Identification of conserved Mycoplasma agalactiae surface antigens by immunoproteomics

Contagious agalactia represents one of the most relevant infectious diseases dairy sheep, with Mycoplasma agalactiae being primary etiological agent. The early, sensitive, and specific identification infected animals, as well development efficient prophylactic tools, remain challenging. Here, we present a comprehensive characterization M. antigens focusing on those shared among different isolates. Leveraging previous proteomic data obtained individual strains, adopted strategy entailing sample pooling to optimize conserved proteins that induce an immune response. liposoluble from previously characterized field isolates type strain PG2T were enriched by Triton X-114 fractionation, pooled, analysed one-dimensional (1D) two-dimensional (2D) electrophoresis, subjected western immunoblotting against sheep sera collected during natural infection agalactiae. Immunodominant identified Matrix-Assisted Laser Desorption-Time-Of-Flight-Mass Spectrometry (MALDI-TOF-MS). This combined immunoproteomic approach confirmed role several known immunogens, including P80, P48, P40, variable surface (Vpmas), unveiled novel immunodominant, antigens, MAG_1000, MAG_2220, MAG_1980, phnD, MAG_4740, MAG_2430. Genomic context, functional prediction, subcellular localization, invariable expression these in all suggest their possible involvement bacterial pathogenicity metabolism. Moreover, elicit host humoral response since early stages infection, persisting for at least 270 days. antigen panel this work supports effective vaccines diagnostic tools higher sensitivity specificity stages.