An evolutionary population genomics approach to determine the genetic basis of virulence in the pathogenic fungus Cryptococcus neoformans

感染性真菌正在世界范围内造成越来越多的疾病负担。真菌新生隐球菌感染免疫系统受损的人，如艾滋病毒/艾滋病患者，通常会导致脑膜炎。这种病原体已成为全球疾病和死亡的主要真菌原因之一，每年造成100万人感染，占所有与艾滋病有关的死亡人数的15%-20%。研究表明，成功感染和致病(毒力)的能力在很大程度上取决于感染隐球菌菌株的DNA序列(基因)，并且特定的谱系(称为VNI)已经经历了从与树木相关的腐烂生物到与鸟类相关的人类病原体的生态转变。我们断定这种大流行的血统出现在非洲之外，并在过去5000年内殖民世界。现在有机会确定这种出现是如何发生的，方法是对许多环境菌株的基因组进行测序，并将它们的遗传多样性与在全球艾滋病毒患者中发现的导致疾病的菌株进行比较。我们的研究将描述撒哈拉以南非洲和东南亚受影响地区感染艾滋病毒患者的隐球菌菌株的基因组多样性。非洲患者正在参加一项新资助的MRC试验，该试验旨在研究抗真菌药物对隐球菌病的疗效。我们将从东南亚(主要是泰国)和撒哈拉以南非洲地区的环境宿主树木中收集新生葡萄球菌的分离株，并将它们的基因组序列与已知的导致人类患者疾病的基因组序列进行比较。这将使我们能够识别与人类毒力相关的基因组区域，并将确定新生隐孢子虫的主要进化事件，包括通过有性重组进行基因改组，从非洲涌现出多少血统来殖民世界，以及这种情况发生的时间。重要的是，我们将介绍我们的环境与临床感染菌株的关键毒力决定因素，包括它们的衣壳的大小和脱落，它们在人类巨噬细胞内入侵和生长的能力，以及它们杀死模式昆虫物种Galeria mellonella的能力。对于毒力有显著差异的菌株，我们将分析在模拟感染条件下培养的菌株的基因表达模式。这些结果将为各种毒力决定因素提供新的见解，其中一些可能为这种难以治疗的感染提供新的治疗靶点。

Infectious fungi are causing an increasing burden of disease worldwide. The fungus Cryptococcus neoformans infects individuals with damaged immune systems, such as those suffering from HIV/AIDS, commonly resulting in meningitis. This pathogen has become one of the leading fungal causes of disease and death worldwide, causing a million infections per year and 15-20% of all AIDS-related deaths. Research has shown that the ability to successfully infect, and to cause disease (virulence), largely depends on the DNA sequence (genotype) of the infecting Cryptococcus strain, and that a specific lineage (known as VNI) has undergone an ecological transition from a tree-associated wood-decaying organism to a bird-associated human pathogen. We have dated the emergence of this pandemic lineage out of Africa to colonise the world within the last 5,000 years. There is now an opportunity to determine how this emergence occurred, by sequencing the genomes of many environmental strains of C. neoformans and comparing their genetic diversity to that found causing disease in HIV patients worldwide. Our study will characterise the the genomic diversity of infecting Cryptococcus strains from HIV patients in affected regions of sub-saharan Africa and Southeast Asia. The African patients are being enrolled in a newly-funded MRC trial on the efficacy of antifungal drugs against cryptococcosis. We will collect isolates of C. neoformans from their environmental reservoir, trees, across South East Asia (principally Thailand) and sub-Saharan Africa, and compare their genome sequences to those known to cause disease in human patients. This will allow us to identify the genomic regions that are associated with virulence in humans, and will date major evolutionary events in C. neoformans, including the shuffling of genes through sexual recombination, how many lineages emerged from Africa to colonise the world, and when this occurred. Importantly, we will profile our environmental versus clinical infectious isolates with respect to key virulence determinants including size and shedding of their capsule, their ability to invade and grow within human macrophage cells and their ability to kill the model insect species Galleria mellonella. For isolates that show significant differences in virulence, we will profile patterns of gene expression for isolates cultured under conditions that mimic infection. These results will provide novel insights into diverse virulence determinants, some of which may provide new therapeutic targets against this difficult to treat infection.