Natural and acquired fenhexamid resistance in Botrytis spp :What's the difference ?

Antifungal compounds such as ergosterol biosynthesis inhibitors are widely used to control crop diseases. Among them, one of the most recent, the hydroxyanilide fenhexamid, is efficient principally against Botrytis cinerea, the major causal agent of grey mould. Fenhexamid is a new type of ergosterol biosynthesis inhibitor affecting the sterol C4 demethylation processes due to its specific interaction with one of the four proteins of the enzymatic complex, the 3-ketoreductase. Our regular monitoring conducted on French vineyards allowed the identification of the first isolates of B. cinerea with acquired resistance. Two types of resistant isolates named HydR3- and HydR3+ were distinguished by their resistance level. This acquired resistance is due to point mutations in the erg27 gene leading to target modifications. These modifications induce a reduced in affinity of fenhexamid towards its target, the 3-ketoreductase. Because of their high resistant level, the HydR3+ strains have to be considered relative to the risk of resistance phenomenom occurrence in vineyards. Fitness studies conducted in vitro on isogenic mutants showed altered "overwintering" capacities of HydR3+ mutants suggesting that they probably do not impact fenhexamid's field efficacy. While B. cinerea's acquired resistance could be explained only by target modifications, as in most cases of fungicide resistance, the situation is different for the related species Botrytis pseudocinerea naturally resistant to fenhexamid. We show that erg27 polymorphism only slightly contributes to resistance whereas fenhexamid detoxification by a cytochrome P450 named cyp68.4 is the major mechanism responsible for the resistance. This is the first case of a functional validation of fungicide detoxification involved in resistance.