The seafood consumption and trade over the years and its expected expansion pose major challenges to the seafood industry and government institutions. In particular, the global trade in fish products and the consequent consumption are linked to reliable authentication, necessary to guarantee lawful trade and healthy consumption. Alterations or errors in this process can lead to commercial fraud and/or health threats. Consequently, the development of new investigative tools became crucial in ensuring unwanted scenarios. Here we used NGS techniques through targeted metagenomics approach on the V3-V4 region of the 16S rRNA genes to characterize the gill bacterial communities in wild-caught seabream (Sparus aurata) and seabass (Dicentrarchus labrax) within different fisheries areas of the "Costa degli Etruschi" in the Tuscan coast. Our challenge involved the possibility of discriminating between the microbiota of both fish species collected from three different fishing sites very close to each other (all within 100 km). Our results showed a significant difference in the assembly of gills’ bacterial communities in terms of diversity (alpha and beta diversity) of seabass and seabream in accordance with the three fishing areas. These differences were represented by a unique site -related bacterial signature, more evident in seabream compared to the seabass. We highlighted how the targeted NGS approach can therefore be used to control the traceability of fish species collected in fishing areas very close to each other. Furthermore, our results contribute to increasing knowledge relating to the gills’ bacterial communities of two fish species of central interest in the fish supply chain and validating a functional methodological approach for tracking fish according to their area of origin.