Streptococcal virulence factors in extra-oral disease

感染性心内膜炎每年影响约22，000人， 杀死了2000多人。这种疾病主要是由草绿色细菌引起的 在草绿色链球菌中，主要是链球菌 sanguis。以往关于链球菌对人的毒力的研究 心内膜炎的诊断主要集中在对可疑毒力因子的检查上。 拟议的研究采取了一种完全不同的方法，即直接 鉴定新的毒力因子。这将通过调整技术来实现 的签名标记诱变（STM）用于与S. sanguis。这种方法 不需要事先假设任何特定基因的重要性 或致病的活动。相反，它依赖于感染过程 以确定疾病所需的细菌基因及其相关基因， 捐款.色葡萄这也是一个单独的主题， 正在进行的基因组测序项目将使用STM随机诱变 体外转座方法。将合并产生的突变体， 接种到作为心内膜炎模型的大鼠中。STM方法将 允许识别库中具有增加或 毒性降低。这些突变体的毒力明显改变， 通过体内竞争测定验证。突变体的进一步表征 将包括DNA序列分析，以确定突变的基因，进一步 遗传操作，以验证突变基因的贡献，在体内 感染性测定和体外功能测定。预计该 对具有改变的毒力的突变体的表征将导致更好的 了解心内膜炎的发病机制，并将确定有前途的 用于疫苗或治疗开发的新靶点。

Infective endocarditis affects about 22,000 people per year in the U.S. and kills over 2,000. This disease is caused primarily by viridans streptococci, and among viridans streptococci, primarily by Streptococcus sanguis. Previous studies concerning the virulence of streptococci for endocarditis have centered upon the examination of suspected virulence factors. The proposed research takes an entirely different approach, that of directly identifying new virulence factors. This will be done by adapting the technique of signature-tagged mutagenesis (STM) for use with S. sanguis. This approach requires no prior assumptions concerning the importance of any particular gene or activity in disease causation. Instead, it relies on the infection process to identify the bacterial genes needed for disease and their relative contributions. An S. sanguis strain that is also the subject of a separate ongoing genome sequencing project will be randomly mutagenized by STM using an in vitro transposition approach. The resulting mutants will be pooled and inoculated into rats serving as models for endocarditis. The STM approach will allow for the identification of mutants within the pool possessing increased or decreased virulence. The apparent altered virulence in these mutants will be verified by in vivo competition assays. Further characterization of mutants will include DNA sequence analysis to identify the mutated genes, further genetic manipulation to verify the contribution of the mutated genes, in vivo infectivity assays, and in vitro functional assays. It is anticipated that the characterization of mutants with altered virulence will result in a better understanding of the pathogenesis of endocarditis and will identify promising novel targets for vaccine or therapeutic development.