Assessment of an LC coupled with tandem MS (LC-MS/MS) tool in the detection of antimicrobial resistance mechanisms in Enterobacterales.

To assess a new MS approach in detection of antimicrobial resistance (AMR) mechanisms in Enterobacterales and its usefulness in the routine practice of a national reference centre for carbapenemase-producing enterobacteria (CPE). A tool utilizing LC coupled with tandem MS (LC-MS/MS) in multiple reaction monitoring mode was tested in the direct and concurrent detection of 13 AMR enzymes. These comprised carbapenemases, extended-spectrum β-lactamases (ESBLs), AmpC-like cephalosporinases, aminoglycoside-modifying enzymes (AMEs) and 16S rRNA methylases. The study included 357 Enterobacterales isolates largely from Polish hospitals (2000-23), including 292 CPE, collected by the National Reference Centre for Susceptibility Testing. The isolates were short-read sequenced to determine their resistomes, and subjected to MIC evaluation. The number of target AMR genes in the isolates was 1128 without defective and duplicated copies. When possible to deduce, the MIC patterns indicated activity of most of the genes/enzymes. The LC-MS/MS assay showed >95% sensitivity in the identification of KPC-, NDM-, VIM-, IMP- and OXA-48-type carbapenemases, CTX-M-like ESBLs, CMY-2- and DHA-type AmpCs, Aac(3)-II- and Aac(6')-Ib-like AMEs and the ArmA 16S rRNA methylase, but only 66.7% and 44.4% of GES-type ESBLs/carbapenemases and RmtB/C-like methylases, respectively. Detection specificity was almost 100% for all of the enzymes. The study demonstrated good performance of the assay against multiple clinically-relevant AMR mechanisms, including main carbapenemase types, and its usefulness in a CPE reference centre was confirmed. However, further studies, e.g. with other bacterial populations, AMR mechanisms and clinical samples, are necessary to characterize its full diagnostic potential.