Anaerobic denitrification is accompanied by biogas emissions, which may orient greenhouse gases or air pollutants. Ferrous iron play an important role in wastewater treatment. However, the effect of ferrous ion on biogas emission and the relationship between microbial community and nitrogen removal performance are not fully understood, especially in up-flow anaerobic sludge blanket (UASB) reactors. Here, the results revealed that nitrogen gas increased rapidly with Fe(II) addition. And ferrous ions enhanced denitrification and COD removal efficiency with more than 60% and 10%, respectively. Moreover, Ferrous ions addition evidently increased Hydrogenophaga, Methylotenera, Zoogloea and Fluviicola during the whole UASB reactor operation progress. The correlation coefficient analysis further confirmed that Fe(II) was positively correlated with the abundances of Hydrogenophaga, Methylotenera and Fluviicola. Based on these results of chemicals transformation kinetics, microbial community and the correlations coefficient analysis, a hypothetical mechanism is proposed: In the UASB system with Fe(II) and NH4+, firstly Paludibacter made a contribution to NH4+ oxidation and generated NO3–, and then Hydrogenophaga, played an important role in NO3– reduction, Zoogloea coupled NO3– reduction to Fe(II) oxidation, Fluviicola combined with Methylotenera conducted NO2– reduction together with Fe(II) oxidation. This study will improve our understanding of ferrous ion’s influence on biogas emission, denitrification process and corresponding microbial community.