Multicenter Performance Assessment of Carba NP Test

Abstract

Eighty Gram-negative bacilli (54 Enterobacteriaceae and 26 nonfermenting Gram-negative bacilli) obtained from multiple institutions in the United States were distributed in a blinded manner to seven testing laboratories to compare their performance of a test for detection of carbapenemase production, the Carba NP test. The Carba NP test was performed by all laboratories, following the Clinical and Laboratory Standards Institute (CLSI) procedure. Site-versus-site comparisons demonstrated a high level of consistency for the Carba NP assay, with just 3/21 site comparisons yielding a difference in sensitivity (P Ͻ 0.05). Previously described limitations with bla OXA-48-like carbapenemases and bla OXA carbapenemases associated with Acinetobacter baumannii were noted. Based on these data, we demonstrate that the Carba NP test, when implemented with the standardized CLSI methodology, provides reproducible results across multiple sites for detection of carbapenemases.KEYWORDS carbapenemase, antimicrobial resistance, screening, Gram-negative bacilli C linical microbiology laboratories around the globe have instituted a range of rapid screening tests for detection of carbapenemase production in the rising tide of carbapenem-nonsusceptible Gram-negative bacilli (GNB). One method, the Carba NP test, offers a rapid and simple colorimetric method for the detection of carbapenemase activity in isolates of GNB. In principle, the test is similar to the classic ␤-lactamase detection assays described by Escamilla and by Skinner and Wise in the 1970s, in which benzylpenicillin acidimetrically produced a red color after hydrolysis by Haemophilus influenzae ␤-lactamase (1, 2). With the Carba NP test, hydrolysis of imipenem produces acid and drives the pH indicator, phenol red, from red to yellow.The Carba NP test has been studied by several investigators (3-14), including evaluations and comparisons to a commercial version (15)(16)(17)(18)(19)(20). However, the literature lacks consistency in the Carba NP methodology. The test has evolved several times, including alterations to reagents, and has gone from a microtiter plate-based format performed on bacterial protein extracts to a rapid tube-based test with abbreviated processing. Most studies have been single-laboratory evaluations; there is a lack of multicenter performance assessments utilizing a single, harmonized method.As part of an effort to standardize the method, the Clinical and Laboratory Standards Institute (CLSI) Subcommittee on Antimicrobial Susceptibility Testing

Multicenter Evaluation of the Modified Carbapenem Inactivation Method and the Carba NP for Detection of Carbapenemase-Producing Pseudomonas aeruginosa and Acinetobacter baumannii

“…As the performance characteristics for the mCIM among A. baumannii were not favorable in our study, the question arose as to whether the Carba NP test was sufficiently evaluated by CLSI against carbapenem-resistant P. aeruginosa and A. baumannii isolates before it was first endorsed for use with these nonfermenters in the 2015 CLSI M100 document. At the time of the evaluation of the Carba NP test by a CLSI working group, there were fewer well-characterized isolates of P. aeruginosa (n ϭ 12) and A. baumannii (n ϭ 14) available to challenge the assay and less information available regarding CHDL in A. baumannii (8). The mean sensitivity, specificity, and kappa coefficient of the Carba NP in our present study were 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100), 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100) and 0.91 for P. aeruginosa and 18.8% (95% CI, 10.4 to 30.1; range, 8.7 to 26.1), 100% (95% CI, 83.9 to 100; range, 100), and 0.83 for A. baumannii.…”

“…Carba NP multisite evaluation. The Carba NP method was performed and interpreted as defined by CLSI (8,10). The recommended QC strains were set up with each day of testing.…”

“…Many phenotypic methods for detection of carbapenemase producers have been described for the Enterobacteriaceae, some of which can be applied to the nonfermenters, such as P. aeruginosa and A. baumannii (2,7). Phenotypic methods defined by the Clinical and Laboratory Standards Institute (CLSI) that clinical microbiology laboratories can apply to detect carbapenemase producers include the modified Hodge test (to be removed from the M100 in 2018; CLSI, January 2017 meeting minutes), the Carba NP test, and most recently the modified carbapenem inactivation method (mCIM) (8,9). To date, only the Carba NP test has been endorsed for the detection of carbapenemase production among P. aeruginosa and A. baumannii strains (10).…”

The Clinical and Molecular Epidemiology of Noncarbapenemase-Producing Carbapenem-Resistant Enterobacteriaceae: A Case-Case-Control Matched Analysis

“…Results were visualized using GrapeTree (v1.5) [ 33 ]. The assemblies were searched for antimicrobial resistance genes using c-SSTAR (v1.2c, with the ResGANNOT_srst2.fasta db—07082018) ( https://github.com/tomdeman-bio/Sequence-Search-Tool-for-Antimicrobial-Resistance-SSTAR-/blob/master/ARG-ANNOT.srst2_July-12-2016.fasta ) [ 34 , 35 ] and ABRicate (v0.8.13, with the CARD database from July 2019) [ https://github.com/tseemann/abricate ¡ [ 36 ].…”

Biographical Feature: Stephen G. Jenkins, Ph.D., D(ABMM), F(AAM)

“…He has studied the emergence and spread of antibiotic-resistant organisms (9)(10)(11)(12)(13)(14) as well as the methods for and utility of detecting such organisms in the microbiota (15)(16)(17)(18). Dr. Jenkins has published on methods of susceptibility testing throughout his career, most recently on detection of carbapenem resistance in Gram-negative bacilli (19)(20)(21). His interest in rapid diagnostic testing has been driven by a desire to improve patient care by providing timely results, with his research including studies of molecular and phenotypic tests to detect the spread of clonal organisms in the hospital setting, resistant organisms, and respiratory virus infections (22)(23)(24)(25)(26).…”