GENETICS OF ABSCESS FORMATION BY BACTEROIDS FRAGILIS

脆弱类杆菌脓肿形成的遗传学

• 批准号: 6124228
• 负责人: LAURIE E COMSTOCK
• 金额: $ 22.93万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-12-01 至 2001-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6124228
• 关键词: Bacteroides; bacterial capsules; bacterial cytopathogenic effect; bacterial genetics; bacterial polysaccharides; carbohydrate biosynthesis; carbohydrate structure; enzyme activity; gene deletion mutation; gene expression; host organism interaction; inflammation; laboratory rat; mutant; nuclear magnetic resonance spectroscopy; protein purification; reporter genes; virulence

DESCRIPTION (Adapted from the Applicant's Abstract): Bacteroides fragilis is the anaerobic pathogen most frequently isolated from intraabdominal sepsis. The capsular polysaccharides of B. fragilis have been identified as the primary virulence determinants for this organism and purified capsule is able to induce abscesses in animal models. Elucidation of the structures of the capsular polysaccharides of one strain has demonstrated that the polysaccharides contain a charge motif that confers the ability to cause abscesses. This charge motif is not commonly found in bacterial polysaccharides. The long term goal of this proposal is to understand the genetic factors responsible for the ability of the B. fragilis capsular polysaccharides to induce abscesses. The experiments outlined in the specific aim will study the genetic factors underlying the formation and expression of these virulence determinants. Preliminary data suggest that the product of one of the genes of the capsular biosynthesis locus may be responsible for the free amino group on the capsular polysaccharide. Because this group is essential for the abscess-inducing capabilities of the polysaccharide, the product of this gene will be studied by creating a mutant and studying the effect on abscess formation, and by purifying the product and assaying for enzymatic activity. The structure of the capsular polysaccharides have been determined for only one strain of B. fragilis. Studies will be conducted to determine the genetic heterogeneity of the polysaccharide biosynthesis loci for both clinical and fecal isolates, with an emphasis on the presence of genes that will enable the bacteria to produce a polysaccharide that is able to induce abscesses. Additionally, experiments will be performed that will address the regulation of expression of these capsular polysaccharides both in vitro and in vivo using transcriptional fusion reporter strains with the promoters that regulate transcription of the polysaccharide biosynthesis loci. The involvement of regulatory genes demonstrated to be involved in the expression of one of the capsular polysaccharides will be studied by creating in-frame deletion mutants of each of the genes. These will be the first studies to address the regulation of expression of B. fragilis polysaccharides and may explain why these bacteria are able to reside in the intestine as normal flora but produce disease when released to extraintestinal sites.

描述（改编自申请人的摘要）：拟杆菌 脆弱杆菌是最常分离到的厌氧病原体 腹腔内败血症。脆弱拟杆菌的荚膜多糖具有 被确定为该生物体的主要毒力决定因素 纯化的胶囊能够在动物模型中诱发脓肿。 阐明一种荚膜多糖的结构 菌株已证明多糖含有电荷基序 这赋予了引起脓肿的能力。这个充电主题不是 常见于细菌多糖。本次活动的长远目标 提议的目的是了解导致这一现象的遗传因素 脆弱拟杆菌荚膜多糖诱导脓肿的能力。 具体目标中概述的实验将研究遗传 这些毒力形成和表达的潜在因素 决定因素。初步数据表明，其中之一的产品 荚膜生物合成位点的基因可能负责游离 荚膜多糖上的氨基。因为这个群体是 对于多糖诱导脓肿的能力至关重要， 将通过创建突变体来研究该基因的产物 研究对脓肿形成的影响，并纯化产品 并测定酶活性。囊膜的结构 仅测定了一种脆弱拟杆菌的多糖。 将进行研究以确定遗传异质性 临床和粪便分离物的多糖生物合成位点， 强调使细菌能够生存的基因的存在 产生能够诱发脓肿的多糖。 此外，还将进行实验来解决 这些荚膜多糖的体外表达调节 并在体内使用转录融合报告菌株 调节多糖生物合成转录的启动子 基因座。调节基因的参与被证明参与其中 将研究其中一种荚膜多糖的表达 通过创建每个基因的框内缺失突变体。这些将 是第一个解决 B 表达调节问题的研究。 脆弱菌多糖，可能解释为什么这些细菌能够 作为正常菌群存在于肠道中，但当 释放到肠外部位。