Cryptococcus neoformans and Cryptococcus gattii are closely related pathogenic basidiomycetous yeasts in which six haploid genotypic groups have been distinguished. The two haploid genotypic groups of C. neoformans have been described as variety grubii and variety neoformans. The four C. gattii genotypic groups have, however, not been described as separate taxa. One hundred and seventeen isolates representing all six haploid genotypic groups were selected for multi-locus sequence typing using six loci to investigate if the isolates consistently formed monophyletic lineages. Two monophyletic lineages, corresponding to varieties grubii and neoformans, were consistently present within C. neoformans, supporting the current classification. In addition, four monophyletic lineages corresponding to the previously described genotypic groups were consistently found within C. gattii, indicating that these lineages should be considered different taxa as well.