© 2007 Her Majesty the Queen in right of Canada

(Canadian Food Inspection Agency), all rights reserved. Use without permission is prohibited.

The Future of Post Entry Quarantine: Canadian National Clean Plant Program

Dr. Michael Rott, Canadian Food Inspection Agency (CFIA)

Sidney, BC, Canada.

Overview

• What is a National Clean Plant Program (NCPP)

• A model: US Clean National Clean Plant Network

• Canadian Imported material: what is required

• Time Line – illustration and explanations

• Current approaches – Testing technologies

• Advances in testing

• Model for the implementation of NCPP

• What we need to get there

• Benefits

2

What is a National Clean Plant Program

(NCPP)

Goal:

To promote the production and use of pathogen-free, healthy plant material for food crops in Canada

Framework:

A set of rules/regulations to govern the maintenance, testing, propagation and distribution of pest and disease free plant material from nuclear generation 1 material to generation 4 material grown for retail.

Enabler: A lab network and testing technology to support the Program

3

A good model: US National Clean Plant

Network (NCPN)

4

• 2005: Idea initiated by grape and tree fruit industries

• 2008: Grape and fruit tree networks developed by stakeholders,

industry members, scientists and other interested parties

• 2009: NCPN created as a voluntary association of specialty crop

networks to promote pathogen-tested, healthy plant material

United States Department of Agriculture –Animal and Plant

Health Inspection Service (APHIS)

Agricultural Research Service (ARS)

National Institute of Food and Agriculture (NIFA)

• 2010: berries, citrus and hops programs added

• 2016: US Farm Bill: invested XXXX$ in program

US NCPN: Grapes

New, state-of-the-art facility, Foundation Plant Service, Davis CA

• Regulates importation of grapevine material

• Provides tested, clean material, using latest equipment and

techniques

Manage Russell Ranch: foundation material for pathogen-tested

US grapevine material

5

http://ncpngrapes.org/

The Canadian Situation: Imported material

Currently, propagative tissues of tree fruit, grapevines and small fruit imported into Canada must be held in post entry quarantine to ensure they are free from disease and pests of concern.

This includes:

• Foreign material from non-certified sources

• Audit samples from approved certified sources

• Domestic material from plant breeding programs and nurseries

Time line: Import of new cultivar from a non-

certified source

7

6 – 12 years!!

8

Why so long?

Current Testing Methods

Molecular and Serological Tests: • Virus specific PCR and ELISA

1-3 days to complete

Bioassay Tests:• Herbaceous bioassay

Nonspecific, 2-3 weeks to complete

• Woody field and greenhouse bioassay*

Nonspecific, up to 3 years to complete

*International “Gold Standard”

Until now: Canadian Situation:

9

• Canada does not currently have a NCPP due to several challenges:

• Additional costs:

• to the public• to growers

• Reluctance to increase regulatory burden

• Requires a willingness from industry stakeholders and government

What is different now?

Technological Advances and Novel Applications

10

DNA sequencing: today a generally accepted a way to ID an organism

e.g. “DNA Barcoding”

1st generation sequencing technology:

- laborious, slow, $$$

human genome: 13yrs, $3B

Next Generation Sequencing (NGS)

- faster, cheaper, automatable

human genome: 1-2 days, <$5K

Metagenomics: use of NGS technology to ID multiple species in a single sample

detect multiple viral infections in plants!

Small samples produce big data!

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Tissue Types: bark/cambium, leaf

Tissue Amount: 1 - 5 g sample (individual/composite)

DNA extraction: manual/automated

Genomic “Libraries” – DNA that has been “Prepped”

for sequencing – all genetic material from the sample

Next Generation Sequencing: various systems

Reads 25-40 Gigabytes of data in <12h

Cost: $150/sample/test – includes above steps

Data analysis: Bioinformatics processes –

computational advances allow for

Computer facilitated analysis of data

i.e. easier to “find” detection targets!

Other Benefits of NGS:

• Samples processed in months vs years

• Less expensive: 1/10th the cost/sample.

• Or, 10x as many samples for the same cost.

• As sensitive or more sensitive than conventional

• More accurate

• Greater specificity

• Greater ability to detect new viruses

• Replace the bioassay?

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Example: 2013 grapevine audit imports

• Approximately 2.5 million vines imported

• Center for Plant Health in Sidney receives 1800 vines for testing (0.07%)

• Samples are:

• inoculated to 5 indicator plants

• rub inoculated onto 3 herbaceous indicators

• tested by PCR for 12 virus

• tested by ELISA for 5 virus

• > 5200 individual tests are performed on these samples over 3 years!!

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vines imported: 2.5 million

vines tested by CPH: 1800

Traditional tests: >5200 tests in 3 years

The future: using NGS, metagenomics and bioinformatics approaches:

90 -180 NGS tests

+ Confirmatory testing (PCR or ELISA)

IN 4 – 8 months

14

Example: 2013 grapevine audit imports

CFIA Programs that could be modernized

through a NCPP:

Export Certification Programs:

• Canadian Fruit Tree Export Program (CFTEP) for Malus,

Pyrus, Chaenomeles and Prunus (D-08-05)

• Plant Protection Export Certification Program for Grapevine

Nursery Stock, Vitis spp. (D-97-06)

Import Programs: Post Entry Quarantine

• To prevent introduction of plant viruses/virus-like diseases in

tree fruit, grapevines & small fruit.

Other activities:

• Viral elimination, maintenance of clean stock

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NCPP Vision:

Harmonize with Canadian National Import and Export Programs

• new test methods based on NGS

Harmonize with USA

• North American Plant Protection Organization (NAPPO)

• USA National Clean Plant Network (NCPN)

Harmonize Internationally

• International Plant Protection Convention (IPPC)

• EUPHRESCO Network for Phytosanitary research

coordination and funding

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www.cscr.in

Supporting Research: Current, Next, Future….

NGS validation & Generation 1 nuclear repository

• Re-test current grapevine and tree fruit export repository by NGS

• Validate NGS for plant viruses

• Develop NGS methods for grape fungal pathogens

NGS Tech transfer to 3rd party diagnostic lab

• Testing need for strawberry plant exports

Methods for Curating Gen 1 nuclear material

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www.biw.kuleuven.be

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Plant Propagation

• In vitro cultures

• micropropagation

• Micrografting

• Others

Viral Elimination: • Heat therapy

• Meristem culture

• Chemotherapy

• Cryotherapy

• Micrografting

• Others

Research: Current, Next, Future….

Imported material to

CPH.

Viruses testing

program

(4-8 months)

Negative

released to importer

(4 plants)

Positive

Virus elimination program

(1-2 years)

Virus free material released

to importer

(4 plants)

Micro-propagation

(1-1/2 years)

First crop

(2 years)

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6–12 years

4-6 years

Go from this:

…….to this:

Benefits to Canada establishing a NCPP

• Help maintain the highest standards for Canadian food production

while improving competitiveness of the Canadian Agro-Foods Sector

• Facilitate access to new material to Industry while reducing the

reliance on imported and potentially infected material

• Provide protection to production systems and to farms, nurseries,

and orchards while promoting trade

• Help prevent the introduction and spread of potentially harmful plant

pests and diseases in Canada

• An integrated consortium of laboratory expertise linking university,

AAFC, CFIA and the various industry sectors

Acknowledgements Funding

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Genomic R&D Initiative:

“Protection of Canadian biodiversity

and trade from the impacts of global

change through improved ability to

monitor invasive alien and

quarantine species”

Canadian Safety and Security

Programs:

CSSP-2014-TA- 2123

CSSP-2015-TA-2124

22© 2007 Her Majesty the Queen in right of Canada

(Canadian Food Inspection Agency), all rights reserved. Use without permission is prohibited.