Abstract Nitrous oxide (N2O) is a potent climate gas, with its strong warming potential and ozone-depleting properties both focusing research on N2O sources. Although a sink for N2O through biological fixation has been observed in the Pacific, the regulation of N2O-fixation compared to canonical N2-fixation is unknown. Here we show that both N2O and N2 can be fixed by freshwater communities but with distinct seasonalities and temperature dependencies. N2O fixation appears less sensitive to temperature than N2 fixation, driving a strong sink for N2O in colder months. Moreover, by quantifying both N2O and N2 fixation we show that, rather than N2O being first reduced to N2 through denitrification, N2O fixation is direct and could explain the widely reported N2O sinks in natural waters. Analysis of the nitrogenase (nifH) community suggests that while only a subset is potentially capable of fixing N2O they maintain a strong, freshwater sink for N2O that could be eroded by warming.
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