The efficacy of iron(II) sulfide (FeS)-based autotrophic denitrification in simultaneous nitrogen and phosphorus removal from wastewater was studied with batch experiments. It was efficient at a wide pH range of 5–9, and temperature range of 10–40 °C. The concentrations of NH4+-N, Mg2+ and HCO3− in the wastewater should be kept over 7.8, 0.24 and 30 mg L−1 for efficient nitrate (NO3−-N) reduction, respectively. The NO3−-N removal rate increased from 0 to 82 mg L−1 d−1 and then leveled off when the NO3−-N concentration increased from 0 to 415 mg L−1 and then to 700 mg L−1, respectively. The NO3−-N removal rate quickly increased, leveled off, and then sharply decreased when the PO43−-P concentration increased from 0 to 0.1 mg L−1, then to 114.0 mg L−1, and further to 683.8 mg L−1, respectively. The PO43−-P removal was over 98% when the PO43−-P concentration ranged 0–683.3 mg L−1. During treatment of the secondary effluent of a local municipal wastewater treatment plant containing NO3−-N of 14.9 mg L−1 and total phosphorus (TP) of 3.9 mg L−1, NO3−-N was reduced to 1.1 mg L−1 and TP was completely removed.