Sporothrix schenckii is a dimorphic fungus that is pathogenic for humans. No sexual cycle has been reported for this fungus and little is known of its genetic constitution. To inquire into the ploidy state of Sporothrix schenckii, different approaches were followed: DNA content during transition from conidia to yeast, survival to ultraviolet irradiation, chemical mutagenesis, and induction of mitotic recombination. No change in ploidy was detected between the conidia and yeast phases of the fungi. Resistance to cell inactivation by UV irradiation was higher in S. schenckii and in Sacharomyces cerevisiae in its diploid state than in isogenic haploids a and alpha from S. cerevisiae that were inactivated at lower doses. Two mutant phenotypes, auxotrophy and albinism, were screened after chemical mutagenesis. One-step mutagenesis with either nitrous acid or ultraviolet light was unsuccessful in inducing auxotrophy but was sufficient to induce albino colonies. Two-step mutagenesis with nitrous acid in combination with UV light was necessary to attain two auxotrophic requirements: adenine and methionine. Prototrophic and pigmented revertants behaved as heterozygotes; after exposure to UV light they gave rise to derivatives which resemble the original mutation. The experiments presented in this work suggest that S. schenckii is a diploid, although aneuploidy cannot be excluded. Copyright 1999 Academic Press.