Rapid direct detection of carbapenemase-producing Enterobacteriaceae in clinical urine samples by MALDI-TOF MS analysis

Abstract

Objectives: Development of an automated MALDI-TOF MS-based method for the rapid, direct detection of carbapenemase-producing Enterobacteriaceae in clinical urine samples within 90 min of sample reception.

Methods: A total of 3041 urine samples were processed by flow cytometry, and a cut-off value of ≥1.5 × 10 5 bacteria/mL was used to select samples for the study. Following these criteria, 608 samples were selected for direct bacterial identification. Detection of carbapenemase activity by MALDI-TOF MS analysis was only performed after reliable direct identification of Gram-negative bacilli. A novel protocol was developed for extracting bacteria from urine samples by using the Sepsityper Kit (Bruker Daltonik, Germany). Carbapenem resistance was detected with imipenem as an antibiotic marker and the results were automatically interpreted using the STAR-BL module of MALDI-TOF Biotyper Compass software (Bruker Daltonik, Germany).

Results: The MALDI-TOF MS-based assay yielded direct reliable identification of 91% (503/553) of the samples. The assay showed 100% sensitivity (30/30) and specificity (454/454) for detecting carbapenemase activity within 90 min of sample reception. Isolates included in the study were further characterized by PCR and sequencing, and bla OXA-48 was detected from all isolates that tested positive in the MALDI-TOF MS-based resistance assay.

Conclusions: The proposed protocol for the direct analysis of urine samples by MALDI-TOF MS is suitable for use in clinical laboratories to identify bacteria and detect carbapenemase activity, thus saving at least 24-48 h relative to current routine methods.