Novel Peptide Biomarker Discovery for Detection and Identification of Bacterial Pathogens by LC-ESI-MS/MS

Abstract

The increase of foodborne diseases in the past years, especially emergent pathogens is a great concern for authorities, industry and consumers. It is important the early detection and identification of the microbiota in order to evaluate the safety of the product and avoid health issues and economic losses for either the producer or the distributor. Consequently, there is a critical need to develop rapid and sensitive methods for detection and accurate identification of food-borne pathogens. Our approach is the description of peptide/protein markers unique for a bacterial species, strain or genera. The identification of bacterial peptide/protein markers was performed after protein extraction, in-solution tryptic digestion and analysizing its tryptic peptides by liquid chromatographyelectrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Subsequent analysis with database search software and comparison with amino acid sequences available in public databases (Uniprot and NCBI) allowed the identification of several peptide biomarkers. In this study we have focused on two proteins of the species Staphylococcus aureus, and compared them with the public databases and with the data obtained from LC-ESI-MS/MS of the trysinized proteins from Gram-positive and Gram-negative bacteria such as Streptococcus spp., Enterococcus spp., Bacillus spp., Listeria spp, Salmonella spp., Enterobacteriaceae. As a result, cysteine synthase and cysteine proteases stophapain A and stophapain B, and 1-pyrroline-5-carboxylate dehydrogenase proteins may be used for S. aureus differentiation. Furthermore, cysteine protease peptides AQKPVDNITQIIGGTPVVK, TESIPTGNNVTQLK, MTTYNEVDNLTK, YTINVSSFLSK and KTGSPDYLLHFLEQKV resulted specific for S. aureus, being potentially useful for a fast detection of bacterial pathogens directly in food products and not requiring previous culture of the bacteria