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High-Resolution Melting-Curve Analysis of Ligation-Mediated Real-Time PCR for Rapid Evaluation of an Epidemiological Outbreak of Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli
Kalmar County Hospital.
Swedish Institute for Communicable Disease Control, Solna.
Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Center for Diagnostics, Department of Clinical Microbiology.
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2011 (English)In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 49, no 12, 4032-4039 p.Article in journal (Refereed) Published
Abstract [en]

Methods for the confirmation of nosocomial outbreaks of bacterial pathogens are complex, expensive, and time-consuming. Recently, a method based on ligation-mediated PCR (LM/PCR) using a low denaturation temperature which produces specific melting-profile patterns of DNA products has been described. Our objective was to further develop this method for real-time PCR and high-resolution melting analysis (HRM) in a single-tube system optimized in order to achieve results within 1 day. Following the optimization of LM/PCR for real-time PCR and HRM (LM/HRM), the method was applied for a nosocomial outbreak of extended-spectrum-beta-lactamase (ESBL)-producing and ST131-associated Escherichia coli isolates (n = 15) and control isolates (n = 29), including four previous clusters. The results from LM/HRM were compared to results from pulsed-field gel electrophoresis (PFGE), which served as the gold standard. All isolates from the nosocomial outbreak clustered by LM/HRM, which was confirmed by gel electrophoresis of the LM/PCR products and PFGE. Control isolates that clustered by LM/PCR (n = 4) but not by PFGE were resolved by confirmatory gel electrophoresis. We conclude that LM/HRM is a rapid method for the detection of nosocomial outbreaks of bacterial infections caused by ESBL-producing E. coli strains. It allows the analysis of isolates in a single-tube system within a day, and the discriminatory power is comparable to that of PFGE.

Place, publisher, year, edition, pages
American Society for Microbiology , 2011. Vol. 49, no 12, 4032-4039 p.
National Category
Medical and Health Sciences
URN: urn:nbn:se:liu:diva-74435DOI: 10.1128/JCM.01042-11ISI: 000298113400002OAI: diva2:484424

Funding Agencies|Kalmar County Hospital||FORSS (The Research Council of Southeast Sweden)||

Available from: 2012-01-27 Created: 2012-01-27 Last updated: 2015-01-07

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Tärnberg, MariaRyberg, AnnaMonstein, Hans-JurgNilsson, Lennart E
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