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World J Gastroenterol. Sep 7, 2008; 14(33): 5217-5222
Published online Sep 7, 2008. doi: 10.3748/wjg.14.5217
Efflux pump gene hefA of Helicobacter pylori plays an important role in multidrug resistance
Zhi-Qiang Liu, Peng-Yuan Zheng, Ping-Chang Yang
Zhi-Qiang Liu, Peng-Yuan Zheng, Department of Gastroenterology, the Second Affiliated Hospital of Zhengzhou University, Zhengzhou 450014, Henan Province, China
Ping-Chang Yang, Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario L8N 4A6, Canada
Author contributions: Liu ZQ, Zheng PY contributed equally to this work; Liu ZQ, Zheng PY, Yang PC designed the research; Liu ZQ, Zheng PY, Yang PC performed the research; Zheng PY contributed to the new reagents/analytic tools; Zheng PY, Yang PC analyzed the data; Liu ZQ, Zheng PY wrote the paper.
Correspondence to: Dr. Peng-Yuan Zheng, Department of Gastroenterology, the Second Affiliated Hospital of Zhengzhou University, 2 Jingba Road, Zhengzhou 450014, Henan Province, China. medp7123@yahoo.com
Telephone: +86-371-65261035 Fax: +86-371-63934118
Received: April 26, 2008
Revised: August 18, 2008
Accepted: August 25, 2008
Published online: September 7, 2008
Abstract

AIM: To determine whether efflux systems contribute to multidrug resistance of H pylori.

METHODS: 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 (ΔH 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.

RESULTS: The MIC of six multidrug-resistant strains (including 5 clinical isolates and H pylori NCTC11637) increased significantly (≥ 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 ΔH 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.

CONCLUSION: The efflux pump gene hefA plays an important role in multidrug resistance of H pylori.

Keywords: Efflux pump, Helicobacter pylori, Multidrug resistance, Fluorescence real-time quantitative PCR, Knockout mutant