Genetic diversity as a major driver of phenotypic variability and virulence in local outbreak strains of the emerging pathogen Candida auris

遗传多样性是新发病原体耳念珠菌局部爆发菌株表型变异和毒力的主要驱动因素

• 批准号: 2302750
• 金额: --
• 依托单位: University of Aberdeen
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2302750
• 关键词: Genetic; diversity; major; driver; phenotypic

Candida auris has emerged as a nosocomial pathogen only a decade ago, and has developed into a global health threat with a mortality rate of ~50%. C. auris is often multidrug resistant, sticks to skin and medical devices, and is difficult to eradicate from high-dependency hospital units. Consequently, national health authorities, including the Centre for Disease Control in the USA (CDC) and Public Health England (PHA) have posted alerts on C. auris.The Centre for Genome Biology and Medicine at Aberdeen undertook the whole genome sequencing of ten C. auris isolates covering the 4 main clades; this provides a major resource for answering these key questions. We also have been gifted unrestricted access to outbreak strains from PHE and the CDC.(1) Understanding how genetic diversity evolves in C. auris and drives the outbreak of specific genotypes around the world Our recent genome sequencing and analysis revealed that C. auris, contrary to previous report, has a complete mating type locus (MAT; the sex determination system in fungi). To elucidate whether C. auris is capable of sexual reproduction including meiosis, we will test whether the MATa-strain VPCI479/P/13 is sexually compatible with any of the characterized MATx-strains. We have already established that the C. auris genome is haploid. Using pulsed-field gel electrophoresis we will also determine whether - in addition to the genetic diversity revealed by the whole genomic sequencing - C. auris isolates display chromosome number and size differences. Differences in karyotype between individuals of the same species introduce a reproductive barrier, but can be a driver of genetic variability. This will guide our choice of mating combinations of isolates to be tested, and will contribute to fully assembled genomes of different C. auris isolates.(2) Characterization of the cell wall's phenotypic diversity in clinical isolates of C. auris and determining how this underpins virulenceTo elucidate why this species is hypervariable in its phenotype and why various isolates show significant differences in their virulence, this project will examine the genetic and physiological variation in cell wall composition and the response of the cell wall to antibiotic stress. This is relevant to the aggregating properties of the cells, as well as to immune recognition and altered sensitivity to echinocandin antifungal drugs which this species is often resistant to. Cell adhesion is as established virulence factor of Candida species and, intriguingly, C. auris infections are passed readily from person to person and are hard to eradicate. This suggests that adherence and virulence correlate well in C. auris. This objective has significant potential for elucidating aspects of the epidemiology of C. auris infections that are not yet understood. (3) Characterization of the genetic diversity of mult-drug resistance determinants of C. auris clinical isolates and determining how these affect virulenceC. auris has a massively expanded family of genes encoding MDR drug efflux pumps, and it is crucially important to understand how these contribute to its overall multi-drug resistance phenotype. In this project bioinformatics analysis will reveal which drug efflux factors are present in which isolates. This information will be used to select representatives of each of the 4 clades whose drug efflux factors will be subjected to realtime-PCR analysis to determine which are upregulated in response to exposure to different classes of antifungals (azoles, echinocandins, amphothericin). Those genes that are upregulated in response to antifungal exposure will be selectively oblated by CRISPR-Cas9 technology, for which we now have extensive experience for C. albicans. This project has great potential to elucidate the origin of differences in drug resistance between clinical isolates of C. auris, and ultimately will provide guidance for treating patients infected with C. auris strains

念珠菌在十年前才作为医院病原体出现，并已发展成为全球健康威胁，死亡率约为50%。耳念珠菌通常具有多重耐药性，粘附在皮肤和医疗设备上，很难从高依赖性的医院单位根除。因此，包括美国疾病控制中心（CDC）和英国公共卫生部（PHA）在内的国家卫生当局已发布了关于金黄色葡萄球菌的警报。阿伯丁基因组生物学和医学中心对10个金黄色葡萄球菌分离株进行了全基因组测序，涵盖4个主要分支；这为回答这些关键问题提供了主要资源。我们也被授予无限制地接触公共卫生部门和疾病控制中心的爆发菌株的权利。(1)了解C. auris的遗传多样性进化和驱动特定基因型在世界范围内爆发的机制。我们最近的基因组测序和分析表明，与之前的报道相反，C. auris具有完整的交配型位点（MAT，真菌中的性别决定系统）。为了阐明金黄色葡萄球菌是否能够有性生殖包括减数分裂，我们将测试mata菌株VPCI479/P/13是否与任何特征的matx菌株有性相容。我们已经确定金黄色葡萄球菌的基因组是单倍体。利用脉冲场凝胶电泳，我们还将确定——除了由全基因组测序揭示的遗传多样性之外——金黄色葡萄球菌分离物是否显示出染色体数量和大小的差异。同一物种个体之间的核型差异引入了生殖障碍，但也可能是遗传变异的驱动因素。这将指导我们选择要测试的分离株的交配组合，并将有助于完全组装不同金黄色葡萄球菌分离株的基因组。(2)临床分离的金黄色葡萄球菌细胞壁表型多样性的特征，并确定这是如何支持毒力的。为了阐明为什么这个物种在表型上是高度可变的，为什么不同的分离株在其毒力上表现出显著的差异，本项目将研究细胞壁组成的遗传和生理变化以及细胞壁对抗生素应激的反应。这与细胞的聚集特性有关，也与免疫识别和对棘白菌素抗真菌药物的敏感性改变有关，这种药物通常对棘白菌素有耐药性。细胞粘附是念珠菌种的确定毒力因素，有趣的是，耳念珠菌感染很容易在人与人之间传播，很难根除。这表明在金黄色葡萄球菌中，粘附性和毒力密切相关。这一目标对于阐明尚不了解的金黄色葡萄球菌感染的流行病学方面具有重要的潜力。(3) auris临床分离株多药耐药决定因素的遗传多样性特征及其对毒力的影响。auris具有大量扩展的编码耐多药药物外排泵的基因家族，了解这些基因如何促进其整体多药耐药表型至关重要。在这个项目中，生物信息学分析将揭示哪些药物外排因子存在于哪些分离株中。这些信息将用于选择4个分支的代表，这些分支的药物外排因子将进行实时pcr分析，以确定哪些在暴露于不同类别的抗真菌药物（唑类、棘白菌素、两性霉素）时被上调。那些因抗真菌暴露而上调的基因将被CRISPR-Cas9技术选择性地清除，我们现在对白色念珠菌有丰富的经验。本项目对阐明临床分离的金黄色葡萄球菌耐药差异的来源具有很大的潜力，并最终为金黄色葡萄球菌感染患者的治疗提供指导