Scedosporium prolificans mutants lacking the ability to synthesize melanin were selected after ultraviolet light (UV) irradiation. UV exposure of S. prolificans conidia resulted in a high frequency of melanin-deficient (mel(-)) mutants. Stable and non-stable morphological variants were found in the population: reversion of the mutant phenotype was always to the wild-type phenotype. Based on their morphological differences, these variants were classified into five different groups that were phenotypically characterized. The mel(-) mutants plus the wild-type strain were examined for in vitro susceptibility to antifungal agents with different and/or the same mechanism of action. There was no apparent difference in minimum inhibitory concentrations when comparing the wild-type and the mel(-) mutants. Therefore, melanin does not appear to confer protection against the more important antifungal agents in S. prolificans.