BASIS OF DETERMINANT RECOGNITION BY THE IMMUNE SYSTEM

本提案的长期目标是了解 脆弱拟杆菌的致病性和毒力， 最重要的厌氧病原体在化脓性腹腔内， 人类的盆腔感染因为这需要基因和 生理分析的毒力因子，我们的方法已经 开发新的遗传系统用于B。fragilis以识别 潜在的毒力决定因素 我们关注的是B的空气动力学耐受性。脆弱性作为一个重要因素 生物体在含氧组织中生存的能力。我们将分析 一套B。fragilis蛋白仅在细胞中高度表达， 氧气的存在。我们将研究这些合成的控制 氧反应蛋白（ORP），并试图确定全球和基因 具体的监管机构。我们还将研究关闭 大多数B。在厌氧条件下产生的脆弱菌蛋白， 当生长中的细胞受到氧气的挑战时。我们将继续 B超氧化物歧化酶的研究fragilis并确定其作用 脆弱B.fragilis在体外和体内存活。 既然我们已经确定脆弱B杆菌在 神经氨酸酶对脆弱B.fragilis生长能力的影响 系统，我们将集中精力定义nanH的控制区域 结构基因，并了解其表达是如何调节的。 我们将使用一个改编的TnphoA系统，该系统已用于 大肠杆菌鉴定输出蛋白，以研究表面蛋白 的B。脆弱的由于许多细胞被膜蛋白在细菌中起作用， 致病性，我们将使用TnCef来确定 B。fragilis表面或输出的蛋白质。突变体， 潜在的毒力因子将被测试它们在 B的两个模型系统。fragilis感染，大鼠肉芽肿袋，和 CHO或Monika P15动物细胞培养。 我们去找B。fragilis基因，只有在 细菌在体内生长。这些基因很可能在 在建立和维持B中的作用。脆弱菌感染 进一步明确脆弱B杆菌感染大鼠的初始步骤 袋，我们将进行组织学分析的感染袋， 感染后早期。我们将确定 感染B。fragilis细胞在建立厌氧 这是细菌生长所需的环境。

The long term objective of this proposal is to understand the factors that contribute to the virulence and pathogenicity of Bacteroides fragilis, the most important anaerobic pathogen in suppurative intra-abdominal and pelvic infections in human beings. Since this requires a genetic and physiological analysis of the virulence factors, our approach has been to develop new genetic systems for use in B. fragilis in order to identify potential virulence determinants. We are focusing on the aerotolerance of B. fragilis as an important factor in the organisms' ability to survive in oxygenated tissue. We will analyze a set of B. fragilis proteins that are highly expressed only in the presence of oxygen. We will study the control of the synthesis of these oxygen response proteins (ORPs) and attempt to identify global and gene specific regulators. We will also study the process of the shut-off of most B. fragilis proteins normally made during anaerobiosis that occurs when the growing cells are challenged with oxygen. We will continue our studies on the superoxide dismutase of B. fragilis and determine its role in B.fragilis survival in vitro and in vivo. Since we have determined an important role for the B.fragilis neuraminidase in the ability of B.fragilis to grow in two in vivo test systems, we will concentrate on defining the control region of the nanH structural gene and in understanding how its expression is regulated. We will use an adaptation of the TnphoA system that has been used in E.coli to identify exported proteins, in order to study surface proteins of B. fragilis. Since many cell envelope proteins play a role in bacterial pathogenicity, we will use TnCef to identify virulence determinants among B. fragilis surface or exported proteins. Mutants with alterations in potential virulence factors will be tested for their ability to grow in two model systems of B. fragilis infection, the rat granuloma pouch, and CHO or Monika P15 animal cells in culture. We will search for B. fragilis genes that are only expressed when the bacteria are growing in vivo. These genes are likely to play an important role in the establishment and maintenance of B. fragilis infections. To further define the initial steps in B.fragilis infection of the rat pouch, we will perform a histological analysis of the infected pouch at early times after infection. We will determine the location of the infecting B. fragilis cells during the establishment of the anaerobic environment that is required for bacterial growth.