Combined supplementation of L-carnitine and stevioside enhances performance, stress tolerance, immune competence, and molecular responses in Nile tilapia

Abstract This study evaluated the individual and combined effects of dietary L-carnitine (500 mg/kg) and stevioside (450 mg/kg) on growth performance, physiological responses, stress tolerance, immunity, antioxidant status, and molecular regulation in Nile tilapia ( Oreochromis niloticus ; initial weight 3.84 ± 0.12 g) over 90 days. Fish were fed a control diet or diets supplemented with L-carnitine, stevioside, or their combination. Two-way ANOVA showed significant main effects of both additives on growth performance, feed consumption, and feed conversion efficiency ( P < 0.05), with the combined diet producing the highest numerical responses. No significant interaction was detected for growth or feed utilization, indicating additive effects. Supplementation enhanced intestinal digestive enzyme activities, with significant interaction effects for protease activity ( P = 0.007), and improved intestinal and liver histological integrity, including a significant interaction for villus height ( P < 0.001). Serum biochemical profiles were improved, with significant interactions for total protein, glucose, post-hypoxia glucose, and total cholesterol ( P < 0.05). Antioxidant status was enhanced through increased SOD, CAT, and GPx activities and reduced lipid peroxidation, with additive effects. Supplementation also improved hypoxia tolerance and innate immune responses. A significant interaction was observed for hypoxia tolerance ( P = 0.007), whereas immune responses were predominantly additive. At the molecular level, dietary treatments modulated genes associated with growth, metabolism, immunity, and antioxidant defense. Significant L-carnitine × stevioside interactions were detected for IGF-1 , GLUT4 , IL-1β , IgH , IFN1-α , and CAT expression ( P < 0.05), while SOD and Gsr expression showed additive responses. Overall, L-carnitine and stevioside improved growth, physiological performance, stress resilience, immune competence, and antioxidant capacity in Nile tilapia. Their combination produced the greatest overall responses, with effects ranging from additive to interactive depending on the biological endpoint.