幽门螺杆菌对6种抗生素的耐药性及其机制的研究

Prevalence and mechanism of antibiotic resistance in helicobacter pylori

目的了解幽门螺杆菌(Helicobacter pylori,Hp)对6种常用抗生素的耐药状况,探讨耐甲硝唑rdxA基因突变与耐药性的关系。方法分离培养76株Hp,以琼脂稀释法检测Hp对甲硝唑、替硝唑、四环素、克拉(红)霉素、阿莫西林、利福平的最小抑菌浓度(Mininum Inhibitory Concentration,MIC),再用PCR方法扩增甲硝唑耐药相关基因rdxA,DNA测序分析敏感株和耐药株耐药基因的序列与耐药性的关系。免疫印记(West-ern Blotting)分析耐药基因的表达与耐药性的关系。结果武汉地区人群Hp对甲硝唑、替硝唑、四环素、克拉(红)霉素、阿莫西林、利福平的耐药率分别为67.1%、43.4%、3.9%、11.8%、17.1%和10.5%;甲硝唑耐药基因序列分析表明rdxA基因有插入或点突变;Western Blot显示耐药株和敏感株的rdxA基因的表达有差异。结论幽门螺杆菌对6种抗生素有较高的耐药率,rdxA基因突变导致基因表达水平改变,导致Hp对甲硝唑耐药。

[Objective ] To investigate the prevalence of helicobacter pylori resistance to six antibiotics and study the relationship between the rdxA gene mutation and the metronidazole resistance. [Methods] 74 H. pylori isolates were collected from the gastric mucosa of patients with peptic ulcer and gastritis. Agar-diffusion method was used for assaying H. pylori susceptibility to metronidazole, nitronidazole, tetracycline, clarithromycin, amoxieillin and rifamipicin, rdxA gene was amplified by PCR. Target DNA were cloned and sequenced. The expression level of the rdxA gene was detected by Western Blot. At last analyzed the relationship between the rdxA gene expression and the metronidazole resistance. [Results] The prevalence of antibiotic resistance to metronidazole, nitronidazole, tetracycline, clarithromycin, amoxicillin and rifamipicin in H. pylori isolates were 67.1%, 43.4%, 3.9%, 11.8%, 17.1% and 10.5%, respectively. DNA sequencing showed that there were mutations in rdxA gene of metronidazole resistance isolates. Western Blot showed there were differences of rdxA gene expression between metronidazole susceptibility and resistance strains. [Conclusion] The prevalence of six antibiotic resistances in H. pylori isolates is high. The gene rdxA mutation is the cause of resistance of H.pylori to metronidazole.