Biogeochemical controls and isotopic signatures of nitrous oxide production by a marine ammonia-oxidizing bacterium

Nitrous oxide (N2O) is a trace gas that contributes to the greenhouse effect and stratospheric ozone depletion. The N2O yield from nitrification (moles N2O-N produced per mole ammonium-N consumed) has been used to estimate marine N2O production rates from measured nitrification rates and global estimates of oceanic export production. However, the N2O yield from nitrification is not constant. Previous culture-based measurements indicate that N2O yield increases as oxygen (O-2) concentration decreases and as nitrite (NO2-) concentration increases. Here, we have measured yields of N2O from cultures of the marine beta-proteobacterium Nitrosomonas marina C-113a as they grew on low-ammonium (50 mu M) media. These yields, which were typically between 4 x 10(-4) and 7 x 10(-4) for cultures with cell densities between 2 x 10(2) and 2.1 x 10(4) cells ml(-1), were lower than previous reports for ammonia-oxidizing bacteria. The observed impact of O-2 concentration on yield was also smaller than previously reported under all conditions except at high starting cell densities (1.5 x 10(6) cells ml(-1)), where 160-fold higher yields were observed at 0.5% O-2 (5.1 mu M dissolved O-2) compared with 20% O-2 (203 mu M dissolved O-2). At lower cell densities (2 x 10(2) and 2.1 x 10(4) cells ml(-1)), cultures grown under 0.5% O-2 had yields that were only 1.25- to 1.73-fold higher than cultures grown under 20% O-2. Thus, previously reported many-fold increases in N2O yield with dropping O-2 could be reproduced only at cell densities that far exceeded those of ammonia oxidizers in the ocean. The presence of excess NO2- (up to 1mM) in the growth medium also increased N2O yields by an average of 70% to 87% depending on O-2 concentration. We made stable isotopic measurements on N2O from these cultures to identify the biochemical mechanisms behind variations in N2O yield. Based on measurements of delta N-15(bulk), site preference (SP = delta N-15(alpha)-delta N-15(beta)), and delta O-18 of N2O (delta O-18-N2O), we estimate that nitrifier-denitrification produced between 11% and 26% of N2O from cultures grown under 20% O-2 and 43% to 87% under 0.5% O2. We also demonstrate that a positive correlation between SP and delta O-18-N2O is expected when nitrifying bacteria produce N2O. A positive relationship between SP and delta O-18-N2O has been observed in environmental N2O datasets, but until now, explanations for the observation invoked only denitrification. Such interpretations may overestimate the role of heterotrophic denitrification and underestimate the role of ammonia oxidation in environmental N2O production.