HP0318基因在幽门螺杆菌适应性定植中的作用研究

Role of HP0318 gene for adaptive colonization of H. pylori in Mongolian gerbils

目的通过构建幽门螺杆菌(Hp)HP0318缺失突变株,初步探明该基因在Hp适应性定植中的作用。方法通过基因克隆和重组技术在HP0318基因上下游片段之间连入氯霉素抗性基因(catGC)作为筛选标记,并克隆到质粒pBluescript SK中构建成缺失突变的打靶载体,将其转化沙鼠适应株M13,经过同源重组,对出发菌的HP0318基因进行置换突变,然后用经氯霉素抗性筛选的突变菌株感染蒙古沙鼠,观察其与未突变的M13出发菌在定植能力和致病性上的差异。结果经PCR鉴定成功构建1株缺失HP0318基因的Hp菌株。突变株与非突变株对蒙古沙鼠的感染率相差显著(P<0.05),2个菌株在胃内的定植密度有显著性差异(P<0.05),突变株定植密度有约5倍的减少。结论HP0318基因在Hp的沙鼠适应性定植过程中发挥了重要的作用。

Objective To construct and identify a deficiency mutant of H. pylori strains without HP0318 gene. Methods Two cloning PCR fragments containing two-end regions of HP0318 and a selectable chloramphenicol resistance marker between them were engineered into pBluescript plasmid. Then the HP0318-deleted derivative plasmid was electrotransformed into adaptive strain MI3 and constructed a HP0318-negative mutant of H. pylori via homologous recombination resulting in allelic exchange. M13 and the mutant were inoculated to Mongolian gerbils, respectively. The level of colonization and pathological change were assessed at 4 weeks after infection. Results An HP0318 negative mutant of H. pylori strain has been constructed successfully. The in- fectious rate and colonization levels of the HP0318 mutant in Mongolian gerbils were significantly lower as compared with M,3 （P 〈 0.05 ）. Conclusion These results demonstrate the importance of HP0318 gene for adaptive colonization of H. pylori in Mongolian gerbils.