Denitrification mode management selects resource-conserving consortia for low-carbon municipal wastewater treatment

Low-carbon municipal wastewater treatment increasingly relies on carbon-limited denitrification, yet how carbon limitation reorganizes denitrifying communities and their greenhouse-gas footprint remains poorly resolved. We implemented denitrification mode management in a municipal wastewater sequencing batch reactor by switching from anaerobic-aerobic (AO) to anaerobic-aerobic-anoxic (AOA) operation. This shift moved denitrification from an pre-anaerobic stage with higher carbon availability to a post-anoxic stage where readily available carbon was limited. We combined metagenomics and metatranscriptomics to link process performance with microbial traits and gene expression. The mode switch improved nitrogen removal from 67.1 ± 1.8% to 88.5 ± 3.9% and reduced carbon requirement from 4.9 ± 0.5 to 3.3 ± 0.4 mg COD per mg N removed, while decreasing the N₂O emission factor from 0.024 to 0.005 mg N₂O-N per mg NO₃⁻-N and lowering CO₂ and CH₄ emissions by 20-30%. Carbon-limited post-denitrification selected taxa with smaller genomes, reduced metabolic redundancy and a pronounced shift from broad extracellular carbon catabolism and complete denitrification towards intracellular carbon storage and truncated denitrification. Intracellular carbon pools insulated organics at the single-cell level and buffered electron delivery across denitrification steps, enabling a division-of-labour network that prevents electron imbalance and suppresses N₂O build-up. Together, these findings link denitrification mode management to trait-based community restructuring and offer a process-level framework for understanding low-carbon, low-emission nitrogen removal under carbon-limited conditions.