Influence of Vibrio anguillarum culture conditions on the efficacy of bacterin-based vaccines in lumpfish (Cyclopterus lumpus).

Lumpfish (Cyclopterus lumpus) is used as cleaner fish to control sea lice infestations in Atlantic salmon (Salmo salar) farms across the North Atlantic. Vibrio anguillarum, the causative agent of vibriosis, is a recurrent bacterial pathogen affecting lumpfish. Bacterin-based vaccines are frequently used to control vibriosis in finfish, but their efficacy is not always consistent. Culture conditions significantly influence synthesis of bacterial outer membrane and secreted proteins, which are critical antigens, and thus impact the immunogenicity of bacterin-based vaccines. In this study, we assessed the effect of V. anguillarum culture conditions on vaccine efficacy in lumpfish. V. anguillarum was cultured under iron-limited at 15 °C, and iron-rich or iron-limited conditions at 28 °C with 2 % NaCl, and these cultures were used to prepare bacterins. A commercial vaccine was used as positive control, while PBS and PBS adjuvant were negative controls. Lumpfish were intraperitoneally immunized and challenged 12 weeks post-immunization with 10-100 times the LD50 dose of V. anguillarum. Bacterins prepared from V. anguillarum grown under iron-limited conditions at 28 °C with 2 % NaCl and mixed with adjuvant conferred the highest protection compared to other preparations and commercial vaccines. In contrast, bacterins derived from V. anguillarum cultured under iron-limited conditions at 15 °C conferred the lowest protection. Reverse vaccinology and transcriptomic analyses of V. anguillarum grown under optimal immunogenic conditions revealed 323 upregulated genes, of which 211 were high-antigenicity proteins suitable for subunit vaccines. This study provides critical knowledge for effective vaccine formulation against V. anguillarum and identifies potential antigens for subunit vaccine development.