Phosphorus availability mediates pathway-specific nitrogen cycling in stratified peatland microbiomes

Peatland microbiomes regulate nitrogen (N) cycling processes that control nutrient retention and greenhouse gas emissions in carbon-rich ecosystems. Although depth-driven redox gradients structure microbial communities, how physicochemical stratification shapes the functional versus taxonomic organization of N-cycling microorganisms remains unclear. We used shotgun metagenomics to characterize N-cycling gene distributions, taxonomic affiliations, and metagenome-assembled genomes (MAGs) across depth and vegetation gradients in a temperate blanket bog. Depth emerged as the primary structuring factor, creating functional-taxonomic decoupling. Surface peat (0-20 cm) harbored functionally diverse but taxonomically constrained communities assembled deterministically around nitrification and labile N acquisition, while subsurface peat (20-40 cm) supported taxonomically richer but functionally-simpler communities assembled stochastically and enriched in denitrification and dissimilatory nitrate reduction. Linear mixed-effects models revealed pathway-specific controls on N cycling. Denitrification increased with depth (β = 11.53, P < .05), whereas organic N transformation declined (β = -5.81, P < .05); depth effects on nitrification and N fixation became non-significant after accounting for environmental variables. Phosphorus (P) emerged as the strongest environmental predictor, regulating nitrification (β = 95.40, P < .01), N fixation (β = 128.33, P < .01), organic N transformation (β = 80.53, P < .01), and denitrification (β = -109.63, P < .05), highlighting the importance of P availability in structuring microbial N cycling. This challenges traditional N-limitation paradigms in ombrotrophic systems. MAGs revealed Pseudomonadota as the dominant N-cycling lineage, while incomplete denitrification capacity indicated genetic potential for N₂O accumulation in subsurface layers. These findings demonstrate that P availability, rather than N content alone, regulates microbial N transformation capacity in peatlands, with implications for predicting nutrient dynamics under altered hydrological and nutrient deposition regimes.