Process-level design of engineered microalgal-bacterial systems for carbon-efficient nitrogen removal from low C/N...

Carbon scarcity in low carbon-to-nitrogen (C/N) wastewater limits electron donor availability and constrains biological nitrogen removal. Although microalgal-bacterial symbiosis (MBS) is a promising low-input alternative, the mechanisms that sustain nitrogen removal under carbon-limited conditions remain unclear. Here, process-level characterization and metagenomic analysis were combined to investigate community assembly and carbon/electron redistribution in engineered MBS systems. Under the tested conditions, a balanced algae-to-bacteria ratio (1:1) created the most stable niche and achieved >97% NH₄⁺-N removal with minimal nitrate accumulation, indicating effective coupling of nitrification, denitrification, and assimilation. Extracellular polymeric substances (EPS) dynamics showed a shift from accumulation to reutilization during prolonged carbon limitation: polysaccharides decreased in the later stage as external chemical oxygen demand (COD) was depleted, suggesting mobilization of EPS as an internal carbon source. Consistently, tricarboxylic acid (TCA) cycle genes (e.g., IDH, OGDH, mdh) were enriched whereas glycolysis-related genes (e.g., GAPDH, PGK) declined, indicating a shift in metabolic potential toward greater generation of reducing equivalents. Overall, the results suggest that EPS functions as a dynamic carbon reservoir and that algae-bacteria interactions promote carbon/electron redistribution under carbon-limited conditions. This study provides a process-level basis for designing carbon-efficient wastewater treatment systems.