Ironing out mycobacterial persistence: Iron access and utilization in chronic mycobacterial infection

消除分枝杆菌的持久性：慢性分枝杆菌感染中铁的获取和利用

• 批准号: 10444899
• 负责人: Alexandra Haley Miller
• 金额: $ 4.89万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10444899
• 关键词: Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Bacteria; Bacterial Genes; Biochemical Reaction; Biological Assay; Cause of Death; Cells; Chronic; Communicable Diseases; Disease; Disease model; Drug resistance; Drug resistance in tuberculosis; Environment; Equilibrium; Genes; Genetic; Genetic Recombination; Genome; Genus Mycobacterium; Goals; Homeostasis; Immune system; In Vitro; Infection; Infectious Agent; Integration Host Factors; Interferons; Iron; Lesion; Libraries; Lung; Macrophage Activation; Maintenance; Methods; Modeling; Modification; Mus; Mutagenesis; Mutation; Mycobacterium Infections; Mycobacterium tuberculosis; Oxidation-Reduction; Phagosomes; Phenotype; Physicians; Proteins; Reporter; Rest; Scientist; Siderophores; Starvation; Stress; System; Testing; Tuberculosis; acute infection; base; career; chronic infection; combat; deprivation; design; drug development; gene synthesis; genetic approach; genetic technology; improved; in vivo; insight; interest; macrophage; mouse model; mutant; mycobacterial; new technology; novel; pathogen; promoter; recombinase; response; skills; tool; uptake

PROJECT SUMMARY/ABSTRACT Tuberculosis disease, caused by the bacterium Mycobacterium tuberculosis (Mtb) is the leading cause of death from a single infectious disease worldwide. Mtb requires iron in order to perform essential biochemical reactions and for the maintenance of redox balance to survive and persist within host cells. During infection, the host encloses Mtb within the macrophage phagosome and actively restricts bacterial access to iron, resulting in Mtb contained within iron-deficient lung lesions. We hypothesize that Mtb confronts an iron-restricted environment as infection persists and requires accessory iron-acquisition molecules to survive inside the host. To test this hypothesis, a low iron-inducible recombination-based reporter system has been developed in mycobacteria, which provides a unique tool to identify iron-starved mycobacteria both in vitro and in vivo. This proposal seeks to i) develop a genetic method to characterize the iron status of mycobacteria, ii) probe the iron available in differentially activated macrophage environments and iii) assay mycobacterial genes required for iron acquisition and utilization in vivo. These aims will employ phenotypically-relevant models of Mtb infection, including macrophage and mouse models of disease. This proposal will also develop a novel technology to probe the mycobacterial genome, as we will generate a library of transposon mutants in the background of the Mtb reporter strain such that recombined bacteria will reflect mutations in genes required to access or utilize iron. These recombined bacteria will be sequenced directly from the inserted transposon to identify the gene of interest. Therefore, successful completion of this proposal will advance the field by providing new insight into the mechanisms by which the host restricts iron availability during chronic infection and the mycobacterial genetic requirements to survive iron depletion in vivo. This proposal will develop cutting-edge technologic approaches to interrogate the mycobacterial genome in vivo, which can be used to identify new bacterial vulnerabilities and avenues for treatment.

项目摘要/摘要 结核菌菌菌菌血细菌引起的结核病（MTB）是死亡的主要原因 来自全世界的一种传染病。 MTB需要铁才能执行必需的生化反应 并维持氧化还原平衡以在宿主细胞内生存并持续。在感染期间，宿主 将MTB封闭在巨噬细胞吞噬体中，并积极限制细菌进入铁，导致MTB 包含铁缺陷型肺部病变。我们假设MTB面对铁限制的环境 随着感染的持续存在并需要辅助铁进取的分子才能在宿主内生存。测试这个 假设是在分枝杆菌中开发了一种低铁诱导的基于重组的记者系统， 它提供了一种独特的工具，可以在体外和体内识别饥饿的分枝杆菌。该建议寻求 i）开发一种遗传方法来表征分枝杆菌的铁状态，ii）探测可用的铁 差异激活的巨噬细胞环境和iii）取得铁所需的分生不清基因 和体内利用。这些目标将采用与表型相关的MTB感染模型，包括 巨噬细胞和小鼠疾病模型。该建议还将开发一种新型技术来探究 分枝杆菌基因组，因为我们将在MTB记者的背景下生成一个转座子突变体库 重新组合细菌的菌株将反映获得或利用铁所需的基因突变。这些 重组细菌将直接从插入的转座子进行测序，以鉴定感兴趣的基因。 因此，该提案的成功完成将通过提供新的见解来推进该领域 宿主在慢性感染和分枝杆菌遗传期间限制铁的可用性的机制 在体内生存铁耗尽的要求。该建议将开发最先进的技术方法 在体内询问分枝杆菌基因组，可用于识别新的细菌脆弱性和 治疗途径。