Stimulatory and immunomodulatory effects of Lagenaria siceraria flower on growth performance, metabolic profile, intestinal integrity, and cytokine/immune-related gene expression in Nile tilapia (Oreochromis niloticus).

Natural immunomodulatory compounds have emerged as vital tools in sustainable aquaculture for their capacity to boost growth and immunity. This study, for the first time, investigates the dietary effects of Lagenaria siceraria flower (LSF) in Nile tilapia (Oreochromis niloticus), specifically analyzing growth performance, metabolic profiles, intestinal integrity, and the expression of cytokine and immune-related genes. In a 70-day feeding trial, 240 Nile tilapia were divided into four groups (60 fish per group). Each group received a basal diet supplemented with varying concentrations of LSF: 0, 1, 2, or 3 g/kg (designated as LSF0, LSF1, LSF2, and LSF3, respectively). The HPLC analysis of LSF indicates that hesperidin (49.75%) was the primary constituent, followed by quercetin (11.6%), ellagic acid (11.56%), gallic acid (4.97%), and quercetin dihydrate (2.2%). Our results demonstrated that the supplementation of 2 or 3 g/kg LSF significantly improved growth performance indices (final body weight, weight gain, average weight gain, and specific growth rate) compared to the other groups (P < 0.05). Feed intake remained unaffected (P > 0.05), and feed conversion ratio (FCR) was significantly lower in the LSF2 and LSF3 groups (P < 0.05). Erythrocyte and leukogram indices were significantly improved (P < 0.05); however, the percentages of lymphocytes and basophils showed a notable decline in the LSF-supplemented groups (P < 0.05). LSF supplementation significantly increased serum protein levels and digestive enzymes (lipase and amylase), while reducing liver enzyme activities (AST and ALT) and nitrogenous metabolites (urea and creatinine) compared to the control group (P < 0.05). At a dosage of 3 g/kg, LSF supplementation effectively minimized lipid peroxidation (lowered MDA) and maximized the antioxidant defenses (SOD, CAT, and GPx) compared to the control (P < 0.05). Dietary LSF demonstrated significant immunomodulatory effects by increasing lysozyme activity, total immunoglobulin (Ig) levels, phagocytic activity, and the phagocytic index (P < 0.05) as well as preserved the gut integrity. Furthermore, LSF modulated immune-related genetic markers, as evidenced by the significantly higher expression of CXC chemokine, IL-1β, IL-8, IL-6, MHC-II, TNF-alpha, TLR9, and TLR3 in fish, with maximum values observed in the LSF3 group (P < 0.05). After a challenge with Aeromonas sobria, the survival rates significantly improved from 30% in the control group (LSF0) to 55%, 65%, and 70% in the LSF1, LSF2, and LSF3 groups, respectively (P < 0.05). Molecular docking analysis further supported these findings, showing that hesperidin and quercetin had strong binding affinities to GSH (ranging from -7.54 to -6.91 kcal/mol and -5.00 to -4.66 kcal/mol, respectively). Notably, hesperidin had a superior docking score with Keap1 (-10.53 kcal/mol) compared to quercetin (-6.13 to -5.95 kcal/mol). In conclusion, LSF supplementation significantly boosts the health and growth of Nile tilapia, specifically through improved gut health and antioxidant defenses. This study underscores the viability of repurposing agricultural waste into high-value natural compounds, paving the way for more sustainable and resilient aquaculture practices.