S. aureus virulence factor expression during kidney abscess formation

肾脓肿形成过程中金黄色葡萄球菌毒力因子的表达

• 批准号: 10370868
• 负责人: Kim Davis
• 金额: $ 24.56万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-18 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10370868
• 关键词: 3-Dimensional; Abscess; Antibiotic Resistance; Antibiotics; Bacteremia; Bacteria; Bacterial Adhesion; Bacterial Antibiotic Resistance; Bacterial Infections; Behavior; Binding; Biological Models; Blood Circulation; Cells; Clinical Trials; Communities; Cues; Diffuse; Disease; Disease Progression; Encapsulated; Fluorescence Microscopy; Future; Gene Expression Regulation; Generations; Genetic Transcription; Growth; Hospitals; Human; Immune; Immune response; Immune system; Immunofluorescence Microscopy; In Vitro; Individual; Infection; Kidney; Knowledge; Lesion; Lesion by Stage; Liver; Modeling; Molecular; Mus; Necrosis; Pathway interactions; Pattern; Phagocytes; Pharmacotherapy; Play; Population; Population Density; Process; Production; Proteins; Regulation; Reporter; Research; Role; Sepharose; Skin Tissue; Soft Tissue Infections; Staphylococcus aureus; Staphylococcus aureus infection; Structure; Surface; System; Systemic infection; Targeted Toxins; Testing; Therapeutic; Tissues; Toxin; United States; Vaccines; Virulence Factors; adaptive immune response; alternative treatment; antimicrobial; antimicrobial peptide; base; efficacious treatment; fluorescence imaging; human pathogen; immunoregulation; in vitro Model; in vivo; macrophage; methicillin resistant Staphylococcus aureus; mouse model; neutrophil; novel; pathogen; prevent; promoter; quorum sensing; recurrent infection; renal abscess; spatiotemporal; treatment strategy; vaccine efficacy; vaccine failure; vaccine platform; vaccine strategy

PROJECT SUMMARY Antibiotic-resistant bacterial infections are becoming increasingly more prevalent, and Staphylococcus aureus infections in particular have high rates of antibiotic resistance, prompting research into alternative treatment strategies. S. aureus vaccines have been developed, but all have failed in clinical trials to date, likely due to the ability of S. aureus to blunt the immune system and block protective immune responses. S. aureus produces many virulence factors that promote disease, including an arsenal of toxins, which directly target and kill host immune cells. Several S. aureus toxins specifically bind to human cells, and are not active in the mouse, rendering it difficult to dissect the specific role of toxins in the disease process. Few models exist to study interactions between S. aureus communities, the toxins they produce, and their human cell targets. S. aureus causes a wide range of disease manifestations in the human host, from skin and soft tissue infections to bacteremia and systemic spread to deep tissues. When S. aureus enters the bloodstream, bacteria are trapped in the liver, and then spread to the kidney to form large lesions called abscesses. Abscesses contain a central core of tightly clustered bacteria, with concentric layers of necrotic and live neutrophils, and an outer layer of macrophages. Because it is difficult to penetrate abscesses with antibiotics, they can persist follow drug treatment, and may represent one of the reservoirs responsible for recurrent infections. Better understanding of the interactions between bacteria within these structures, and the interactions with surrounding host cells, will be critical in developing future therapeutics to more efficiently eliminate these structures. This proposal will utilize fluorescent transcriptional reporters to determine the spatiotemporal expression patterns of S. aureus toxin expression within kidney abscesses, and will also develop an in vitro system to study host-pathogen interactions within abscesses. We hypothesize that direct interactions with neutrophils, and diffusible antimicrobials from macrophages, promotes expression of virulence factors specifically at the periphery of abscesses. We will utilize fluorescent reporter strains and immunofluorescence microscopy to determine whether toxin expression patterns change over the course of kidney abscess formation in our mouse model, and determine if this is dictated by the presence of neutrophils and macrophages. We will also develop an in vitro model of abscess formation using 3D agarose droplets to encapsulate bacteria and adhere host cells to the droplet surface. In this model, we will visualize the dynamics of virulence factor expression in the presence and absence of mouse and human primary phagocytes using live imaging fluorescence microscopy. Establishing these robust systems to study S. aureus spatial patterning and host-pathogen interactions will enable us to uncover key S. aureus vulnerabilities, which could be exploited to generate more efficacious treatments against this important human pathogen.

项目概要 抗生素耐药性细菌感染变得越来越普遍，葡萄球菌 金黄色葡萄球菌感染尤其具有很高的抗生素耐药性，促使人们研究替代品 治疗策略。金黄色葡萄球菌疫苗已经开发出来，但迄今为止所有疫苗都在临床试验中失败了，很可能 由于金黄色葡萄球菌能够削弱免疫系统并阻止保护性免疫反应。金黄色葡萄球菌 产生许多促进疾病的毒力因子，包括直接靶向和治疗的毒素库。 杀死宿主免疫细胞。几种金黄色葡萄球菌毒素与人体细胞特异性结合，并且在人体细胞中不活跃。 小鼠，使得很难剖析毒素在疾病过程中的具体作用。很少有模型存在 研究金黄色葡萄球菌群落、它们产生的毒素及其人类细胞靶标之间的相互作用。 金黄色葡萄球菌在人类宿主中引起多种疾病表现，包括皮肤和软组织 感染导致菌血症并全身扩散至深层组织。当金黄色葡萄球菌进入血液时， 细菌被困在肝脏中，然后扩散到肾脏，形成称为脓肿的大病变。 脓肿包含紧密聚集的细菌的中央核心，具有同心的坏死层和活细菌层 中性粒细胞和外层巨噬细胞。因为抗生素很难穿透脓肿， 它们可以在药物治疗后持续存在，并且可能是导致复发的储库之一 感染。更好地了解这些结构内细菌之间的相互作用，以及 与周围宿主细胞的相互作用，对于开发未来更有效的治疗方法至关重要 消除这些结构。 该提案将利用荧光转录报告基因来确定时空 肾脓肿内金黄色葡萄球菌毒素的表达模式，并且还将开发体外 研究脓肿内宿主与病原体相互作用的系统。我们假设与 中性粒细胞和巨噬细胞的扩散性抗菌剂促进毒力因子的表达 特别是在脓肿的周围。我们将利用荧光报告菌株和 免疫荧光显微镜以确定毒素表达模式是否在过程中发生变化 在我们的小鼠模型中形成肾脓肿，并确定这是否是由中性粒细胞的存在决定的 和巨噬细胞。我们还将使用 3D 琼脂糖液滴开发脓肿形成的体外模型 封装细菌并将宿主细胞粘附到液滴表面。在这个模型中，我们将可视化动态 使用小鼠和人类原代吞噬细胞存在和不存在时的毒力因子表达 实时成像荧光显微镜。建立这些强大的系统来研究金黄色葡萄球菌空间模式 宿主与病原体的相互作用将使我们能够发现金黄色葡萄球菌的关键漏洞，这可能是 用于针对这种重要的人类病原体产生更有效的治疗方法。