Simultaneous Detection of Multiple Benzimidazole-Resistant β-Tubulin Variants of Botrytis cinerea using Loop-Mediated Isothermal Amplification

A - Papers appearing in refereed journals

Duan, Y. B., Yang, Y., Wang, J . X., Chen, C. J., Steinberg, G., Fraaije, B. A. and Zhou, M. G. 2018. Simultaneous Detection of Multiple Benzimidazole-Resistant β-Tubulin Variants of Botrytis cinerea using Loop-Mediated Isothermal Amplification. Plant Disease. 102 (10), pp. 2016-2024.

AuthorsDuan, Y. B., Yang, Y., Wang, J . X., Chen, C. J., Steinberg, G., Fraaije, B. A. and Zhou, M. G.
Abstract

Optimal disease management depends on the ability to monitor the development of fungicide resistance in plant pathogen populations. Benzimidazole resistance is caused by the point mutations of the β-tubulin gene in Botrytis cinerea, and three mutations (E198A, E198K, and E198V) at codon 198 account for more than 98% of all resistant strains. Although traditional methods remain a cornerstone in monitoring fungicide resistance, molecular methods that do not require the isolation of pathogens can detect resistance alleles present at low frequencies, and require less time and labor than traditional methods. In this study, we present an efficient, rapid, and highly specific method for detecting highly benzimidazole-resistant B. cinerea isolates based on loop-mediated isothermal amplification (LAMP). By using specific primers, we could simultaneously detect all three resistance-conferring mutations at codon 198. The LAMP reaction components and conditions were optimized, and the best reaction temperatures and times were 60 to 62°C and 45 min, respectively. When B. cinerea field isolates were assessed for benzimidazole resistance, similar results were obtained with LAMP, minimal inhibition concentration, and sequencing. The LAMP assay developed in the current study was highly suitable for detection of highly benzimidazole-resistant field isolates of B. cinerea.

Year of Publication2018
JournalPlant Disease
Journal citation102 (10), pp. 2016-2024
Digital Object Identifier (DOI)doi:10.1094/PDIS-03-18-0542-RE
Web address (URL)https://doi.org/10.1094/PDIS-03-18-0542-RE
Open accessPublished as non-open access
FunderNational Natural Science Foundation of China
Output statusPublished
Publication dates
Online22 Aug 2018
Publication process dates
Accepted04 Apr 2018
Copyright licensePublisher copyright
PublisherAmerican Phytopathological Society (APS)
ISSN0191-2917

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