Introduction. Rapid bacterial antibiotic susceptibility test (AST) and minimum inhibitory concentration (MIC) measurement are important to improve clinical care but also to help reduce both the widespread misuse of antibiotics and the growing drug-resistance problem. In recent years, matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) has become established as a first-line diagnostic tool in the identification of microorganisms, including those producing human infections. Rapid detection of antimicrobial resistance is one of the future applications of this technique with the greatest likelihood of success. In this study, different protocols of meropenem hydrolysis assay (MHA) by MALDI-TOF MS were applied in order to develop a valid tool for the phenotypic discrimination between carbapenemases- producing and -non-producing Gram-negative bacteria. Furthermore, we evaluate the performance of the innovative Accelerate PhenoTM System (APS, Accelerate Diagnostics) for ID and AST directly from positive blood cultures (BCs) in comparison to conventional methods (CM). Materials and Methods. MHA was applied to 1185 Enterobacteriaceae strains for the phenotypic detection of carbapenemases. In addition, a preliminary study on 24 Enterobacteriaceae strains was performed to evaluate the possibility of detecting carbapenemase-producing strains by directly analysis of the MALDI-TOF spectra obtained for the identification of microorganisms, avoiding the meropenem hydrolysis assay. Finally, the performances of the APS were evaluated on 23 positive blood cultures and 10 challenge isolates. Results. The MHA was successfully applied to detect carbapenemase activity in 981 well- characterized Enterobacteriaceae strains producing KPC or VIM carbapenemases, and in 146 carbapenem fully susceptible strains. This assay, applied also to NDM and OXA-48-producing strains and to CRE with resistance mechanisms other than carbapenemase production, has proved to be able to distinguish between carbapenemase-producing and -nonproducing Enterobacteriaceae. APS performed on 10 challenge isolates showed an ID concordance of 100% as well as 9 AST, in one case AST wasn’t provided due to very high clones count by APS. The evaluation on 23 positive blood cultures showed a ID sensitivity and specificity of 94.7% and 99,7%, respectively. The AST performance showed 88.3% of essential agreement, 85.6% of categorical agreement, 0% of very major error, 5.8% of major error, and 9.9% of minor error. Discussion and Conclusions. In our experience, MHA demonstrated to be able to reveal the hydrolyzation of meropenem in all KPC or VIM carbapenemases, whereas no false positive results were obtained from fully susceptible Enterobacteriaceae strains. In our hands APS showed noteworthy errors both for ID and AST, despite the high sensitivity and specificity observed for ID, and the AST performance is encouraging by the absence of very major error. However, the reduction in time-to-result represents one of the main advantage of this assay.
Rapid detection of bacteria antimicrobial resistance by MALDI-TOF mass spectrometry and Accelerate phenoTM system / Calderaro, Adriana; Martinelli, Monica; Buttrini, Mirko; Covan, Silvia; Ruggeri, Alberto; Montecchini, Sara; Arcangeletti, Maria Cristina; Chezzi, Carlo; DE CONTO, Flora. - (2018), p. 108. ((Intervento presentato al convegno 46° Congresso Nazionale della Società Italiana di Microbiologia tenutosi a Palermo nel 26-29 settembre 2018.