Dietary caffeic acid mitigates Streptococcus agalactiae pathogenesis in Asian seabass (Lates calcarifer): antibacterial efficacy, immunomodulatory gene regulation, and hepatoprotection

Abstract

Caffeic acid (CA) has recently gained attention as a natural compound with antimicrobial and immunomodulatory potential relevant to aquaculture health management. However, its functional efficacy, optimal dietary inclusion level, and protective capacity against Streptococcus agalactiae infection in Asian seabass (Lates calcarifer) remain insufficiently characterized. This study evaluated the in vitro antibacterial activity of CA, assessed its short-term dietary safety (n = 150 fish), examined growth and immune responses (n = 400 fish) during a 4-week feeding trial, and determined its protective effects against S. agalactiae. In vitro antibacterial assays demonstrated dose-dependent inhibition of Streptococcus agalactiae, with a sigmoidal dose-response model yielding an EC50 of 9.90 mu g/mL. Acute toxicity testing showed no detrimental effects at dietary inclusion levels up to 100 mg/kg. Feeding trials demonstrated that 25-50 mg/kg CA enhanced weight gain, specific growth rate, villus length, and immune-related gene expression. Following bacterial challenge, dietary CA significantly improved survival, with CA-50 yielding the highest survival rate (70.67%) and relative percent survival (47.62%), accompanied by attenuated hepatic degeneration, necrosis, and vascular congestion. Gene expression and histopathological analyses further confirmed enhanced immune activation and reduced lesion severity in fish treated with CA. These findings demonstrate that CA functions as an effective antimicrobial agent, growth promoter, and immunostimulatory dietary additive for Asian seabass.