Hydrogen (H₂) is an important electron donor in deep-sea reducing environments, yet its role in regulating nitrate reduction remains poorly understood, particularly across different chemosynthetic ecosystems. Here, we investigated the interactions between H₂ metabolism and nitrate reduction in sediments collected from deep-sea cold seeps, hydrothermal vents, and transitional habitats. Geochemical profiles, metagenomic sequencing, 16S rRNA amplicon analysis, and ¹⁵N isotope-tracer incubation experiments were integrated to quantify nitrate reduction pathways, including denitrification, dissimilatory nitrate reduction to ammonium (DNRA), anammox, and N₂O production. Distinct nitrate reduction patterns were observed among habitats, indicating substantial variability in nitrogen transformation processes under different geochemical settings. By linking H₂ distributions with nitrogen transformation rates and microbial community composition, we identified potential microbial groups and metabolic pathways involved in H₂-dependent nitrate reduction. Genomic evidence further suggested that hydrogen oxidation is coupled to multiple nitrate reduction pathways through diverse hydrogenases and nitrogen cycling genes, highlighting H₂ as an important energy source supporting nitrogen turnover in deep-sea reducing sediments. Our study provides a process-based framework for understanding how hydrogen availability regulates microbial nitrogen cycling in chemosynthetic ecosystems and offers new insights into the mechanisms controlling nitrogen retention, nitrogen loss, and N₂O production in the deep ocean.

 

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