In Silico Characterization of Conserved Epitopes in Alphavirus E2 Proteins: A Promising Approach for Pan-vaccine Design.

Alphaviruses infect a wide range of hosts, including humans and domestic animals, and they represent an increasing public health concern. Among them, arthritogenic Chikungunya virus (CHIKV) and encephalitogenic Eastern equine encephalitis virus (EEEV) stand out for their epidemic potential and clinical severity. Developing effective and licensed vaccine models against these viruses remains a significant challenge. Rational epitope design, supported by immunoinformatics, offers a promising route for next-generation effective vaccine development. In this study, we utilized the POA pipeline to assist in the selection and prioritization of predicted epitopes from the E2 glycoproteins of CHIKV and EEEV. A total of 39 conserved linear epitopes were selected, comprising 8 B-cell, 2 T-cell, and 29 Th-cell epitopes. These epitopes were characterized for allergenicity, toxicity, and physicochemical properties, including polarity and hydrogen-bonding potential. Structural mapping onto the quasi-3-fold (q3) symmetry unit enabled assessment of their solvent accessibility and spatial organization in the native quaternary context. Our result provides a basis for the rational design of a multiepitope vaccine targeting conserved antigenic regions in alphaviruses with high translational relevance. This integrative approach aligns with the One Health perspective, highlighting its potential for developing biotechnological solutions that address human, animal, and environmental health. The POA pipeline is available on GitHub (https://github.com/UbiratanBatista/POA_Project).