Abstract: In order to investigate the biofilm characteristics of different fillers, four different fillers, including coral stone, suspended ball filler, elastic filler and combined filler, were used to build biofilters. The mature time of biofilms was compared, and the bacterial community structure of filler-biofilms and seawater in biofilters were analyzed using Illumina Miseq high-throughput sequencing technology. Our results show that the mature time of biofilms using coral stone, suspended ball, elastic, or combined filler were 15 d, 18 d, 21 d and 21 d, respectively. The bacterial abundance and diversity on biofilms of the filler were higher than the seawater of their corresponding biofilter. The diversity index of the suspended ball biofilter was the highest among different biofilters. At the taxonomic level, Proteobacteria was the dominant bacteria in all biofilters. At the genus classification level, different biofliters showed different characteristics of denitrifying functional bacteria. The relative abundance of Nitrosomonas and Nitrospina in elastic filler biofilter was the highest among different biofilters. Bacillus was the dominant genus of coral stone biofilter. The dominant denitrifying bacteria of sulfitobacter, maribacter and phaeodactylibacter were the characteristic bacteria of the suspended ball biofilter. In general, biofilters using different fillers showed a relatively complete nitrification and denitrification microbial system, which played an important role in the denitrification treatment of mariculture wastewater.