Detection of mixed clarithromycin-resistant and -susceptible Helicobacter pylori using nested PCR and direct sequencing of DNA extracted from faeces

J Med Microbiol. 2007 Sep;56(Pt 9):1174-1180. doi: 10.1099/jmm.0.47302-0.

Abstract

The major cause of chemotherapy failure in patients with chronic gastritis and peptic ulcers caused by Helicobacter pylori is clarithromycin (CAM) resistance due to a mutation in the 23S rRNA gene. This study describes a non-invasive and accurate method for the detection of mixed CAM-resistant and -susceptible H. pylori by sequencing of the H. pylori 23S rRNA gene. Faeces were crushed with beads and the 23S rRNA gene was amplified using a nested PCR on the extracted DNA. Mutation analysis of this gene using this method showed that 20.4 % of patients carried mixed CAM-susceptible (wild type) and -resistant (A2142G or A2143G mutant) H. pylori. Furthermore, it was found that 66.6 % of patients who had been treated unsuccessfully carried one of these mutations in the 23S rRNA gene (including the mixed type), whilst standard culture detected CAM-resistant isolates in only 22.2 % of patients with unsuccessful treatment. These data suggest that, for successful therapy, the diagnosis method described here would more accurately detect CAM-resistant H. pylori, including mixed infections.

Publication types

  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Aged, 80 and over
  • Anti-Bacterial Agents / pharmacology
  • Clarithromycin / pharmacology*
  • DNA, Bacterial / genetics
  • DNA, Ribosomal / genetics
  • Drug Resistance, Bacterial / genetics*
  • Feces / microbiology*
  • Female
  • Helicobacter pylori / drug effects*
  • Helicobacter pylori / genetics*
  • Helicobacter pylori / isolation & purification
  • Humans
  • Male
  • Middle Aged
  • Point Mutation
  • Polymerase Chain Reaction / methods*
  • RNA, Ribosomal, 23S / genetics
  • Sensitivity and Specificity
  • Sequence Analysis, DNA / methods*

Substances

  • Anti-Bacterial Agents
  • DNA, Bacterial
  • DNA, Ribosomal
  • RNA, Ribosomal, 23S
  • Clarithromycin