Resumo

Current knowledge about sheep and goat caseous lymphadenitis indicates that the resistance to Corynebacterium pseudotuberculosis (Cp) infection is a complex process in which humoral and cellular immune responses are both operative. Considering that no ideal antigen candidates have been identified to date for the development of efficient vaccines, this study aimed to identify overlapping T and B cells linear epitopes within selected Cp proteins through an in silico prediction approach. A reverse vaccinology strategy was applied to predict protein targets based on genome sequences of the Cp strains 1002, C231, FRC41 and I19. The prediction criteria included the extracellular location, presence of transmembrane helices, adhesin probability higher than 0.51 and high conservation degree among the investigated Cp strains. After targets selection, the T cell epitopes were identified using prediction tools located at both the Vaxign pipeline and the Immune Epitope Database (IEDB); these epitopes were classified based on their binding affinity for mouse major histocompatibility complex (class I and II MHC) alleles using the consensus method. ElliPro was used to predict linear B cell epitopes, using modeled 3D structure templates. Three proteins were identified meeting all the criteria established for the reverse vaccinology: metalloendopeptidase-like membrane protein (ADL20536), putative guanyl-specific ribonuclease (ADL21376) and putative secreted protein with LPxTG motif (ADL21554); however, only the first protein had overlapping T and B cells epitopes predicted. In general, the metalloendopeptidase presented 28 epitopes binding to class I MHC alleles, 51 epitopes binding to class II MHC alleles and 11 linear antibody epitopes. The peptides “TSAKLGDSAPQILAIQEFKPNTNL” and “KAINYNEERALAD” were predicted to be antibody linear epitopes (PI=0.749 and PI=0.742, respectively) located in the outside region of the protein. These peptides present overlapping residues sequences with the T cell epitopes “AINYNEERA”, “APQILAIQE” and “KPNTNLSDQ”, which presented high binding affinity (p<0.05 in Vaxign and IC50 <50 in IEDB-AR) for both class I and II MHC molecules. Although in silico data requires in vitro validation, the Cp metalloendopeptidase has already shown to be reactive in an immunoblot using serum from infected goats. These overlapping epitopes might be used as part of a designed vaccine model to stimulate both humoral and cellular immune responses.