Synergistic effect of imp/ostA and msbA in hydrophobic drug resistance of Helicobacter pylori.
BACKGROUND
Contamination of endoscopy equipment by Helicobacter pylori (H. pylori) frequently occurs after endoscopic examination of H. pylori-infected patients. In the hospital, manual pre-cleaning and soaking in glutaraldehyde is an important process to disinfect endoscopes. However, this might not be sufficient to remove H. pylori completely, and some glutaraldehyde-resistant bacteria might survive and be passed to the next patient undergoing endoscopic examination through unidentified mechanisms. We identified an Imp/OstA protein associated with glutaraldehyde resistance in a clinical strain, NTUH-C1, from our previous study. To better understand and manage the problem of glutaraldehyde resistance, we further investigated its mechanism.


RESULTS
The minimal inhibitory concentrations (MICs) of glutaraldehyde andexpression of imp/ostA RNA in 11 clinical isolates from the National Taiwan University Hospital were determined. After glutaraldehyde treatment, RNA expression in the strains with the MICs of 4-10 microg/ml was higher than that in strains with the MICs of 1-3 microg/ml. We examined the full-genome expression of strain NTUH-S1 after glutaraldehyde treatment using a microarray and found that 40 genes were upregulated and 31 genes were downregulated. Among the upregulated genes, imp/ostA and msbA, two putative lipopolysaccharide biogenesis genes, were selected for further characterization. The sensitivity to glutaraldehyde or hydrophobic drugs increased in both of imp/ostA and msbA single mutants. The imp/ostA and msbA double mutant was also hypersensitive to these chemicals. The lipopolysaccharide contents decreased in individual imp/ostA and msbA mutants and dramatically reduced in the imp/ostA and msbA double mutant. Outer membrane permeability assay demonstrated that the imp/ostA and msbA double mutation resulted in the increase of outer membrane permeability. Ethidium bromide accumulation assay demonstrated that MsbA was involved in efflux of hydrophobic drugs.


CONCLUSIONS
The expression levels of imp/ostA and msbA were correlated with glutaraldehyde resistance in clinical isolates after glutaraldehyde treatment. Imp/OstA and MsbA play a synergistic role in hydrophobic drugs resistance and lipopolysaccharide biogenesis in H. pylori.
Chiu
Hung-Chuan
HC
Department of Microbiology, National Taiwan University College of Medicine, Taipei City 10051, Taiwan, Republic of China. gaki98@yahoo.com.tw
Lin
Tzu-Lung
TL
Yang
Jyh-Chin
JC
Wang
Jin-Town
JT
D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015139:Blotting, Southern; D002463:Cell Membrane Permeability; D004202:Disinfectants / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics; D004996:Ethidium / Q000378:metabolism; D005798:Genes, Bacterial / Q000187:drug effects; D016680:Genome, Bacterial / Q000187:drug effects; D005976:Glutaral / Q000494:pharmacology*; D016480:Helicobacter pylori / Q000187:drug effects* / Q000302:isolation & purification / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D008070:Lipopolysaccharides / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D012329:RNA, Bacterial / Q000378:metabolism
D018528:ATP-Binding Cassette Transporters; D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004202:Disinfectants; D008070:Lipopolysaccharides; C079726:MsbA protein, Bacteria; D012329:RNA, Bacterial; D004996:Ethidium; D005976:Glutaral