Overflow metabolism drives high-rate carbon, nitrogen and phosphorus sequestration in an enriched aerobic microbial...

Waste stabilisation ponds (WSPs) are globally ubiquitous for low-cost wastewater treatment, yet the functional potential of their microbiomes remains largely underutilised. This study investigates the kinetic and metabolic capabilities of an aerobic microbial community enriched from a WSP, revealing a phenotype defined by "unbalanced growth" that fundamentally restructures remediation stoichiometry. Under pulse-fed conditions, the enriched community exhibited high nutrient removal rates (10, 0.7, and 0.3 mmol gMLSS^-1·h^-1 for glucose, NH₄⁺-N, and PO₄³⁻-P, respectively) at an operational dissolved oxygen concentration of 6 mg L^-1, exceeding conventional activated sludge rates by an order of magnitude. Comprehensive metabolic profiling was consistent with carbon uptake being decoupled from oxidative phosphorylation, an overflow-like phenotype not directly confirmed by respirometry, NAD(P)H, or proteomic data. This kinetic mismatch between glycolytic flux and respiratory capacity triggered several metabolic relief valves: excess carbon flux was shunted toward pyruvate excretion and delayed polyhydroxybutyrate (PHB) synthesis, while surplus ATP is proposed to have driven the luxury uptake of phosphate into intracellular polyphosphate (Poly-P), evidenced by Methylene Blue staining and the concurrent depletion of phosphate from solution. Uniquely, nitrogen assimilation and intracellular sequestration were also decoupled from biomass formation, reflecting a complex, process-sustained partitioning among assimilatory pathways, cyanophycin storage, and transient intracellular pooling driven by carbon limitation. Operationally, this overflow-like phenotype diverted ∼61% of incoming carbon into harvestable biomass and storage polymers, reducing on-site oxidative carbon loss relative to conventional activated sludge. This strategy suits high-COD, low-nutrient waste streams, requiring ∼4-fold higher COD than typical municipal wastewater. These findings offer a mechanistic blueprint for process intensification, demonstrating that exploiting the thermodynamic trade-offs between kinetic rate and energetic yield can transform wastewater treatment from a liability-management process into a high-rate, carbon-conserving resource-recovery strategy.