Urban drainage from wastewater treatment plants (WWTPS) and stormwater runoff is a major source of fecal bacteria, antibiotics, and heavy metals entering waterways. Antibiotic and metal pollution may facilitate antibiotic resistance gene persistence and dissemination. Metals are known to be co-selecting factors for antibiotic resistance genes perhaps because of co-occurrence of metal and antibiotic resistance genes on plasmids. Because the spread of antibiotic resistance genes may adversely affect human health, it is necessary to understand processes that drive antibiotic resistance gene occurrence in aquatic ecosystems. Biofilm samples were collected after large rain events from multiple sites along an urbanized stream with multiple wastewater treatment plants in Northeast Ohio and two reference streams. Next-generation sequencing was used to examine bacterial community structure, and quantitative polymerase chain reaction was used to determine distribution of genes contributing to antibiotic (tetracycline [tetW
] and sulfonamide [sul
II]) and metal (copper [copA
], lead [pbrT
], and chromium, cadmium, and zinc [czcA
]) resistance. Additionally, stream physicochemical properties were measured.
Results/Conclusions: Bacterial community profiles of biofilms were significantly different among dates and sites (P ≤ 0.05). Spatiotemporal differences in community structures were driven by differences in discharge, dissolved oxygen, carbon, nitrogen species, and metal concentrations (P ≤ 0.05). Moreover, antibiotic and metal resistance genes were readily detected, and significantly differed over time (tetW, sulI, sulII, copA, and czcA [P < 0.001]) and space (tetW, sulI, and sulII [P ≤ 0.04]). Antibiotic gene (tetW, sulI, and sulII) abundances were significantly and positively correlated to metal resistance genes (pbrT, copA, and czcA) and dissolved and sediment extracted metals (P ≤ 0.05). Collectively, these results suggest that activities associated with urbanization, including storm and waste water effluent, significantly alter physicochemical conditions, which in turn impact the composition of bacterial communities. These findings highlight the potential of metals as a selective pressure for maintenance and spread of antibiotic resistance genes in aquatic environments.