GENETIC SYSTEMS TO STUDY VIRULENCE IN B FRAGILIS

研究脆弱拟杆菌毒力的遗传系统

• 批准号: 2060940
• 负责人: MICHAEL H MALAMY
• 金额: $ 23.25万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-01-01 至 1998-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2060940
• 关键词: Bacteroides; CHO cells; anaerobiosis; bacterial genetics; bacterial proteins; electron microscopy; enzyme activity; exo alpha sialidase; gene expression; genetic regulation; histology; laboratory rat; structural genes; superoxide dismutase; transposon /insertion element; virulence

DESCRIPTION (Adapted from applicant's abstract): 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, the principal investigator's approach has been to develop new genetic systems for use in B. fragilis in order to identify potential virulence determinants. The investigator is focusing on the aerotolerance of B. fragilis as an important factor in the organisms' ability to survive in oxygenated tissue. He will analyze a set of B. fragilis proteins that are highly expressed only in the presence of oxygen. He will study the control of the synthesis of these oxygen response proteins (ORPs) and attempt to identify global and gene specific regulators. He 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. He will continue his studies on the superoxide dismutase of B. fragilis and determine its role in B. fragilis survival in vitro and in vivo. Since the investigator has determined an important role for the B. fragilis neuraminidase (nanH)in the ability of B. fragilis to grow in two in vivo test systems, he will concentrate on defining the control region of the nanH structural gene and in understanding how its expression is regulated. He 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, he 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. He 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, he will perform a histological analysis of the infected pouch at early times after infection. He will determine the location of the infecting B. fragilis cells during the establishment of the anaerobic environment that is required for bacterial growth.

描述（根据申请人的摘要改编）：长期目标 该提议的内容是了解导致的因素 细菌片的毒力和致病性，是最重要的 化脓性腹内和骨盆感染中的厌氧病原体 人类。 由于这需要对 毒力因素，主要研究者的方法是 开发新的遗传系统，以识别脆弱的B. fragilis 潜在的毒力决定因素。 研究人员将重点放在脆弱的B. fragilis的空气上 生物体在氧化组织中生存的能力的重要因素。 他将分析一组高度表达的脆弱蛋白 仅在存在氧气的情况下。 他将研究合成的控制 在这些氧反应蛋白（ORP）中，并试图识别全球和 基因特异性调节剂。 他还将研究关闭的过程 通常在厌氧菌中生产的大多数脆弱蛋白质蛋白 当生长细胞被氧气挑战时。 他将继续他的 关于脆弱芽孢杆菌的超氧化物歧化酶的研究，并确定其作用 在脆弱的B. fragilis生存中，体外和体内。 由于研究人员已经确定了脆弱的。 神经氨酸酶（NANH）具有脆弱芽孢杆菌在两个体内生长的能力 测试系统，他将集中精力定义 NANH结构基因并了解其表达的调节方式。 他将使用已在E中使用的Tnphoa系统的改编。 大肠杆菌鉴定出口蛋白，以研究 B. Fragilis。 由于许多细胞包膜蛋白在细菌中起作用 致病性，他将使用TNCEF识别毒力决定因素 B.脆弱的表面或导出的蛋白质。 变化的突变体 潜在的毒力因子将测试其在两个中生长的能力 脆弱芽孢杆菌感染的模型系统，大鼠肉芽瘤和CHO或CHO或 Monika P15培养物中的动物细胞。 他将搜索仅在 细菌在体内生长。 这些基因可能发挥重要作用 在脆弱芽孢杆菌感染的建立和维持中的作用。 进一步定义大鼠脆弱的B. fragilis感染的初始步骤 小袋，他将对受感染的小袋进行组织学分析 感染后的早期。 他将确定 在厌氧的建立过程中感染脆弱的细胞 细菌生长所需的环境。