The species constituting the genus Malassezia are considered to be emergent opportunistic yeasts of great importance. Characterized as lipophilic yeasts, they are found in normal human skin flora and sometimes are associated with different dermatological pathologies. We have isolated seven Malassezia species strains that have a different Tween assimilation pattern from the one typically used to differentiate M. furfur, M. sympodialis, and M. slooffiae from other Malassezia species. In order to characterize these isolates of Malassezia spp., we studied their physiological features and conducted morphological and molecular characterization by PCR-restriction fragment length polymorphism and sequencing of the 26S and 5.8S ribosomal DNA-internal transcribed spacer 2 regions in three strains from healthy individuals, four clinical strains, and eight reference strains. The sequence analysis of the ribosomal region was based on the Blastn algorithm and revealed that the sequences of our isolates were homologous to M. furfur sequences. To support these findings, we carried out phylogenetic analyses to establish the relationship of the isolates to M. furfur and other reported species. All of our results confirm that all seven strains are M. furfur; the atypical assimilation of Tween 80 was found to be a new physiological pattern characteristic of some strains isolated in Colombia.The genus Malassezia comprises lipophilic yeasts found in the normal flora of human skin and other mammals. These yeasts were described by Eichstedt in 1848 as being associated with pityriasis versicolor (PV) lesions (13). The taxonomy and nomenclature of the genus Malassezia was controversial for many decades. Indeed, until 1990 only three species were recognized: M. furfur, M. sympodialis, and M. pachydermatis, a non-lipid-dependent species (17,21,38 (4,6,11,17,18,20,21,26,34,38,(40)(41)(42).Malassezia species have been associated with diverse dermatological pathologies, including PV, seborrheic dermatitis dandruff, atopic dermatitis, folliculitis, psoriasis, onicomycosis, and blepharitis. M. furfur and M. pachydermatis have been associated with systemic infections in patients with underlying diseases and those receiving intravenous lipid emulsions (6, 7, 9-11, 16, 29, 33, 34).Although the role of Malassezia species in the development of these diseases is not clear, some authors suggest that M. globosa is the causal agent of PV, while others have found a greater percentage of isolates of M. sympodialis associated with the disease. Differences in diagnosis might be due to sampling methods and differences between the culture media used, leading to controversies in clinical studies of these dermatological pathologies (1, 7, 9, 10). New physiological patterns for identification have been described, and recently the availability of molecular biology and sequencing techniques has allowed the species to be distinguished more clearly (17-21). Despite the difficulty in isolating, maintaining, and identifying these yeasts, different characteristics of ...