Oxygen isotope fractionation during anaerobic ammonium oxidation by the marine representative <i>Candidatus</i> Scalindua sp.

Analysing the nitrogen (15ε) and oxygen (18ε) isotope effects of anaerobic ammonium oxidation (anammox) is essential for accurately assessing its potential contribution to fixed-N losses in the ocean, yet the 18ε of anammox remains unexplored. Here, we determined the previously unexplored 18ε of anammox using a highly enriched culture of the marine anammox species "Ca. Scalindua sp". Because Scalindua significantly accelerated oxygen isotope exchange between NO2- and H2O, we introduced a new rate constant for anammox-mediated oxygen isotope exchange (keq, AMX = 8.44 ~ 13.56 × 10-2 h-1), which is substantially faster than abiotic oxygen isotope exchange (keq, abio = 1.13 × 10-2 h-1), into a numerical model to estimate the 18ε during anammox. Based on our experimental results, we successfully determined the 18ε associated with: (1) conversion of NO2- to N2 (18εNO2- → N2 = 10.6 ~ 16.1‰), (2) NO2- oxidation to NO3- (18εNO2- → NO3- = -2.9 ~ -11.0‰, inverse fractionation), (3) incorporation of oxygen from water during NO2- oxidation to NO3- (18εH2O = 16.4 ~ 19.2‰). Our study underscores the possibility that unique anammox oxygen isotope signals may be masked due to substantial anammox-mediated oxygen isotope exchange between NO2- and H2O. Therefore, careful consideration is required when utilizing δ18ONO3- and δ18ONO2- as geochemical markers to assess the potential contribution of anammox to fixed-N losses in the ocean.