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. 2010 Jan;48(1):277-80.
doi: 10.1128/JCM.01411-09. Epub 2009 Oct 28.

Multiplex PCR-ligation detection reaction assay for simultaneous detection of drug resistance and toxin genes from Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium

K Granger  1 , M S Rundell, M R Pingle, R Shatsky, D H Larone, L M Golightly, F Barany, E D Spitzer
Affiliations

Multiplex PCR-ligation detection reaction assay for simultaneous detection of drug resistance and toxin genes from Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium

K Granger et al. J Clin Microbiol. 2010 Jan.

Abstract

A multiplex PCR-ligation detection reaction (PCR-LDR) assay was developed for rapid detection of methicillin, tetracycline, and vancomycin resistance, as well as toxic shock toxin and Panton-Valentine leukocidin. The assay was tested on 470 positive blood culture bottles containing Staphylococcus aureus or enterococci. PCR-LDR exhibited a sensitivity and specificity of > or = 98% for all components except tetracycline resistance, which had a sensitivity of 94.7%. Rapid and sensitive detection of antimicrobial resistance and virulence genes could help guide therapy and appropriate infection control measures.

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Figures

FIG. 1.
FIG. 1.
(A) Capillary electrophoresis (CE) trace for the antibiotic resistance, and toxin PCR-LDR assay of a vancomycin-resistant E. faecium isolate. The x axis shows the LDR product size, and the y axis shows fluorescence intensity of the bands. Red arrows indicate signals that are unique to the 16S rRNA genes of E. faecium and the vanA gene. (B) The CE data are displayed as a reconstructed gel image generated by a software program developed in our laboratory. The left panel represents the size and fluorescence of the 16S rRNA gene LDR products which act as a positive control and differentiate between the bacteria E. faecalis, E. faecium, and Staphylococcus species. The middle panel represents the LDR signals that identify the presence of methicillin resistance (mecA), vancomycin resistance (vanA, vanB, vanD), tetracycline resistance (tetL tetK, tetM), the toxic shock toxin gene (tst), and the PVL toxin genes (lukS-lukF). Data in the right panel represent two blood cultures. In one, the 16S rRNA gene control identified E. faecium, and the vanA gene was detected, indicating a VRE; in the other the 16S rRNA gene control identified S. aureus, and the mecA and lukS-lukF genes were detected, indicating an MRSA gene encoding the PVL toxin.
See this image and copyright information in PMC

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