title: Rapid genotyping of CTX-M extended-spectrum beta-lactamases by denaturing high-performance liquid chromatography.
abstract: Denaturing high-performance liquid chromatography (dHPLC) is a powerful technique which has been used extensively to detect genetic variation. This is the first report of the application of dHPLC for rapid genotyping of bacterial beta-lactamase genes. The technique was specifically developed to genotype members of all blaCTX-M DNA homology groups. Thirteen well-defined blaCTX-M extended-spectrum beta-lactamase (ESBL)-producing strains were used to develop and optimize the dHPLC genotyping assay. Further evaluation was carried out with a blinded panel of 62 clinical isolates. The results of blaCTX-M genotyping achieved by dHPLC were comparable to the typing results obtained by DNA sequencing. Applying the newly developed dHPLC-based genotyping method, we successfully genotyped all 73 blaCTX-M ESBL-producing strains from the 4-month survey study. Furthermore, we found the first reported cases in the United Kingdom of clinically significant disease caused by CTX-M-14- and CTX-M-1-producing Escherichia coli strains. We conclude that the novel dHPLC assay is highly accurate, rapid, and cost-effective for the genotyping of blaCTX-M-producing ESBLs and has great potential for determining the clinical relevance of different and new blaCTX-M genotypes, as well as for epidemiological studies and surveillance programs.
authors.affiliation: Health Protection Agency, West Midlands Public Health Laboratory, Heart of England NHS Foundation Trust, Bordesley Green East, Birmingham B9 5SS, United Kingdom.
authors.forename: Li
authors.initials: L
authors.lastname: Xu
authors.forename: Jason
authors.initials: J
authors.lastname: Evans
authors.forename: Thomas
authors.initials: T
authors.lastname: Ling
authors.forename: Kathy
authors.initials: K
authors.lastname: Nye
authors.forename: Peter
authors.initials: P
authors.lastname: Hawkey
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*; D005821:Genetic Techniques; D005838:Genotype; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D001618:beta-Lactamases / Q000145:classification / Q000378:metabolism*
chemical_list: D001618:beta-Lactamases