2020; 40(3): 209-215
Ann Lab Med 2020; 40(3): 259-263
Published online May 1, 2020 https://doi.org/10.3343/alm.2020.40.3.259
Copyright © Korean Society for Laboratory Medicine.
Rapid Identification of OXA-48-like, KPC, NDM, and VIM Carbapenemase-Producing Enterobacteriaceae From Culture: Evaluation of the RESIST-4 O.K.N.V. Multiplex Lateral Flow Assay
Wonkeun Song, M.D.1 , Min-Jeong Park, M.D.1 , Seri Jeong, M.D.1 , Dong Hoon Shin, M.D.1 , Jae-Seok Kim, M.D.1 , Hyun Soo Kim, M.D.1 , Han-Sung Kim, M.D.1 , Nuri Lee, M.D.1 , Jun Sung Hong, Ph.D.2 , and Seok Hoon Jeong, M.D.2
1Department of Laboratory Medicine, Hallym University College of Medicine, Chuncheon, Korea; 2Department of Laboratory Medicine and Research Institute of Antimicrobial Resistance, Yonsei University College of Medicine, Seoul, Korea
Correspondence to: Wonkeun Song, M.D.
Department of Laboratory Medicine, Hallym University College of Medicine, Kangnam Sacred Heart Hospital, 1 Singil-ro, Youngdeungpo-gu, Seoul 07441, Korea
Tel: +82-2-829-5259 Fax: +82-2-847-2403 E-mail: firstname.lastname@example.org
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There is an urgent need for accurate and rapid diagnostic assays capable of identifying carbapenemase-producing Enterobacteriaceae (CPE). We assessed the performance of the RESIST-4 O.K.N.V. (OKNV) assay (Coris BioConcept, Gembloux, Belgium) for the identification of oxacillinase (OXA)-48-like-,
Keywords: Carbapenemase-producing Enterobacteriaceae, OXA-48-like, KPC, NDM, VIM, Evaluation, Performance, RESIST-4 O.K.N.V. assay
The global dissemination of carbapenemase-producing Enterobacteriaceae (CPE) is a major public health concern. Accurate and rapid detection of CPE is essential for patient management and for implementation of appropriate infection control measures [1,2]. Globally,
Recently, lateral flow immunochromatographic assays based on monoclonal antibodies generated by immunization of mice have been developed for rapid and easy identification of OXA-48-like, KPC, and NDM carbapenemases. This technology exhibited excellent accuracy (100% sensitivity and 100% specificity) in the identification OXA-48-like, KPC, and NDM producers directly from bacterial colonies within 15 minutes [6,7]. We evaluated the new RESIST-4 O.K.N.V. (OKNV) multiplex lateral flow assay developed by Coris BioConcept (Gembloux, Gembloux, Belgium), which identifies specific antibodies against OXA-48, KPC, NDM, and VIM carbapenemases directly from different culture media such as sheep blood agar (SBA) and a chromogenic medium. Different types of chromogenic media, such as CHROMagar KPC, chromID CARBA SMART (BioMérieux, Marcy-I'Etoile, France), and Brilliance CRE (ThermoFisher Scientific, Illkirch, France), are often used in rectal surveillance culture for identifying carbapenem-resistant Enterobacteriaceae (CRE) and/or CPE [8,9].
Globally, the OKNV assay shows 97–100% sensitivity and 100% specificity, with unequivocal results [10,11,12]. Therefore, additional confirmatory assays were not required. Unlike other studies, the number of carbapenemase co-producing isolates included was relatively high, and this is the first study to include isolates grown on a chromogenic medium.
In total, 65 CRE clinical isolates were included in the study (49 CPE and 16 non-CP-CRE). The CRE isolates were obtained from six Hallym University Medical Centers (two hospitals in Seoul, two hospitals in Gyunggi, and one hospital in Gangwon) in Korea between 2012 and 2018, and all isolates were sent to one institution (Kangnam Sacred Heart Hospital, Seoul) and frozen before study. The study protocol was approved by the Institutional Review Board of each institution, which waived the need for informed consent. All isolates were tested for carbapenemase by PCR and DNA sequencing according to previously described methods [13,14]. The CPE isolates included nine KPC, eight NDM-1, seven OXA-48-like, five VIM, two IMP, and two Guiana extended-spectrum β-lactamase (GES)-5,carbapenemase variants. In addition, 16 Enterobacteriaceae isolates co-producing carbapenemases (OXA-48-like and NDM [N=9], KPC and NDM [N=5], and VIM and NDM [N=2]) were included. Sixteen non-CP-CRE isolates were also included (Table 1).
The OKNV assay is a multiplex lateral flow immunochromatographic assay for the detection of OXA-48-like, KPC, NDM, and VIM carbapenemases in two lateral flow cassettes (one for OXA-48-like and KPC and the other one for NDM and VIM). The assayed isolates were grown on SBA (SPL Life Sciences, Gyeonggi-do, Korea) and CHROMagar KPC medium (CHROMagar, Paris, France) for 16–24 hours at 37℃, and the assays were performed according to the manufacturer's instructions. An example of the identification results for OXA-48, KPC, NDM, and VIM using the OKNV assay is shown in Fig. 1. The sensitivity and specificity of the assay were calculated for all isolates. Sensitivity was calculated from the number of true-positive isolates, whereas specificity was calculated from the number of true-negative isolates.
All isolates that grew on both SBA and CHROMagar KPC media showed the same results in the OKNV assay. The OKNV assay could identify all isolates that produced one of the four targeted carbapenemase families, including eight OXA-48-like (OXA-48 [N=1], OXA-181 [N=3], and OXA-232 [N=4]), nine KPCs (KPC-2 [N=7] and KPC-4 [N=2]), eight NDMs (all NDM-1), and five VIMs (VIM-1 [N=2], and VIM-2 [N=3]). Additionally, it correctly identified 15 isolates that co-produced KPC and NDM (N=5), VIM and NDM (N=2), or OXA-48-like and NDM (N=8), but failed to identify one
The OKNV assay performed well, when using colonies grown on both SBA and CHROMagar KPC. It is possible that the negative result obtained for the NDM-1 and OXA-232 co-producing
In Korea, KPC, NDM, and OXA-48-like genes are expressed by 71%, 14%, and 9% of CPE, respectively, while VIM and IMP genes are very rare (<2% of CPE) . As the OKNV assay does not identify certain carbapenemases (mostly those belonging to the IMP family), the use of other diagnostic methods, such as PCR, is still required. Furthermore, another lateral flow assay, including IMP-type monoclonal antibodies, is currently under development . Although a few NDM-positive or VIM-positive isolates showed weak positive results, the OKNV assay was easy to use and provided easy-to-read results after only 15 min of incubation.
A limitation of this study is that a relatively small number of isolates were assayed; however, the sensitivity and specificity of the OKNV assay were similar to those reported in previous studies [10,11,12].
Overall, the speed and ease-of-use of the OKNV assay represent significant technical advances; the assay also shows excellent performance for detecting the OXA-48-like, KPC, NDM, and VIM-type carbapenemases from Enterobacteriaceae isolates cultured on both SBA and CHROMagar KPC medium.
Example of results for a KPC-2 and NDM-1 co-producing
Abbreviations: OXA, oxacillinase; KPC,
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