Most studies of whether and how nanoparticles (NPs) affect antibiotic resistance genes (ARGs) focus on testing single NPs type. In this study, we determined the combined effect of Cu- and ZnO- NPs in the water samples collected from the Yangtze River Estuary and found the effect differs greatly from that produced by individual NPs. The results showed that the Cu- and ZnO- NPs co-exposure resulted in an enrichment of ARGs, whereas individual Cu- and ZnO- NPs exposure decreased the abundance of ARGs. Furthermore, the co-exposure of Cu- and ZnO- NPs induced obvious changes in the microbial communities compared to the control communities. Redundancy analysis suggested that the microbial community contributed the most (43.5%) to the ARG profiles, followed by dissolved metal ions (25.7%), MRGs, (19.4%), and MGEs (4.4%). Network analysis found several potential hosts (such as Mycobacterium and Escherichia coli) and implied the extent of the risk of ARG transmission into various environmental niches by these common microbes.