Direct and indirect costs of dinitrogen fixation in Crocosphaera watsonii WH8501 and possible implications for the nitrogen cycle

The recent detection of heterotrophic nitrogen (N₂) fixation in deep waters of the southern Californian and Peruvian OMZ questions our current understanding of marine N₂ fixation as a process confined to oligotrophic surface waters of the oceans. In experiments with Crocosphaera watsonii WH8501, a marine unicellular diazotrophic (N₂ fixing) cyanobacterium, we demonstrated that the presence of high nitrate concentrations (up to 800μM) had no inhibitory effect on growth and N₂ fixation over a period of 2 weeks. In contrast, the environmental oxygen concentration significantly influenced rates of N₂ fixation and respiration, as well as carbon and nitrogen cellular content of C. watsonii over a 24-h period. Cells grown under lowered oxygen atmosphere (5%) had a higher nitrogenase activity and respired less carbon during the dark cycle than under normal oxygen atmosphere (20%). Respiratory oxygen drawdown during the dark period could be fully explained (104%) by energetic needs due to basal metabolism and N₂ fixation at low oxygen, while at normal oxygen these two processes could only account for 40% of the measured respiration rate. Our results revealed that under normal oxygen concentration most of the energetic costs during N₂ fixation (~60%) are not derived from the process of N₂ fixation per se but rather from the indirect costs incurred for the removal of intracellular oxygen or by the reversal of oxidative damage (e.g., nitrogenase de novosynthesis).Theoretical calculations suggest a slight energetic advantage of N₂ fixation relative to assimilatory nitrate uptake, when oxygen supply is in balance with the oxygen requirement for cellular respiration (i.e., energy generation for basal metabolism and N₂ fixation). Taken together our results imply the existence of a niche for diazotrophic organisms inside oxygen minimum zones, which are predicted to further expand in the future ocean.