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A pyrosequencing‐based method to quantify genetic substitutions associated with resistance to succinate dehydrogenase inhibitor fungicides in Botrytis spp. populations
Author(s) -
GobeilRichard Mélanie,
Tremblay DavidMathieu,
Beaulieu Carole,
Van der Heyden Hervé,
Carisse Odile
Publication year - 2016
Publication title -
pest management science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.296
H-Index - 125
eISSN - 1526-4998
pISSN - 1526-498X
DOI - 10.1002/ps.4026
Subject(s) - pyrosequencing , botrytis cinerea , fungicide , botrytis , biology , genetics , population , gene , computational biology , microbiology and biotechnology , botany , medicine , environmental health
BACKGROUND The genetic underlying resistance mechanisms in the population of the phytopathogenic fungus Botrytis cinerea are well documented. Specifically, several genetic substitutions associated with succinate dehydrogenase inhibitor ( SDHI )‐based fungicide resistance have been identified in the succinate dehydrogenase gene. The objective of the present work was to develop a molecular tool for accurate quantification of these genetic substitutions within Botrytis populations. A test using the PyroMark Q24 instrument was designed to detect and quantify five genetic substitutions associated with SDHI resistance. RESULTS The technique is based on sequencing by synthesis, and it generated quantitative and accurate data with a limit of quantification of a minimum of 500 spores. There was a linear relationship between the known and estimated percentages of spores with the targeted genetic substitutions and wild‐type strains at ratios of 0–100%, with a 20% increment. CONCLUSION With the pyrosequencing assay developed in this study, a large number of Botrytis spp. individuals can be characterised in a timely fashion with greater accuracy than by commonly used methods. Hence, pyrosequencing‐based methods will be useful for improving our understanding of fungicide resistance, detecting the arrival of new genetic substitutions, monitoring shifts in fungal populations and assessing the effectiveness of antiresistance strategies, and for routine monitoring of fungicide resistance. © 2015 Her Majesty the Queen in Right of Canada Pest Management Science © 2015 Society of Chemical Industry