Resistance to Thiophanate-Methyl in Botrytis cinerea Isolates From Californian Vineyards and Pistachio and Pomegranate Orchards

In this study, a mycelial growth assay was used to evaluate the sensitivity to thiophanate-methyl of 144 Botrytis cinerea isolates (collection A) from Californian vineyards and pistachio and pomegranate orchards. Based on the effective concentration that inhibits 50% of growth (EC50) values for mycelial growth inhibition on fungicide-amended media, 3, 28, 10, and 58% of the isolates showed sensitivity (SS; EC50 < 1 mu g/ml), low resistance (LR; 1 < EC50 < 10 mu g/ml), weak resistance (WR; 10 < EC50 < 50 mu g/ml), and high resistance (HR; EC50 > 100 mu g/ml) toward thiophanate-methyl, respectively. The LR and HR phenotypes were observed in pistachio and pomegranate orchards, even though pomegranate was not sprayed with thiophanate-methyl. Sensitivity to thiophanate-methyl of a historical collection of 257 B. cinerea isolates (collection B) isolated from pistachio orchards in 1992, 2005, and 2006 was assessed on potato dextrose agar amended with thiophanate-methyl at the discriminatory concentration of 10 mu g/ml. Average percentages of thiophanate-methyl-resistant isolates were 50, 72, and 64% in the orchards in 1992, 2005, and 2006, respectively. A study of fitness components of selected thiophanate-methyl-resistant (LR, WR, and HR) and -sensitive (SS) isolates from collection A did not reveal any significant difference between them with respect to mycelial growth on fungicide-free media and pathogenicity on cultivar Crimson Seedless berries. Comparison of beta-tubulin sequences from resistant and sensitive phenotypes revealed that a glutamic acid at position 198 was changed to alanine in all HR isolates and three LR isolates. The occurrence of thiophanate-methyl resistance in B. cinerea populations should be considered when designing spray programs against blossom and shoot blight of pistachio and gray mold of grape.