Detection of NDM-1, VIM-1, KPC, OXA-48, and OXA-162 Carbapenemases by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

Abstract

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

Efficient Detection of Carbapenemase Activity in Enterobacteriaceae by Matrix-Assisted Laser Desorption Ionization−Time of Flight Mass Spectrometry in Less Than 30 Minutes

“…MALDI-TOF MS has recently been shown to be a reliable method for KPC and NDM detection in Enterobacteriaceae with high sensitivity and specificity (15,16,(30)(31)(32)(33)(34)(35). Only one study investigated 6 OXA-48 producers (33).…”

Evaluation of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry for Rapid Detection of β-Lactam Resistance in Enterobacteriaceae Derived from Blood Cultures

“…After vortexing, the sample was centrifuged at 14,000 rpm for 2 min, the supernatant was discarded, and the pellet was washed with 1 ml of deionized water. The water was thoroughly removed, and the bacteria (pellets contained 10 7 to 10 9 CFU) were resuspended in 10 l of antibiotic solution (0.5 mg/ml cefotaxime [CTX]) (Serva Electrophoresis GmbH, Heidelberg, Germany) in water supple-mented with 0.01% sodium dodecyl sulfate (SDS) (Sigma-Aldrich, Taufkirchen, Germany) or 10 mg/ml ampicillin (Sigma-Aldrich) in water without SDS (14,15). The suspensions were incubated in an Eppendorf Thermomixer comfort (Eppendorf, Hamburg, Germany) at 37°C with agitation (950 rpm) for 90 min.…”

“…Thus, methods that allow early prediction of antibiotic susceptibility are of great interest. Recently, it has been shown that MALDI-TOF MS can be used to detect resistance against ␤-lactam antibiotics (14)(15)(16)(17). The mass spectrometric ␤-lactamase assay is based on a short coincubation of the Gramnegative bacterium with the antibiotic in question.…”

Carbapenem-resistant Enterobacteriaceae in special populations: Solid organ transplant recipients, stem cell transplant recipients, and patients with hematologic malignancies

“…Two real-time, multiplex polymerase chain reaction systems have recently been approved in the US and can identify carbapenemase genes within 2 hours of blood culture positivity. [101][102][103] Implementation of these tools can decrease the time from blood culture collection until detection of CRE from 3 d to less than 24 hours. Furthermore, these systems also allow clinicians to de-escalate therapy when appropriate and avoid the attendant toxicities of CREactive agents.…”