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Discarded face masks from the global COVID-19 pandemic have contributed significantly to plastic pollution in surface water, whereas their potential as a reservoir for aquatic pollutants is not well understood. Herein, we conducted a field experiment along a human-impacted urban river, investigating the variations of antibiotic resistance genes (ARGs), pathogens, and water-borne contaminants in commonly-used face masks. Results showed that high-biomass biofilms formed on face masks selectively enriched more ARGs than stone biofilm (0.08–0.22 vs 0.07–0.15 copies/16 S rRNA gene copies) from bulk water, which mainly due to unique microbial communities, enhanced horizontal gene transfer, and selective pressure of accumulated contaminants based on redundancy analysis and variation partitioning analysis. Several human opportunistic pathogens (e.g., Acinetobacter , Escherichia - Shigella , Bacillus , and Klebsiella), which are considered potential ARG carriers, were also greatly concentrated in face-mask biofilms, imposing a potential threat to aquatic ecological environment and human health. Moreover, wastewater treatment plant effluents, as an important source of pollutants to urban rivers, further aggravated the abundances of ARGs and opportunistic pathogens in face-mask biofilms. Our findings demonstrated that discarded face masks provide a hotspot for the proliferation and spread of ARGs and pathogens in urban water, highlighting the urgent requirement for implementing stricter regulations in face mask disposal. [Display omitted] • High-biomass biofilms were formed on discarded face masks in urban river. • Distinctive microbial community structure was detected in face-mask biofilm. • Face-mask biofilm simultaneously enriched ARGs and opportunistic pathogens. • Multiple drivers for enhanced antibiotic resistomes in face-mask biofilm were proposed. • WWTP effluents exacerbated the accumulation of pollutants in face-mask biofilm. [ABSTRACT FROM AUTHOR]