From city waste to river risk: tracking AMR and pathogenic potential in urban water gradient of the Kathajodi river

Abstract Background Urban rivers are critical recipients of untreated wastes, contributing to the environmental spread of antimicrobial resistance (AMR) and pathogenic microbes, as is exemplified in the Kathajodi River gradient. Methods Shotgun metagenomic sequencing using Illumina HiSeq was employed to profile community diversity, antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), metal resistance genes (MRGs), and virulence factors (VFs) across spatially distinct sites; upstream (US), catchment (CM), and downstream (DS) of the Kathajodi River. Functional pathways were annotated using the KEGG and COG databases, and Canonical Correspondence Analysis (CCA) connected functional shifts to environmental gradients. Results Relatively pristine upstream area hosts stable microbiomes predominately; Proteobacteria (80.32%) and Cyanobacteria (6.70%), has low species richness (138), and a modest resistome (33 ARGs), adapted to natural biogeochemical cycles. In contrast, the catchment, impacted by urban wastewater, shows elevated Bacteroidetes (31.11%), Firmicutes (3.63%), high species richness (663), and diverse ARGs (98) and MGEs (141), driven by heavy metals (Cu, Zn, Cd) and pollutants (TDS, BOD). It emerges as a hotspot for ARGs, MGEs, and virulence factors, reflecting intense selective pressure from faecal and industrial inputs. Downstream with intermediate diversity (704 species, 53 ARGs, 88 VFs), with persistent Bacteroidetes (25.33%) exhibits partial ecological recovery, yet persistent high-risk ARGs and elevated virulence genes signal ongoing contamination and microbial adaptation to fluctuating stressors. Prediction of energy production and primary metabolism in upstream exhibited core functions. Whereas, membrane transport, defence mechanisms, and mobile element activity in catchment and downstream, as well as pronounced signal transduction, xenobiotic degradation, and bacterial chemotaxis in Catchment indicates stress response. CCA highlights the role of environmental gradients: high dissolved oxygen upstream versus elevated pollutants at the catchment in shaping these microbial shifts. Network analysis further revealed strong co-occurrence between ARGs, MGEs, and MRGs, identifying MGEs as key connectors driving horizontal gene transfer and co-selection processes. The study underscores the co-selection of ARGs and virulence traits under pollution stress, facilitated by MGE-mediated horizontal gene transfer, amplifying public health risks. Conclusion First river metagenomic study from eastern India emphasizes that human activities reshape microbial landscapes. The research advocates for interdisciplinary strategies to mitigate urban pollution, safeguard environmental health, and curb the global spread of antimicrobial resistance, calling for proactive environmental management and policy interventions.