Solid and liquid organic substances as carbon sources for denitrification process were deeply explored. In this study, the effect of three carbon sources, referred to as poly (3-hydroxybutyrate-co-3-hydroxyvalerate)/poly (lactic acid) (PHBV/PLA) polymer, glucose and CH3COONa, on denitrification performance, microbial community and functional genes were investigated. It was found that maximum denitrification rates of 0.37, 0.46 and 0.39gN/(L·d) were achieved in PHBV/PLA, glucose and CH3COONa supported denitrification systems, respectively. Meanwhile, Illumina MiSeq sequencing revealed that three carbon sources led to different microbial community structures. It can be seen that Brevinema/Thauera/Dechloromonas, Tolumonas/Thauera/Dechloromonas, Thauera dominated in the PHBV/PLA, glucose and CH3COONa supported denitrification systems, respectively. Transcriptome-based analysis further indicated that the glucose supported denitrification system showed the highest FPKM values (the fragments per kilobase per million mapped reads) of the genes participating in the dissimilatory nitrate reduction process, corresponding to the greatest effluent NH4+-N concentration. A better knowledge of effect of different carbon sources on denitrification process will be significant for nitrate removal in practice. Display omitted •The maximum denitrification rate was lowest in PHBV/PLA supported system.•The relative abundance of denitrification bacteria was highest in CH3COONa system.•PHBV/PLA and glucose systems showed abundant hydrolysis and fermenting bacteria.•The FPKM value of functional genes for DNRA process was highest in glucose system.