCInternational Standards for Phytosanitary Measures (ISPMs) of FAO‐IPPC
ISPM 1 to 36www.ippc.int
Challenges for cassava conservation
•Pest matters, irrespective of disease causing potential
Pests
Endophytes
Regulated Not regulated
EndophytesCommon pests of insignificant consequence
Quarantine pestsRegulated non-
quarantine pests
consequence
quarantine pests
Endemic pests, know to cause debilitating impact on host: eg.
Endemic pests, know to cause debilitating impact on host (eg.
b k) Cassava mosaic virusescassava brown streak)
NPPO provides the list of quarantine pests and non‐qurantine pests
Major cassava pests in Africa( l d i )(regulated non‐quarantine pests)
Indigenous pestsIndigenous pests• Cassava mosaic viruses• Whiteflies, fungal diseases, root scales etc
Pests of regional importance• Cassava brown streak virus
, g ,
• Cassava brown streak virus• East African cassava mosaic virus – Uganda (EACMV‐UG)
Exotic pests• Green spider mite (Mononychellus tanajoa)
C l b (Ph ih ti)• Cassava mealy bug, (Phenacoccus manihoti) • Cassava Bacterial Blight (Xanthomonas axonopodis)• Indian cassava mosaic virusIndian cassava mosaic virus
Quarantine pests of cassava in Africa
Pests not present in the continentPests not present in the continent1. Cassava common mosaic virus (??)2 Cassava green mosaic virus2. Cassava green mosaic virus 3. Cassava vein virus 4 Cassava X virus4. Cassava X virus5. Frog Skin Disease (complex etiology)6 Cassava antholysis (phytoplasma like)6. Cassava antholysis (phytoplasma‐like)7. Cassava witches broom (Phytoplasma)8 Super Elongation (Sphaceloma manihoticola)8. Super Elongation (Sphaceloma manihoticola)
Risk of pest spread through planting materials
Pest Stems Sterileti lt
seed Steriletifi d titissue culture certified tissue
culture plants*Bacteria + ‐ ‐ ‐Fungi + ‐ ‐ ‐Virus + + ‐ (?) ‐Insects & other + ‐ ‐ ‐
Stems: carriers of all pests
Ti lt l t O l i ( d h ti b t i )Tissue culture plants: Only viruses (some endophytic bacteria)
Virus elimination requires combination of tissue culture and therapy q pytechniques cryotherapy, etc) and selection of plants following virus indexing.
The viruses of cassava in Africa
African cassava mosaic virus (ACMV)Indian cassava mosaic virus
CMGCMGEast African cassava mosaic virus (EACMV)South African cassava mosaic virus (SACMV) East African cassava mosaic Cameroon virus
CMGsCMGs
East African cassava mosaic Cameroon virus East African cassava mosaic Malawi virusEast African cassava mosaic Zanzibar virus East African cassava mosaic Kenya virusEast African cassava mosaic Kenya virusEACMV-Uganda (recombinant virus)Cassava mosaic Madagascar virus
Cassava brown streak virus (Ipomovirus)Cassava brown streak Uganda virus
Cassa a I o ian bacillifo m i s*
CBSVCBSV
Cassava Ivorian bacilliform virus* Cassava Kumi virus*Cassava ‘Q’ virus*Cassava common mosaic virus* (Potexvirus)Cassava common mosaic virus (Potexvirus)
Challenges to cassava conservation
•Cassava is seriously affected by Wh t i t accumulation of pathogens in the
propagation material. What goes in stays
• Gene banksGene banks conserves cassava
• Cassava conserves•High risk of pathogen spread through stems
Cassava conserves pests, particularly viruses
•Constraint to field gene banks High risk of infection & re‐infection; cause germplasm decline and genetic erosion
•Restrictions on exchanging stems
•International exchanges require in vitro plants
Challenges to cassava conservation:Vi h i iVirus threat to micropropagation
Tissue culture cleanses
•In vitro conservation is emerging as mainstay for conserving clonal crops.
Tissue culture cleanses pests and pathogens, but not viruses.
•Effectiveness of in vitro therapy procedures depends on the genotype, host p g yp ,susceptibility, infection status, virus strains, type of therapy procedure and practice
•Procedures capable of eradicating cassava viruses are time consuming.
•Requires expensive capacity and facilities, and round the clock access to water andand round the clock access to water and power.
Challenges to cassava conservation:Vi di iVirus diagnostics
•ELISA, and PCR‐based tests (conventional and RT‐PCR) are used for the detection of cassava viruses. R tl iRNA b d th d
High diversity is a bane to accurate diagnosis of
Recently even siRNA based methods are used for diagnosis.
accurate diagnosis of viruses •High diversity poses challenges to
the virus detection.
•Generic (broad‐specific) diagnostics b tt f i d t ti fare better for virus detection for
quarantine monitoring, and virus indexing.indexing.
Pest detection: requirements
•Symptomatic stage: Confirmation of pathogen cs
•Asymptomatic stage: Confirmation of infection & pathogen
gnostic
•Undetectable stage Germplasm (dormant)
Diag
Low‐titers of pathogens
Knowledge on pathogens and their distribution
Availability of diagnostic tools
Fund
s
Capacity (human skills and infrastructure)
Choice of methods depends on the need
Identify known pest( i i i d i
Identify new pest ( f(repetitive; e.g. indexing,
surveillance)(one time; e.g. cause of
new disease)
Specific test(target pest of interest)
Non‐specific (generic) test(no prior knowledge)
ELISA, PCR, NASH( p g )Morphometrics, bioassay, EM,
ELISA, PCR, NASH, sRNAs
•PCR‐based assays are commonly used for cassava indexing
Source: A Adkins
•Small RNA‐based approach is gaining popularity as a useful broad‐specific tools
Guidelines for use of diagnostics
International Plant Protection Convention (IPPC) WTO‐SPS guidelinesWTO SPS guidelines
Regional and Country‐specific guidelines
Product quality For monitoring infectious diseasescertification
Pl ti t i l
For monitoring infectious diseases
Pests ‐Planting material
‐Product quality
Pests
Regulated Not regulated Product quality(inputs as well as plant products)
Quarantine pests Regulated non-quarantine pests
Plenty of options for cassava pest diagnostics
•Diagnostics is a very well established field, but not saturated
•Tremendous progress over last 3 years
•Low‐tech to very high‐tech tools
•Simple cost‐effective tools to most expensive tools
•Availability is not an issue application and practice matters•Availability is not an issue, application and practice matters.
Sustainability of diagnostics
Diagnostic labs 2. Establishment‐Infrastructure ‐Equipment‐Supplies and reagentsMaintenance
1. ScopeD fi h f h l b ‐Maintenance
‐Human skills‐Water and power
Define the scope of the lab•Pests, pathogens, pesticides, food‐borne toxins etc
3 Sustainability
Water and power‐Maintenance strategy
food‐borne toxins, etc.•Specific or generic lab•Indexing & certification, 3. Sustainability
‐Regular use‐Funding supportFunding support
Cassava Phytosanitation
Total plants tested
Number that tested negative to CBSV
Varietytested to CBSV
1 MM06/0011 20TC 15
2 MM06/0024 20TC 10
3 MM06/0138 20TC 3
4 MM06/0131 20TC 3
5 MM06/0019 20TC 9
6 MM06/0079 20TC 116 MM06/0079 20TC 11
7 MM06/0013 20TC 1
8 MM06/0045 20TC 20
9 MM06/0012 20TC 20
10 MM06/0023B
20TC
8
11 MM06/0139
20TC
2
12 MM06/0124
20TC
4
13 MM06/0112
20TC
3
14 MM06/0076
20TC
6
IPCC guidelines for cassava germplasm exchangeg g p g
V t ti (i it ) t i lVegetative (in vitro) material: • Import permit and phytosanitary certificate
• Declaration certifying the absence of virus, bacteria, and nematodes (particular those quarantine/regulated pests (based on country pest lists)
• Compulsory treatment of consignment before shipment into Africa.
• Exchange only virus tested virus‐free materials
• Cassava seed: Seed sanitation and in vitro plants derived from meristems of t d l t
• Exchange only virus tested, virus‐free materials.
regenerated plants
Country specific guidelines can differ from IPPC and IAPSC guidelines
IITA Germplasm Health Unit
•Adjunct to NPPO•Prevent the spread of pests and pathogens through germplasm exchangePrevent the spread of pests and pathogens through germplasm exchange•Produce healthy planting material & safe conservation and exchange procedures•Ensure compliance with national and international regulations
Indexing for insect pests, fungi and bacteria: Visual inspection, blotter
test, and sedimentation assays (Optional seed treatment / fumigation)
Health Testing(For pathogens under
Release of pathogen free‐G l
seed treatment / fumigation)
Import / Export (For pathogens under containment facilities)
pathogen freematerial
Germplasm
Indexing for viruses: Grow‐out tests, evaluation for target
virus by ELISA, PCR or EM
Requirements for germplasm export
Material owner(GRC, Breeding Units, etc.)
*Requirements:-Accession details-Geographic origin of the material (site of nt
atio
n
Submit request to GHU with necessary details*
g p g (production) -Year of propagation/production-Previous indexing history (if any)-Import permit from the recipient-Any other instructions D
ocum
e
-Any other instructions
Indexing at GHUfor fungal and bacterial virus, as
Dh
per the standard procedures, as well as importing country
requirements.
disp
atch
Pest-free material selected for dispatch
catio
n &
NAQS certificate and GHU health statement on health status
Pack and dispatch
certi
fic
Pack and dispatch pest-free material
Procedure for importing
l ti t i l
Importer at IITA (GRC, breeders, etc.)
Routed throughGermplasm Health Unit
atio
n
planting material
Importer to sign MTA as applicable;
NAQS may put specific conditions depending on the crop species/ material and country of origin.
Submit request to NAQS for import permitPermit issued
ocum
ent
Consignment arrives in Nigeria
Exporter to comply with the importer’s phytosanitary needs. Communicate any difficulties to requester. A solution will be worked out to get the material.
Submit to exporter Do
e
Process germplasm (seed/ in vitro material / vegetative
propagules) as per the standard procedure
Material that meets importing country standards
Material that do not satisfy importing country standards
Mandator health testing rele
asestandard procedure
Certified consignments Uncertified consignments
y
Mandatory health testing •Material held in post-entry quarantine facility•Visual inspection•Biochemical analysis for quarantine pathogens
ectio
n &
Depends on the
Grow-out in post-entry quarantine isolation
Weekly inspections Inse
pe
Depends on the NAQS advise
Release ‘clean’ material to requisitioner
Depends on the NAQS advise
Distribution of imported material
Certified Clones that meets the importer NPPO guidelines are released
International distribution In‐country distribution
Follow established regulations.
Depends on NPPO regulations:
•This could include certification of clones by authorized labs/agencies present in‐country
Or Depends on NPPO regulations:•Material can be supplied to end‐users for further propagation and distribution.
or
•Distribute directly
•Plant in ‘quarantine plots’ for certification by local agencies and release to end‐users.
Germplasm exchange with a t ith l ticountry with regulations
Regulator:• Create awareness about the policy and standards• Create awareness about the policy and standards• Assess the producer constantans and find solutions (policy/technical)
Producer / client•Strictly comply with the regulatory guidelinesStrictly comply with the regulatory guidelines•Discuss any constrains with the regulators
Germplasm exchange with a t ith t l ticountry without regulations
Regulator:g•Follow the IPPC standards •Identify gaps and establish a feasible model
Producer/client• Adopt international standards• Adopt international standards • Work with regulators to establish guidelines
Other issues
• Phytosanitary permits are issued based on the NPPO analysis of risk priority end use etcof risk, priority, end use, etc.
• Permit to import prohibited (quarantine) plant material for research purposeresearch purpose
• Permit to import plant material for research purpose
• Permit to import plant material for processing and industrial use (not meant for propagation)
• Most stringent regulations apply for the material imported or exported for ‘propagation purpose’.
• Retrospective indexing
Risk of transferring material
• Within Africa, biggest risk is CBSD, which is already present in TanzaniaWithin Africa, biggest risk is CBSD, which is already present in Tanzania• Requires a perfect phytosanitary system for re‐distribution
Endnote
• Risk assessment, analysis and phytosanitary system should be kept inRisk assessment, analysis and phytosanitary system should be kept in place prior to commissioning collection missions for conservation of germplasm in a central facility (perform in consultation with NPPO)
• Create awareness about the advantages (benefits) of the process, facility and phytosanitary procedures in place, to the participatory y p y y p p , p p ycountries.
• Establish a virus indexing certification and documentation system• Establish a virus indexing, certification and documentation system (traceability), and obtain accreditation from TPPC.
• Establish a safe measures to harden virus‐free plants for exchange with countries with inadequate capacity to handle TC plants.