Project: Do Rhizosolenia mats obtain N from cryptic co-occurring diazotrophs?

NSF Award Abstract:
Ocean ecosystems are reliant on tiny, microscopic phytoplankton that form the base of the marine food web, yet vast regions of sunlit open ocean waters also have chronically low concentrations of dissolved nitrogen (N), a nutrient that limits photosynthesis and growth. These ecosystems are highly regenerative, meaning that the organisms are adapted to low concentrations of nitrogen and recycle it efficiently. While this nitrogen recycling sustains growth, addition from other sources (or 'new' nitrogen) is important for fueling new growth and is ultimately linked to the ocean's ability to remove carbon dioxide from the atmosphere and store it in the deep ocean. New nitrogen is introduced into the surface ocean through the movement of deep waters with high nitrogen concentrations to the surface or through the process of biological nitrogen fixation -- the microbial conversion of nitrogen gas into a biologically available form. Diatoms are one of the most important phytoplankton groups in modern oceans and form the base of the food web in the most productive ocean ecosystems. In the low nutrient open ocean they can sometimes form dense aggregates (or 'mats') that can descend into deeper waters to obtain the nitrogen needed for their growth using buoyancy regulation. The investigators recently observed and sampled mats of multiple Rhizosolenia diatom species in the North Pacific Subtropical Gyre (NPSG) and showed that their microbiome contained a diverse array of microbes capable of nitrogen fixation. This discovery calls into question where Rhizosolenia mats acquire nitrogen and suggests that they may obtain it from living in symbiosis with nitrogen-fixing microbes. Importantly, fragile Rhizosolenia mats are not well sampled using traditional oceanographic techniques, as such we know very little about these microbial ecosystems and their contribution to oceanic productivity. This project is characterizing Rhizosolenia mat ecosystems, determining whether they are growing on nitrogen from nitrogen-fixers, and assessing their contribution to the nitrogen cycle in the NPSG. The investigators are combining traditional microscopy techniques, as well as modern multi-omics, imaging, and stable isotope tracer techniques. They are using deployable optical instrumentation and satellite data to track the location of mats during a research cruise to the NPSG, and using blue-water diving to sample and incubate the fragile mats. This project is having an impact beyond advancing discovery by providing professional development opportunities for early career ocean researchers, including exposure to a broad array of transferable skills, from scientific diving to molecular techniques. The investigators are also developing a hands-on educational module about marine phytoplankton, symbioses, and ocean nutrient cycles to be featured at the Moss Landing Marine Labs Open House, a free public outreach event held annually each spring.

Diazotrophy, the microbial fixation of dinitrogen gas into ammonia, supports a significant amount of primary production in the chronically nitrogen-limited oligotrophic ocean. However, the relative importance of different diazotrophs to primary production is not clear, and ongoing discoveries of novel diazotrophs highlight our incomplete understanding of marine nitrogen-fixers. Phytoplankton vertical migration is an additional source of new nitrogen to surface waters in oligotrophic systems, and multispecies, migrating Rhizosolenia aggregates (or 'mats') have been reported to contribute significantly to both primary production and carbon export fluxes due to their ability to transport deep nitrogen into surface waters. The investigators encountered Rhizosolenia mats on a research cruise in the North Pacific Subtropical Gyre (NPSG) in 2022, which led to the discovery that they contain a varied assemblage of diazotrophs, but not the heterocyst-forming Richelia known to form associations with some Rhizosolenia sp. These findings, along with a historical observation of dinitrogen gas fixation in Rhizosolenia mats, suggest that these mats may acquire some of their needed nitrogen from diazotrophy, and mat-associated dinitrogen gas fixation constitutes an unrecognized important source of nitrogen to the NPSG. This project is assessing the composition, activity, and symbiotic nature of Rhizosolenia mat communities, as well as determining their significance to the nitrogen inventory in the NPSG. The investigators are providing the first detailed characterization of mat-forming Rhizosolenia and their associated diazotroph communities by using a combination of traditional microscopy techniques (light microscopy, Scanning Electron Microscopy, Transmission Electron Microscop), molecular and 'omics tools (metagenome-assembled genomes, Rhizosolenia barcoding using voucher isolate strains, fluorescence-based visualization, amplicon High Throughput Sequencing) and stable isotope-based approaches at both whole mat and sub-mat scales (using nanoscale secondary ion mass spectrometry). Demonstrating that Rhizosolenia mats obtain diazotroph-derived nitrogen would transform the current paradigm about the role of these mats in nitrogen and carbon biogeochemical cycles and identify a novel diazotroph niche that is missed with conventional sampling. This project is also opening avenues to explore fundamental questions of diatom evolution and characterization of diatom strategies for metabolic adaptation to low nutrient environments through the isolation of mat-forming diatoms and generation of metagenome-assembled genomes. Additionally, morphological and molecular characterization of these fragile and cryptic Rhizosolenia mats is significantly contributing to illuminating the unseen pelagic microbiome.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.