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

The purpose of this study was to develop the modified carbapenem inactivation method (mCIM) for the detection of carbapenemase-producing Pseudomonas aeruginosa (CP-PA) and carbapenemase-producing Acinetobacter baumannii (CP-AB) and perform a multicenter evaluation of the mCIM and Carba NP tests for these nonfermenters. Thirty P. aeruginosa and 30 A. baumannii isolates previously characterized by whole-genome sequencing from the CDC-FDA Antibiotic Resistance Isolate Bank were evaluated, including CP isolates (Ambler class A, B, and D), non-carbapenemase-producing (non-CP) carbapenem-resistant isolates, and carbapenem-susceptible isolates. Initial comparison of a 1-µl versus 10-µl loop inoculum for the mCIM was performed by two testing sites and showed that 10 µl was required for reliable detection of carbapenemase production among P. aeruginosa and A. baumannii Ten testing sites then evaluated the mCIM using a 10-µl loop inoculum. Overall, the mean sensitivity and specificity of the mCIM for detection of CP-PA across all 10 sites were 98.0% (95% confidence interval [CI], 94.3 to 99.6; range, 86.7 to 100) and 95% (95% CI, 89.8 to 97.7; range, 93.3 to 100), whereas the mean sensitivity and specificity among CP-AB were 79.8% (95% CI, 74.0 to 84.9; range, 36.3 to 95.7) and 52.9% (95% CI, 40.6 to 64.9; range, 28.6 to 100), respectively. At three sites that evaluated the performance of the Carba NP test using the same set of isolates, the mean sensitivity and specificity of the Carba NP test were 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100) and 97.8% (95% CI, 88.2 to 99.9; range, 93.3 to 100) for P. aeruginosa and 18.8% (95% CI, 10.4 to 30.1; range, 8.7 to 26.1) and 100% (95% CI, 83.9 to 100; range, 100) for A. baumannii Overall, we found both the mCIM and the Carba NP test to be accurate for detection of carbapenemase production among P. aeruginosa isolates and less reliable for use with A. baumannii isolates.

High-Stringency Evaluation of the Automated BD Phoenix CPO Detect and Rapidec Carba NP Tests for Detection and Classification of Carbapenemases.

There is an urgent need for rapid, accurate detection and classification of carbapenemases. The current study evaluated the automated BD Phoenix CPO Detect and the manual bioMérieux Rapidec Carba NP tests for meeting these needs. Both tests were challenged with 294 isolates of Enterobacteriaceae spp., Pseudomonas aeruginosa, and Acinetobacter baumannii chosen to provide extreme diagnostic difficulty. Carbapenemases such as KPC, NMC-A, IMI, SME, NDM, SPM, IMP, VIM, and OXA-23, 40, 48, 58, 72, 181, and 232 were produced by 243 isolates and 51 carbapenemase-negative isolates included porin mutants and producers of extended-spectrum ß-lactamases (ESBLs), AmpCs, K1, and broad-spectrum ß-lactamases. Both tests exhibited high sensitivity of carbapenemase detection (>97%). Due to the highly challenging carbapenemase-negative isolates, specificities were lower than typical for evaluations involving mostly routine clinical isolates. BD Phoenix CPO Detect was 68.6% specific and Rapidec Carba NP was 60.8% to 78.4% specific, depending on how borderline results were interpreted. Only BD Phoenix CPO Detect classified carbapenemases. It correctly classified 85.0% of class A, 72.4% of class B, and 88.6% of class D carbapenemases. Importantly with respect to empirical therapy with new ß-lactamase inhibitor combinations such as ceftazidime/avibactam, no class B carbapenemases were misclassified as class A carbapenemases. Both tests offer advantages. Used alone, without initial susceptibility tests, Rapidec Carba NP can provide positive results for some isolates after only 10 to 30 min incubation. BD Phoenix CPO Detect provides novel advantages such as automated carbapenemase detection, inclusion in susceptibility panels to eliminate delays and subjectivity in initiating carbapenemase tests, and classification of most carbapenemases.

Modified Carbapenem Inactivation Method for Phenotypic Detection of Carbapenemase Production among Enterobacteriaceae.

The ability of clinical microbiology laboratories to reliably detect carbapenemase-producing carbapenem-resistant Enterobacteriaceae (CP-CRE) is an important element of the effort to prevent and contain the spread of these pathogens and an integral part of antimicrobial stewardship. All existing methods have limitations. A new, straightforward, inexpensive, and specific phenotypic method for the detection of carbapenemase production, the carbapenem inactivation method (CIM), was recently described. Here we describe a two-stage evaluation of a modified carbapenem inactivation method (mCIM), in which tryptic soy broth was substituted for water during the inactivation step and the length of this incubation was extended. A validation study was performed in a single clinical laboratory to determine the accuracy of the mCIM, followed by a nine-laboratory study to verify the reproducibility of these results and define the zone size cutoff that best discriminated between CP-CRE and members of the family Enterobacteriaceae that do not produce carbapenemases. Bacterial isolates previously characterized through whole-genome sequencing or targeted PCR as to the presence or absence of carbapenemase genes were tested for carbapenemase production using the mCIM; isolates with Ambler class A, B, and D carbapenemases, non-CP-CRE isolates, and carbapenem-susceptible isolates were included. The sensitivity of the mCIM observed in the validation study was 99% (95% confidence interval [95% CI], 93% to 100%), and the specificity was 100% (95% CI, 82% to 100%). In the second stage of the study, the range of sensitivities observed across nine laboratories was 93% to 100%, with a mean of 97%; the range of specificities was 97% to 100%, with a mean of 99%. The mCIM was easy to perform and interpret for Enterobacteriaceae, with results in less than 24 h and excellent reproducibility across laboratories.

Correlation of cefoxitin MICs with the presence of mecA in Staphylococcus spp.

This report describes the results of an 11-laboratory study to determine if a cefoxitin broth microdilution MIC test could predict the presence of mecA in staphylococci. Using breakpoints of < or = 4 microg/ml for mecA-negative and > or = 6 or 8 microg/ml for mecA-positive isolates, sensitivity and specificity based on mecA or presumed mecA for Staphylococcus aureus at 18 h of incubation were 99.7 to 100% in three cation-adjusted Mueller-Hinton broths tested. For coagulase-negative strains at 24 h of incubation, breakpoints of < or = 2 microg/ml for mecA-negative and > or = 4 microg/ml for mecA-positive isolates gave sensitivity and specificity of 94 to 99% and 69 to 80%, respectively.

Detection of inducible clindamycin resistance in staphylococci by broth microdilution using erythromycin-clindamycin combination wells.

A study conducted by 11 laboratories investigated the ability of four combinations of erythromycin (ERY) and clindamycin (CC) (ERY and CC at 4 and 0.5, 6 and 1, 8 and 1.5, and 0.5 and 2 microg/ml) in a single well of a broth microdilution panel to predict the presence of inducible CC resistance. Each laboratory tested approximately 30 Staphylococcus aureus isolates and 20 coagulase-negative staphylococcus (CoNS) isolates in a panel using cation-adjusted Mueller-Hinton broth from three different manufacturers. Only the strains resistant to ERY and those susceptible or intermediate to CC were included in the analysis (S. aureus, n = 333; CoNS, n = 97). Results of the D-zone test were used as the gold standard. After an 18-h incubation, the combination of 4 microg/ml ERY and 0.5 microg/ml CC performed the best, with 98 to 100% sensitivity and 100% specificity for both organism groups. After a 24-h incubation, the ERY-CC combinations of 4 and 0.5, 6 and 1, and 8 and 1.5 microg/ml correlated well with the D-zone test.