Immunoactive alginate-eugenol microbeads driving synchronized drug delivery and host immune reprogramming in...

Nanotechnology-driven feed additives provide an effective approach to enhance aquaculture sustainability through the improved delivery and bioavailability of bioactive compounds. This study aimed to formulate eugenol-loaded sodium alginate beads (E-SABs) as a biopolymer-mediated sustained-delivery system and to evaluate their effects on growth performance, antioxidant and immune status, and resistance to Aeromonas hydrophila infection in Nile tilapia (Oreochromis niloticus). The formulated E-SABs had a mean particle (Z-average) diameter of 79.66 nm, a polydispersity index of 0.413 and a zeta potential of -9.18 mV, and released eugenol in a sustained manner in vitro (20-25% burst release within 2 h, reaching 96% cumulative release at 24 h; Korsmeyer-Peppas model, R² = 0.999, release exponent n = 0.69, indicating non-Fickian diffusion). Juvenile Nile tilapia (initial weight approximately 20-100 g; n = 100) were randomly allotted to four dietary groups (a basal-diet control and E-SABs at 0.25, 0.50 and 1.0 mg kg^-1 feed) and fed for 30 days. Final body weight, specific growth rate and survival were significantly higher, and feed conversion ratio significantly lower, in all E-SAB groups than in the control (P < 0.05), with the greatest improvement at 1.0 mg kg^-1 (e.g., final weight 69.90 ± 0.45 g vs. 59.20 ± 0.52 g; survival 98.8 ± 0.85% vs. 90.0 ± 1.5% in the control). E-SAB-fed fish also showed higher serum total protein, albumin and globulin, and lower AST, ALT, ALP, cholesterol and glucose, than the control (P < 0.05). Innate immunity was enhanced, with dose-dependent increases in phagocytic activity and phagocytic index, lysozyme activity and serum IgM (P < 0.05), together with higher activities of the antioxidant enzymes SOD, CAT and GPx and lower malondialdehyde levels (P < 0.05). These physiological changes were paralleled by up-regulation of genes governing somatic growth (GH, IGF-1 and IGF1-R), immune signalling (IL-1β and IL-8) and antioxidant defence (SOD, CAT and GPx). Following experimental A. hydrophila challenge, cumulative survival at day 15 was significantly higher in E-SAB-fed fish (60-85%, dose-dependent) than in the control (30%) (P < 0.05). Collectively, these findings establish alginate-eugenol microbeads as an effective biomacromolecular delivery platform that couples sustained release with immunomodulation, and support the dietary inclusion of E-SABs, particularly at 1.0 mg kg^-1 feed, as a practical, antibiotic-free strategy for improving growth, immune competence and disease resilience in Nile tilapia aquaculture; further evaluation under commercial farm conditions is recommended.