Development and application of a simple, rapid and sensitive method for detecting moderately carbendazim-resistant isolates in Botrytis cinerea

作者：Duan, Y. B.^*; Yang, Y.; Li, M. X.; Li, T.; Fraaije, B. A.; Zhou, M. G.^*

来源：Annals of Applied Biology, 2018, 172(3): 355-365.

DOI：10.1111/aab.12426

• Botrytis cinerea
• carbendazim
• fungicide resistance
• loop-mediated isothermal amplification
• beta-tubulin

Sustainable disease management depends on the ability to monitor the development of fungicide resistance in pathogen populations. A point mutation resulting in an alteration (F200Y) at codon 200 of the target protein beta-tubulin leads to a moderate level of resistance to carbendazim in Botrytis cinerea. Although traditional methods remain a cornerstone in detection of fungicide resistance, molecular methods that do not require the isolation of pathogens, can detect the presence of resistance alleles at low frequencies, and require less time and labour than traditional methods. In this study, we present an efficient, rapid, and highly specific method for detecting the moderately carbendazim-resistant isolates in B. cinerea based on loop-mediated isothermal amplification (LAMP). By using specific LAMP primers, we detected the resistance-conferring mutation underlying beta-tubulin F200Y. The concentrations of LAMP components and LAMP parameters were optimised, resulting in reaction temperatures and times of 61-65 degrees C and 45 min, respectively. The feasibility of the LAMP assay was verified by assaying the diseased samples with artificial inoculation in the different hosts. The LAMP assay developed in the current study was specific, stable, repeatable and sensitive, and was successfully applied for detection of moderately carbendazim-resistant isolates of B. cinerea in plant samples.

• 出版日期2018-5
• 单位南京农业大学