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Antimicrobial Agents and Chemotherapy, April 2007, p. 1446-1454, Vol. 51, No. 4
0066-4804/07/$08.00+0     doi:10.1128/AAC.01088-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Rapid Genotyping of CTX-M Extended-Spectrum ß-Lactamases by Denaturing High-Performance Liquid Chromatography

Health Protection Agency West Midlands Public Health Laboratory, Heart of England NHS Foundation Trust, Birmingham B9 5SS, United Kingdom,¹ Division of Immunity and Infection, University of Birmingham, Birmingham B15 2TT, United Kingdom,² Prince of Wales Hospital, Hong Kong³

Received 29 August 2006/ Returned for modification 27 October 2006/ Accepted 21 December 2006

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 ß-lactamase genes. The technique was specifically developed to genotype members of all bla_CTX-M DNA homology groups. Thirteen well-defined bla_CTX-M extended-spectrum ß-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 bla_CTX-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 bla_CTX-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 bla_CTX-M-producing ESBLs and has great potential for determining the clinical relevance of different and new bla_CTX-M genotypes, as well as for epidemiological studies and surveillance programs.

Published ahead of print on 8 January 2007.