Multiple Fungicide Resistance in Botrytis cinerea from Greenhouse Strawberries in Hubei Province, China.

Two hundred and forty isolates of Botrytis cinerea were collected during the early summer of 2012 and 2013 from strawberry greenhouses in 10 locations in Hubei Province and examined for sensitivity to five fungicides, most of which were commonly used to control this fungus. High frequency of resistance to carbendazim (Car, 63.63%) and cyprodinil (Cyp, 42.42%) was detected. Boscalid-resistant (Bos^R) isolates were detected for the first time in China, whereas no fludioxonil-resistant isolates were identified. Dual resistance to carbendazim and diethofencarb (Die) was also detected. There were six phenotypes of resistance profile (i.e., Car^RDie^SBos^SCyp^S, Car^RDie^RBos^SCyp^S, Car^RDie^SBos^SCyp^R, Car^RDie^SBos^RCyp^S, Car^RDie^RBos^SCyp^R, and Car^RDie^SBos^RCyp^R). Car^RDie^SBos^SCyp^S and Car^RDie^SBos^SCyp^R were the most common phenotypes, occurring at eight and seven locations, respectively. After 10 successive transfers on fungicide-free potato dextrose agar, tested resistant isolates retained levels of resistance similar to or comparative with the initial generation, indicating the stability of these resistances. Fitness evaluations based on investigation of mycelial growth, osmotic sensitivity, sporulation in vitro and in vivo, and virulence revealed the uncompromising fitness in resistant isolates, except that decreased virulence was observed in Bos^R isolates. The molecular basis of carbendazim, diethofencarb, and boscalid resistance was investigated. Results showed that all 13 sequenced carbendazim-resistant isolates harbored the mutation E198V or E198A in the β-tubulin gene and the five isolates with dual resistance to carbendazim and diethofencarb showed the mutation E198K in the same gene. Bos^R isolates possessed the H272R mutation in succinate dehydrogenase subunit B gene. The results achieved in this study challenge the current management strategies for B. cinerea, which largely depend on applications of these fungicides.