Reverse vaccinology-driven construction and bioinformatics validation of a multi-epitope vaccine against Brucella spp.

Brucella spp., the causative agent of zoonotic brucellosis, poses a significant threat to both animal husbandry and human health. Given the limitations of current antibiotic treatments for human brucellosis, there is an urgent need to develop effective human vaccines. Using reverse vaccinology and bioinformatics approaches, we designed a multi-epitope vaccine (MEV) specifically targeting the CcmA, ccmC, and BepC membrane proteins, with the aim of providing robust protection against human infection.We screened 11 CTL, 9 HTL, 2 conformational B cell epitopes, and 9 linear B cell epitopes, linked them with GGGS,GPGPG and KK. and incorporated HMGN1 (high mobility group nucleosome binding protein 1) as an adjuvant and PADRE (a universal HTL epitope to enhance cross-population immunogenicity) to boost immunogenicity. Analyses showed the 638-amino-acid MEV had high antigenicity (1.2542, exceeded the threshold of 0.4), was non-toxic(ToxinPred2) and non-allergenic(AllergenFP), had low human homology(BLASTP e-value < 0.005), Instability index 26.80 (< 40),and good solubility(0.896, > 0.5).Molecular docking and dynamics confirmed its stability. Codon optimization for E. coli enhanced expression potential. Immunoinformatics simulations showed the MEV effectively increased antibody level,T cell counts, and proinflammatory cytokines.In conclusion, the MEV shows promise for brucellosis vaccine development. To fully assess its efficacy and safety, a series of in vivo experiments will be conducted using animal models, which will serve as a critical step towards validating its potential as a viable vaccine candidate.