In this study, the short- and long-term effects of 1 mg L-1 of CuO nanoparticles (NPs), Cu NPs, and Cu(II) on the Anammox process were comparatively investigated. Results showed that short-term exposure to both CuO NPs and Cu NPs decreased the Anammox rate, while it slightly increased with Cu(II) (p < 0.05). Long-term exposure to CuO, Cu NPs, and Cu(II) all significantly suppressed nitrogen removal of Anammox, and the inhibition level decreased in the following order: CuO NPs > Cu NPs > Cu(II). The relative abundance of Candidatus Kuenenia decreased to 1.72%, 4.07%, and 3.93%, respectively, from their abundance of 8.29% in the control and that of Candidatus Brocadia decreased to 0.80%, 1.18%, and 1.87%, respectively, from 4.08%. The presence of Cu NPs elevated the expression of the copA metal resistance gene (p < 0.05) to 12.4 +/- 3.2 x 10(-4) from 3.56 +/- 0.39 x 10(-4) in the control, while CuO NPs and Cu(II) induced no significant variation. The copper content of sludge showed high negative correlation with nitrogen removal, and the nanoeffect was also attributed to inhibition by NPs. Cu NPs induced the highest expression to help the Anammox resist NPs when compared with CuO NPs. As a result, Cu NPs inhibited Anammox to a slightly greater extent than CuO NPs. The high accumulation of copper in the Cu(II) exposed sludge also led to Anammox suppression after long-term exposure.