Staying alive: sinking Trichodesmium fixes nitrogen in the dark ocean

Sunlight extinguishes about 150 m in the ocean, retaining photosynthetic microbes such as the diazotrophic cyanobacteria in the surface ocean layer. Here we find that Trichodesmium, a key photosynthetic diazotroph, fixes nitrogen while sinking into the deep ocean down to 1000 m depth.

Life in the oceans is sustained by nitrogen. In about 60% of the global ocean surface, nitrogen is provided by "diazotrophs", small planktonic cells capable of reducing atmospheric dinitrogen (N2) into reactive forms. Diazotrophs provide the nitrogen that fuels as much as half of the global primary production and balance the global nitrogen reservoir. Hence, diazotrophs are crucial for global climate regulation.

N2 fixation was long attributed solely to photosynthetic diazotrophic cyanobacteria that inhabit the warm and oligotrophic surface waters of the (sub)tropical oceans, such as Trichodesmium and Crocosphaera (Zehr and Capone, 2020). However, several studies have found active N2 fixation and nifH genes (encoding for N2 fixation enzymes) below 200 m in the dark ocean, where there is no light available to do photosynthesis. This suggested that non-cyanobacterial diazotrophs were the resident microorganisms responsible for those signals in the dark ocean (Moisander et al., 2017). However, after years of dark ocean N2 fixation and nifH gene measurements in the Mediterranean Sea, North and South Pacific Oceans (Benavides et al., 2018), we came to observe a regular pattern: cyanobacteria nifH genes were regularly detected in the dark ocean. What were they doing there? Were these diazotrophs active? How could they cope with the dark, cold and high-pressure conditions of the deep ocean?

So we went out to sea to find answers. In 2019 during the TONGA cruise (http://tonga-project.org/) in the western tropical South Pacific, we deployed sediment traps at two locations (Fig. 1). The traps were filled with 15N2-enriched brine, which allowed measuring N2 fixation activity by any diazotroph sinking into the traps at in situ conditions. Replicate traps were filled with a nucleic acid preservative, which allowed us to get intact RNA from diazotrophs sinking into the traps. We found that Trichodesmium, the surface ocean dweller, was the majoritary diazotroph in the traps and… that they were actively fixing N2 and expressing nifH!

Fig. 1: Sediment trap deployment during the TONGA cruise (10.17600/18000884) onboard the R/V L'Atalante in 2019. Copyright H. Bataille, IRD. More about the TONGA cruise: https://www.youtube.com/watch?v=UeABf-cVR-k

Trichodesmium is a filamentous colonial cyanobacterium common in the low latitude bands of the ocean, thought to provide alone 60-80 Tg N per year globally Trichodesmium uses sunlight and nutrients to do photosynthesis, producing the high energy amounts needed to sustain the expensive process of N2 fixation. How could sinking Trichodesmium cope with the harsh conditions experienced while sinking into the dark ocean? To explore that further, we took Trichodesmium cultures and submitted them to a nifty experiment: a sinking particle simulator (Fig. 2). These are titanium bottles where temperature and pressure can be respectively decreased and increased to simulate the conditions cells experience while sinking from the surface into the dark ocean. These experiments showed that pressure decreased but did not completely impair N2 fixation in Trichodesmium

Fig. 2: Sinking particle simulator. Photo courtesy of Christian Tamburini.

Still, the mystery of the light remained to be solved. Trichodesmium relies on photosynthesis to generate energy and fuel N2 fixation. If there's no light, how can they fix N2 in the dark ocean? To address this question, we adapted a previously developed Trichodesmium metabolism computational model (Inomura et al., 2019). The model is written in a few hundred lines of a programming language. It reads environmental information (e.g., light, oxygen concentration and temperature), and calculates metabolic fluxes (e.g., photosynthesis, respiration and N2 fixation). Applying the model to our field site conditions we found that carbon storage accumulated via photosynthesis before sinking allows Trichodesmium to fuel N2 fixation down to 1000 m into the dark ocean.

Diazotrophically active sinking Trichodesmium can provide mesopelagic bacteria and archaea with a more labile source of nitrogen (ammonium and amino acids) than the more abundant nitrate, contributing to microbial organic matter remineralization or chemoautotrophy in the ocean’s twilight zone. While the role of microbes in controlling organic matter export by respiration is well established, our results point towards a role of sinking surface microbes in anabolic processes in the deep ocean.