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Sensitivity of Botrytis cinerea to seven fungicide chemical classes was determined for 888 isolates collected in 2016 from 47 California strawberry fields. Isolates were collected early season (minimum fungicide exposure) and late season (maximum fungicide exposure) from the same planting block in each field. Resistance was determined using a mycelial growth assay, and variable frequencies of resistance were observed to each fungicide at both sampling times (early season %, late season %): boscalid (12, 35), cyprodinil (12, 46), fenhexamid (53, 91), fludioxonil (1, 4), fluopyram (2, 7), iprodione (25, 8), isofetamid (0, 1), penthiopyrad (8, 25), pyraclostrobin (77, 98), and thiophanate-methyl (81, 96). Analysis of number of chemical class resistances (CCRs) revealed an increasing shift in CCR from the early to late season. Phenotypes of 40 isolates that were resistant or sensitive to different chemical classes were associated with presence or absence of mutations in target genes. Fungicide-resistance phenotypes determined in the mycelial growth assay closely matched (93.8%) the genotype observed. Previously described resistance-conferring mutations were found for each gene. A survey of fungicide use from 32 of the sampled fields revealed an average of 15 applications of gray mold-labeled fungicides per season at an average interval of 12 days. The most frequently applied fungicides (average number of applications during the 2016 season) were captan (7.3), pyraclostrobin (2.5), cyprodinil (2.3), fludioxonil (2.3), boscalid (1.8), and fenhexamid (1.4). Multifungicide resistance is widespread in California. Resistance management tactics that reduce selection pressure by limiting fungicide use, rotating among Fungicide Resistance Action Committee codes, and mixing/rotating site-specific fungicides with multisite fungicides need to be improved and implemented.

Gray mold is a devastating disease on strawberry, and may be caused by several species of Botrytis. The goal of this study was to better understand and characterize the species of Botrytis with reduced sensitivity to the fungicide Polyoxin D, particularly Botrytis fragariae. In total, 78 Botrytis isolates of unknown species that were sensitive (28 isolates; S), moderately sensitive (22 isolates; MS), or reduced sensitive (28 isolates; RS) to Polyoxin-D were collected from commercial strawberry fields of five states in the United States, identified to the species level, and characterized. The majority (75%) of S isolates were Botrytis cinerea and the majority (79%) of RS isolates were the recently described species B. fragariae, indicating an innate ability of B. fragariae to tolerate Polyoxin-D. B. fragariae produced fluffy, white mycelium and was less likely to sporulate on potato dextrose agar than B. cinerea. Isolates from a commercial field recovered from blossoms in early spring were all B. fragariae, those from leaves of the same plants in late spring were a mixture of B. fragariae and B. cinerea, and those from fruit in early summer were all B. cinerea, indicating that B. fragariae may preferentially colonize blossom tissue. A polymerase chain reaction-based assay was developed based on NEP2 sequence variability to distinguish B. fragariae from other Botrytis spp. that have been reported on strawberry, including B. cinerea, B. mali, B. caroliniana, and B. ricini. None of the isolates collected from Canada, California, or North Carolina nurseries were B. fragariae, indicating that the newly described species may not exist or not be widely distributed in planting stock.