Phenotypic switching in the human pathogenic fungus Cryptococcus neoformans is associated with changes in virulence and pulmonary inflammatory response in rodents

High-frequency reversible changes in colony morphology were observed in three strains of Cryptococcus neoformans. For one strain (SB4, serotype A), this process produced three colony types: smooth (S), wrinkled (W), and serrated (C). The frequency of switching between colony types varied for the individual colony transitions and was as high as 10−3. Mice infected with colony type W died faster than those infected with other colony types. The rat inflammatory response to infection with colony types S, W, and C was C > S > W and ranged from intense granulomatous inflammation with caseous necrosis for infection with type C to minimal inflammation for infection with type W. Infection with the various colony types was associated with different antibody responses to cryptococcal proteins in rats. Analysis of cellular characteristics revealed differences between the three colony types. High-frequency changes in colony morphology were also observed in two additional strains of C. neoformans. For one strain (24067A, serotype D) the switching occurred between smooth and wrinkled colonies. For the other strain (J32A, serotype A), the switching occurred between mucoid and nonmucoid colonies. The findings indicate that C. neoformans undergoes phenotypic switching and that this process can affect virulence and host inflammatory and immune responses. Phenotypic switching may play a role in the ability of this fungus to escape host defenses and establish chronic infections.

The second STE12 homologue of Cryptococcus neoformans is MATa-specific and plays an important role in virulence

Cryptococcus neoformans STE12α, a homologue of Saccharomyces cerevisiae STE12, exists only in MATα strains. We identified another STE12 homologue, STE12a, which is MATa specific. As in the case with Δste12α, the mating efficiency for Δste12a was reduced significantly. The Δste12a strains surprisingly still mated with Δste12α strains. In MATα strains, STE12a functionally complemented STE12α for mating efficacy, haploid fruiting, and regulation of capsule size in the mouse brain. Furthermore, when STE12a was replaced with two copies of STE12α, the resulting MATa strain produced hyphae on filament agar. STE12a regulates mRNA levels of several genes that are important for virulence including CNLAC1 and CAP genes. STE12a also modulates enzyme activities of phospholipase and superoxide dismutase. Importantly, deletion of STE12a markedly reduced the virulence in mice, as is the case with STE12α. Brain smears of mice infected with the Δste12a strain showed yeast cells with a considerable reduction in capsule size compared with those infected with STE12a strains. When the disrupted locus of ste12a was replaced with a wild-type STE12a gene, both in vivo and in vitro mutant phenotypes were reversed. These results suggest that STE12a and STE12α have similar functions, and that the mating type of the cells influences the alleles to exert their biological effects. C. neoformans, thus, is the first fungal species that contains a mating-type-specific STE12 homologue in each mating type. Our results demonstrate that mating-type-specific genes are not only important for saprobic reproduction but also play an important role for survival of the organism in host tissue.

Dimorphism and haploid fruiting in Cryptococcus neoformans: association with the alpha-mating type.

Cryptococcus neoformans is a major opportunistic fungal pathogen in AIDS and other immunosuppressed patients. We have shown that wild-type haploid C. neoformans can develop an extensive hyphal phase under appropriate conditions. Hyphae produced under these conditions are monokaryotic, possess unfused clamp connections, and develop basidia with viable basidiospores. The ability to undergo this transition is determined by the presence of the alpha-mating type locus and is independent of serotype. The association of the hyphal phase with the alpha-mating type may explain the preponderance of this mating type in the environment and the nature of the infectious propagule of C. neoformans.