Susceptibility testing of Helicobacter pylori: Comparison of E-test and Disk Diffusion for Metronidazole and Mutations in rdxA gene sequences of Helicobacter pylori strains

Metronidazole is a main stay of modern multidrug therapies for Helicobacter pylori (H. pylori) infection. Metronidazole resistance reduces the effectiveness of these combination therapies. Various methods have been used for the determination of the sensitivity of H. pylori to metronidazole, that have shown conflicting results. The aims of this study are: 1) Comaring E-Test and disk diffusion methods for determining the susceptibility of H. pylori to metronidazole; and 2) As metronidazole resistance in H. pylori has been found to be associated with mutations in rdxA, the role of this gene in metronidazole resistance in H. pylori has been examined in this study. A total of 46 H. pylori strains from 223 consecutive patients were examined. The E-Test was performed according to the manufacturer’s guidelines, and the disk diffusion test, according to standard procedure, using 5 ?g metronidazole disks. Extraction of DNA was done from all H. pylori isolates by boiling and the use of phenol-chloroform methods, and afterwards Polymerase Chain Reaction (PCR) was performed. Metronidazole resistance as determined by E-test and disk diffusion methods, was 64.3% and 47.6% respectively. None of the resistant or sensitive samples possessed rdxA gene deletion. Disk diffusion method is not reliable in determining metronidazole resistance in H. pylori. An intact rdxA gene has also been reported in metronidazole-resistant H. pylori, suggesting that additional metronidazole resistance mechanisms exist in H. pylori and even molecular methods are not reliable for the detection of this resistance.