Overexpression of Multidrug ABC Transporter Is Associated with Reduced Sensitivity to Propiconazole and Boscalid in Sclerotinia Homoeocarpa Field Isolates.

Dollar spot, caused by Sclerotinia homoeocarpa, is a prevalent turfgrass disease on North American golf courses and sterol demethylation inhibitor (DMI) fungicides are commonly used for disease control. Fungicide resistance to the benzimidazole, dicarboximide and DMI classes has been confirmed in S. homoeocarpa, however resistance has not been confirmed to the succinate dehydrogenase inhibitor (SDHI) fungicide class.  In vitro sensitivity assays of isolates from five sites (four golf courses with exposed and one baseline) were conducted with DMI (propiconazole) and SDHI (boscalid) fungicides using minimal growth medium (MM). The isolates that were resistant to propiconazole showed significantly higher EC₉₅ values than propiconazole-sensitive isolates using MM amended with a range of concentrations (1, 1000, 3000, 5000 and 10000 μg ml^-1) of boscalid. Recently, we identified an ABC transporter gene homolog associated with multidrug efflux, ShMR1, from RNA-Seq transcriptome data and sought to determine if it might be involved in reduced sensitivity to DMI and SHDI fungicides. Real-Time PCR experiments were conducted for the relative expression (R.E.) of ShMR1 before (constitutive) and after (induced) treatment of propiconazole (0.1 μg ml^-1) or boscalid (10 μg ml^-1) for 1h in a panel of eight isolates from the five sites. We further assayed constitutive R.E. of ShMR1 in two sets of 8 isolates sampled from two of the golf courses with previously confirmed practical propiconazole field resistance to a DMI fungicide. The ShMR1 showed significantly higher constitutive and induced R.E. for propiconazole-insensitive isolates, and induced expression of ShMR1 gene after propiconazole and boscalid was significantly higher for propiconazole-insensitive isolates, as well. The linear relationships between log₁₀ R.E. of ShMR1 and EC₅₀ values of propiconazole and EC₉₅ values of boscalid were highly significant (P = 0.0062) and significant (P = 0.0135), respectively. These results indicate that overexpression of ShMR1 is associated with reduced sensitivity to propiconazole and to a lesser extent, boscalid, suggesting that this ABC transporter may play a role in multidrug resistance to fungicides.