Liu Zhi-Qiang, Zheng Peng-Yuan, Yang Ping-Chang
Department of Gastroenterology, the Second Affiliated Hospital of Zhengzhou University, 2 Jingba Road, Zhengzhou 450014, Henan Province, China.
World J Gastroenterol. 2008 Sep 7;14(33):5217-22. doi: 10.3748/wjg.14.5217.
To determine whether efflux systems contribute to multidrug resistance of H pylori.
A chloramphenicol-induced multidrug resistance model of six susceptible H pylori strains (5 isolates and H pylori NCTC11637) was developed. Multidrug-resistant (MDR) strains were selected and the minimal inhibitory concentration (MIC) of erythromycin, metronidazole, penicillin G, tetracycline, and ciprofloxacin in multidrug resistant strains and their parent strains was determined by agar dilution tests. The level of mRNA expression of hefA was assessed by fluorescence real-time quantitative PCR. A H pylori LZ1026 knockout mutant (delta H pylori LZ1026) for (putative) efflux protein was constructed by inserting the kanamycin resistance cassette from pEGFP-N2 into hefA, and its susceptibility profiles to 10 antibiotics were evaluated.
The MIC of six multidrug-resistant strains (including 5 clinical isolates and H pylori NCTC11637) increased significantly (>or=4-fold) compared with their parent strains. The expression level of hefA gene was significantly higher in the MDR strains than in their parent strains (P=0.033). A H pylori LZ1026 mutant was successfully constructed and the delta H pylori LZ1026 was more susceptible to four of the 10 antibiotics. All the 20 strains displayed transcripts for hefA that confirmed the in vitro expression of these genes.
The efflux pump gene hefA plays an important role in multidrug resistance of H pylori.
确定外排系统是否与幽门螺杆菌的多重耐药性有关。
建立了六种敏感幽门螺杆菌菌株（5个分离株和幽门螺杆菌NCTC11637）的氯霉素诱导多重耐药模型。筛选出多重耐药（MDR）菌株，并通过琼脂稀释试验测定多重耐药菌株及其亲本菌株中红霉素、甲硝唑、青霉素G、四环素和环丙沙星的最低抑菌浓度（MIC）。通过荧光实时定量PCR评估hefA的mRNA表达水平。通过将来自pEGFP-N2的卡那霉素抗性盒插入hefA构建了（假定的）外排蛋白的幽门螺杆菌LZ1026敲除突变体（delta幽门螺杆菌LZ1026），并评估了其对10种抗生素的敏感性谱。
六种多重耐药菌株（包括5个临床分离株和幽门螺杆菌NCTC11637）的MIC与其亲本菌株相比显著增加（≥4倍）。MDR菌株中hefA基因的表达水平显著高于其亲本菌株（P = 0.033）。成功构建了幽门螺杆菌LZ1026突变体，delta幽门螺杆菌LZ1026对10种抗生素中的4种更敏感。所有20株菌株均显示hefA的转录本，证实了这些基因的体外表达。
外排泵基因hefA在幽门螺杆菌的多重耐药性中起重要作用。
