Metagenome-resolved evidence that soluble factors in granular activated carbon-amended reactor effluent reprogram propionate metabolism and methanogenic pathways.

Granular activated carbon (GAC) enhances anaerobic digestion performance, yet the mechanisms underlying reactor-scale improvements remain incompletely understood, particularly how GAC affects biomass not attached to its surface. Here, sludge from a non-GAC up-flow anaerobic sludge blanket reactor was incubated with 0.45-μm-filtered effluents from non-GAC and GAC-amended reactors under repeated propionate loading, followed by genome-resolved metagenomics. GAC-reactor effluent increased methane yield from 64 ± 3% to 76 ± 3% (p < 0.01) in the absence of GAC particles. A non-redundant catalog of 170 quality-filtered metagenome-assembled genomes (MAGs) was recovered, enabling pathway- and gene-set quantification. Genomic potential for both major propionate-oxidation routes increased in the GAC-effluent group relative to the non-GAC group, with a larger increase for the methylmalonyl-CoA (MMC) route than for the dismutation route (1.289- versus 1.221-fold). Accordingly, the MMC-to-dismutation preference ratio was 5.60% higher in the GAC-effluent group, alongside a broader carrier base. Cobamide potential shifted toward remodeling and cobamide-dependent use rather than increased de novo corrin-ring synthesis. Candidate electron-transfer architectures were also rebalanced: PilA-associated carriers became less prominent, whereas maturation-supported multiheme cytochrome carriers increased from 22.96% to 34.90% of community abundance, although H2/formate-module carriers remained prevalent. Quorum-sensing systems underwent pathway- and carrier-specific redistribution, while all eight curated extracellular-polysaccharide modules showed higher mean gene abundance in the GAC-effluent composite. These findings show that a filter-passing effluent fraction can extend GAC-associated effects beyond direct particle contact and link enhanced methanogenesis to a broader, redistributed network of metabolic, redox, and coordination capacities. This expands the mechanistic framework of conductive-material-assisted anaerobic digestion and provides a basis for harnessing GAC-derived functions throughout the reactor.