title: Rapid detection of clarithromycin resistant Helicobacter pylori strains in Spanish patients by polymerase chain reaction-restriction fragment length polymorphism.
abstract: BACKGROUND
  The aim of this study was to characterize the mutations types present in the 23S rRNA gene related to H. pylori clarithromycin-resistance strains in Spain and evaluate a novel PCR-RFLP method for detection of the most frequent point mutation in our population.
  
  
  METHODS
  Gastric biopsies were obtained by endoscopy from patients with gastric symptoms. H. pylori was cultured according to standard microbiological procedures and clarithromycin resistance was determined by E-test. DNA extraction was performed by NucliSens platform with the NucliSens magnetic extraction reagents (bioMérieux) according to the manufacturer instructions. Analyses for point mutations in 23S rRNA gene strains were performed by sequence analysis of amplified polymerase chain reaction products. Restriction fragment length polymorphism was performed using BsaI enzyme to detect restriction sites that correspond to the mutation (A2143G).
  
  
  RESULTS
  We found 42 out of 118 (35.6%) strains resistant to clarithromycin by E-test. E-test results were confirmed for the presence of point mutation in 34 (88.1%) of these strains. Mutation A2143G was found in 85.3% of the strains. Analyses with the restriction enzyme BsaI was able to confirm the presence of A2143G mutation. There were 8 H. pylori strains resistant to clarithromycin by E-test but without any point mutation in the 23 rRNA gene.
  
  
  CONCLUSIONS
  We conclude that PCR-RFLP is a reliable method to detect clarithromycin-resistance H. pylori strains in countries with a high prevalence of clarithromycin-resistance as Spain. It may be useful before choosing regimens of H. pylori eradication.
authors.affiliation: Hospital de la Princesa, Madrid, Spain. soniaagu@hotmail.com
authors.forename: S
authors.initials: S
authors.lastname: Agudo
authors.forename: G
authors.initials: G
authors.lastname: Pérez-Pérez
authors.forename: T
authors.initials: T
authors.lastname: Alarcón
authors.forename: M
authors.initials: M
authors.lastname: López-Brea
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002648:Child; D017291:Clarithromycin / Q000494:pharmacology*; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial; D005260:Female; D016481:Helicobacter Infections / Q000382:microbiology*; D016480:Helicobacter pylori / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D008297:Male; D008826:Microbial Sensitivity Tests; D008875:Middle Aged; D017354:Point Mutation / Q000235:genetics; D012150:Polymorphism, Restriction Fragment Length; D012338:RNA, Ribosomal, 23S / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013030:Spain; D055815:Young Adult
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D012338:RNA, Ribosomal, 23S; D017291:Clarithromycin