Isotopic overprinting of nitrification on denitrification as a ubiquitous and unifying feature of environmental nitrogen cycling

Natural abundance nitrogen and oxygen isotopes of nitrate (δ¹⁵NNO3 and δ¹⁸ONO3) provide an important tool for evaluating sources and transformations of natural and contaminant nitrate (NO₃⁻) in the environment. Nevertheless, conventional interpretations of NO₃⁻ isotope distributions appear at odds with patterns emerging from studies of nitrifying and denitrifying bacterial cultures. To resolve this conundrum, we present results from a numerical model of NO₃⁻ isotope dynamics, demonstrating that deviations in δ¹⁸ONO3 vs. δ¹⁵NNO3 from a trajectory of 1 expected for denitrification are explained by isotopic over-printing from coincident NO₃⁻ production by nitrification and/or anammox. The analysis highlights two driving parameters: (i) the δ¹⁸O of ambient water and (ii) the relative flux of NO₃⁻ production under net denitrifying conditions, whether catalyzed aerobically or anaerobically. In agreement with existing analyses, dual isotopic trajectories >1, characteristic of marine denitrifying systems, arise predominantly under elevated rates of NO₂⁻ reoxidation relative to NO₃⁻ reduction (>50%) and in association with the elevated δ¹⁸O of seawater. This result specifically implicates aerobic nitrification as the dominant NO₃⁻ producing term in marine denitrifying systems, as stoichiometric constraints indicate anammox-based NO₃⁻ production cannot account for trajectories >1. In contrast, trajectories <1 comprise the majority of model solutions, with those representative of aquifer conditions requiring lower NO₂⁻ reoxidation fluxes (<15%) and the influence of the lower δ¹⁸O of freshwater. Accordingly, we suggest that widely observed δ¹⁸ONO3 vs. δ¹⁵NNO3 trends in freshwater systems (<1) must result from concurrent NO₃⁻ production by anammox in anoxic aquifers, a process that has been largely overlooked.