Genotypic diversity in clinical and environmental isolates of Cryptococcus neoformans from India using multilocus microsatellite and multilocus sequence typing.

Abstract

Cryptococcus neoformans is the leading cause of cryptococcal meningitis in HIV/AIDS patients. As infections in humans are predominantly caused by the inhalation of basidiospores from environmental sources, therefore, analysing the population structure of both clinical and environmental populations of Cneoformans can increase our understanding of the molecular epidemiology of cryptococcosis. To investigate the genotypic diversity and antifungal susceptibility profile of a large collection of Cneoformans isolates (n=523) from clinical and environmental sources in India between 2001 and 2014. Cryptococcus neoformans isolates were genotyped by AFLP, microsatellite typing (MLMT) and MLST. In vitro antifungal susceptibility for standard antifungals was undertaken using CLSI M27-A3. All isolates were Cneoformans, AFLP1/VNI and exhibited mating-type MATα. MLMT revealed that the majority of isolates belonged to microsatellite cluster (MC) MC3 (49%), followed by MC1 (35%), and the remaining isolates fell in 11 other MC types. Interestingly, two-thirds of clinical isolates were genotype MC3 and only 17% of them were MC1, whereas majority of environmental strains were MC1 (54%) followed by MC3 (16%). Overall, MLST assigned 5 sequence types (STs) among all isolates and ST93 was the most common (n=76.7%), which was equally distributed in both HIV-positive and HIV-negative patients. Geometric mean MICs revealed that isolates in MC1 were significantly less (P<.05) susceptible to amphotericin B, 5-flucytosine, itraconazole, posaconazole and isavuconazole than isolates in MC3. The study shows a good correlation between MLMT and MLST genotyping methods. Further, environmental isolates were genetically more diverse than clinical isolates.