Comparison of 11 Phenotypic Assays for Accurate Detection of Carbapenemase-Producing Enterobacteriaceae

Abstract

Early identification of carbapenemase-producing Enterobacteriaceae (CPE) is essential to prevent their dissemination within health care settings. Our objective was to evaluate the accuracy of 11 phenotypic assays for the detection of CPE. Two collections of carbapenem-resistant Enterobacteriaceae (CRE) isolates were evaluated, including 191 retrospective isolates (122 CP-CRE and 69 non-CP isolates) as well as 45 prospective clinical isolates (15 CP-CRE and 30 non-CP-CRE) obtained over a 3-month period. The sensitivity and specificity of each test was determined, with molecular genotype serving as the gold standard. Among the retrospective cohort, sensitivities ranged from 72% for the boronic acid synergy test for the detection of KPC producers to Ն98% for the modified Carba NP, the Rapidec Carba NP, the manual Blue Carba, and the modified carbapenem inactivation method for the detection of any CPE. Sensitivity differed among tests across enzyme classes. All assays had excellent specificity exceeding 95%, with the exception of the boronic acid synergy test (88%) and modified Hodge test (91%). Prospectively, 45 CRE isolates were encountered over a 3-month period, including 15 CPE (33%) and 30 non-CP-CRE (67%). Results from the prospective cohort were similar. However, a decrease in specificity was observed across most tests, likely due to restricted inclusion of non-CP-CRE to assess the specificity of the assays. Overall, accuracy of CPE detection varied across phenotypic tests. Local epidemiology of CP genotypes, turnaround time, and ease of incorporation into the laboratory workflow should be considered when selecting a phenotypic assay for clinical use.KEYWORDS carbapenem-resistant organisms, carbapenemase-producing organisms, prevalence C arbapenemase-producing carbapenem-resistant Enterobacteriaceae (CP-CRE) are associated with significant morbidity and mortality (1, 2). A variety of carbapenemase genes have been described that are either plasmid or chromosomally encoded, including bla KPC , bla SME , bla IMI , bla NDM , bla VIM , bla IMP , and bla OXA-48-type (2). Early and accurate identification of CP-CRE is essential to prevent their dissemination within health care settings.Detection of CP-CRE in clinical laboratories is challenging, as isolates may only have moderate reductions in susceptibilities to carbapenems (3), and resistance may be mediated by other mechanisms, such as extended-spectrum-␤-lactamase (ESBL) and/or AmpC ␤-lactamase producers with decreased membrane permeability. Molecular methods for the detection of carbapenemase genes are costly, may require significant expertise, and are limited by the targets included. Thus, rapid and affordable phenotypic assays to broadly classify CRE into CP-CRE versus non-CP-CRE have been developed.

Carbapenemase Detection among Carbapenem-Resistant Glucose-Nonfermenting Gram-Negative Bacilli

“…To the best of our knowledge, this is the most comprehensive evaluation of phenotypic assays for CPNF identification. The sensitivity of carbapenemase detection for P. aeruginosa was 93 to 100% for most assays and was comparable to the accuracy of these assays for CPE detection (4). In contrast, all tests had compromised sensitivity when used to identify carbapenemase-producing A. baumannii.…”

“…Of note, CLSI is reevaluating their endorsement of the Carba NP assay for detecting carbapenemase producers among carbapenem-resistant A. baumannii isolates. For more details on the relative pros and cons of the various phenotypic assays tested in this study, we refer the reader to previous work from these investigators (4). A notable limitation to this work is that there were small numbers of isolates producing any particular carbapenemase enzyme.…”

“…K. pneumoniae ATCC 1705 and ATCC 1706 were included daily as positive and negative controls, except for the metallo-␤-lactamase (MBL) Etest, where Stenotrophomonas maltophilia ATCC 13636 was included as the positive control. (18); (x) the carbapenem inactivation method (CIM; set up with a 10 l loopful of CRNF) (23); and (xi) the modified carbapenem inactivation method (mCIM; set up with a 10-l loopful of CRNF as described in the CLSI January 2017 AST Subcommittee meeting minutes [http://clsi.org/standards/micro/microbiology-files/]) (4,18). All methods were performed as previously described or according to the instructions in the package insert for commercially available assays.…”

“…Multiple phenotypic methods have been described for the detection of carbapenemase production among carbapenem-resistant Enterobacteria-ceae (CRE) (4). However, the performance of these assays among carbapenem-resistant glucose-nonfermenting isolates is not well described.…”

Is This the Carbapenemase Test We've Been Waiting for? A Multicenter Evaluation of the Modified Carbapenem Inactivation Method

“…In the original description of the assay, the authors reported 100% correlation with carbapenemase gene PCR. However, subsequent studies show reduced sensitivity of the CIM assay for the detection of both OXA-48 and NDM carbapenemases (21,22). To improve sensitivity for the detection of carbapenemases that are expressed at low levels or that naturally have weaker hydrolytic activities, the mCIM assay uses less organism (1-l versus 10-l loopful), a longer incubation period of the disk-bacterial suspension (4 h versus 2 h), and a different suspension matrix (tryptic soy broth versus water) than the original CIM assay.…”