Phenotypic Variability Correlates with Clinical Outcome in Cryptococcus Isolates Obtained from Botswanan HIV/AIDS Patients

Pathogenic species of Cryptococcus cause hundreds of thousands of deaths annually. Considerable phenotypic variation is exhibited during infection, including increased capsule size, capsule shedding, giant cells (≥15 μm), and micro cells (≤1 μm). We examined 70 clinical isolates of Cryptococcus neoformans and Cryptococcus tetragattii from HIV/AIDS patients in Botswana to determine whether the capacity to produce morphological variants was associated with clinical parameters. Isolates were cultured under conditions designed to simulate in vivo stresses. Substantial variation was seen across morphological and clinical data. Giant cells were more common in C. tetragattii, while micro cells and shed capsule occurred in C. neoformans only. Phenotypic variables fell into two groups associated with differing symptoms. The production of “large” phenotypes (greater cell and capsule size and giant cells) was associated with higher CD4 count and was negatively correlated with intracranial pressure indicators, suggesting that these are induced in early stage infection. “Small” phenotypes (micro cells and shed capsule) were associated with lower CD4 counts, negatively correlated with meningeal inflammation indicators, and positively correlated with intracranial pressure indicators, suggesting that they are produced later during infection and may contribute to immune suppression and promote proliferation and dissemination. These trends persisted at the species level, indicating that they were not driven by association with particular Cryptococcus species. Isolates possessing giant cells, micro cells, and shed capsule were rare, but strikingly, they were associated with patient death (P = 0.0165). Our data indicate that pleomorphism is an important driver in Cryptococcus infection.