Resistance to azole antifungals in Ustilago maydis was associated with a leaky defect in sterol delta 5(6)desaturase. This defect resulted in reduced accumulation of 14 alpha-methylergosta-24(28)-diene-3 beta,6 alpha-diol and an increase in the proportion of 14 alpha-methylfecosterol in treated cells when compared to the parent strain. The results demonstrate the importance of this mechanism in pathogenic fungi.