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.

Determination of organic acids in aerosol particles from a coniferous forest by liquid chromatography-mass spectrometry.

LC-MS methods with use of ion-trap and time-of-flight mass spectrometers were developed for the determination of organic acids in aerosol samples collected by a high-volume sampler in a Finnish coniferous forest. Comparison was made of the composition of samples collected during atmospheric formation of new aerosol particles and on days when this formation did not occur. A dynamic sonication-assisted solvent extraction system was developed for fast and quantitative extraction of the filter samples. Several organic acids, including pinonic acid, pinic acid, and homologous series of n-alkanoic acids, n-alkenoic acids, and aliphatic dicarboxylic acids, were identified. In samples collected between 08:00 and 16:00 hours the concentration of pinonic acid ranged from 0.5 ng m(-3) to 3.7 ng m(-3) and that of pinic acid from 0.2 ng m(-3) to 1.5 ng m(-3). For most of the compounds identified, the trends in concentration could be explained by the differences in temperature during collection. However, concentrations of short-chain n-alkanoic acids were clearly higher on the days when new aerosol particle formation occurred.