Inhibition of Sterol 14α-Demethylase Activity in Penicillium italicum Does Not Correlate with Resistance to the DMI Fungicide Imazalil

Abstract

Sensitivity of sterol 14α-demethylase activity in cell-free extracts of Penicillium italicum isolates E300-3, H17, and I33 with increasing degrees of resistance to imazalil was studied and compared with that of the wild-type isolate W5. Incorporation of [2-14C]mevalonate into C4-desmethyl sterols in cell-free extracts of medium- and high-resistant isolates H17 and I33 was about 33% of nonsaponifiable lipids. This was slightly higher than activity in cell-free extracts of wild-type isolate W5 (26%) and low-resistant isolate E300-3 (24%). Ergosterol appeared to be the only C4-desmethyl sterol synthesized in cell-free extracts of all isolates. Imazalil inhibited incorporation of [2-14C]mevalonate into ergosterol in cell-free extracts of all isolates tested, resulting in accumulation of radioactivity in eburicol. These results indicate that the sterol biosynthetic pathway and the target site for imazalil in cell-free extracts of both wild-type and DMI-resistant isolates are the same. IC50 values of imazalil (concentrations which inhibit incorporation of [2-14C]mevalonate into ergoserol by 50%) for isolates E300-3, H17,and I33 ranged from 1.6 ± 0.4 × 10−8 to 2.1 ± 0.4 × 10−8M. These values were not significantly different from that of the wild-type isolate W5 (1.6 ± 0.4 × 10−8M). The results suggest that affinity of sterol 14α-demethylase of wild-type and DMI-resistant isolates to imazalil is similar and that decreased affinity of the target enzyme to imazalil does not play a major role as a mechanism of resistance in P. italicum.