This study aimed to investigate the short- and long-term effects of Cu2+ on the activity and performance of denitrifying bacteria. The short-term effects of various concentrations,of Cu2+ on the denitrifying bacteria were evaluated using batch assays. The specific denitrifying activity (SDA) decreased from 14.3 +/- 2.2 (without Cu2+) to 6.1 +/- 0.1 mg N h(-1) g(-1) VSS (100 mgCu(2+) L-1) when Cu2+ increased from 0 to 100 mg L-1 with an increment of 10 mgCu(2+) L-1. A non-competitive inhibition model was used to calculate the 50% inhibition concentration (IC50) of Cu2+ on denitrifying sludge (30.6 +/- 2.5 mg L-1). Monod and Luong models were applied to investigate the influence of the initial substrate concentration, and the results suggested that the maximum substrate removal rate would be reduced with Cu2+ supplementation. Pre-exposure to Cu2+ could lead to an 18.2-46.2% decrease in the SDA and decreasing percentage of the SDA increased with both exposure time and concentration. In the continuous-flow test, Cu2+ concentration varied from 1 to 75 mg L-1; however, no clear deterioration was observed in the reactor, and the reactor was kept stable, with the total nitrogen removal efficiency and total organic carbon efficiency greater than 89.0 and 85.0%, respectively. The results demonstrated the short-term inhibition of Cu2+ upon denitrification, and no notable adversity was observed during the continuous-flow test after long-term acclimation. (C) 2015 Elsevier B.V. All rights reserved.