Microbiome Mechanisms of Composite Carbon Sources for Enhancing Denitrification and Reducing N2O Emissions

• • Composite carbon source (sodium acetate: sodium succinate: ethanol = 2:1:3) increased denitrification rate by 22.7% (from 6.822±0.141 to 8.370±0.186 mg/(L·h)) and reduced N2O accumulation by ~55%, directly addressing carbon limitation in WWTPs and lowering operational costs. • • Metatranscriptomic analysis revealed that composite carbon sources upregulated key denitrification gene transcription: nirS by 37.8%, norB by 27.4%, and nosZ by 48.6%, with nosZ upregulation being the primary driver of N2O emission reduction. • • Metagenomic analysis identified Ottowia, Rubrivivax, Thauera, and Zoogloea as dominant denitrifying genera, indicating that composite carbon sources activate low-abundance denitrifiers and enhance transcriptional activity in dominant ones. • • Composite carbon sources induced complementary carbon metabolic strategies: Dechloromonas exhibited carbon-source inhibition, while Thauera and Zoogloea showed multi-substrate synergistic metabolism, improving reducing equivalent supply and enabling rapid, complete denitrification.