Survey and Genetic Analysis of Demethylation Inhibitor Fungicide Resistance in From Michigan Orchards.

Resistance to sterol demethylation inhibitor (DMI) fungicides in , causal agent of brown rot of stone fruit, has been reported in the southeastern and eastern United States and in Brazil. DMI resistance of some isolates, in particular those recovered from the southeastern United States, is associated with a sequence element termed "Mona" that causes overexpression of the cytochrome demethylase target gene . In this study, we conducted statewide surveys of Michigan stone fruit orchards from 2009 to 2011 and in 2019, and we determined the sensitivity to propiconazole of a total of 813 isolates of . A total of 80.7% of Michigan isolates were characterized as resistant to propiconazole by relative growth assays, but the Mona insert was not uniformly detected and was present in some isolates that were not characterized as DMI resistant. Gene expression assays indicated that elevated expression of was only weakly correlated with DMI resistance in isolates from Michigan, and there was no obvious correlation between the presence of the Mona element and elevated expression of . However, sequence analysis of from 25 DMI-resistant isolates did not reveal any point mutations that could be correlated with resistance. Amplification and sequencing upstream of resulted in detection of DNA insertions in a wide range of isolates typed by DMI phenotype and the presence of Mona or other unique sequences. The function of these unique sequences or their presence upstream of cannot be correlated to a DMI-resistant genotype at this time. Our results indicate that DMI resistance was established in Michigan populations of by 2009 to 2011, and that relative resistance levels have continued to increase to the point that practical resistance is present in most orchards. In addition, the presence of the Mona insert is not a marker for identifying DMI-resistant isolates of in Michigan.