Award: OCE-1458095

Determining the taxonomic identity of marine single-cell eukaryotic microorganisms, the extent to which they contribute to carbon and nutrient cycling, and how they respond to environmental change remain among the major challenges in oceanography. Answering these questions is fundamental to understanding marine ecosystem functioning and to predicting how the marine environment will respond to global changes. Although the abundance, distribution, and ecological roles of eukaryotic microorganisms (i.e. those, unlike bacteria, having a separate compartment for storing genetic materials) have been the focus of many studies, there is still much to be learned, particularly for the smaller-sized members of eukaryotes, that are increasingly being recognized as important to oceanic biogeochemical cycling. This project filled major gaps in our understanding of the diversity and function of <20 µm microbial eukaryotes, with emphasis on their ecological role as predators (e.g. eating bacteria) and competitors with the picocyanobacteria Prochlorococcus and Synechococcus, the most abundant phototrophic cells in the ocean. We used a set of field- and culture-based experiments in combination with state-of-the-art methodologies, including fluorescence-activated cell sorting, isotope and fluorescent stain labeling, and next-generation molecular sequencing. We determined that despite their low abundance relative to picocyanobacteria, small microbial eukaryotes have a disproportionate contribution to carbon cycling. For example, in the western subtropical North Atlantic, although small photosynthetic eukaryotes (< 5 μm) were only ~ 5% of the microbial phytoplankton cell abundance in surface layers of the ocean, they represented at least two thirds of the microbial phytoplankton carbon biomass and fixed more CO2 than picocyanobacteria, accounting for roughly half of the volumetric CO2 fixation by the microbial phytoplankton, or a third of the total primary production. We also learned that phosphate assimilation rates of very small eukaryotes (< 5 μm) were generally higher than of picocyanobacteria but, when normalized to biovolumes, picocyanobacteria assimilated roughly four times more phosphate than small eukaryotes, indicating different strategies to cope with phosphate limitation. This outcome has consequences for ocean biology and biogeochemistry, since our results underline an imbalance in the CO2 to phosphate uptake rate ratios, which may be explained by bacterial predation providing microbial eukaryotes with their largest source of phosphate. Our work also significantly advanced our understanding of the ecological role of small microbial eukaryotes as predators of bacteria and the factors regulating their mode of nutrition. For example, in the phosphate-depleted western subtropical North Atlantic we found that bacterivory by photosynthetic microbial eukaryotes, less than 5 μm in size, was reduced when phosphate was added during experimental incubations, indicating that their feeding rate is regulated by phosphate availability. In laboratory culture experiments, we further confirmed the role of phosphate availability in regulating bacterial consumption by the small-sized marine green alga known as Cymbomonas tetramitiformis. Our team also screened small-sized green algae available in culture collections for their potential to eat bacteria using color-tagged live bacteria, a protocol developed for the project. From this effort, several marine green algae that previously assumed to be simply photosynthetic, were identified to be capable of eating bacteria as food source. We also conducted series of culture and feeding experiments to identify environmental factors that influence their bacterial feeding behavior. Overall, it appears that their eat bacteria more when the soluble nutrients become scarce. Further, we have gathered molecular sequence data from these algae in order to better understand their feeding behavior at the molecular level. Finally, our work significantly advanced our knowledge of small microbial eukaryotes? diversity in oligotrophic seawater. For example, we sequenced samples from eight tropical lagoon sites of the South Pacific and revealed a surprising number of novel eukaryotic microorganisms. Overall, this project furthered knowledge regarding the influence of understudied, but key, marine microbes in the functioning of marine food webs and biogeochemical cycling. This project supported the work of two early career scientists, two graduate students, a post-doctoral researcher as well as helped train several undergraduate and high school students, including women and minorities in the fields of science, technology, engineering, and mathematics (STEM). Results from the project have been published in peer-reviewed journals (five manuscripts as of August 2019, plus several in preparation), and disseminated through presentations at scientific meetings and databases. This research also stimulated new international collaborations. Our findings have also been shared at many public outreach events, including sciences fairs, science festivals and open houses in Museums and Universities. Last Modified: 08/30/2019 Submitted by: Eunsoo Kim