Rapid Bacterial Identification, Resistance, Virulence and Type Profiling using Selected Reaction Monitoring Mass Spectrometry

Abstract

Mass spectrometry (MS) in Selected Reaction Monitoring (SRM) mode is proposed for in-depth characterisation of microorganisms in a multiplexed analysis. Within 60–80 minutes, the SRM method performs microbial identification (I), antibiotic-resistance detection (R), virulence assessment (V) and it provides epidemiological typing information (T). This SRM application is illustrated by the analysis of the human pathogen Staphylococcus aureus, demonstrating its promise for rapid characterisation of bacteria from positive blood cultures of sepsis patients.

Deciphering Multifactorial Resistance Phenotypes in Acinetobacter baumannii by Genomics and Targeted Label-free Proteomics

“…If needed, more targeted proteins could be easily monitored; the method was very far from the multiplexing capabilities of SRM. Moreover, the robustness of SRM hyphenated with standard chromatography was shown to be compatible with clinical applications (19,51). A targeted strategy seems therefore a good compromise to accurately measure a reasonable number of proteins and to address biological questions at the protein level.…”

“…As in previous studies (18,19), the quality of the protein measurements was assessed by verifying the preservation of the ratio between transitions belonging to the same protein. The quantities of quantotypic peptides from a given protein should be equimolar if they are not impaired by posttranslational modifications, stochastic digestion, or affected by contaminants.…”

“…peptides that are specific for the resistance proteins inferred from WGS, were selected in silico as described (18,19,37). Briefly, proteins of interest from consensual UniProtKB sequences were digested in silico by trypsin using Skyline software (38).…”

“…On the contrary, targeted MS provides a robust and accurate quantification but at the cost of the detection of a lower number of proteins in a single multiplex. We reported the adequacy of a targeted approach to assess resistance in Pseudomonas aeruginosa (18) and Staphylococcus aureus (19), using a liquid chromatography hyphenated to an electrospray triple quadrupole mass spectrometer (LC-ESI-QqQ-MS) in Selected Reaction Monitoring mode (SRM). This technology is compatible with analytical requirements of accurate, sensitive, and selective assays for quantitation of dozens of resistance traits.…”

Identification of Microorganisms Grown in Blood Culture Flasks using Liquid Chromatography–Tandem Mass Spectrometry

“…Recent publications demonstrated that nontargeted LC-MS/MS-based approaches could analyze the susceptibility against β-lactam-based future science group www.futuremedicine.com antibiotics [32,33] in Gram-negatives. Next, by using a targeted approach (SRM mass spectrometry) was possible to detect methicillin resistance in S. aureus and virulence factors directly from positive blood cultures [34]. Furthermore, by the detection of amino acid substitutions in the GyrA protein using SRM it was possible to detect decreased quinolone susceptibility in typhoidal Salmonella isolates [35].…”