Implications of mycoviral infection in Talaromyces marneffei: an analysis of human patient samples, RNAi, and hypermutation

马尔尼菲踝节菌中真菌病毒感染的影响：对人类患者样本、RNAi 和超突变的分析

• 批准号: 10381581
• 负责人: JOSEPH HEITMAN
• 金额: $ 24.15万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-02 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10381581
• 关键词: AIDS/HIV problem; Agrobacterium; Alleles; Animal Model; Automobile Driving; Biological; Biological Assay; Biology; Blood; Cessation of life; Clinical; Collection; Congenic Strain; DNA Transposable Elements; Data; Development; Diagnostic; Disease Management; Double Stranded RNA Virus; Double-Stranded RNA; Down-Regulation; Drug resistance; Environment; Exposure to; FK506; Flucytosine; Foundations; Gene Deletion; Gene Expression Regulation; Genes; Genetic; Genetic Determinism; Genetic study; Genome; Genomic approach; Geographic Locations; Goals; Growth; Health; Human; Immunocompetent; Immunocompromised Host; Incubated; Infection; Insertional Mutagenesis; Investigation; Laboratories; Life; Link; Malassezia; Mediating; Methods; Microbiology; Modality; Modeling; Modification; Molecular Genetics; Morbidity - disease rate; Movement; Mus; Mutation; Mycoses; Neomycin; Nucleic Acids; Organism; Outcome; Parasites; Pathogenesis; Pathogenicity; Pathologic Mutagenesis; Pathway interactions; Patient Isolators; Patient-Focused Outcomes; Patients; Pharmaceutical Preparations; Phenotype; Plasmids; Play; Population; Process; Publishing; RNA Interference; Rattus; Recurrent disease; Reporting; Research; Resistance; Risk; Role; Sampling; Severity of illness; Sirolimus; Southeastern Asia; Talaromyces; Temperature; Testing; The science of Mycology; Therapeutic; Untranslated RNA; Viral; Viral Load result; Virulence; Virulence Factors; Virus; Work; antimicrobial drug; base; clinical diagnostics; disorder control; drug isolation; experience; experimental study; follow-up; fungus; gel electrophoresis; genetic approach; genome sequencing; homologous recombination; human disease; hygromycin A; improved; insight; mortality; mouse model; mutant; novel; particle; pathogenic fungus; pathogenic microbe; patient prognosis; prognostic; prospective; relapse patients; sample collection; screening; treatment response; whole genome

Abstract Talaromyces marneffei (Tm) is one of seven dimorphic human fungal pathogens that are causative agents of substantial global morbidity and mortality in both immunocompetent and immunocompromised patients. Tm is endemic throughout Southeast Asia in a geographic region that encompasses more than half of the global population, and is a frequent cause of infection and death in HIV/AIDS patients. The organism is present in the environment and in association with bamboo rat species and there is substantial risk of frequent and repeated exposure. Treatment options are limited and suboptimal, diagnostic modalities remain to be optimized, and our understanding of virulence determinants is limited. Although studies have shown that Tm is amenable to transformation and gene deletion via homologous recombination through Agrobacterium mediated trans-kingdom (AMT) conjugation, little investigation has been done to elucidate virulence factors in Tm. A recent study reported the presence of a novel dsRNA mycovirus in some clinical isolates, and that presence of this virus is correlated with enhanced virulence in animal models and the down regulation of the expression of genes involved in RNAi, a process which can target and degrade dsRNAs. In our proposal, we seek to capitalize upon these advances in order to dissect the mycoviral and genetic determinants of Tm virulence. In aim 1, we will analyze the presence or absence of the recently discovered mycovirus in a large collection (N=293) of well-validated Tm human clinical isolates with robust mycological and clinical outcome data and for which whole-genome data is available, and to assess correlations of the presence of this mycovirus determinant with disease severity and patient outcomes. We will develop approaches to cure and reintroduce this mycovirus in order to construct congenic strains with and without the virus for direct laboratory analyses of virulence. In aim 2, we will determine if the supression of RNAi caused by the mycovirus results in a hypermutator phenotype in virus-infected strains. Using a transposon trap assay, we will screen the Tm patient sample collection for isolates that produce a larger number of drug-resistant colonies, and determine if drug resistance is due to the insertion of transposable elements into the gene responsible for resistance. We will then assess any correlations of the hypermutator phenotype with virus-infected and virus-uninfected Tm patient samples. We will use AMT to delete components of the RNAi pathway and we will determine the consequences of these mutations on hypermutation, viral load, and if virulence is altered. These studies will advance our understanding of Tm mycoviral and genetic virulence determinants and provide insights with possible implications for patient prognosis and treatment.

抽象的 马尔尼菲踝节菌 (Tm) 是七种二态性人类真菌病原体之一 在免疫功能正常和免疫功能低下的人群中造成大量全球发病率和死亡率的因素 患者。 Tm 在整个东南亚地区流行，该地理区域覆盖了一半以上的人口 全球人口，是艾滋病毒/艾滋病患者感染和死亡的常见原因。该生物体是 存在于环境中并与竹鼠物种相关，并且存在频繁发生的巨大风险 并反复曝光。治疗选择有限且欠佳，诊断方式仍有待改进 优化，并且我们对毒力决定因素的理解是有限的。尽管研究表明 Tm 易于通过农杆菌介导的同源重组进行转化和基因删除 跨界（AMT）接合，很少有研究来阐明 Tm 的毒力因子。最近的一个 研究报告了一些临床分离株中存在一种新型 dsRNA 真菌病毒，并且该病毒的存在 与动物模型中毒力增强和基因表达下调相关 参与 RNAi，这是一个可以靶向并降解 dsRNA 的过程。 在我们的提案中，我们寻求利用这些进展来剖析真菌病毒和遗传 Tm毒力的决定因素。在目标 1 中，我们将分析最近发现的是否存在 大量经过充分验证的 Tm 人类临床分离株（N = 293）中的真菌病毒，具有强大的真菌学和 临床结果数据和可用的全基因组数据，并评估存在的相关性 这种真菌病毒与疾病严重程度和患者结果的决定因素。我们将开发治疗方法 并重新引入这种真菌病毒，以构建有或没有该病毒的同源株，用于直接 毒力的实验室分析。在目标 2 中，我们将确定是否由真菌病毒引起的 RNAi 抑制 导致病毒感染菌株出现超突变表型。使用转座子陷阱测定，我们将筛选 收集产生大量耐药菌落的患者样本，并确定 如果耐药性是由于转座元件插入到导致耐药性的基因中造成的。我们 然后将评估超变子表型与病毒感染和病毒未感染的 Tm 的任何相关性 患者样本。我们将使用 AMT 删除 RNAi 途径的成分，然后确定 这些突变对超突变、病毒载量以及毒力是否改变的后果。这些研究将 增进我们对 Tm 真菌病毒和遗传毒力决定因素的理解，并提供见解 对患者预后和治疗的可能影响。

项目成果

Talaromyces marneffei promotes M2-like polarization of human macrophages by downregulating SOCS3 expression and activating the TLR9 pathway.

马尔尼菲踝节菌通过下调 SOCS3 表达和激活 TLR9 通路来促进人类巨噬细胞的 M2 样极化。

• 发表时间: 2021
• 影响因子: 5.2
• 作者: Wei, Wudi;Ning, Chuanyi;Huang, Jiegang;Wang, Gang;Lai, Jingzhen;Han, Jing;He, Jinhao;Zhang, Hong;Liang, Bingyu;Liao, Yanyan;Le, Thuy;Luo, Qiang;Li, Zhen;Jiang, Junjun;Ye, Li;Liang, Hao
• 通讯作者: Liang, Hao