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Multicenter Study
. 2014 Jan 14:14:27.
doi: 10.1186/1471-2334-14-27.

Multi-centre evaluation of real-time multiplex PCR for detection of carbapenemase genes OXA-48, VIM, IMP, NDM and KPC

Anneke van der Zee  1 , Lieuwe Roorda, Gerda Bosman, Ad C Fluit, Mirjam Hermans, Paul H M Smits, Adri G M van der Zanden, René Te Witt, Lesla E S Bruijnesteijn van Coppenraet, James Cohen Stuart, Jacobus M Ossewaarde
Affiliations
Multicenter Study

Multi-centre evaluation of real-time multiplex PCR for detection of carbapenemase genes OXA-48, VIM, IMP, NDM and KPC

Anneke van der Zee et al. BMC Infect Dis. .

Abstract

Background: Resistance to carbapenem antibiotics is emerging worldwide among Enterobacteriaceae. To prevent hospital transmission due to unnoticed carriage of carbapenemase producing micro-organisms in newly admitted patients, or follow-up of patients in an outbreak setting, a molecular screening method was developed for detection of the most prevalent carbapenemase genes; blaOXA-48, blaVIM, blaIMP, blaNDM and blaKPC.

Methods: A real-time multiplex PCR assay was evaluated using a collection of 86 Gram negative isolates, including 62 carbapenemase producers. Seven different laboratories carried out this method and used the assay for detection of the carbapenemase genes on a selection of 20 isolates.

Results: Both sensitivity and specificity of the multiplex PCR assay was 100%, as established by results on the strain collection and the inter-laboratory comparisons.

Conclusions: In this study, we present a multiplex real-time PCR that is a robust, reliable and rapid method for the detection of the most prevalent carbapenemases blaOXA-48, blaVIM, blaIMP, blaNDM and blaKPC, and is suitable for screening of broth cultured rectal swabs and for identification of carbapenemase genes in cultures.

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Figures

Figure 1
Figure 1
Standard curve of multiplex PCR. Inferred efficiencies of multiplex amplification range from 82% for IMP, to 85% for VIM.
Figure 2
Figure 2
A) Amplification plot of VIC labelled PCR products of VIM, NDM, and KPC (protocol 1), and B) OXA-48/CTX-M and VIM/IMP (protocol 2). Reactions were performed in the presence of the internal control PhHV. For clarity, a selection of strains is shown. OXA-48 positives with lower plots were also positive for CTX-M.
See this image and copyright information in PMC

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