Genomic diversity of Candida bloodstream infections

项目摘要 念珠菌属是美国医院血流感染（BSI）的第四大原因。到期BSI 梭白色念珠菌和C.光滑念珠菌是最常见的导致侵袭性感染的念珠菌属，占20%-40% 尽管用一线抗真菌剂棘白菌素治疗。棘白菌素耐药性日益 描述于C.白色念珠菌和C.光滑的临床分离株。然而，大多数棘白菌素治疗 失败与阻力的出现无关。棘白菌素耐受，其中念珠菌生长 抑制但细胞仍存活，可能易于随后产生耐药性，但具有临床意义 这一表型的特征尚不清楚。长期以来，几乎所有念珠菌血症和其他BSI都源于 一个单一的、无性繁殖的有机体。然而，在初步研究中，我们通过全基因组测序（WGS） 和系统发育分析表明，C.白色念珠菌和C.光滑型BSI是由遗传学上 临床微生物学实验室通常无法识别的不同菌株。经过深入分析 梭白色念珠菌WGS数据，我们发现在多个BSI患者中鉴定的基因变异富集于 已知在棘白菌素应答和毒力中重要的生物学过程。我们的目标 该项目将更详细地表征血流C的遗传和表型多样性。白色 和c.光滑菌株，特别注意与持续性或复发性感染相关的菌株 尽管棘白菌素治疗，并涉及棘白菌素中的特定念珠菌基因和基因变体 耐受性、抗性和毒力。我们假设，通过研究纵向BC和外BC的应变， 持续性或复发性丙型肝炎患者的血液部位。 albicans)和非白念珠菌(C.光滑血流感染 棘白菌素治疗后，我们将鉴定出与棘白菌素相关的新基因或基因变异体 耐受性/抗性和毒力。在我们的第一个目标中，我们将完成WGS和C.白色念珠菌和C. 与基线BC相比的光滑（各10例患者）。然后，我们将进行WGS和菌株分析，从阳性 在棘白菌素处理期间或之后收集的纵向血液和血外培养物。最后我们将 确定来自BC的遗传多样性菌株的表型，包括棘白菌素耐受性和抗性。 在第二个目标中，我们将构建同基因突变体和互补C.白色念珠菌和C.光滑菌株 在目标1中鉴定和优先排序的基因和基因变体。我们将在体外测试菌株的表型 以及使用小鼠血源性播散的棘白菌素治疗反应和毒力 感染.结果将确认C的范围和类型。白色念珠菌和C.光滑的遗传多样性，提供 在持续性或复发性BSI期间，念珠菌对棘白菌素反应的新见解，并识别新的 棘白菌素治疗失败和毒力的遗传决定因素。我们的发现将挑战当前临床 和微生物学实验室实践，并通过以下方式为研究BSI期间的遗传多样性提供了基础： 其他念珠菌属和细菌，以及抗真菌剂耐受性、耐药性和发病机制。

Project Abstract Candida species are fourth leading cause of bloodstream infections (BSIs) in United States hospitals. BSIs due to C. albicans and C. glabrata, the most common Candida species to cause invasive infections, are 20%-40% despite treatment with echinocandins, the frontline antifungal agents. Echinocandin resistance is increasingly described among C. albicans and C. glabrata clinical isolates. Nevertheless, most echinocandin treatment failures are not associated with emergence of resistance. Echinocandin tolerance, in which Candida growth is inhibited but cells remain viable, may predispose to subsequent development of resistance, but clinical relevance of this phenotype is unclear. The longstanding paradigm is that almost all candidemia and other BSIs stem from a single, clonal organism. In preliminary studies, however, we showed by whole genome sequencing (WGS) and phylogenetic analyses that C. albicans and C. glabrata BSIs are caused by mixed populations of genetically diverse strains, which are not typically recognized by the clinical microbiology laboratory. Upon deeper analysis of C. albicans WGS data, we found that gene variants identified in multiple patients with BSIs were enriched for biological processes that are known to be important in echinocandin responses and virulence. Our objectives in this project are to characterize in greater detail the genetic and phenotypic diversity of bloodstream C. albicans and C. glabrata strains, with particular attention to strains associated with persistent or recurrent infections despite echinocandin treatment, and to implicate specific Candida genes and gene variants in echinocandin tolerance, resistance and virulence. We hypothesize that by studying strains from longitudinal BCs and extra- blood sites of patients with persistent or recurrent C. albicans or C. glabrata bloodstream infections despite echinocandin treatment, we will identify novel genes or gene variants that are responsible for echinocandin tolerance/resistance and virulence. In our first aim, we will complete WGS and analyses of C. albicans and C. glabrata from baseline BCs (10 patients each). Then, we will perform WGS and analyses of strains from positive longitudinal blood and extra-blood cultures collected during or after echinocandin treatment. Finally, we will determine phenotypes of genetically diverse strains from BCs, including echinocandin tolerance and resistance. In our second aim, we will construct isogenic mutant and complemented C. albicans and C. glabrata strains for genes and gene variants that are identified and prioritized in aim 1. We will test strains for phenotypes in vitro and for echinocandin treatment responses and virulence using mouse models of hematogenously disseminated infections. Results will affirm the extent and type of C. albicans and C. glabrata genetic diversity in BCs, afford new insights into Candida responses to echinocandins during persistent or recurrent BSIs, and identify novel genetic determinants of echinocandin treatment failure and virulence. Our findings will challenge current clinical and microbiology laboratory practices, and provide a foundation for studies of genetic diversity during BSIs by other Candida species and bacteria, and mechanisms of antifungal tolerance, resistance and pathogenesis.