Virulence Determinants in Helicobacter hepaticus

描述（由申请人提供）： H.小鼠肝脏中的肝球菌与肝炎和肝癌的发展相关，这些症状可以通过用该细菌接种小鼠来研究。这项工作的具体目标是开始鉴定一些可能是H.通过靶向诱变方法对Hepaticus进行诱变。模式菌株的全基因组序列预计将在2003年的某个时候获得，一些可疑的毒力因子（超氧化物歧化酶，烷基过氧化氢还原酶和氢化酶）已经通过H.肝提取物（主要研究者实验室）。上述三种酶已被证明影响相关细菌幽门螺杆菌（H. pylori），通过本文提出的相同方法（靶向诱变）。在全基因组序列发布之前，主要研究者可获得三种基因（sodB、ahpC和hydA）的序列，以便在这三种潜在毒力因子中产生靶向突变体。然而，突变体H.从未在任何基因中发现过肝细胞。因此，需要一个初步的试验研究，以确定这种方法的可行性，使用卡那霉素抗性盒alMic交换诱变（目前用于H。pylori）。一旦产生突变体，将其与亲本菌株进行小鼠定殖和感染比较，并由委员会认证的兽医病理学家评估所得肝组织的疾病。如果能产生特异性突变体，那么毒性决定因子就可以开始被系统地研究。

DESCRIPTION (provided by applicant): The presence of H. hepaticus in the liver of mice is correlated with the development of hepatitis and liver carcinomas, and these symptoms can be studied by inoculating mice with the bacterium. The specific aim of the proposed work is to begin to identify some of the bacterial factors that may be virulence components in H. hepaticus via a targeted mutagenesis approach. A complete genome sequence for the type strain is expected to be available sometime in 2003, and some suspected virulence factors (superoxide dismutase, alkyl hydroperoxide reductase, and hydrogenase) have been identified by enzyme assays on H. hepaticus extracts (in the Principal Investigator's lab). The above three enzymes have been shown to affect virulence in the related bacterium Helicobacter pylori (H. pylori), by the same approach (targeted mutagenesis) proposed here. The sequences for the three genes (sodB, ahpC, and hydA) are being made available to the Principal Investigator prior to the full genome sequence release so that targeted mutants can be generated in these three potential virulence factors. However, a mutant H. hepaticus has never been recovered in any gene. Therefore, an initial pilot study is needed to ascertain the feasibility of this approach, using kanamycin-resistance cassette alMic exchange mutagenesis (the method that is currently used for H. pylori). Once mutants are made they will be compared to the parent strain for colonization and infections of mice, and the resulting liver tissue will be assessed for disease by a board certified veterinary pathologist. If specific mutants can be created, then virulence determinants can begin to be systematically studied.