Analyses of the Global Multilocus Genotypes of the Human Pathogenic Yeast Cryptococcus neoformans Species Complex

Cryptococcus neoformans species complex (CNSC) is a globally distributed human opportunistic yeast pathogen consisting of five major molecular types (VNI, VNII, VNB, VNIII and VNIV) belonging to two species, C. neoformans (VNI, VNII and VNB, collectively called serotype A) and C. deneoformans (VNIV, commonly called serotype D), and their hybrids (VNIII, serotype AD). Over the years, many studies have analyzed the geographical distribution and genetic diversity of CNSC. However, the global population structure and mode of reproduction remain incompletely described. In this study, we analyze the published multilocus sequence data at seven loci for CNSC. The combined sequences at the seven loci identified a total of 657 multilocus sequence types (STs), including 296 STs with known geographic information, representing 4200 non-redundant isolates from 31 countries and four continents. Among the 296 STs, 78 and 52 were shared among countries and continents, respectively, representing 3643 of the 4200 isolates. Except for the clone-corrected serotype D sample among countries, our analysis of the molecular variance of the 4200 isolates revealed significant genetic differentiations among countries and continents in populations of CNSC, serotype A, and serotype D. Phylogenetic analyses of the concatenated sequences of all 657 STs revealed several large clusters corresponding to the major molecular types. However, several rare but distinct STs were also found, representing potentially novel molecular types and/or hybrids of existing molecular types. Phylogenetic incompatibility analyses revealed evidence for recombination within all four major molecular types-VNI, VNII, VNIV and VNB-as well as within two VNB subclades, VNBI and VNBII, and two ST clusters around the most common STs, ST5 and ST93. However, linkage disequilibrium analyses rejected the hypothesis of random recombination across most samples. Together, our results suggest evidence for historical differentiation, frequent recent gene flow, clonal expansion and recombination within and between lineages of the global CNSC population.