Type VI Protein Secretion in an Emerging Multidrug-Resistant Pathogen

描述（由申请方提供）：多重耐药菌引起的医院获得性感染是免疫力低下患者发病率和死亡率增加的主要原因。这种感染的主要病原体是细菌鲍氏不动杆菌。这种微生物会导致包括肺炎和败血症在内的侵袭性感染，近年来这些感染变得越来越严重，越来越耐药，这让人们担心它们可能有一天无法治愈。预防和治疗A.鲍曼不动杆菌是迫切需要的，但令人震惊的是，人们对这些细菌如何引起疾病知之甚少。最近的证据表明，这些细菌诱导粘附依赖性细胞毒性对培养的上皮细胞;鲍曼不动杆菌在粘附时致毒细胞的作用尚不清楚。拟议研究的目标是了解这些微生物如何与人类细胞相互作用，以及这种相互作用如何破坏正常的细胞过程，促进感染。为了实现这一目标，本提案旨在详细描述接触依赖性蛋白质递送系统，称为VI型分泌系统（T6 SS），其存在于A.鲍曼不动杆菌导致了最近的流行病。这些系统在一系列微生物的宿主感染中发挥作用，但该系统在A.鲍曼不动杆菌。拟议的工作将确定A.鲍曼不动杆菌T6 SS，确定它发送到这些细胞的效应蛋白，并检查系统及其效应器在宿主相互作用和发病机制中的功能作用。这些目标将通过分析报告基因融合体向靶细胞的转移、对效应子进行定向全基因组筛选以及检查T6 SS和效应子缺失突变体在与宿主细胞相互作用期间对细菌毒性和宿主先天免疫反应的影响来实现。在培养物和小鼠肺炎模型中。这些研究将为A.鲍曼不动杆菌促进感染，并将解决我们对T6 SS病理生物学理解的关键差距。最终，这些信息将揭示新型抗感染治疗的目标 以及针对高抗药性细菌的疫苗。培训经验将拓宽我在分子生物学方面的专业知识，教我哺乳动物细胞培养和动物模型的新技术，并提高我对传染病发病机制的理解。 公共卫生相关性：鲍曼不动杆菌是侵袭性医院获得性感染的常见原因，近年来变得更具耐药性和侵袭性。拟议的研究旨在了解细菌如何与受感染宿主内的细胞相互作用，从而促进细菌生长和宿主细胞的失调。这些知识将揭示针对这些高度耐药微生物的新型抗感染治疗的潜在靶点。

DESCRIPTION (provided by applicant): Hospital-acquired infections due to multidrug-resistant bacteria are a major cause of increased morbidity and mortality in patients with weakened immunity. A leading agent of such infections is the bacterium Acinetobacter baumannii. This microbe causes invasive infections, including pneumonia and sepsis, that in recent years have become increasingly severe and antibiotic-resistant, raising the specter that they may one day be untreatable. New ways to prevent and treat infections with A. baumannii are urgently needed, but alarmingly little is known about how these bacteria cause disease. Recent evidence indicates that these bacteria induce adherence-dependent cytotoxicity toward cultured epithelial cells; however, the mechanisms used by A. baumannii to intoxicate cells upon adhesion are unclear. The goal of the proposed studies is to understand how these microbes interact with human cells and how such interactions undermine normal cellular processes, promoting infection. Towards this goal, this proposal aims to characterize in detail a contact-dependent protein delivery system, known as a Type VI Secretion System (T6SS) that is present in clinical strains of A. baumannii responsible for recent epidemics. These systems play a role in host infections by a range of microbes, but the system is undefined in A. baumannii. The proposed work will determine which cells are targeted by the A. baumannii T6SS, identify the effector proteins it sends into these cells, and examine the functional roles of the system and its effectors in host interactions and pathogenesis. These aims will be accomplished by analyzing the transfer of reporter fusions into target cells, conducting directed genome-wide screens for effectors, and examining the effects of T6SS- and effector-null mutants on bacterial toxicity and the host innate immune response during interactions with host cells in culture and in a murine model of pneumonia. These studies will break new ground on the mechanisms used by A. baumannii to promote infection and will address key gaps in our understanding of T6SS pathobiology. Ultimately, this information will uncover targets for novel anti-infective treatments and vaccines against highly drug- resistant bacteria. The training experience will broaden my expertise in molecular biology, teach me new techniques in mammalian cell culture and animal models, and advance my understanding of the pathogenesis of infectious diseases. PUBLIC HEALTH RELEVANCE: Acinetobacter baumannii, a frequent cause of invasive, hospital-acquired infections, has become more drug resistant and aggressive in recent years. The proposed research is aimed at understanding how the bacterium interacts with cells within the infected host in a way that promotes bacterial growth and misregulation of host cells. This knowledge will expose potential targets for novel anti-infective therapeutics against these highly resistant microbes.!