In vivo persistence and immuno-pathogenesis of Mycobacterium abscessus in a new Xenopus tadpole model

脓肿分枝杆菌在新爪蟾蝌蚪模型中的体内持久性和免疫发病机制

• 批准号: 10350750
• 负责人: Martin S. Pavelka
• 金额: $ 23.1万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-11 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10350750
• 关键词: Acids; Address; Alleles; Amphibia; Animal Model; Antibiotic Resistance; Antibiotic Therapy; B-Lymphocytes; Bacteria; Biological Assay; Breathing; Bronchiectasis; Burkholderia cepacia; Chronic; Chronic lung disease; Comparative Biology; Cystic Fibrosis; Development; Disease; Embryo; Exhibits; Fertilization; Flow Cytometry; Gene Mutation; Genes; Genetic Determinism; Genome; Genus Mycobacterium; Histology; Homologous Gene; Human; Immune; Immune response; Immune system; Immunity; Immunocompetent; Immunocompromised Host; Infection; Interferons; Investigation; Knock-out; Knockout Mice; Label; Laboratory Organism; Leukocytes; Liver; Lower respiratory tract structure; Lung; Lung diseases; Lung infections; Mammals; Microbial Biofilms; Microscopy; Modeling; Morphology; Multi-Drug Resistance; Mus; Mycobacterium abscessus; Mycobacterium marinum; Myeloid Cells; Pathogenesis; Pathogenicity; Pathology; Patient Isolators; Phenotype; Play; Positioning Attribute; Prevalence; Production; Proliferating; Pseudomonas aeruginosa; Refractory; Research Proposals; Resistance; Role; Skin; Soil; System; T-Lymphocyte; Tadpoles; Tissues; Transgenic Organisms; Virulence Factors; Visualization; Water; Xenopus; Xenopus laevis; Zebrafish; adaptive immunity; arm; cell envelope; chronic infection; emerging pathogen; host colonization; human disease; human tissue; immunopathology; in vivo; insight; intravital microscopy; lung pathogen; macrophage; mutant; non-tuberculosis mycobacteria; novel; pathogen; prevent; promoter; respiratory pathogen; response; transcriptomics

Owing to its increasing prevalence, persistence, multidrug resistance, pathogenicity and treatment challenges, the non-tuberculosis mycobacterium (NTM) Mycobacterium abscessus (Mab) has become a serious threat to humans with chronic lung diseases such as bronchiectasis and cystic fibrosis, as well as immunocompromised patients. However, Mab is still understudied compared to other pathogenic mycobacteria. In addition, there is a limitation in current animal models that poses a real challenge for investigating the respective roles of two Mab morphotypes, smooth (S) and rough (R), in lung immunopathology, persistence and host immune response. The clearance of Mab in immunocompetent and even immunocompromised mice prevent the study of chronic infection, and while for zebrafish embryos are instrumental for investigating Mab infection, they cannot be used to model a pulmonary infection and T cell involvement. Thus, there is a need for complementary animal models. Since like zebrafish embryos, X. laevis tadpoles are transparent, allowing visualization of pathogen dissemination, but that in addition they do have functional lungs and exhibit an immune system including T cells remarkably similar to humans, we propose to develop a X. laevis tadpole model of Mab infection that mimics human pulmonary disease. Relying on an established comparative biology approach to study immunity to mycobacteria in tadpoles and our recent demonstration that both S and R Mab morphotypes readily infect tadpoles, disseminate in tadpole lungs and persist up to 50 days, the objective of the exploratory research proposal is to investigate in vivo for over 40 days Mab persistence, and immuno-pathogenesis. Specifically, to address the hypothesis that persistent R Mab is more actively proliferating and immunopathogenic than S Mab in X. laevis tadpoles we propose: , by comparing the host immune response and persistence of R and S Mab in the lung and other tissues using intravital (1) Characterize immune-pathogenesis in tadpole lungs during persisting Mab infection microscopy flow cytometry and transcriptomics as well as fluorescently labeled Mab and transgenic tadpoles with fluorescent macrophages. (2) Investigate the relevance of S to R morphotype switch in vivo for pathogenesis and persistence , by using a promoter system to control the S to R switch following infection and examining effect of switches on persistence, pathogenicity and host immune response. (3) Investigate genetic determinants of Mab immune-pathogenesis , using selected Mab deletion mutants generated by allelic exchange of Mab-specific genes as well as gene homologs shared by Gram- negative lung pathogens. We anticipate that novel insights relevant to human will be gathered. Our collaborative team is uniquely positioned to carry these studies due to our background and complementary expertise.

由于其流行率、持久性、多药耐药性、致病性和治疗日益增加， 挑战，非结核分枝杆菌（NTM）结核分枝杆菌（Mab）已成为 严重威胁人类的慢性肺部疾病，如支气管扩张和囊性纤维化，以及 免疫功能低下的患者。然而，单克隆抗体与其他致病性 分枝杆菌此外，目前的动物模型存在局限性，这对免疫学研究构成了真实的挑战。 研究两种Mab形态型，光滑（S）和粗糙（R），在肺中的各自作用， 免疫病理学、持久性和宿主免疫应答。单抗在免疫活性和 甚至免疫功能低下的小鼠也阻止了慢性感染的研究，而斑马鱼胚胎则是 虽然它们是研究Mab感染的工具，但它们不能用于模拟肺部感染和T细胞 参与。因此，需要补充的动物模型。因为像斑马鱼胚胎一样，X.爪蟾 蝌蚪是透明的，可以看到病原体的传播，但除此之外，它们还有 功能正常的肺，并表现出免疫系统，包括与人类非常相似的T细胞，我们建议 发展出一个X。Mab感染的光滑蝌蚪模型，其模拟人类肺部疾病。援用 建立了比较生物学方法来研究蝌蚪对分枝杆菌的免疫力， 证明S和R Mab形态型都容易感染蝌蚪，在蝌蚪肺中传播， 持续长达50天，探索性研究提案的目的是在体内研究超过40 天单抗持久性，和免疫发病机制。具体地说，为了解决这个假设， 单克隆抗体在X.我们建议： 通过比较 使用活体内注射的方法，观察R和S Mab在肺和其他组织中的宿主免疫应答和持久性。 (1)Mab持续感染期间蝌蚪肺中免疫发病机制的特征 显微镜流式细胞术和转录组学以及荧光标记单抗和转基因蝌蚪 荧光巨噬细胞。 (2)研究体内S至R形态型转换与发病机制和持续性的相关性 得子茶杯. 使用启动子系统来控制感染后的S到R转换，并检查转换对 持久性、致病性和宿主免疫反应。 (3)研究单克隆抗体免疫发病机制的遗传决定因素 ，使用选定的Mab缺失 通过单克隆抗体特异性基因的等位基因交换产生的突变体以及革兰氏阴性菌共有的基因同源物， 阴性肺部病原体。 我们期待与人类相关的新见解将被收集。我们的合作团队是独一无二的 由于我们的背景和互补的专业知识，我们有能力进行这些研究。