Abstract: In recent years, the research on enhanced microbial fuel cell (MFC) treatment of nitrogen-containing wastewater has attracted extensive attention at home and abroad. FeS₂ was introduced into MFC anode to construct a FeS₂-enhanced microbial fuel cell (Pyr-MFC) system for wastewater with different carbon to nitrogen ratios (C/N). The blank control group without FeS₂ (C-MFC) was used as a control to explore its effects on nitrogen removal and electro-generation. High throughput sequencing, X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM) were used to investigate the changes of microbial abundance, sulfur and iron elements in the system, and the mechanism of nitrogen removal under low C/N in FeS₂-enhanced system was obtained. The results showed: 1) The denitrification efficiency and power generation density of Pyr-MFC were higher than those of the control group, the nitrate nitrogen removal rate was increased by 15.7%, and the maximum voltage increase was as high as 0.274 V. 2) NO₃ ⁻-N removal rates of Pyr-MFC at different C/N ratios (4, 3, 2 and 1) were 100%, 97.8%, 58.4% and 49.7%, respectively, all higher than those of the control group, indicating that FeS₂ could effectively reduce the dependence of the system on carbon sources. 3) The microbial community test results showed that FeS₂ increased the species abundance of electrogenic microorganisms (Thauera, Thiobacillus and Geobacter) by about 9.43%. 4) The results of material transfer analysis showed that S⁻ provided electrons for the denitrification process, and Fe²⁺ enhanced the electron transfer and improved the electro-generation performance of the system as an electron shuttle.