plexor multiplex qpcr tool as a way to quantify esca...

Plexor multiplex qPCR tool as a way to quantify esca-associated

fungal agents in grapevine wood.

Plexor multiplex qPCR tool as a way to quantify esca-associated

fungal agents in grapevine wood.

Photo: Nancy Wenner, Penn State

Alban JACQUES, INP-EI PURPAN, Toulouse Lisbon, 2014, 20th February

ESCA Without symptoms Chronic symptoms Apoplectic symptoms

a.k.a. « tiger stripes »

5-10 yrs

Death occurs within days

Introduction: esca

Epidemiology of P.ch and P.al

Penetration Pruning wounds Roots

Sporulation All the year or only in vegetative period Perithecia located in crevices

Inoculum sources Air Soil Plant material grafted in nurseries Insects

Introduction: esca

Rolshausen et al. (2010); Whiting et al. (2006); Larignon et al. (1997)

Goals

Development of tools to understand plant microbes interactions in an early model of esca

Diagnosis development

Introduction

Goals

Development of tools to understand plant microbes interactions in an early model of esca

Introduction

Development of a molecular tool to detect and quantify esca-associated fungal agents

Introduction: available tools

In vitro plate screening PCR (qPCR ?)

Speed - +

False negative + (competition) +

False positive - +

Exhaustive + -

Identification complex Known before

Sensitivity -/+ +

Our developped tool

qPCR multiplex

Quantitative Multi-fungal Specific

Chemistry ? Primers ? Test ? Purposes we developped

Plexor chemistry

Plexor chemistry

Plexor chemistry

i.e. P.al

Plexor chemistry

i.e. P.al

Plexor chemistry

i.e. P.al

copies

Plexor chemistry

i.e. P.al

copies

Plexor chemistry

Primers

Targeted gene

ITS ? rDNA ? Encoding gene ?

b-tubulin

Primers

Targeted species

Species for specificty tests

Mostert et al (2006) Taxonomy and Pathology of Togninia (Diaporthales) and its Phaeoacremonium Anamorphs. Studies in mycology 56

Ex. Phaeoacremonium aleophilum

Phylogenetic studies

Primers

b-tubulin

Test

In lab infected plants Nursery samples

Test

In lab infected plants

Test

Nursery samples

TAMRA P.ch

ROX P.al

FAM VvActin

Test

Nursery samples

Test

Nursery samples

Test

Nursery samples

RBE are more contaminated than GU

More contamination by P.al than P.ch

Fungal pressure is higher in GU

In few words

Purposes

Conclusion

Plexor multiplex qPCR esca pathosystem Plexor multiplex qPCR BDA pathosystem (unpublished) Plexor qPCR Eutypa dieback In lab model to test early efficiency of putative solutions Tool to measure defence efficiency of grapevine

Conclusion

Team

R.Pierron J.Pouzoulet N.Mailhac C.Couderc F.Violleau M.Gaucher

Thanks for your attention!!!