Rapid MRSA detection via tandem mass spectrometry of the intact 80 kDa PBP2a resistance protein

Treatment of antibiotic-resistant infections is dependent on the detection of specific bacterial genes or proteins in clinical assays. Identification of methicillin-resistant Staphylococcus aureus (MRSA) is often accomplished through the detection of penicillin-binding protein 2a (PBP2a). With greater dependence on mass spectrometry (MS)-based bacterial identification, complementary efforts to detect resistance have been hindered by the complexity of those proteins responsible. Initial characterization of PBP2a indicates the presence of glycan modifications. To simplify detection, we demonstrate a proof-of-concept tandem MS approach involving the generation of N-terminal PBP2a peptide-like fragments and detection of unique product ions during top-down proteomic sample analyses. This approach was implemented for two PBP2a variants, PBP2amecA and PBP2amecC, and was accurate across a representative panel of MRSA strains with different genetic backgrounds. Additionally, PBP2amecA was successfully detected from clinical isolates using a five-minute liquid chromatographic separation and implementation of this MS detection strategy. Our results highlight the capability of direct MS-based resistance marker detection and potential advantages for implementing these approaches in clinical diagnostics.

Validation of the Method for Determination of Melamine and Investigation its Trace in Milk from Vietnam by LC-MS/MS

This work describes a rapid, selective, and sensitive method by using liquid chromatography-tandem mass spectrometry (LC-MS/MS) to detect melamine (MEL) in milk and dairy products. The optimal conditions of liquid chromatographic separation extraction and mass spectroscopy of MEL have also been examined. The linear range for analyte detected by the method was 0.5÷100.0 ng/mL, with correlation coefficients was 0.999. Mean recoveries of the method in the real samples at three spike levels (low, medium, and high) were within the range of 98.5% ÷102.5% (n =7). LOD, LOQ values of the method were 10 and 30 ng/mL, respectively. The influence of the matrix effect on the accuracy, repeatability, and recovery of the process was insignificant. The proposed method was used to quantify the content of this compound in various real samples, which were collected in Ho Chi Minh City-Vietnam in 2020.