Anaerobic ammonia oxidizing bacteria (AnAOB) have poor retention capacity in actual denitrification processes, which is one of the main technical bottlenecks restricting the large-scale application of this technology. Compared with traditional biological fillers, high-temperature pyrolysis tires can be used to prepare a new type of biological filler with conductive properties. This not only recycles fixed waste, but also achieves a dual effect of accelerating the formation of biofilm in AnAOB and improving denitrification efficiency. The experimental results indicate that 800 ℃ is the most suitable for pyrolysis tire fillers, while having higher electrical conductivity and more favorable characteristics for fixed microbial growth. Compared to the Anammox group alone, the average removal rates of NH4+-N, NO2--N, and TN with Anammox+conductive filler increased by 9.12%, 13.20%, and 15.69%, respectively, compared to the Anammox group alone, and were significantly higher than the strengthening effect of polypropylene (PP) filler. The Q-PCR analysis results showed that adding conductive fillers can promote an increase in the abundance of denitrifying bacteria, with AnAOB, nitrite reducing bacteria, and nitrate reducing bacteria increasing by 50.21%, 69.70%, and 145.59%, respectively. The development of this new conductive filler technology can provide new ideas and data basis for the promotion of Anammox technology.