Improved Blue Carba test and Carba NP test for detection and classification of major Class A and B carbapenemases, including dual producers, among Gram-negative bacilli.

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Rapid and accurate identification of carbapenemase-producing organisms is of vital importance in guiding appropriate clinical antibiotic treatments and curbing their transmission. The emergence of negative bacilli carrying multiple carbapenemase combinations during and after the severe acute respiratory syndrome coronavirus 2 pandemic has posed a challenge to the conventional biochemical tests typically used to determine the specific carbapenemase type in the isolated strains. Several initiatives have aimed to enhance colorimetric methods, enabling them to independently identify the presence of Class A or Class B carbapenemases. Notably, no previous efforts have been made to distinguish both classes simultaneously. Additionally, these modifications have struggled to differentiate between carriers of multiple carbapenemases, a common occurrence in many Latin American countries. In this study, we introduced specific Class A and Class B carbapenemase inhibitors into the Blue Carba test (BCT) and Carba NP-direct (CNP) colorimetric assays to identify the type of carbapenemase, even in cases of multiple carbapenemase producers within these classes. These updated assays demonstrated exceptional sensitivity and specificity (≥ 90%) all within a rapid turnaround time of under 2 hours, typically completed in just 45 minutes. These in-house enhancements to the BCT and CNP assays present a rapid, straightforward, and cost-effective approach to determining the primary carbapenemase classes. They could serve as a viable alternative to molecular biology or immuno-chromatography techniques, acting as an initial diagnostic step in the process.