Development of a qPCR assay for specific quantification of Botrytis cinerea on grapes

The aim of this study was to develop a system for rapid and accurate real‐time quantitative PCR (qPCR) identification and quantification of Botrytis cinerea, one of the major pathogens present on grapes. The intergenic spacer (IGS) region of the nuclear ribosomal DNA was used to specifically detect...

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Bibliographic Details
Published in:FEMS microbiology letters 2010-12, Vol.313 (1), p.81-87
Main Authors: Diguta, Camélia Filofteia, Rousseaux, Sandrine, Weidmann, Stéphanie, Bretin, Nicolas, Vincent, Béatrice, Guilloux‐Benatier, Michèle, Alexandre, Hervé
Format: Article
Language:English
Subjects:
Assaying
Biological and medical sciences
Botrytis
Botrytis - genetics
Botrytis - isolation & purification
Botrytis - physiology
Botrytis cinerea
Calibration
Deoxyribonucleic acid
DNA
DNA, Fungal - analysis
Food Handling - standards
Fundamental and applied biological sciences. Psychology
Fungal plant pathogens
Fungi
Grapes
Microbiology
Miscellaneous
Mycology
Organic Chemicals - chemistry
Phytopathology. Animal pests. Plant and forest protection
Polymerase Chain Reaction
qPCR
quantification
Ribosomal DNA
Sensitivity and Specificity
Spacer
Spores
Vineyards
Vitaceae
Vitis - microbiology
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Summary:The aim of this study was to develop a system for rapid and accurate real‐time quantitative PCR (qPCR) identification and quantification of Botrytis cinerea, one of the major pathogens present on grapes. The intergenic spacer (IGS) region of the nuclear ribosomal DNA was used to specifically detect and quantify B. cinerea. A standard curve was established to quantify this fungus. The qPCR reaction was based on the simultaneous detection of a specific IGS sequence and also contained an internal amplification control to compensate for variations in DNA extraction and the various compounds from grapes that inhibit PCR. In these conditions, the assay had high efficiency (97%), and the limit of detection was estimated to be 6.3 pg DNA (corresponding to 540 spores). Our method was applied to assess the effects of various treatment strategies against Botrytis in the vineyard. Our qPCR assay proved to be rapid, selective and sensitive and may be used to monitor Botrytis infection in vineyards.
ISSN:0378-1097
1574-6968
DOI:10.1111/j.1574-6968.2010.02127.x