Evolution of Cryptococcus neoformans strains from patients with HIV/AIDS

描述（由申请人提供）：目前有超过一百万的HIV/AIDS患者患有真菌性脑膜脑炎，由此导致的发病率和死亡率非常显著。这些感染绝大多数是由新型格鲁比隐球菌变种引起的，该菌株具有荚膜血清型a，这种酵母在环境中普遍存在，与鸟类栖息地和植物残骸有关。感染不是传染性的，但通过吸入干燥的酵母细胞或担子孢子获得外源性感染。虽然感染哺乳动物的能力对真菌没有明显的进化优势，但大多数天然分离物都能够在哺乳动物宿主中定植，这表明致病性是在环境中进化的。在初步研究中，我们开发了强大的多位点基因分型方法，并发现了两个独特的、遗传分离的血清型A亚群：(i)一个由普遍存在的、与野鸽排泄物相关的高度克隆的单型菌株组成的全球亚群；（ii）一个独特的、高度变异的、重组的非洲南部亚群，与非洲本土树木相关。该南部非洲人群的遗传多样性是前所未有的，这表明该人群可能代表了现存的全球a血清型菌株的祖先起源。我们还分析了临床和环境样本中多位点基因型的频率，发现并非所有菌株（或基因型）都同样可能引起人类隐球菌疾病。尽管非洲人口的基因型存在巨大差异，但博茨瓦纳所有患者中几乎有一半（46%）和南非大部分患者（17%）被具有三种密切相关基因型的菌株感染。相比之下，非洲环境分离株的基因型分布具有可比性，没有一个是显性的。在全球分离株中，大多数基因型在患者和主要的非非洲生态位鸽子粪便中同样流行，尽管一种基因型（A2）在环境中高度流行，但从未从患者中分离出来。本研究将在种群遗传和基因组水平上研究新生弧菌致病性的进化。这个项目涉及两个独立但互补的目标。目标1将使用群体遗传和系统发育分析的方法来检验血清型a起源于非洲的假设。目标2将使用比较基因组学、全基因组关联图谱和表型分析的方法来确定区分经常引起感染的菌株与很少（如果有的话）从人类身上分离出来的菌株的基因组区域。将全基因组方法应用于自然菌株的新生芽孢杆菌将鉴定出与人类致病性显著相关的基因或基因组区域。

DESCRIPTION (provided by applicant): Over a million patients with HIV/AIDS are currently afflicted with fungal meningoencephalitis, and the resultant morbidity and mortality are profoundly significant. The vast majority of these infections are caused by strains of Cryptococcus neoformans var. grubii, which have been recognized by possessing capsular serotype A. This yeast is ubiquitous in the environment, where it is associated with avian habitats and vegetative debris. Infection is not contagious but acquired exogenously by the inhalation of desiccated yeast cells or basidiospores. Although the ability to infect mammals offers no obvious evolutionary advantage to the fungus, most natural isolates are capable of colonizing mammalian hosts, which suggests that pathogenicity evolved in the environment. In preliminary studies, we developed robust multilocus genotyping methods and discovered two unique, genetically isolated subpopulations of serotype A: (i) a global subpopulation comprised of ubiquitous, highly clonal monomorphic strains associated with the excreta of feral pigeons and (ii) a unique, highly-variable, recombining subpopulation in southern Africa that is associated with indigenous African trees. The genetic diversity of this southern African population is unprecedented and suggests that this population may represent ancestral origin of the extant, global strains of serotype A. We also analyzed the frequencies of multilocus genotypes among clinical and environmental samples and discovered that not all strains (or genotypes) are equally likely to cause cryptococcal disease in humans. Despite the enormous diversity of genotypes in the African population, almost half of all patients in Botswana (46%) and a major proportion of patients in South Africa (17%) were infected by strains with three closely related genotypes. In contrast, the distribution of genotypes among environmental isolates in Africa is comparable, and none are dominant. Among global isolates, most genotypes were equally prevalent in both patients and the predominant non-African ecological niche, pigeon feces, although one genotype (A2) was highly prevalent in the environment but never isolated from patients. This proposal will investigate the evolution of pathogenicity in C. neoformans at the population genetic and genomic levels. This project involves two independent but complementary aims. Aim 1 will use methods of population genetic and phylogenetic analysis to test the hypothesis of the African origin of serotype A. Aim 2 will use methods of comparative genomics, genome-wide association mapping and phenotypic assays to identify genomic regions that differentiate strains that frequently cause infection from strains that are rarely, if ever, isolated from humans. This application of whole-genome methods to natural strains of C. neoformans will identify genes or genomic regions that are significantly associated with pathogenicity for humans.