Subproject 4: Role of Pumps in Resistance, Physiology, and Infection

摘要 该项目的长期目标是确定葡萄球菌外排泵的完整阵列 金黄色葡萄球菌和肠球菌在多重耐药中的作用及阐明 它们表达的决定因素、在微生物生理学中的作用和对细菌的影响 感染时对抗菌药的反应。工作重点将放在结构和功能分析上 Tet38外排泵可实现多药耐药、在脓肿中存活和侵袭 在上皮细胞中，多个泵对周围细胞亚群的潜在贡献 抗菌素的存在，扩大了对肠球菌外排耐药的理解，以及 协作性评价新型抗菌化合物对外排、组织的影响 耐受性和体内疗效。有四个具体目标：1)评估结构性决定因素 金黄色葡萄球菌Tet38外排泵的阻力、生理和致病机制的研究 对细胞生理的不同影响机制；2)探索促进细胞生长的条件 和金黄色葡萄球菌外排泵在单细胞组中表达的异质性；3)评估其作用 肠球菌耐药情况；4)评价金黄色葡萄球菌外排泵的效果 来自合作项目的新化合物。这项工作将利用基因操作，等位基因 在金黄色葡萄球菌和粪肠球菌中的交换和表达载体，基因/蛋白的测定 用荧光记者表达，并建立了小鼠感染模型(皮下 脓肿，肠道定植)利用基因组定义的菌株。该计划的总体目标 计划项目是采用综合的、多学科的方法来理解抗生素 细胞生理学中的耐药发展和外排结合研究 通过抑制新的靶点和途径来危害抗药性病原体的化合物。这 该项目将增加对多药外排泵相关耐药机制的了解 提供菌株，用于测试这种泵对新化合物的影响 目标和路径。它还将利用哺乳动物的感染模型来筛选新的化合物 耐受性和体内活性。

Summary The longterm goals of the project are to identify the full array of efflux pumps of Staphylococcus aureus and Enterococcus spp that contribute to multiple antimicrobial resistance and to elucidate the determinants of their expression, their role in microbial physiology and their effect on bacterial response to antimicrobials in infection. The work will focus on a structural and functional analysis of the Tet38 efflux pump that enables multidrug resistance, survival in abscesses, and invasion of epithelial cells, the potential contribution of multiple pumps to persister subpopulations in the presence of antimicrobials, expanding the understanding of efflux resistance in enterococci, and evaluating collaboratively novel novel antimicrobial compounds for effects of efflux, tissue tolerability, and in vivo efficacy. There are four specific aims: 1) assess the structural determinants of the S. aureus Tet38 efflux pump in resistance, physiology, and pathogenesis and determine the mechanism of its diverse effects on cellular physiology; 2) explore the conditions promoting and heterogeneity of S. aureus efflux pump expression in single-cell groups; 3) evaluate the role of effux pumps in resistance in enterococci; and 4) evaluate the effect of S. aureus efflux pumps on novel compounds from collaborating projects. The work will utilize genetic manipulation, allelic exchange, and expression vectors in S. aureus and E. faecalis, measurements of gene/protein expression with fluorescent reporters, and established murine models of infection (subcutaneous abscess, intestinal colonization) utilizing a genomically defined strains. The overall goal of the program project is to take a well‐integrated, multi‐disciplinary approach to understanding antibiotic resistance development and efflux within cellular physiology combined with the search for compounds that compromise resistant pathogens by inhibiting novel targets and pathways. This project will add to understanding of resistance mechanisms related to multidrug efflux pumps and provide strains for testing the effect of such pumps on novel compounds active against new targets and pathways. It will also utilize a mammalian model of infection to screen new compound tolerability and in vivo activity.