Antibacterial, growth-promoting, and immunostimulatory effects of dietary caffeic acid in Asian seabass (Lates calcarifer) challenged with Streptococcus agalactiae

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, examined growth and immune responses during a 4-week feeding trial, and determined its protective effects against S. agalactiae . Antibacterial assays quantified dose-dependent inhibition of S. agalactiae , with a sigmoidal dose-response model estimating an EC₅₀ of 9.90 µg/mL. Acute toxicity testing showed no adverse effects at dietary concentrations 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 immuno-protective feed additive for Asian seabass.