INTRODUCTION:Within the Cryptococcus neoformans species complex, two species and five serotypes are recognized: C. neoformans (var. grubii, serotype A; var. neoformans, serotype D and a hybrid, serotype AD) and C. gattii (serotypes B and C). Mating types a and alpha are designated by a single locus, with the mating type alpha being most prevalent in serotype A and D strains. OBJECTIVE:To evaluate the ability of Colombian isolates of the C. neoformans species complex to mate in vitro with tester strains of the opposite mating type. MATERIALS AND METHODS:Fifty three clinical isolates were included in this study, 33 C. neoformans var. grubii serotype A, 4 C. neoformans var. neoformans serotype D, all mating type alpha, and 16 C. gattii, 13 serotype B (mating type a) and 3 serotype C (mating type alpha), were mixed on V8 juice agar, using a modified method, with the appropriate tester strains to determine the mating types in vitro. RESULTS:Mating studies revealed that 9 of 33 (27.3%) serotype A isolates and 6 of 13 (46.2%) serotype B isolates were able to mate. Clamp connections and basidia with basidiospores were observed microscopically, indicating that the mating process had occurred. All mating competent serotype A strains were mating type alpha and the serotype B mating competent strains were mating type a. CONCLUSION:This is the first report of the determination of the mating ability of Colombian Cryptococcus neoformans isolates to mate in vitro with appropriate tester strains, which is of great importance to study the propagation of the fungus around the globe.