Melatonin supplementation alleviates stocking-density stress and enhances growth, immune, and physiological performance of Sparus aurata cultured in groundwater-based systems

The intensive mariculture of S. aurata can promote aquaculture development, although it can also pose challenges because of predatory behavior and environmental stressors caused by intensification. The objective of this study is to alleviate stress and prevent cannibalism, thereby improving the fish's performance and welfare. A 90-day growth experiment investigated the dietary inclusion of melatonin (ML) on Sparus aurata performance, physiology, immunity, histopathology, and welfare when grown in saline groundwater (36 ppt) at stocking densities (SD) of 50 fish/500 L‒tank (SD₅₀) and 100 fish/500 L‒tank (SD₁₀₀). Six fish groups in triplicate were fed three increasing levels of ML (0, 25, and 50 mg/kg^{- 1} diet) expressed as T1, T2 and T3 under SD₅₀, and T4, T5, and T6 under SD₁₀₀. Fish (n = 1350) weighing 16.46 ± 0.18 g/fish were grown using a flow-through system and a daily feeding rate of 4%. The results revealed significant declines in nitrogen by-product (NH₃ and NO₂) levels of fish fed ML in a dose density-dependent manner. Fish fed ML under both densities showed significantly improved growth indices, FCR, growth hormone, and crude protein content compared with those fed the control diet, with T2 and T6 groups achieving the highest values. ML augmentation enhanced serum lipase, protease, immunoglobulin, albumin, globulin, antioxidants, and proinflammatory cytokine interleukin-1β and lowered serum cortisol, glucose, ALT, AST, and ALP. Under both densities, the ML-fish presented healthier livers and intestines than those fed the control diet, which showed marked liver vacuoles and steatotic changes. The ML-SD interaction showed superior fish performance, particularly in groups T2 and T6; thus, it is recommended to administer dietary melatonin at 25 mg/kg for seabream reared at 50 fish per 500-liter tank and 50 mg/kg for those cultivated at 100 fish per 500-liter tank using groundwater.