Metagenomics Reveals Microbial Community Shifts Associated With Contrasting Anthropogenic Impacts in Freshwater Sources of A Coastal Protected Area in Southeastern Brazil

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Abstract
This study aimed to characterize freshwater microbial communities, environmental drivers, and anthropogenic impact patterns across three sites on Marambaia Island (southeastern Brazil) using metagenomics. Samples collected from freshwater sources used for human consumption were processed through concentration, nucleic acid extraction, and sequencing on the Illumina NextSeq 2000 platform. A total of 67.2 million reads were assembled into 89,230 bacterial contigs, mostly attributed to Gammaproteobacteria, Alphaproteobacteria, and Betaproteobacteria. Sites under lower anthropogenic influence exhibited higher microbial diversity, whereas impacted sites showed enrichment of opportunistic and fecal‐associated genera. A heterogeneous anthropogenic impact profile was observed across sites, corroborated by the proposed Anthropogenic Impact Index (AII). Fourteen antimicrobial resistance genes conferring resistance to beta ‐lactams, quinolones, sulfonamides, tetracyclines, and macrolides were detected predominantly in sewage‐impacted areas, indicating potential diffuse contamination. Redundancy analysis revealed that environmental variables explained 88.1% of microbial community variation, with conductivity, salinity, and turbidity as key drivers. These findings demonstrate the applicability of metagenomics as a powerful tool for assessing microbial diversity, ecological dynamics, and contamination risks in vulnerable freshwater systems.

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