Detection of Flavescence dorée and Bois noir phytoplasmas, Grapevine leafroll associated virus‐1 and ‐3 and Grapevine virus A from the same crude extract by reverse transcription‐RealTime Taqman assays

摘要: Multiple detection of the phytoplasmas associated with Flavescence doree (FD) and Bois noir (BN) diseases viruses Grapevine leafroll virus -1 -3 (Ampelovirus) A (Vitivirus) is described, using same crude extract as template. Sap was prepared by semi-automatic maceration requiring minimal time effort, consisting a tissue grinding step in carbonate buffer boiling glycine buffer; two microlitres were used template each pathogen-specific assay. RealTime reverse transcription (RT)-PCR for FD phytoplasma found to be five orders magnitude more sensitive than RT-PCR method described previously. However, assay BN needed nested achieve high sensitivity, suggesting low concentration host. The detected nested-PCR, which ELISA assays previously described. methods presented here have been successfully monitor infections field nursery samples during 2008 grapevine growing season.