title: Real-time PCR screening for 16S rRNA mutations associated with resistance to tetracycline in Helicobacter pylori.
abstract: The effectiveness of recommended first-line therapies for Helicobacter pylori infections is decreasing due to the occurrence of resistance to metronidazole and/or clarithromycin. Quadruple therapies, which include tetracycline and a bismuth salt, are useful alternative regimens. However, resistance to tetracycline, mainly caused by mutations in the 16S rRNA genes (rrnA and rrnB) affecting nucleotides 926 to 928, are already emerging and can impair the efficacies of such second-line regimens. Here, we describe a novel real-time PCR for the detection of 16S rRNA gene mutations associated with tetracycline resistance. Our PCR method was able to distinguish between wild-type strains and resistant strains exhibiting single-, double, or triple-base-pair mutations. The method was applicable both to DNA extracted from pure cultures and to DNA extracted from fresh or frozen H. pylori-infected gastric biopsy samples. We therefore conclude that this real-time PCR is an excellent method for determination of H. pylori tetracycline resistance even when live bacteria are no longer available.
authors.affiliation: Nationales Referenzzentrum für Helicobacter pylori, Abteilung Medizinische Mikrobiologie und Hygiene, Institut für Medizinische Mikrobiologie und Hygiene, Universitätsklinikum Freiburg, Hermann-Herder-Strasse 11, D-79104 Freiburg, Germany.
authors.forename: Erik
authors.initials: E
authors.lastname: Glocker
authors.forename: Marco
authors.initials: M
authors.lastname: Berning
authors.forename: Monique M
authors.initials: MM
authors.lastname: Gerrits
authors.forename: Johannes G
authors.initials: JG
authors.lastname: Kusters
authors.forename: Manfred
authors.initials: M
authors.lastname: Kist
mesh_terms: D004269:DNA, Bacterial / Q000032:analysis; D004275:DNA, Ribosomal / Q000235:genetics; D016481:Helicobacter Infections / Q000382:microbiology*; D016480:Helicobacter pylori / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D012680:Sensitivity and Specificity; D013752:Tetracycline / Q000494:pharmacology; D013753:Tetracycline Resistance / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S; D013752:Tetracycline