Encapsulin nanoparticle-conjugated p54 protein boosts immune responses against African swine fever virus

African swine fever virus (ASFV) poses a severe threat to global swine production, with no commercially approved vaccine. The ASFV p54 protein is critical for viral entry and trafficking, interacting with the dynein light chain 8 (DLC8) to facilitate replication and spread within host cells. Despite advances in vaccine development, current strategies struggle to induce sustained immune responses. In this study, we utilized SpyTag/SpyCatcher technology to covalently conjugate the ASFV p54 protein to ferritin and encapsulin nanoparticles (F/E-p54), aiming to enhance immune recognition and optimize antigen presentation. The conjugated nanoparticles exhibited a 60-fold enhancement in receptor binding affinity over the soluble p54 monomer, improving targeted delivery to lymphoid tissues and stimulating T follicular helper (Tfh) cells and germinal center B (GCB) cells. The encapsulin-based nanoparticle approach elicited strong B- and T-cell responses, promoting prolonged immune activation and the production of neutralizing antibodies. Additionally, antibodies generated by the F/E-p54 conjugates effectively inhibited both genotype II and recombinant I/II ASFV strains. These findings highlight the potential of nanoparticle-based vaccines, particularly encapsulin-conjugated systems, as promising platforms for advancing ASFV vaccine development and guiding future immunization strategies.