一种新的幽门螺杆菌克拉霉素耐药相关基因

A novel gene associated with ciarithromycin resistance of Helicobacter pylori

目的:分析不同克拉霉素浓度下H pylori菌株耐药的发生机制,以期发现新的耐药基因．方法:以H pylori 26695株为初发菌株,经抗生素压力筛选、转座子Tn5插入失活、E-test检测等实验方法探讨H pyolri对克拉霉素的耐药机制．结果:经12次传代,在0．5 mg／L克拉霉素浓度下培养出的耐药株经-80℃冻存30 d复苏,仍为抗性菌株;克拉霉素抗性的转座插入株于 700 bp左右出现条带;经96-168 h培养,单纯经抗生素选择压力筛选的低浓度耐药株E-test 出现椭圆形抑菌环,而经Tn5插入的克拉霉素抗性菌株无抑菌环出现．对插入位点基因的测序发现该片段与H pylori 26695菌株中“H pylori 1469”完全同源,为编码Omp3l的基因．结论:H pylori对克拉霉素的低浓度耐药,经转座后可转化为高浓度耐药,其耐药性的发生与23S rRNA点突变以外的Omp31基因有关．

AIM： To investigate the mechanism of H pylori resistance to clarithromycin in different concentrations, and to look for a novel gene associated with clarithromycin resistance. METHODS： H pylori 26695 were used as the primary strain. Antibiotic selection pressure test, mutation by transponson （TnS） containing chloromycetin marker and E-test were performed for the analysis of H pylori resistance to clarithromycin in different concentrations. RESULTS： After 12 generations, the 0.5 mg/L clarithromycin resistance strains remained resistant after the storage in -80℃ for 30 d. Clarithromycin-restant H pylori which had been inserted by transponson showed a band of 700 bp. After culturing for 98-168 h, the resistant H pylori from clarithromycin selection pressure test showed an elliptic inhibition ring. On the other hand, clarithromycin-resistant H pylori inserted by trans- ponson showed no inhibition ring. After sequencing, the gene at the insertion site was homogenous with H py/or/1469, which coded Omp31. CONCLUSION： The resistant H pylori from lower concentrations of clarithromycin can be changed into higher concentration resistance strain through transponson insertion, indicating that the mechanism of H pylori resistance to clarithromycin may be associated with Omp31 apart from 23S rRNA mutation.