Genetic Systems to Study Virulence in Bacteroides

描述（由申请方提供）：本研究将重点关注允许专性厌氧菌脆弱拟杆菌（虽然是正常结肠微生物群的组成部分）成为成功病原体的因素。这些包括在感染的早期阶段耐受有氧环境（空气耐受性）的能力;存在将血红素输入细胞的系统，用于血红素依赖性途径的中枢代谢和对活性氧的防御; B的能力。fragilis从宿主成分中去除唾液酸残基的能力，以及它在从复杂的寡糖和糖蛋白中获得体内生长所需的营养素方面的精湛技艺。具体目标包括：1、继续研究允许B的因素。fragilis承受长时间的氧气挑战（耐氧性）：我们建议，活动在B。fragilis周质作为抵抗活性氧（ROS）形成的初始防线，保护敏感靶标免受ROS攻击并逆转ROS损伤。此外，我们已经确定了特定的功能（超氧化物歧化酶，SOD），和广泛的基因簇（蝙蝠操纵子）所需的耐氧性。我们将测试的假设，蝙蝠操纵子形成一个多蛋白质复合物在细胞膜中发挥重要作用，从细胞质输出的还原电位的周质。2.铁和血红素的获得对B很重要。体外和体内脆弱细胞的生长。我们将研究血红素在B中的摄取过程。我们还将继续研究中心代谢双途径中的血红素依赖性和含Fe-S簇的酶，以确定它们在耐氧性和在氧挑战期间提供能量中的作用。3.研究操纵子的组成、功能和控制，以从受感染的宿主获得生长底物。我们将集中在操纵子含有神经氨酸酶（nanH 1）基因和其他几个糖水解酶能够转换复杂的刘易斯抗原发现在许多人类细胞表面的单个单糖。我们将继续分析NANA利用的操纵子，NanLET操纵子，并确定NanR结合的三个NanR阻遏启动子中的位点。我们将确定神经氨酸酶是否是一种毒力因子，因为它提供NANA生长，或因为它的活性改变宿主细胞的表面，或两者兼而有之。

DESCRIPTION (provided by applicant): This study will focus on factors that allow the obligate anaerobe Bacteroides fragilis, although a component of the normal colonic microbiota, to be a successful pathogen. These include its ability to withstand an aerobic environment (aero-tolerance) during early stages of infection; the presence of systems to import heme into the cell for the heme-dependent pathways of central metabolism and defense against reactive oxygen species; the ability of B. fragilis to remove sialic acid residues from host components, and its virtuosity in obtaining nutrients for growth in vivo from complex oligosaccharides and glycoproteins. Specific aims include: 1, to continue to study factors that allow B. fragilis to withstand prolonged oxygen challenge (aerotolerance): We propose that activities in the B. fragilis periplasm serve as the initial line of defense to combat the formation of reactive oxygen species (ROS), protect sensitive targets from ROS challenges and to reverse ROS damage. In addition we have identified specific functions (superoxide dismutase, SOD), and an extensive gene cluster (the Bat operon) that are required for aerotolerance. We will test the hypothesis that the Bat operon forms a multi-protein complex in the cell membrane that plays an important role in exporting reducing potential from the cytoplasm to the periplasm. 2. Acquisition of iron and heme is important for B. fragilis growth in vitro and in vivo. We will study the process of heme uptake in B. fragilis by the heme permease systems whose genes and functions we have described We will also continue to study the heme-dependent, and Fe-S cluster-containing enzymes in the dual pathways of central metabolism to establish their roles in aerotolerance and in providing energy during oxygen challenge. 3. to investigate the composition, functions and control of operons for the acquisition of growth substrates from the infected host. We will focus on the operon containing the neuraminidase (nanH1) gene and several other glycohydrolases capable of converting the complex Lewis antigen found on the surface of many human cells to individual monosaccharides. We will continue to analyze the operon for NANA utilization, the NanLET operon and to define the sites in the three NanR repressed promoters for NanR binding. We will determine if neuraminidase is a virulence factor because it supplies NANA for growth, or because its activity alters the surface of host cells, or both.