In order to understand the relationships between human pathogenic fungi and their close, nonpathogenic relatives, we compared small-subunit ribosomal DNA sequences among four closely related pathogens, Histoplasma capsulatum, Blastomyces dermatitidis, Trichophyton rubrum, and Coccidioides immitis, and seven nonpathogenic fungi expected on morphological grounds to be their nearest relatives. We sequenced small-subunit RNA genes from these fungi and used both genetic distance and parsimony algorithms to evaluate their evolutionary relationships to the pathogens. We show that the pathogens are not a monophyletic group, but rather are interspersed among nonpathogenic fungi; thus it is likely that pathogenicity has arisen multiple times within this group. The saprobic fungi Chrysosporium parvum and Uncinocarpus reesii are the closest known relatives of the highly pathogenic Blastomyces dermatitidis and Coccidioides immitis, respectively, and thus may be considered primary candidates for model systems for researchers studying these fungi. The branching order suggests that the conidium (asexual spore) types aleurioconidia and arthroconidia do not define monophyletic groups and may be less distinct than their names suggest. Fungi with a complete life cycle are shown to have closest relatives that lack a known sexual cycle. Such analyses offer a means by which the sexual and asexual fungi could be integrated into a single classification system.