Lack of an Intron in Cytochrome and Overexpression of Sterol 14α-Demethylase Indicate a Potential Risk for QoI and DMI Resistance Development in spp. on Grapes.

Asian grapevine leaf rust, caused by and , is often controlled by quinone outside inhibitor (QoI) and demethylation inhibitor (DMI) fungicides in Brazil. Here, we evaluated the sensitivity of 55 spp. isolates to pyraclostrobin (QoI) and tebuconazole (DMI). To elucidate the resistance mechanisms, we analyzed the sequences of the cytochrome (CYTB) and cytochrome P450 sterol 14α-demethylase (CYP51) target proteins of QoI and DMI fungicides, respectively. The expression levels were also determined in a selection of isolates. In leaf disc assays, the mean 50% effective concentration (EC) value for pyraclostrobin was about 0.040 µg/ml for both species. sequences were identical among all 55 isolates, which did not contain an intron immediately after codon 143. No amino acid substitution was identified at codons 129, 137, and 143. The mean EC value for tebuconazole was 0.62 µg/ml for and 0.46 µg/ml for , and no sequence variation was identified among isolates of the same species. However, five isolates grew on leaf discs treated at 10 µg/ml tebuconazole, and these were further exposed to tebuconazole selection pressure. Tebuconazole-adapted laboratory isolates of showed an eight- to 25-fold increase in resistance after four rounds of selection that was not associated with CYP51 target alterations. In comparison with sensitive isolates, expression was induced in the presence of tebuconazole in three out of four tebuconazole-adapted isolates tested. These results suggest a potential risk for QoI and DMI resistance development in spp.