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

Si Hyun Kim; Il Kwon Bae; Na Young Kim; Sae Am Song; Sunjoo Kim; Joseph Jeong; Jeong Hwan Shin

doi:10.5145/ACM.2019.22.1.9

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Kim, Bae, Kim, Song, Kim, Jeong, and Shin: Development and Evaluation of Multiplex PCR for the Detection of Carbapenemase-Producing Enterobacteriaceae

Original Article

Annals of Clinical Microbiology 2019; 22(1): 9-13.

Published online: 21 March 2019

DOI: https://doi.org/10.5145/ACM.2019.22.1.9

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

Si Hyun Kim^1,^*, Il Kwon Bae^2,^*, Na Young Kim^3,⁴, Sae Am Song³, Sunjoo Kim⁵, Joseph Jeong⁶, Jeong Hwan Shin^3,⁴

¹Department of Clinical Laboratory Science, Semyung University, Jecheon, Korea.

²Department of Dental Hygiene, Silla University, Busan, Korea.

³Department of Laboratory Medicine, Inje University College of Medicine, Inje University College of Medicine, Busan, Korea.

⁴Paik Institute for Clinical Research, Inje University College of Medicine, Busan, Korea.

⁵Department of Laboratory Medicine, Gyeongsang National University College of Medicine, Jinju, Korea.

⁶Department of Laboratory Medicine, Ulsan University Hospital, University of Ulsan College of Medicine, Ulsan, Korea.

Correspondence: Jeong Hwan Shin, Department of Laboratory Medicine, Busan Paik Hospital, Inje University College of Medicine, 75 Bokjiro, Busanjin-gu, Busan 47392, Korea. (Tel) 82-51-890-6475, (Fax) 82-51-890-8615, jhsmile@inje.ac.kr

Equal contributor:

*These authors contributed equally to this work.

Received 3 August 2018 Revised 3 October 2018 Accepted 4 October 2018

The Korean Society of Clinical Microbiology

(open-access):

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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.

Keywords: Carbapenem, Carbapenem-resistant Enterobacteriaceae, Carbapenemase-producing Enterobacteriaceae, Enterobacteriaceae, Multiplex PCR

Figures and Tables

Fig. 1

Multiplex PCR for the detection of seven carbapenemase-producing 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).

Table 1

Clinical strains of carbapenemase-producing Enterobacteriaceae used in this study

Table 2

Carbapenem-susceptible clinical strains tested (n=71)

Table 3

Primers used in this study

ACKNOWLEDGMENTS

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

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