Molecular characterization of Anaplasma ovis Msp4 protein in strains isolated from ticks in Turkey: A multi-epitope synthetic vaccine antigen design against Anaplasma ovis using immunoinformatic tools

Background: Tick-borne pathogens increasingly threaten animal and human health as well as cause great economic loss in the livestock industry. Among these pathogens, Anaplasma oviscausing a decrease in meat and milk yield is frequently detected in sheep in many countries including Turkey. Unfortunately a vaccine has not been developed against Anaplasma ovis yet. Msp4 is a immunogenic surface protein of Anaplasma bacteria. This study aimed to reveal potential vaccine candidate epitopes in Msp4 protein using sequence data belonging to Msp4 gene of Anaplasma ovis isolates and then to design a multi-epitope protein to be used in vaccine formulations against Anaplasma ovis. Results: For this purpose, Msp4 gene was sequenced from Anaplasma ovis isolates (n:6) detected in ticks collected from sheep in Turkey and the sequence data was compared with previous sequences from different countries in order to detect the variations of Msp4 gene/protein. Later, potential vaccine candidate and diagnostic epitopes were predicted using various immunoinformatics tools including physicochemical parameters, secondary structures, antigenicity, solubility, subcellular localization, transmembrane helix, signal peptide, allergenicity, BetaWrap motif, homology checking, postranslational modifications, B/T cell epitopes, toxicity, solvent exposure, and epitope conservancy. Among the discovered vaccine candidate epitopes, antigenic and conserved were selected, and then a multi-epitope protein was designed. The designed vaccine protein was tested for the assessment of TLR-2, IgG, and IFN-g responses by molecular docking and immune simulation analyses. Finally, among the discovered epitopes of Anaplasma ovis, EVASEGSGVM and YQFTPEISLV epitopes that have properties of high antigenicity, non-allergenicity, and non-toxicity were proposed for the development of peptide-based vaccines and serodiagnostic assays. Conclusions: This study provides prominent epitopes and a candidate vaccine model to be used for the tick-borne pathogen Anaplasma ovis in further serodiagnostic and vaccine studies.