The performance and microbial community of activated sludge from a sequencing batch reactor (SBR) were investigated under long-term exposure of magnetic Fe3O4 nanoparticles (Fe3O4 NPs). The COD removal showed a slight decrease at 5-60 mg/L Fe3O4 NPs compared to 0 mg/L Fe3O4 NPs, whereas the NH4+-N removal had no obvious variation at 0-60 mg/L Fe3O4 NPs. It was found that 10-60 mg/L Fe3O4 NPs improved the denitrification process and phosphorus removal of activated sludge. The microbial enzymatic activities of activated sludge could be affected by Fe3O4 NPs, which had similar variation trends to the nitrogen and phosphorus removal rates of activated sludge. The reactive oxygen species (ROS) production and lactate dehydrogenase (LDH) release demonstrated that Fe3O4 NPs led to the toxicity to activated sludge and destroyed the integrity of microbial cytomembrane. High throughput sequencing indicated that Fe3O4 NPs could obviously affect the microbial richness and diversity of activated sludge. (C) 2016 Elsevier Ltd. All rights reserved.