Carbon flow allocation patterns of CH4, CO2, and biomass production vary with sewage and sediment microbial and biochemical factors in the anaerobic sewer environment

Understanding the carbon (C) fate in municipal sewers is imperative for optimizing current sewer-C-degradation control and treatment efficiency, aligning with China's C-neutrality strategy in determining the exact C budget of the wastewater system. This study used laboratory batch tests mimicking the anaerobic sewer environment and sewage-sediment stratification to evaluate C flow allocation (CFA) patterns in response to biotic and abiotic variables. We quantified the C equivalent mass (CEM) and used absolute quantitative 16S rRNA gene amplicon sequencing to characterize the microbiome. The substantial methane production (CH₄, 17.2%–18.8%) required both activated sediment and exogenous C, while biomass production (BP, 63.1%–74.9%) formed C sink predominated as the main CFA direction under the stratified state. This was supported by the high diversity, interspecific interactions, and metabolic capacity of the sediment microbiome. However, CH₄ and BP patterns demonstrated non-synchronicity and opposite dynamic characteristics. Carbon dioxide (CO₂, 64.0%–81.3%) production dominated the sewage CFA. The absolute abundance of the sediment microbiome, which was 5.6 times higher than that of the sewage, exhibited a strong increase in magnitude across the phases. It was primarily associated with biomass growth and N metabolism, whereas sewage showed differentiated and competing communities and appeared to act mainly as the exogenous C sources. We constructed a binary quadratic linear model revealing the non-linear relationship between ACK activity, DOC degradation rate, and CEM_CH4 rate; the former maintained low CH₄ production when the available substrate was insufficient. The influence of N and S factors on the CFA is complex and multi-faceted. These findings highlight the importance of further investigations into the process-based framework of the sewer C budget, focusing on the C source-emission-sink functions and mass balance.