A multiplex nested-PCR assay for sensitive and simultaneous detection and direct identification of phytoplasma in the Elm yellows group and Stolbur group and its use in survey of grapevine yellows in France

Flavescence dorée and Bois noir (or Vergilbungskrankheit), are two main yellows diseases of grapevines in Europe. The two diseases cannot be distinguished on the basis of symptoms but they are associated with two different phytoplasmas which belong to the Elm yellows (16SrV) group and Stolbur (16SrXII) group, respectively. Their spreading areas are overlapping in France, Italy and Spain but they have different vector insects. Flavescence dorée is an epidemic disease and a quarantine organism. National surveys conducted annually in France require straightforward and sensitive assays to detect phytoplasma that sometimes occur in grapevine with a low titre and to characterize them readily. A bi-specific multiplex nested-PCR procedure was developed, to amplify simultaneously two non-ribosomal DNA fragments, 1150 bp and 720 bp in length, specific for Elm yellows-group and Stolbur-group phytoplasmas, respectively. They were identified using agarose gel electrophoresis of amplification products. The procedure is quick, sensitive and reliable. It was used on 2,525 grapevine samples from the field, in the frame of the French survey in 2002. Mixed samples containing both phytoplasmas displayed a mixed profile in the gel. It was confirmed that the nested-PCR amplimer obtained in the FD9 DNA region with Elm yellows-group phytoplasmas, though shorter than the initial FD9 fragment, nevertheless contained the restriction sites that permit the RFLP identification of geographic phytoplasma isolates already characterized in former studies.

Triplex real-time PCR assay for sensitive and simultaneous detection of grapevine phytoplasmas of the 16SrV and 16SrXII-A groups with an endogenous analytical control

Flavescence dorée (FD) and Bois noir (BN) are the two main yellows of grapevine in Europe and are caused by phytoplasmas of the 16SrV and 16SrXII-A groups respectively. A new triplex real-time PCR assay was developed in order to detect simultaneously the FD and BN phytoplasmas as well as grapevine chloroplastic DNA with TaqMan minor groove binder probes. Each set of designed primers and probes specifically detected the map gene of the FD and BN phytoplasmas, respectively and did not detect phytoplasmas from other phylogenetic groups. PCR efficiencies varied from 90 to 110 %. The PCR assay showed good intra-test and inter-test reproducibility. Triplex real-time PCR was compared to the conventional biplex nested-PCR method. The sensitivity of the real-time PCR, tested on several infected periwinkle and grapevine samples, was up to 5 and 100 times higher for the BN-P and the FD-P targets, respectively. Out of 109 grapevine samples analysed 10, which were negative with the nested PCR, turned to be FD-P positive with the real-time PCR. A decision scheme was set up according to the Ct values of the FD-P, BN-P and grapevine targets in order to assess the routine detection results

Viroses du houblon en Alsace

Trois virus ont été décelés par la méthode sérologique ELISA sur houblon en Alsace : le « Hop Mosaic Virus» (HMV), le « Prunus Necrotic Ringspot Virus » (PNRSV), l’« Arabis Mosaic Virus » (AMV) dans respectivement 44, 13,8 et 12,2 p. 100 des cas. Les plantes infectées n’extériorisent pas de symptômes foliaires caractéristiques. L’incidence de ces virus sur les rendements en cônes séchés et en acide oc est mise en évidence dans les houblonnières fortement contaminées.Three damaging viruses were detected in hops in Alsace by the ELISA technique. Hop Mosaic Virus (HMV), prunus Necrotic Ringspot Virus (PNRSV) and Arabis Mosaic Virus (AMV) infected 44, 13.8 and 12.2 % of plants respectively. Infected plants did not show typical leaf symptoms. A reduction in the yield of dried cones and -acid was obvious in highly contaminated hop gardens

Evolution de la flore adventice du vignoble de Cote-d'Or sous la pression des techniques d'entretien des cultures

National audienceThe present situation of the weed flora in Côte-d’Or vineyards is reviewed in relation to weeding methods. Comparison of this situation with that existing 10 years ago shows that the proportion of the different species has been affected by weed control methods. In the absence of chemical weeding, the weed flora has not changed since 1969. However, the introduction of weed killers after this date has allowed colonization by certain ruderal or forest species like Cornus sanguinea, Epilobium tetragonum, Hedera helix, Rubia peregrina... and has facilited the spread of all the perennial species like Allium spp., Muscari spp., Rubus spp., as well as species resistant to herbicides like Senecio vulgarisLes auteurs présentent la situation actuelle de la flore adventice du vignoble de Côte-d’Or en fonction du mode de désherbage. La comparaison de cette situation avec celle qui existait 10 ans plus tôt montre que l’importance relative des différentes espèces a évolué en fonction des techniques d’entretien. En l’absence de désherbage chimique, comme c’était le cas en 1969, la flore adventice n’a pratiquement pas varié. En revanche, l’introduction du désherbage chimique depuis cette date a permis l’implantation d’espèces rudérales ou forestières comme Cornus sanguinea, Epilobium tetragonum, Hedera helix, Rubia peregrina... et facilité l’extension de l’ensemble des espèces pérennes comme Allium spp., Muscari spp., Rubus spp. ainsi que de quelques espèces aujourd’hui résistantes aux herbicides comme Senecio vulgari

A multiplex nested-PCR assay for sensitive and simultaneous detection and direct identification of phytoplasma in the Elm yellows group and Stolbur group and its use in survey of grapevine yellows in France

International audienc

Establishment of a relationship between grapevine leafroll closteroviruses 1 and 3 by use of monoclonal antibodies

International audienc

Epidemiological survey of the grapevine leafroll disease in french wine growing regions

International audienc

Enquête d'épidémiologie sur la maladie de l'enroulement de la vigne en France

National audienc

Triplex real-time PCR assay for sensitive and simultaneous detection of grapevine phytoplasmas of the 16Srv and 16SrXII-A groups with an endogenous analytical control

International audienc