Pyrosequencing analysis for rapid and accurate detection of clarithromycin resistance-associated mutations in Iranian Helicobacter pylori isolates

Abstract

Background Treatment of Helicobacter pylori (H. pylori) infection has become challenging following the development of primary antibiotic resistance. A primary therapeutic regimen for H. pylori eradication includes clarithromycin; however, the presence of point mutations within the 23S rRNA sequence of H. pylori contributes to clarithromycin resistance and eradication failure. Thus, we aimed to develop a rapid and precise method to determine clarithromycin resistance-related point mutations using the pyrosequencing method. Methods and results H. pylori was isolated from 82 gastric biopsy samples and minimal inhibitory concentration (MIC) was evaluated using the agar dilution method. Clarithromycin resistance-associated point mutations were detected by Sanger sequencing, from which 11 isolates were chosen for pyrosequencing. Our results demonstrated a 43.9% (36/82) prevalence in resistance to clarithromycin. The A2143G mutation was detected in 8.3% (4/48) of H. pylori isolates followed by A2142G (6.2%), C2195T (4.1%), T2182C (4.1%), and C2288T (2%). Although the C2195T mutation was only detected by Sanger sequencing, the overall results from pyrosequencing and Sanger sequencing platforms were comparable. Conclusions Pyrosequencing could be used as a rapid and practical platform in clinical laboratories to determine the susceptibility profile of H. pylori isolates. This might pave the way for efficient H. pylori eradication upon detection.

A reconstructed genome‐scale metabolic model of Helicobacter pylori for predicting putative drug targets in clarithromycin and rifampicin resistance conditions

“…Helicobacter pylori ( H. pylori ) is a Gram‐negative pathogen that can persistently colonize the gastric epithelium of more than half of the world's population, especially in developing countries 1 . The presence of H. pylori evokes various diseases including chronic gastritis, duodenal ulcer, gastric ulcer, mucosa‐associated lymphoid tissue (MALT) lymphoma, and gastric adenocarcinoma 2,3 . The type and severity of H. pylori ‐associated diseases vary greatly depending on bacterial strain features, host characteristics, and environmental parameters 4 .…”

“…1 The presence of H. pylori evokes various diseases including chronic gastritis, duodenal ulcer, gastric ulcer, mucosa-associated lymphoid tissue (MALT) lymphoma, and gastric adenocarcinoma. 2,3 The type and severity of H. pylori-associated diseases vary greatly depending on bacterial strain features, host characteristics, and environmental parameters. 4 Despite the availability of multiple eradication regimens to combat H. pylori infection, treatment fails in over 20% of infected patients, primarily due to antibiotic resistance.…”

Editorial: Helicobacter pylori infection and antibiotic resistance: clinical, translational and experimental studies

“…Although analysis of 2109 H. pylori clinical isolates presented an overall reduction of primary resistance rate over the past years, H. pylori strains are as yet highly resistant to metronidazole (67.2%), clarithromycin (36.0%), and levofloxacin (24.2%). Considering the recommendation that clarithromycin susceptibility testing should be performed prior to prescribing any clarithromycin containing H. pylori first-line therapy ( Malfertheiner et al., 2022 ), developing simple and more rapid techniques for detecting H. pylori clarithromycin resistance seems essential ( Alavifard et al., 2023 ).…”