Screening of acute Vibrio anguillarum-resistant turbot (Scophthalmus maximus) families and integrated immune expression profiling

Background Vibrio anguillarum is a significant pathogen of turbot (Scophthalmus maximus), characterized by rapid disease progression and high mortality rates. Infection in juvenile turbot can result in mortality exceeding 80% within 2-3 days, posing a severe threat to the turbot aquaculture industry. Objective Screening protocol development for Edwardsiellosis resistance in S. maximus pedigrees.Methods This study selected eight different families of turbot (S. maximus) (A-H) and compared their survival rates after artificial infection with V. anguillarum. Additionally, the roles of 12 immune-related genes (il-1β, mx, tnf-α, tnf-α, il8r, tlr5, mhc-Ⅰ, mhc-Ⅱ, IgM,tcr, cd4, cd8) and three immune factors (complement C3, LZM, ACP) in the early stages of V. anguillarum infection were analyzed. Results The pathogen challenge results revealed that the majority of infected fish died within 2-3 days. By 10 days post-infection with V. anguillarum, Family A exhibited a survival rate of 13.15%, Family G showed 10.52% survival, while all other families suffered complete mortality. Immunological factor analysis demonstrated that at 24 hours post-challenge: Families D and F showed significant increases in complement C3 levels (P<0.05). Families A and G displayed significantly elevated acid phosphatase (ACP) activity (P<0.05). qRT-PCR analysis of immune-related genes at 24 hours post-challenge showed followings, Family A: Significant upregulation of tnf-α, tlr5, tcr, cd4, and cd8, with tlr5 showing 7-fold increase and cd8 exhibiting 20-fold increase; Family G: Marked upregulation of mx, tnf-α, tlr5, tcr, cd4 and cd8, with tlr5 increasing 2-fold and tcr rising ~8-fold. In summary, the robust synergistic response of the immune systems in pedigrees A and G following challenge suggests their capacity to swiftly recognize pathogens, recruit immune cells, and initiate specific cytotoxicity. Integrating with the group survival rate data post-challenge, pedigrees A and G exhibit the highest resistance to V anguillarum infection, qualifying them as high disease-resistant potential pedigrees against V anguillarum. For pedigrees C and E, significant upregulation of il-1β, tcr, and cd8 was observed; pedigree F showed notable elevation in C3 content, il-1β, mx,and tnf-α; pedigree H presented significant upregulation of il-1β, tlr5, and tnf-α genes. However, other pathways (e.g., il8r, cd4) displayed mild responses, indicating that immune activation in these pedigrees was characterized by local pathway dominance. Relying on single or local pathways to combat infection during the early stage of V anguillarum challenge, these pedigrees exhibited immune responses with narrower breadth and shallower depth compared to pedigrees A and G, suggesting moderate disease resistance to V anguillarum. Pedigrees B and D showed only feeble responses in a few genes (e.g., il-1β in pedigree D) post-challenge, with overall delayed activation of immune pathways, demonstrating the lowest resistance to V anguillarum infection. Conclusion In conclusion, Families A and G demonstrate superior suitability as candidate populations for resistance against acute V. anguillarum infection. Through this study, we have preliminarily identified six immune resistance-related genes (mx, tnf-α, tlr5, tcr, cd4, cd8) that show particular sensitivity to V. anguillarum infection in these families. These genes may serve as valuable molecular markers for selective breeding of V. anguillarum-resistant turbot (S. maximus) strains, providing critical theoretical data to support the development of disease-resistant varieties in aquaculture.