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Identification and Characterization of an Organic Solvent Tolerance Gene in Helicobacter pylori
Author(s) -
Chiu HungChuan,
Lin TzuLung,
Wang JinTown
Publication year - 2007
Publication title -
helicobacter
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.206
H-Index - 79
eISSN - 1523-5378
pISSN - 1083-4389
DOI - 10.1111/j.1523-5378.2007.00473.x
Subject(s) - glutaraldehyde , mutant , microbiology and biotechnology , biology , polyclonal antibodies , helicobacter pylori , chemistry , gene , antibody , biochemistry , genetics , chromatography
Background: Pre‐cleaning and soaking in glutaraldehyde is the necessary procedure to disinfect endoscopes. However, some chemical‐solvent‐tolerant bacteria may survive after incomplete endoscopic disinfection. The goal of this study was to identify glutaraldehyde resistance‐related genes in Helicobacter pylori . Materials and Methods: λ‐Zap phagemid expression library of H. pylori strain NTUH‐C1 was selected with 0.1% glutaraldehyde. The minimal inhibitory concentration (MIC) of glutaraldehyde‐resistant DNA fragments of H. pylori NTUH‐C1 strain were determined. Imp/OstA recombinant protein was expressed, purified, and used to generate anti‐Imp/OstA polyclonal antibody. Imp/ostA knockout, deletion, and complementation strains were constructed. The function of Imp/OstA was monitored by organic solvent tolerance assay, antibiotics susceptibility test, and n ‐phenylnapthylamine assay. Results: Using Imp/ostA polyclonal antibody against cell lysate of wild‐type and imp/ostA mutant showed that it is not essential in H. pylori . Organic solvent tolerance assay demonstrated the role of Imp/ostA in n‐hexane tolerance. MIC test showed that the mutation of imp/ostA was susceptible to hydrophobic and β‐lactam antibiotics. NPN assay demonstrated that the level of outer membrane permeability was increased by 50% in mutant strain comparing to wild‐type strain ( p < .001). Conclusions: We have identified an Imp/OstA protein that was associated with glutaraldehyde resistance in our clinical strain H. pylori NTUH‐C1 by screening of λ‐Zap expression library. Disruption of this protein results in altering membrane permeability, sensitivity to organic solvent, and susceptibility to antibiotics.