A sequencing batch reactor was operated to study the effects of influent Ca2+ on the efficiency, bacterial population, and microbial metabolism of denitrifying phosphorus removal system. Results showed that high Ca2+ loading (>= 80 mg/L) significantly inhibited the performance of simultaneous nitrogen and phosphorus removal. The abundance of phosphorus removal-related organisms (Dechloromonas and Candidatus Accumulibacter) decreased with increasing Ca2+ concentration from 20 to 140 mg/L, while the abundance of glycogen-accumulating organisms and other bacteria increased. Metabolomic analyses revealed that the metabolic profiles of microbial community were also affected by high influent Ca2+ concentrations. 3-Hydroxybutyrate, acetate, alanine, and glutamate were the main differentiated metabolites in the system. An accumulation of amino acids and a reduction of nucleotides and amines were important response to high Ca2+ loading. Long-term Ca2+ loading had a reversible effect on the denitrifying phosphorus removal system as it could revive after a 50-day recovery process. (C) 2017 Elsevier Ltd. All rights reserved.