Bacterial signatures of anthropogenic pressures in a high-mountain river: a One Health study using full-length 16S profiling

Anthropogenic pressures can reshape riverine microbiomes, with implications for water quality and One Health surveillance. Here, we profiled the bacterial composition, diversity, and 16S-based predicted functional potential across five sites along an anthropogenic pressure gradient in a high-mountain Andean system (Chicamocha River, Boyacá, Colombia), influenced by wastewater discharge, thermoelectric cooling, and agro-livestock and municipal activities. Full-length 16S rRNA gene sequencing (PacBio HiFi) generated 737,344 high-quality reads and 5,036 Amplicon Sequence Variant (ASVs). Community composition differed significantly among sites, characterized by high β-diversity and a notable association between ammonium levels and community structure. The phyla Pseudomonadota and Bacteroidota dominated most sites, whereas the wastewater outfall was enriched in phyla Bacillota and Campylobacterota (genus Arcobacter ). The cooling pond site showed enrichment of genus Sphingorhabdus , genus Flavobacterium was most abundant at agro-livestock influenced sites, and the genera Limnohabitans/Polynucleobacter dominated downstream oxygen-rich, low-nutrient reaches. 16S-based functional inference suggested site-specific metabolic profiles; the wastewater treatment plant outfall showed higher predicted representation of pathways associated with aromatic/heterocyclic compound degradation and predicted functional categories linked to motility, membrane transport, and antimicrobial resistance, whereas downstream sites showed predicted enrichment of xenobiotic-biodegradation pathways. Together, these data provide a high-resolution baseline for an under-sampled high-mountain urban river and support the utility of full-length 16S rRNA profiling for molecular surveillance to inform effluent management.