Metagenomic understanding of the performance enhancement in anaerobic digestion by granular activated carbon coupled with riboflavin under high organic loading.

Anaerobic digestion (AD) often suffers operation failure from ammonia inhibition and volatile fatty acids (VFAs) accumulation under high organic loading rates (OLRs). To overcome these limitations, this study employed granular activated carbon coupled with riboflavin (RFGAC) by stimulating direct interspecies electron transfer (DIET). A semi-continuous AD experiment was conducted for 145 days with OLRs ranging from 2.25 to 11.25 kg COD/(m3·d). The results showed that the RFGAC group achieved the highest methane content of 78%, and maintained a COD removal rate above 95%, outperforming the GAC group and the control. At an OLR of 6.75 kg COD/(m3·d), the control collapsed due to severe acidification when the pH dropped lower than 6.5, while the RFGAC group stably operated with effluent COD of 2200-5300 mg/L and seldom VFAs accumulation. Microbial community analysis revealed that RFGAC selectively shifted microbial community composition especially at high OLR, promoting Methanosarcina to form a synergistic consortium. The Pearson correlation analysis of digestion performance and metagenome revealed that Methanosarcina had a stronger correlation with methanogenesis than Methanothrix, which was enriched in the presence of GAC alone. Metabolic pathway analysis confirmed key DIET-related functional genes, hdrA2 and methyl transfer-associated mtrH, were respectively upregulated by 7-fold and 5-fold. This study offers a viable strategy to improve chicken manure AD, and provides deep mechanistic insights on RFGAC modulation of microbial community succession and functional gene expression.