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Fish host complex intestinal bacterial communities that contribute to a wide range of functions, from nutrient assimilation to modulation of the immune system. Understanding how environmental and host-related factors shape the fish gut microbiota is essential for advancing sustainable aquaculture practices. This study compared the intestinal microbiota of gilthead sea bream (<i>Sparus aurata</i>) between wild and aquaculture populations in western Greece using 16S rRNA gene amplicon sequencing targeting the V3-V4 region, combined with culture-based methods. The analysis was based on a 97% similarity threshold and included 141 gastrointestinal samples of fish collected at two aquaculture facilities and two wild fisheries, representing two different growth phases (150 g and 300 g body weight). High-throughput sequencing data revealed a clear separation of gut microbial communities according to origin (wild vs. aquaculture), geographic location, and body growth phase, with most wild fish groups exhibiting higher microbial diversity than their farmed counterparts, except for group MES_150 which showed similar or lower values. The gut microbiota was dominated by Pseudomonadota (53%), Bacillota (29%), Actinomycetota (7%), Deinococcota (5%), and Bacteroidota (4%). A shared core microbiome, comprising <i>Psychrobacter</i>, <i>Staphylococcus</i>, <i>Geobacillus</i>, <i>Aeromonas</i>, <i>Enterobacter</i>, <i>Pantoea</i>, <i>Bacillus</i>, and <i>Acinetobacter</i>, was detected across all populations. Wild fish were enriched in <i>Psychrobacter</i>, <i>Aeromonas</i>, and <i>Photobacterium</i>, while aquaculture fish displayed higher abundances of <i>Vibrio</i>, <i>Allomeiothermus</i>, and <i>Staphylococcus</i>. Network analysis revealed mostly mutually exclusive interactions in both groups but distinct patterns of co-occurrence, driven mainly by <i>Paenibacillus</i>, <i>Enterobacter</i>, and <i>Staphylococcus</i> in wild samples, and by <i>Vibrio</i>, <i>Aeromonas</i>, and <i>Pseudomonas</i> in farmed fish. Culture-based assays demonstrated greater diversity in wild fish, dominated by <i>Pseudomonas</i>, <i>Staphylococcus</i>, and <i>Vibrio</i> strains, in contrast to the frequent occurrence of <i>Staphylococcus</i> and <i>Psychrobacter</i> in aquaculture samples. The findings suggest that aquaculture practices significantly alter gut microbial community structure and reduce diversity, with potential implications for fish health and disease resistance. The identified core and differentially abundant taxa provide candidates for probiotic development to improve aquaculture sustainability.