The role of anaerobic microbiota in cystic fibrosis airway disease trajectory

项目概要/摘要 尽管最近大量的研究报告了厌氧微生物的流行和丰富， 囊性纤维化（CF）个体气道中的微生物群，它们对 疾病的病理生理学仍然是有争议的和知之甚少。这种知识差距是由 部分原因是流行病学和体外实验数据相互矛盾，缺乏合适的模型系统进行研究 它们与宿主的相互作用，以及缺乏宿主相关厌氧微生物的实验室专业知识， 微生物学.对厌氧菌在CF肺部感染中的作用产生更深入了解的研究， 特别是在早期阶段，有必要确定是否，何时以及如何针对这组生物体 治疗上 我们的研究小组已经表明：（i）厌氧菌是儿童CF气道的早期定植者， 典型病原体的获得，（ii）厌氧菌细菌群落引起促炎反应 当与支气管上皮细胞共培养时，和（iii）厌氧菌介导的粘蛋白降解和交叉- 进食可影响典型CF病原体的生长、毒力和抗生素敏感性。建筑 基于这些数据，本项目建议使用多学科方法来调查个体 厌氧菌彼此、气道上皮和传统病原体（例如，绿脓杆菌， S.金黄色葡萄球菌）在早期CF气道疾病的背景下。首先，使用实验微生物组模型 系统组成的10 CF厌氧物种，我们将确定如何CF细菌群落组装 以及它们的组装动力学是如何被宿主和微生物过程塑造的。第二，使用 创新的双重缺氧-缺氧系统，可实现厌氧菌和气道上皮细胞的共培养， 我们将使用单细胞RNA测序来确定厌氧菌如何与气道上皮相互作用， 以及宿主的反应如何因上皮细胞类型而异。最后，考虑到铜绿假单胞菌、S.金黄色 其他需氧病原体与厌氧微生物群共定植， 年龄，我们将使用体外实验进化方法来测试早期CF微生物群是否 足以驱动典型病原体适应CF气道的营养环境。 在完成这些目标时，我们将确定单个厌氧菌如何与一个厌氧菌相互作用。 另一种是宿主和传统的CF病原体。这样做，这些研究将产生有意义的影响， 我们对气道微生物群在慢性气道疾病中的作用的机制理解， 推动未来针对特定厌氧菌的研究，甚至促进其生长，作为一种手段， 改善临床结果。最后，我们使用CF作为我们的模型系统，但这项工作是广泛适用的 其他急性和气道感染（例如，COPD、鼻窦炎、呼吸机相关性肺炎）， 厌氧微生物也有牵连，但它们的作用机理仍不明确。

PROJECT SUMMARY / ABSTRACT Despite the recent surge in studies reporting on the prevalence and abundance of anaerobic microbiota in the airways of individuals with cystic fibrosis (CF), their mechanistic contributions, if any, to disease pathophysiology remain controversial and poorly understood. This knowledge gap is driven, in part, by conflicting epidemiological and in vitro experimental data, a lack of suitable model systems to study their interactions with the host, and the dearth of laboratory expertise in host-associated anaerobic microbiology. Studies that generate greater insight into the role of anaerobes in CF lung infection, particularly at early stages, are necessary to determine if, when, and how to target this group of organisms therapeutically. Our team has shown that (i) anaerobes are early colonizers of the pediatric CF airways prior to the acquisition of canonical pathogens, (ii) anaerobe bacterial communities elicit pro-inflammatory response when co-cultured with bronchial epithelial cells, and (iii) anaerobe-mediated mucin degradation and cross- feeding can shape the growth, virulence, and antibiotic susceptibility of canonical CF pathogens. Building on these data, this project proposes to use a multi-disciplinary approach to investigate how individual anaerobes interact with one another, the airway epithelium, and traditional pathogens (e.g., P. aeruginosa, S. aureus) in the context of early CF airway disease. First, using an experimental microbiome model system composed of 10 CF anaerobic species, we will determine how CF bacterial communities assemble and how their assembly dynamics are shaped by host and microbial processes. Second, using an innovative dual oxic-anoxic system that enables co-culture of anaerobic bacteria and airway epithelial cells, we will use single-cell RNA sequencing to determine how anaerobes interact with the airway epithelium, and how the host response varies by epithelial cell type. Finally, given that P. aeruginosa, S. aureus, and other aerobic pathogens co-colonize with anaerobic microbiota and rapidly adapt to the airways as patients age, we will use an in vitro experimental evolution approach to test whether early CF microbiota are sufficient to drive the adaptation of canonical pathogens to the nutritional milieu of the CF airways. At the completion of these aims, we will have defined how individual anaerobes interact with one another, the host, and traditional CF pathogens. In doing so, these studies will have a meaningful impact on our mechanistic understanding of the role of airway microbiota in chronic airway disease and may motivate future studies targeting specific anaerobic bacteria, or even promoting their growth, as a means of improving clinical outcomes. Finally, we use CF as our model system, but this work is broadly applicable to other acute and airway infections (e.g., COPD, sinusitis, ventilator associated pneumonias) in which anaerobes have been implicated but their mechanistic contributions remain undefined.