Effects of puerarin on growth performance, digestive ability, immune response, and resistance to ammonia stress in Pacific white shrimp (Litopenaeus vannamei) in low-salinity water

Ammonia stress is a severe environmental problem in shrimp aquaculture, causing oxidative damage, immune suppression, and metabolic disturbance. Puerarin is a bioactive flavonoid possessing antioxidant, anti-inflammatory, antibacterial, and immunomodulatory properties. This study examines puerarin supplementation's impact on immune response, growth performance, oxidative defense system, and ammonia metabolism in Litopenaeus vannamei under ammonia-induced stress. For 56 days, shrimp (designated as groups p0, p15, p30, p60, and p120, respectively) were fed diets supplemented with puerarin at 0, 15, 30, 60, and 120 mg/kg, followed by a 48h acute ammonia challenge. The findings indicated that dietary puerarin significantly improved feed utilization, growth, and survival rates. Analysis of enzyme activity revealed that treatment groups p30, p60, and p120 had higher levels of trypsin (TPS) and amylase (AMS) activity (P < 0.05). After 56 days, acid phosphatase (ACP) and alkaline phosphatase (AKP) activities were significantly increased in all treated groups (P < 0.05), with the highest levels observed in the p60 group. A similar trend was observed following ammonia stress. After 56 days, shrimp exhibited elevated antioxidant enzyme activity, superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) levels rising significantly (P < 0.05), and a significant decline in malondialdehyde (MDA) content (P < 0.05), suggesting improved ability to cope with oxidative stress. p30, p60, and p120 groups showed increased gene expression of target of rapamycin (tor), eukaryotic translation initiation factor 4E1a (eif4e1a, eif4e2, and eif4ebp) (P < 0.05). After the ammonia stress challenge, antioxidant indicators (SOD, T-AOC, CAT, and MDA) exhibited trends consistent with those observed at the feeding trial's conclusion. The expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2), superoxide dismutase (SOD), glutamine synthetase (GS) and glutamate dehydrogenase (GDH) genes were significantly upregulated in p30, p60, and p120 groups compared to p0 (P < 0.05), while heat shock protein 70 (HSP70), tumor necrosis factor alpha (TNF-α), and interleukin 1 beta (IL-1β) were significantly downregulated within these groups relative to p0 (P < 0.05). The catalase (CAT) gene showed its lowest expression in the p30 group (P < 0.05), whereas no significant differences were found among the other treatments (P > 0.05). The findings indicate that puerarin increases L. vannamei growth, digestive utilization, and immunological capacity under ammonia stress, indicating its potential as a functional feed addition. A dietary inclusion level of 30 mg/kg was shown to be optimal.