Xiaohong Liu

Background: Vibrio harveyi is a major bacterial pathogen threatening turbot aquaculture, necessitating the development of more effective vaccines. Bacterial ghosts (BGs), which are empty bacterial envelopes with preserved surface antigens, offer a promising alternative to traditional formaldehyde-killed vaccines that often suffer from reduced immunogenicity. Methods: We developed an optimized BGs vaccine for V. harveyi by combining the nonionic surfactant NP-40 with sodium hydroxide (NaOH). This NP-40/NaOH combination demonstrated a synergistic lytic effect, halving the minimum inhibitory concentration of NaOH required for complete inactivation. Results: The resulting BGs exhibited intact cellular morphology with transmembrane pores, efficient removal of cytoplasmic contents, and significantly better preservation of lipopolysaccharide structure compared to NaOH-alone treatment. Vaccination trials in turbot demonstrated that the NP-40/NaOH BGs provided the highest relative percent survival (RPS = 58.8%) upon challenge, outperforming both NaOH-alone BGs (RPS = 55.0%) and a traditional formaldehyde-killed vaccine (RPS = 34.8%). The superior protection was correlated with the induction of a more robust and sustained immune response, characterized by significantly higher levels of specific IgM antibodies, elevated lysozyme activity, and increased total serum protein. Conclusions: This study establishes the NP-40/NaOH protocol as an effective strategy for producing high-quality BGs with enhanced immunogenicity, presenting a potent vaccine candidate for controlling vibriosis in aquaculture.