Stage-specific biochar addition reprograms microbial metabolism to bridge hybrid anaerobic-aerobic bioprocess for...

The integration of high-solid anaerobic digestion (HSAD) and aerobic composting (AC) to form a hybrid anaerobic-aerobic bioprocess offers a viable route for simultaneous stabilization and resource recovery of organic wastes, such as sewage sludge, food waste, and agricultural wastes. However, the mechanisms by which stage-specific biochar intervention regulates metabolic pathways and microbial succession to optimize HSAD-AC integration remain unclear. Thus, this study investigated the timing and associated mechanisms of biochar addition to simultaneously enhance methane (CH₄) production and compost humification in the hybrid bioprocess. Results show that biochar addition at the beginning of the HSAD stage markedly outperformed that at AC stage to simultaneously increase the maximum daily CH₄ yield by approximately 3.5 times and the humic-to-fulvic acid ratio by 57.4%. Further analysis using Fourier Transform Ion Cyclotron Resonance Mass Spectrometry revealed that biochar addition at the HSAD stage reduced organic direct decarboxylation to 29.7% and increased oxidation by 220.9%. These altered reactions retained essential carbon precursors for subsequent repolymerization toward humification at the AC stage. The improved CH₄ yield and humification could also be related to biochar addition to enrich the keystone methanogens, particularly Methanosaeta to the relative abundance of 45.0%, at the HSAD stage and then obligate aerobes to above 68.8% at the AC stage. Genomic profiling confirmed this metabolic reprogramming via the significant enrichment of key genes (e.g., cdhC and atoB) to directionally enhance organic methanogenesis and humification. In addition, the emissions of greenhouse gases and ammonia at the AC stage could be mitigated by 8.2% - 35.0% to prevent nitrogen loss. As a result, the biochar-amended hybrid system thus synchronously yields intensive bioenergy and a mature compost, providing a mechanistic framework for advancing circular waste management.