The Impact of the Cystic Fibrosis infection environment on biofilm development of nontuberculous mycobacteria

项目摘要/摘要--DePas 非结核分枝杆菌(NTM)作为危险的、耐药的肺病的出现 病原体的研究速度超过了对其致病机制的研究。我们的长期目标是将 NTM在感染环境中的作用，并开发针对体内特定细菌的新治疗方法 活动。本建议的目的是直接评估NTM生物膜的形成和生长速度，并在原位和 确定感染环境如何影响这些过程。我们假设空间和 囊性纤维化患者痰中的化学环境对其形成的支持 耐抗生素、生长缓慢的NTM生物膜通过调节的细胞过程。这样做的理由是 提出了一种特定细菌病原体的生物被膜状态和生长速度可以对 抗生素的疗效和宿主的免疫反应。我们将用两个具体的例子来检验我们的中心假设 目的：1)确定碳纤维化学环境如何调节NTM生物膜的形成以及它是如何 影响抗生素耐受性；2)确定缺氧休眠如何影响生理耐受性 和NTM的生物被膜形成。拟议的工作将结合三项创新的互补技术 形成一个连贯的策略，用于研究感染相关的表型，如生物膜的形成和 休眠。我们将在这两个目标中使用一种新的组织清除/细菌可视化技术MiPACT-HCR。 在目标2中，我们还将利用CF感染环境的3D模型，琼脂块生物膜分析。我们会 使用一种新的体外聚集试验，使我们能够跟踪和量化从浮游细胞到 这两个目标都有生物膜。这项提议意义重大，因为它将准确地描述 人类感染期间NTM的生理状态，以指导抗生素的选择和剂量决定。它也是 重要的是，它将为开发抗生物膜和抗休眠策略提供分子靶点 对NTM的指控。这项工作的预期结果是彻底了解结构和流行率 CF患者感染过程中NTM群落的变化及机制探讨 生物膜的形成和休眠。这些结果将产生积极的影响，帮助医生做出更多 针对NTM感染的适当治疗选择。

Project Summary/Abstract -- DePas The emergence of nontuberculous mycobacteria (NTM) as dangerous, antibiotic resistant pulmonary pathogens is outpacing research into their mechanisms of pathogenesis. Our long-term goal is to characterize NTM in the infection environment and develop new therapeutic approaches aimed at specific in vivo bacterial activities. The objective of this proposal is to directly assess NTM biofilm formation and growth rate in situ and determine how the infection environment impacts these processes. We hypothesize that the spatial and chemical environment of sputum from people with Cystic Fibrosis (CF) sputum supports the formation of antibiotic tolerant, slow-growing NTM biofilms through regulated cellular processes. The rationale for this proposal is that the biofilm state and growth rate of a specific bacterial pathogen can have drastic influences on the efficacy of antibiotics and the host immune response. We will test our central hypothesis with two specific aims: 1) Determine how NTM biofilm formation is regulated by the CF chemical environment and how it impacts antibiotic tolerance and 2) Determine how anoxia-induced dormancy influences physiological tolerance and biofilm formation of NTM. The proposed work will combine three innovative complementary techniques into one coherent strategy for investigating infection-relevant phenotypes such as biofilm formation and dormancy. We will employ a novel tissue clearing/bacterial visualization technique MiPACT-HCR in both aims. In Aim 2, we will also utilize a 3D model of the CF infection environment, the Agar Block Biofilm Assay. We will use a new in vitro aggregation assay that allows us to track and quantify the transition from planktonic cells to biofilms in both Aims. The proposal is significant because it will provide an accurate description of the physiological state of NTM during human infection to inform antibiotic choice and dosage decisions. It is also significant in that it will provide molecular targets for development of anti-biofilm and anti-dormancy strategies against NTM. The expected outcome of this work is a thorough understanding of the structure and prevalence of NTM communities during infection of patients with CF and insight into the mechanistic pathways driving biofilm formation and dormancy. These results will have a positive impact by assisting physicians make more appropriate treatment choices for NTM infections.