resaCPE: a rapid, low-cost colorimetric assay for the detection of carbapenemase-producing Enterobacterales

The use of carbapenem antibiotics is threatened by the global spread of carbapenemase-producing Enterobacterales (CPE), bacterial pathogens which hydrolyze these last-resort antimicrobials. Rapid detection of CPE is vital to ensure timely administration of antimicrobial therapy to infected patients, as well as the implementation of infection control measures to prevent outbreaks in healthcare settings. In this study, we report the development of resaCPE, a rapid, low-cost CPE detection method that couples the inactivation of an imipenem disk with a cell viability assay employing a carbapenem-hypersusceptible Escherichia coli strain. Results are interpreted by a simple colorimetric readout in which CPE-positive samples turn pink, while CPE-negative samples remain purple. The assay was validated with a panel of 116 CPE and non-CPE isolates, and its performance was compared with two widely used CPE detection strategies: the modified carbapenem inactivation method (mCIM) and the CARBA-NP test. The resaCPE test demonstrated a sensitivity of 98.7%, outperforming the CARBA-NP (87.8% sensitivity) and the mCIM (96.3% sensitivity) assays. The resaCPE test provides results within 3.5 h, significantly faster than the mCIM, and demonstrated a high degree of specificity (94.1%). Due to the rapid turnaround time, minimal setup requirements, and low cost (~$1 USD/sample), the resaCPE test is a potentially attractive option for primary CPE screening, particularly in lower resource settings.IMPORTANCECarbapenem antibiotics serve as a last line of defense against severe bacterial infections. However, these antibiotics are endangered by bacterial pathogens which produce carbapenemases, enzymes that inactivate the carbapenem antibiotics, rendering them ineffective. Infections caused by carbapenemase-producing bacteria are associated with high mortality and pose a significant threat to global health. Rapid and reliable detection of these pathogens is critical to ensure timely treatment and prevent their spread in healthcare settings. Current detection methods suffer from long turnaround times, high cost, or the requirement of specialized laboratory equipment. In this study, we developed resaCPE, a simple, inexpensive, and rapid colorimetric test that identifies carbapenemase-producing bacteria in 3.5 h. This test demonstrated excellent performance when compared to currently employed assays. Due to the rapid turnaround time, low cost, and minimal equipment requirements, the resaCPE test could be a promising alternative for routine carbapenemase screening, particularly in laboratories with limited resources.