Chiang S L, Mekalanos J J
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115, USA.
Mol Microbiol. 1998 Feb;27(4):797-805. doi: 10.1046/j.1365-2958.1998.00726.x.
The pathogenesis of cholera begins with colonization of the host intestine by Vibrio cholerae. The toxin co-regulated pilus (TCP), a fimbrial structure produced by V. cholerae, is absolutely required for colonization (i.e. the persistence, survival and growth of V. cholerae in the upper intestinal milieu), but many other aspects of the colonization process are not well understood. In this study, we use signature-tagged transposon mutagenesis (STM) to conduct a screen for random insertion mutations that affect colonization in the suckling mouse model for cholera. Of approximately 1100 mutants screened, five mutants (approximately 0.5%) with transposon insertions in TCP biogenesis genes were isolated, validating the use of STM to identify attenuated mutants. Insertions in lipopolysaccharide, biotin and purine biosynthetic genes were also found to cause colonization defects. Similar results were observed for mutations in homologues of pta and ptfA, two genes involved in phosphate transfer. Finally, our screen identified several novel genes, disruption of which also caused colonization defects in the mouse model. These results demonstrate that STM is a powerful method for isolating colonization-defective mutants of V. cholerae.
霍乱的发病机制始于霍乱弧菌在宿主肠道的定殖。毒素协同调节菌毛（TCP）是霍乱弧菌产生的一种菌毛结构，是定殖（即霍乱弧菌在上消化道环境中的持续存在、存活和生长）所绝对必需的，但定殖过程的许多其他方面尚未得到很好的理解。在本研究中，我们使用签名标签转座子诱变（STM）对影响霍乱乳鼠模型定殖的随机插入突变进行筛选。在大约1100个筛选的突变体中，分离出了5个在TCP生物合成基因中有转座子插入的突变体（约0.5%），验证了使用STM来鉴定减毒突变体的有效性。还发现脂多糖、生物素和嘌呤生物合成基因中的插入会导致定殖缺陷。参与磷酸转移的两个基因pta和ptfA的同源物发生突变也观察到了类似结果。最后，我们的筛选鉴定出了几个新基因，其破坏也会导致小鼠模型中的定殖缺陷。这些结果表明，STM是分离霍乱弧菌定殖缺陷突变体的一种有效方法。
