Immunopotentiating effects of recombinant TNFα in formalin-killed Streptococcus parauberis vaccine for starry flounder

Abstract

To enhance vaccine efficacy in aquaculture species, molecular adjuvants such as cytokines are gaining attention for their immunostimulatory potential. In this study, we evaluated the functional role of recombinant tumor necrosis factor-alpha (rTNF alpha) as a molecular adjuvant in starry flounder (Platichthys stellatus) vaccinated against Streptococcus parauberis. Intraperitoneal administration of rTNF alpha at different concentrations (10-100 mu g/mL) significantly induced pro-inflammatory cytokines (IL-1(3, TNF alpha), apoptotic regulators (Caspase-3, Caspase-8), and cytolytic effectors (Perforin-2), in a dose- and time-dependent manner. Notably, STAT3 expression peaked at 12 h, and Perforin-2 exhibited a similar to 400-fold increase by day 7, indicating sustained immune activation. In vivo, FKC vaccination alone conferred dose-dependent protection, but co-administration with rTNF alpha further improved survival (up to 100 %) and maintained high antigen-specific IgM titers through 6 weeks. These results confirm that rTNF alpha promotes both innate and adaptive immune responses and significantly enhances the protective efficacy of the FKC vaccine. Our findings highlight rTNF alpha as a promising molecular adjuvant for developing nextgeneration vaccines in finfish aquaculture.