Protein biomarker elucidation for the verification of biological agents in the taxonomic group of Gammaproteobacteria using tandem mass spectrometry.

Some pathogenic microbes can be used for nefarious applications and instigate population-based fear. In a bio-threat scenario, rapid and accurate methods to detect biological agents in a wide range of complex environmental and clinical matrices, is of paramount importance for the implementation of mitigation protocols and medical countermeasures. This study describes targeted and shot-gun tandem MS based approaches for the verification of biological agents from the environmental samples. The marker proteins and peptides were elucidated by an exhaustive literature mining, in silico analysis of prioritized proteins, and MS/MS analysis of abundant proteins from selected bacterial species. For the shot-gun methodology, tandem MS analysis of abundant peptides was carried from spiked samples. The validation experiments employing a combination of shot-gun tandem MS analysis and a targeted search reported here is a proof of concept to show the applicability of the methodology for the unambiguous verification of biological agents at sub-species level, even with limited fractionation of crude protein extracts from environmental samples.