Molecular and genetic characterization of a serotype A MATa Cryptococcus neoformans isolate

Cryptococcus neoformans comprises two varieties (neoformans and gattii) and four serotypes (A, B, C and D). Fertile isolates of both mating types have been identified in serotypes B, C and D; however, a fertile serotype A MATa strain has not been confirmed, although serotype A MATalpha strains will mate with serotype D MATa strains. Preliminary analysis of a recent Italian environmental isolate (IUM 96-2828) suggested that this strain was haploid, serotype A and MATa. In this study, IUM 96-2828 has been characterized in detail. A mating reaction between IUM 96-2828 and H99 (serotype A MATalpha) produced abundant spores with an equal distribution of MATa and MATalpha progeny, all of which were serotype A. Karyotypic analysis of F(1) spores revealed evidence of recombination, confirming that IUM 96-2828 was fertile. The MATa pheromone gene from IUM 96-2828 was sequenced and found to be most closely related to the serotype D MATa pheromone gene. Phylogenetic comparisons of other genes not linked to mating type also suggested IUM 96-2828 was most closely related to serotype A strains. Biochemical analysis showed that the carbon assimilation profiles of H99 and IUM 96-2828 were identical for 97 % (30/31) of the substrates while isozyme analysis showed 89 % (17/19) identity. Assays of major virulence factors found no difference between H99 and IUM 96-2828. Virulence studies using the mouse model demonstrated that IUM 96-2828 was virulent for mice, although it was less virulent than H99. These data strongly suggest that IUM 96-2828 is a true haploid serotype A MATa isolate that is fertile.