The long-term effects of Ni(II) on substrate removal and microorganism activities were investigated by operating sequencing batch reactors (SBRs). Compared to the control system lacking Ni(II), the removal efficiencies of total organic carbon (TOC) and ammonium nitrogen (NH4(+)-N) in SBR system loading with 10mgL(-1) Ni(II) decreased drastically from 90.2±3.6 percent to 75.0±8.9 percent, and 99.2±0.6 percent to 50.8±11.5 percent, respectively. As compared to the control system, a inhibitory rate of more than 50 percent for the 2,3,5-triphenyltetrazolium chloride electron transport system (TTC-ETS) and the 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride electron transport system (INT-ETS), and 43 percent for the specific oxygen uptake rate (sOUR) were detected in SBR system loading with 20mgL(-1) Ni(II). TTC-ETS, INT-ETS, and sOUR were significantly correlated with substrate removal efficiencies, suggesting that they could all serve as effective indicators of the performance of activated sludge processes. Additionally, INT-ETS is superior to sOUR and TTC-ETS in detecting the toxic effects of Ni(II) on sludge microorganism activity.