<p>This study evaluates the efficacy of <i>Sargassum cristaefolium</i> against eight strains of Gram-positive and Gram-negative bacterial species. Different solvents including ethanol, methanol, n-hexane, dichloromethane, ethyl acetate, chloroform, benzene and aqueous were used to isolate crude extracts containing bioactive compounds. The effectiveness of crude extract was determined by phytochemical qualitative, phytochemical quantitative (GC-MS), antimicrobial sensitivity test, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), antioxidant activity (2,2-diphenyl-1-picrylhydrazyl assay, Fe²⁺ chelating assay and phenol estimation), in vivo studies followed by hematological parameters. Each extract exhibited distinct inhibitory effects, with the ethanolic extract showing the highest antibacterial activity (17.7 ± 0.13) against <i>Aeromonas hydrophila</i>. The MIC for each extract ranged from 2&#xa0;mg/L to 10&#xa0;mg/L against different pathogenic bacterial strains. The qualitative analysis suggests that compounds like alkaloids, terpenoids, tannins, phenols, steroids, quinones, coumarins, and saponins are mainly responsible for antibacterial activity. Quantitative analysis using GC-MS showed that the ethanolic extract contained highest amount of diethyl phthalate (94.82%). The ethanolic extract of <i>S. cristaefolium</i> demonstrated the highest free radical activity while it showed slightly lower Fe²⁺ chelating activity than other extracts. Statistical significant difference between treatments in antioxidant analysis, survival and haematological parameters had been observed. The in vivo exposure of the ethanolic extract to <i>Labeo rohita</i> fingerling showed antibacterial effects against <i>A. hydrophila</i> via modifying immune response in fish. Hence, present study concludes that <i>S. cristaefolium</i> inclusion in fish helps to mitigate economical loss occurring through bacterial infection and antibiotic drugs application.</p>