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Current impact and future direction of High Throughput Sequencing in plant virus diagnostics

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Current impact and future direction of High

Throughput Sequencing in plant virus diagnostics

Sébastien Massart

Laboratoire de Phytopathologie – Gembloux Agro-Bio Tech Université de Liège – Belgique

sebastien.massart@ulg.ac.be

,Antonio Olmos, Haissam Jijakli and Thierry Candresse

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NGS & Viruses

Etiology Viral ecology Plant-virus interaction Plant resistance Viral population genetics Taxonomy

Current impact

on R&D

Expected impact on diagnostics

Diagnostic & Etiology

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LES VIRUS

More than 50 publications with new virus identification

Diagnostic & Etiology

Diagnostic & Etiology

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Diagnostic & Etiology

New virus

Host range Transmission Plant-virus interaction Plant resistance Viral population genetics Symptomatology

?

?

?

?

?

?

? Threat to the plants & control measures ?

Certification, sanitation and quarantine

Field diagnostic and eradication

Diagnostic & NGS

CLEAN UPSTREAM IMPROVE DOWNSTREAM

Improved and standardized NGS methodologies for indexing

Deluge of sequences to accelerate and improve rapid-testing

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Cost evolution

Technical Challenges

1. Preparation & sequencing strategies ?

Extraction protocol

DNA, total RNA, particle isolation, small RNA, dsRNA

2. Sensitivity ?

Dependent on sequencing depth Similar to PCR

What about a single or a couple of viral reads ?

3. Reproducibility and repeatability ?

Intra and inter-laboratory comparison

Cost evolution

Technical Challenges

4. Contaminations ?

Human & beef contigs -> what about viruses ? Partial or total genome coverage

Strict confinement needed !

5. Adapatation to host species ?

Virus titre and extraction method

Practical and technical challenges requiring

both fundamental and applied research

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Practical impact

Identification of a new virus or a poorly characterized virus by a diagnostician ?

Halt the distribution of the plant material ? Confirmatory assays (Molecular Koch postulate) ? Long-term experiment on virus biology ?

Technical framework and decision schemes are needed

Latent (or symbiotic )viruses discovered ?

Probability of discovering non pathogenic viruses is high Re-think trophic relationships ?

Adaptation of regulatory framework ?

NGS & Viruses

Etiology Viral ecology Plant-virus interaction Plant resistance Viral population genetics Taxonomy

Current impact

on R&D

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COST Action FA1407

Coordinate and raise the European capacity to apply NGS technologies for the study and diagnosis of viral

diseases in vegetatively propagated

plants, seeds and seedlings.

Fundamental virologists Industries Applied virologists Diagnostician Regulatory authorities NPPO/EPPO Technological improvement

New scientific Knowledge

Societal innovations Societal innovations

1.

2.

3.

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Large impact in R&D

Impact ongoing in diagnostic

Scientific & regulatory consequences

Feedback loop to fundamental research

Long term :

Safer movement and trade

Better understanding of the virus threat and

biology (including the Pathovirome)

Conclusion

Merci – Thanks

For more information:

Massart et al., 2014. Virus Research, 188: 90-96

Massart et al., 2014 Virologie, 18(5):247-250

Références

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