Mechanisms of resistance to fenpropimorph and terbinafine, two sterol biosynthesis inhibitors, in Nectria haematococca, a phytopathogenic fungus

The mechanisms of resistance to terbinafine, a squalene epoxidase inhibitor , and to fenpropimorph, a sterol Delta(14)-reductase and/or Delta(8) --> De lta(7)-isomerase inhibitor, were investigated in laboratory mutants of the phytopathogenic fungus Nectria haematococca. Neither modified fungicide upt ake nor fungicide metabolism could explain resistance in the mutants studie d. The terbinafine-resistant mutants contained at least 10 times more squal ene than the wild-type strain, and when cultivated in presence of terbinafi ne, they required a 30-times higher fungicide concentration to obtain a ver y high level of accumulated squalene similar to that of the wild-type strai n. Thus, a reduced affinity of the squalene epoxidase toward both the subst rate and the fungicide could explain terbinafine resistance in these mutant s. In some fenpropimorph-resistant mutants, the effects of fenpropimorph on sterol biosynthesis were similar to those in the wild-type strain, suggest ing that resistance could be due to tolerance to sterol Delta(8,14)-sterol accumulation and to Delta(5,7)-sterol decrease. In one of them, the major s terol in absence of fungicide was not ergosterol, but ergosta-5,7,22,24(24( 1))-tetraenol, indicating reduced Delta(24('241))-reductase activity. The p otential role of a modified Delta(24(241)) reductase in fenpropimorph resis tance has yet to be elucidated. In the remaining fenpropimorph-resistant mu tants, much higher fenpropimorph concentrations were required to cause ergo sterol decreases similar to that in the wild-type strain, suggesting a Delt a(14)-reductase modification. Among them, some mutants accumulated Delta(8) -sterols as well as Delta(8,14)-sterols, indicating that the mechanism of r esistance may be due to reduced affinity of the Delta(14)- reductase toward fenpropimorph, whereas the other mutants accumulated only Delta(8,14)-ster ols. Those last mutants contained much larger amounts of 4,4-dimethyl and 4 alpha-methyl sterols than the wild-type strain, in the absence of fungicid e. The most abundant methylated sterol was 4,4-dimethylfecosterol, the pres umed product of the Delta(14)-reductase, suggesting that resistance may be due to overproduction of the Delta(14)-reductase in these mutants. (C) 1999 Academic Press.