The role of cytochrome bd in uropathogenic Escherichia coli pathogenesis

项目摘要 生物膜是涉及大多数细菌感染的多细胞细菌群落，并且几乎所有细菌感染都是由生物膜引起的。 慢性细菌感染这些社区几乎不可能通过传统的 化疗方法，并代表对人类健康的主要威胁。生物膜细菌分泌一种 细胞外基质（ECM），限制吞噬细胞，补体，抗生素和其他外部 应激源与生物膜细菌相互作用。因此，生物膜对抗生素具有高度抗性， 免疫系统，通常需要生物膜相关感染的患者接受长期 抑制性抗生素或接受手术切除感染组织。除了抑制 由于抗生素和免疫防御，ECM限制了营养物质（如氧气）的扩散， 与常驻细菌的代谢活动，建立氧梯度内的生物膜，使 生物膜内部缺氧。几项研究已经证明，氧梯度在细胞生长中起关键作用。 弹性生物膜社区的发展，以及在生物膜氧的可用性是一个中央调节器， 细菌代谢和ECM组分的表达。尿路致病性大肠埃希菌的前期研究 （UPEC），尿路感染的主要原因，已证明，尽管是一个兼性的， UPEC是厌氧生物，UPEC在感染期间依赖于有氧呼吸并形成生物膜群落。通过我 在论文研究中，我已经表明，UPEC不均匀地表达呼吸酶，并且这些酶 酶在离散的亚群中表达。尽管这种表达的异质性，只有表达 细胞色素bd是一种在缺氧条件下有氧呼吸所必需的高亲和力醌醇氧化酶， 是UPEC发病机制和生物膜形成所必需的。细胞色素bd的损失，但不是其他醌醇氧化酶， 破坏生物膜的发育，改变ECM的产生，增加对抗生素的敏感性， 在小鼠感染模型中的毒力。该提案概述了一系列实验，这些实验将确定 细胞色素bd在尿路感染发病机制中的作用及UPEC形成生物膜的能力 社区能够承受抗生素治疗和免疫攻击。完成本提案将 确定限制细胞色素bd缺陷型UPEC定植的瓶颈， 细胞色素bd在尿路感染的细胞内阶段，并确定机制， 细胞色素BD促进抗生素耐受性生物膜的形成。这些研究将产生基本的 深入了解中央代谢过程的适应如何使细菌适应不同的宿主生态位， 建立弹性生物膜社区，同时还研究细胞色素bd作为潜在的药物靶点， 预防或根除生物膜相关感染。

PROJECT SUMMARY Biofilms are multicellular bacterial communities implicated in the majority of bacterial infections, and nearly all chronic bacterial infections. These communities are nearly impossible to eradicate by traditional chemotherapeutic approaches and represent a major threat to human health. Biofilm bacteria secrete an extracellular matrix (ECM) that limits the ability for phagocytes, complement, antibiotics, and other external stressors to interact with biofilm bacteria. Accordingly, biofilms are highly resistant to antibiotics and the immune system, often necessitating that patients with biofilm-associated infections receive long-term suppressive antibiotics or undergo surgery to remove infected tissues. In addition to inhibiting penetrance of antibiotics and immune defenses, the ECM limits diffusion of nutrients such as oxygen, which, in conjunction with the metabolic activity of resident bacteria, establishes oxygen gradients within biofilms that render the interior of biofilms hypoxic. Several studies have demonstrated that oxygen gradients play a critical role in the development of resilient biofilm communities, and that in biofilms oxygen availability is a central regulator of bacterial metabolism and expression of ECM components. Previous work in uropathogenic Escherichia coli (UPEC), the primary cause of urinary tract infections, has demonstrated that despite being a facultative anaerobe, UPEC relies on aerobic respiration during infection and to form biofilm communities. Through my thesis research, I have shown that UPEC heterogeneously expresses respiratory enzymes, and that these enzymes are expressed in discrete subpopulations. Despite this heterogeneity of expression, only expression of cytochrome bd, a high affinity quinol oxidase necessary for aerobic respiration under hypoxic conditions, is required for UPEC pathogenesis and biofilm formation. Loss of cytochrome bd, but not other quinol oxidases, disrupts biofilm development, alters ECM production, increases susceptibility to antibiotics, and impairs virulence in a murine model of infection. This proposal outlines a series of experiments which will define the role of cytochrome bd in urinary tract infection pathogenesis and the ability for UPEC to form biofilm communities capable of withstanding antibiotic therapy and immune assault. Completion of this proposal will identify bottlenecks that restricts colonization by cytochrome bd deficient UPEC, biochemically define the role of cytochrome bd in the intracellular phase of urinary tract infection, and define mechanisms by which cytochrome bd promotes the formation of antibiotic tolerant biofilms. These studies will yield fundamental insights into how adaptation of central metabolic processes allows bacteria to adapt to diverse host niches and establish resilient biofilm communities, while also investigating cytochrome bd as a potential drug target to aid in the prevention or eradication of biofilm-associated infection.