Dual nitrogen and oxygen isotope fractionation during anaerobic ammonium oxidation by anammox bacteria

Natural abundance of stable nitrogen (N) and oxygen (O) isotopes are invaluable biogeochemical tracers for assessing the N transformations in the environment. To fully exploit these tracers, the N and O isotope effects ((15)epsilon and (18)epsilon) associated with the respective nitrogen transformation processes must be known. However, the N and O isotope effects of anaerobic ammonium oxidation (anammox), one of the major fixed N sinks and NO3- producers, are not well known. Here, we report the dual N and O isotope effects associated with anammox by three different anammox bacteria including Ca. Scalindua japonica, a putative marine species, which were measured in continuous enrichment culture experiments. All three anammox species yielded similar N isotope effects of NH4+ oxidation to N-2 ((15)epsilon(NH4 -> N2)) ranging from 30.9 parts per thousand to 32.7 parts per thousand and inverse kinetic isotope effects of NO2- oxidation to NO3- ((15)epsilon(NO2)(->)(NO3) = -45.3 parts per thousand to -30.1 parts per thousand). In contrast, (15)epsilon(NO2 ->)(N2)(NO2- reduction to N-2) were significantly different among three species, which is probably because individual anammox bacteria species might possess different types of nitrite reductase. We also report the combined O isotope effects for NO2- oxidation (E-18(NO2)->(NO3)) by anammox bacteria. These obtained dual N and O isotopic effects could provide significant insights into the contribution of anammox bacteria to the fixed N loss and NO2- reoxidation (N recycling) in various natural environments.