Application of 3'-mismatched reverse primer PCR compared with real-time PCR and PCR-RFLP for the rapid detection of 23S rDNA mutations associated with clarithromycin resistance in Helicobacter pylori.
Helicobacter pylori clarithromycin (Cla) resistance dramatically reduces efficacy of eradication therapy. In this study, 3'-mismatched reverse primer PCR (3M-PCR), real-time PCR (LightCycler), and PCR-RFLP assays were investigated to determine their sensitivity for detecting clarithromycin resistance associated with 23S rDNA mutations (A2142G, A2142C, and A2143G). For 84.8% (123/145) of isolates, the same allelic type was detected by each method although methods differed in efficiency of detecting mutations in cultures either containing mixtures of two alleles (24 isolates), or that were dual allelic variants (two isolates). The novel 3M-PCR assay format was the most sensitive, detecting all alleles at > or =0.02 ng/microl in DNA mixtures, and thus provides more precise information to guide clinical management of patients at risk of treatment failure.
Elviss
Nicola C
NC
Helicobacter Reference Unit, Laboratory of Enteric Pathogens, Specialist and Reference Microbiology Division, Health Protection Agency, 61 Colindale Avenue, London NW9 5HT, UK.
Lawson
Andrew J
AJ
Owen
Robert J
RJ
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D017291:Clarithromycin / Q000494:pharmacology*; D017931:DNA Primers; D004275:DNA, Ribosomal / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D016480:Helicobacter pylori / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length; D012338:RNA, Ribosomal, 23S / Q000235:genetics*
D000900:Anti-Bacterial Agents; D017931:DNA Primers; D004275:DNA, Ribosomal; D012338:RNA, Ribosomal, 23S; D017291:Clarithromycin