Development and Evaluation of Multiplex PCR for the Detection of Carbapenemase-Producing Enterobacteriaceae

Bareum Gwon†1   Il Kwon  Bae†2   Na Young  Kim34   Sae Am  Son3   Sunjoo  Kim5   Joseph  Jeong6   Jeong Hwan  Shin*34   

1 Department of Clinical Laboratory Science, Semyung University, Jecheon,
2 Department of Dental Hygiene,Silla University
3 Department of Laboratory Medicine, Inje University College of Medicine,
4 Paik Institute for Clinical Research, Inje University College of Medicine, Busan
5 Department of Laboratory Medicine, Gyeongsang National University College of Medicine, Jinju,
6 Department of Laboratory Medicine, Ulsan University Hospital,University of Ulsan College of Medicine, Ulsan,

* Corresponding author: Tel: +82-51-890-6475, Fax: +82-51-890-8615,, E-mail: jhsmile@inje.ac.kr

Abstract

Background: The isolation of carbapenemase-producing Enterobacteriaceae (CPE) has become increasingly common. Continuous surveillance for these organisms is essential because their infections are closely related to outbreaks of illness and are associated with high mortality rates. The aim of this study was to develop and evaluate multiplex PCR as a means of detecting several important CPE genes simultaneously.

Methods: We aimed to develop a multiplex PCR that could detect seven CPE genes simultaneously. The multiplex PCR was composed of seven primer sets for the detection of KPC, IMP, VIM, NDM-1, GES, OXA-23, and OXA-48. We designed different PCR product sizes of at least 100 bp. We evaluated the performance of this new test using 69 CPE-positive clinical isolates. Also, we confirmed the specificity to rule out false-positive reactions by using 71 carbapenem- susceptible clinical strains.

Results: A total of 69 CPE clinical isolates showed positive results and were correctly identified as KPC (N=14), IMP (N=13), OXA-23 (N=12), OXA-48 (N=11), VIM (N=9), GES (N=5), and NDM (N=5) by the multiplex PCR. All 71 carbapenem-susceptible clinical isolates, including Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa, showed negative results.

Conclusion: This multiplex PCR can detect seven CPE genes at a time and will be useful in clinical laboratories. (Ann Clin Microbiol 2019;22:9-13)

Acknowledgements

This research was supported by a fund (2017-ER5402-01) by Research of Korea Centers for Disease Control and Prevension.

Figures & Tables

Fig. 1. Multiplex PCR for the detection of seven carbapenemaseproducing Enterobacteriaceae genes. Abbreviations: M, molecular weight marker; N, negative control. Lane: IMP (180 bp), VIM (306 bp), NDM (410 bp), OXA-23 (505 bp), OXA-48 (602 bp), KPC (747 bp), and GES (855 bp).