Unveiling the hidden nitrogen sink: significant contribution of oxygen-tolerant denitrification in a freshwater river–lake continuum

Denitrification serves as a critical natural buffer against anthropogenic nitrogen inputs in river–lake continuums. While traditionally considered an anaerobic process predominantly occurring in rivers and lakes, our study reveals the remarkable coexistence and substantial potential of both anaerobic and oxygen-tolerant denitrification in the Fu River–Baiyangdian Lake continuum. By integrating helium-based closed incubation with 16S rRNA gene sequencing, we simultaneously quantified these processes, quantified maximum anaerobic and oxygen-tolerant denitrification potentials at 239.84 and 442.99 mg/kg/day, respectively. The anaerobic and oxygen-tolerant denitrification potentials exhibited distinctly different spatiotemporal heterogeneity: anaerobic denitrification potential peaked in June (∼48.07 mg/kg/day), whereas oxygen-tolerant denitrification potential reached its maximum in August (∼62.18 mg/kg/day). Oxygen-tolerant denitrification potential generally exceeded anaerobic during summer but reversed in winter (December). Spatially, anaerobic denitrification potentials dominated upstream river sections with rates 1.5 to 8.2 times higher than oxygen-tolerant denitrification potentials, while oxygen-tolerant denitrification potentials prevailed in the river–lake transition and lake areas, reaching up to 41 times the anaerobic rates. Moreover, abundant nitrogen reservoirs, spatiotemporally dynamic hydrodynamic conditions governing sediment–water interactions along the river–lake continuum, and the distinct environmental preferences within microbial communities likely facilitate the unique coexistence and sustained activity of both anaerobic and oxygen-tolerant denitrification processes in this system. These findings challenge the traditional unified view of denitrification, revealing a functional decoupling of anaerobic and oxygen-tolerant pathways across environmental gradients. Distinguishing these processes is essential for advancing mechanistic understanding and improving hydrologically constrained nitrogen budget estimations across river–lake systems and other global inland waters.