Diversity in the erg27 Gene of Botrytis cinerea Field Isolates from Strawberry Defines Different Levels of Resistance to the Hydroxyanilide Fenhexamid

The hydroxyanilide (Hyd) fenhexamid has played a major role in gray mold management in Florida strawberry fields since 2000. Recent monitoring of the sensitivity of Botrytis cinerea to fenhexamid indicated that resistance has emerged. In this study, mutations in the target gene erg27 encoding the 3-keto reductase enzyme were investigated and the shift in fenhexamid sensitivity over time was evaluated in 630 isolates collected between 2005 and 2013 from locations sprayed with different spray programs. Overall, 227, 155, 48, and 200 isolates were sensitive (Hyd(s)), had reduced sensitivity (HydR2), or were moderately (HydR3(-)) and highly (HydR3(+)) resistant, respectively. Analysis of complete sequences of the erg27 gene from 70 isolates revealed seven and five mutations and one deletion in the HydR3- and HydR3+ isolates, respectively, at eight and five different codons, respectively. In addition to the three mutations (F412S, -I, and -V) reported previously at codon 412, two new mutations from glycine to arginine at codon 170 (G170R, two isolates) and from alanine to glycine at codon 210 (A210G, eight isolates) were detected for the first time in HydR3+ isolates from Florida. These isolates were not controlled on detached fruit sprayed with the recommended field rate of fenhexamid, whereas all Hyds, HydR2, and HydR3- isolates were controlled on detached fruit. Overall, there was no clear correlation between the spray frequency of fenhexamid and the frequency of resistant phenotypes. This study provides an overview of the current distribution of erg27 genotypes in Florida and will serve as a baseline for future studies on shifts in population diversity and resistance. The frequency of fenhexamid-resistant populations has increased progressively in different strawberry fields but has not reached a plateau yet, indicating that the effective life of fenhexamid could be extended if appropriate rotation and management strategies are implemented. Sensitivity to other sterol biosynthesis inhibitors is discussed.