Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is used for the determination of molecular weights of different chemical compounds. We describe here the use of MALDI-TOF mass spectrometry to detect a carbapenem antibiotic, meropenem, and its degradation products. Buffered meropenem solution (0.1 mM Tris-HCl, pH 6.8) was mixed with an overnight culture of bacteria. After 3-h incubation, the reaction mixture was centrifuged, and the supernatant was analyzed by MALDI-TOF mass spectrometry. The presence or absence of peaks representing meropenem and its sodium salts was crucial. The average turnaround time of this test, considering the use of overnight culture, is 4 h. We validated this method for the detection of resistance to carbapenems in Enterobacteriaceae and Pseudomonas aeruginosa mediated by carbapenemase production. A total of 124 strains, including 30 carbapenemase-producing strains, were used in the study. The sensitivity of this method is 96.67%, with a specificity of 97.87%. Our results demonstrate the ability of this method to routinely detect carbapenemases in Enterobacteriaceae and Pseudomonas spp. in laboratories. This assay is comparable with a labor-intensive imipenem-hydrolyzing spectrophotometric assay that is a reference method for the detection of carbapenemase. As demonstrated here, MALDI-TOF mass spectrometry may be used in microbiological laboratories not only for microbial identification but also for other applications, such as studies of mechanisms of antibiotic resistance.

SUMMARYMatrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) has been successfully applied as an identification procedure in clinical microbiology and has been widely used in routine laboratory practice because of its economical and diagnostic benefits. The range of applications of MALDI-TOF MS has been growing constantly, from rapid species identification to labor-intensive proteomic studies of bacterial physiology. The purpose of this review is to summarize the contribution of the studies already performed with MALDI-TOF MS concerning antibiotic resistance and to analyze future perspectives in this field. We believe that current research should continue in four main directions, including the detection of antibiotic modifications by degrading enzymes, the detection of resistance mechanism determinants through proteomic studies of multiresistant bacteria, and the analysis of modifications of target sites, such as ribosomal methylation. The quantification of antibiotics is suggested as a new approach to study influx and efflux in bacterial cells. The results of the presented studies demonstrate that MALDI-TOF MS is a relevant tool for the detection of antibiotic resistance and opens new avenues for both clinical and experimental microbiology.

e Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is a potentially useful tool for the detection of antimicrobial resistance, especially that conferred by ␤-lactamases. Here we describe a modification of a previously reported MALDI-TOF MS meropenem hydrolysis assay. The modified method was validated on 108 carbapenemase-producing members of the Enterobacteriaceae, two NDM-1-producing Acinetobacter baumannii isolates, and 35 carbapenem-resistant enterobacteria producing no carbapenemase. The detection of carbapenemases by MALDI-TOF MS seems to be a powerful, quick, and cost-effective method for microbiological laboratories. Detection of carbapenemases has been one of the challenges in clinical microbiology diagnostics (9). Recently, new matrixassisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) assays for ␤-lactamase activity have been developed independently by at least three groups (2, 7, 12). These techniques are based on the detection of ␤-lactams and their degradation products. A similar assay was validated to detect carbapenemases in Acinetobacter baumannii (8).However, for some ␤-lactams, like meropenem, visualization of degradation products by MALDI-TOF MS seemed to be problematic (7,12). This might be due to binding of the molecules to cell lysate components. Here we describe a modification of one of these protocols (7) that allows the detection of degradation products and shortening of the turnaround time to ca. 2.5 h. The modified assay was validated with NDM-1-, VIM-1-, KPC-2-, KPC-3-, and OXA-48/-162-producing members of the Enterobacteriaceae and NDM-1-producing A. baumannii isolates.(The data included in this article were presented in part as a poster at the 22nd European Congress of Clinical Microbiology