Oral immunization with a DNA vaccine encoding capsid protein and IFN-γ provokes efficient and long-term immunity against nervous necrosis virus.

Viral nervous necrosis (VNN), caused by nervous necrosis virus (NNV), poses a major threat to global aquaculture, causing severe mortality and economic losses in high-value species like grouper. Oral vaccination offers a practical approach for large-scale prevention but faces challenges including gastrointestinal antigen degradation and inadequate mucosal immunity. To overcome these, we developed an oral DNA vaccine by encapsulating a plasmid encoding the NNV capsid protein (CP) and IFN-γ within PLGA microparticles (CPIF@PLGA). Synthesized via double emulsion-solvent evaporation, these spherical microparticles (ca. 8 μm in diameter) exhibited high encapsulation efficiency (84.3 %). In vivo immunization of grouper demonstrated that CPIF@PLGA significantly increased survival to 82.2 % post-NNV challenge versus controls, reduced viral loads in brain and eyes, and enhanced systemic and mucosal immunity. This was evidenced by elevated serum IgM, IgT, lysozyme, SOD, and complement activity, alongside upregulated expression of immune-related genes (IL-1β, TNF-α, CD4, CD8α, MHC-Iα, IgM) in spleen and hindgut. Our findings establish PLGA microparticles as an effective oral delivery platform and IFN-γ as a potent adjuvant, eliciting robust and durable protection. This study provides a promising strategy for developing effective and scalable oral vaccines against NNV in aquaculture.