Genetic Manipulation of Competition for Nitrate between Heterotrophic Bacteria and Diatoms

Diatoms are a dominant group of eukaryotic phytoplankton that contribute substantially to global primary production and the cycling of important elements such as carbon and nitrogen. Heterotrophic bacteria, including members of the gammaproteobacteria, are commonly associated with diatom populations and may rely on them for organic carbon while potentially competing with them for other essential nutrients. Considering that bacterioplankton drive oceanic release of CO₂ (i.e., bacterial respiration) while diatoms drive ocean carbon sequestration vial the biological pump, the outcome of such competition could influence the direction of carbon flux in the upper ocean. A common determining factor for the growth of diatom populations, particularly in coastal and upwelling regions, is the availability of nitrate, which both diatoms and many marine bacteria are able to utilize as a nitrogen source. However, the ability of bacteria to compete with phytoplankton for nitrate may be hindered by carbon limitation. Here we have developed a genetically tractable model system using the diatom Phaeodactylum tricornutum and the widespread heterotrophic bacteria Alteromonas macleodii to study carbon-nitrogen dynamics. A. macleodii can subsist solely on P. tricornutum derived carbon, and does not appear to compete for nitrate. However, allochthonous carbon addition triggers A. macleodii proliferation and nitrate uptake, leading to a suppression of P. tricornutum growth. Nitrate-reductase deficient mutants of A. macleodii (ΔnasA) do not exhibit such explosive growth competitive ability in response to allochthonous carbon when nitrate is the sole N source. However, ΔnasA lines are able to persist in P. tricornutum cultures with nitrate as the sole nitrogen source, suggesting the ability to survive utilizing P. tricornutum-derived nitrogen. We also examine the ability of wild-type A. macleodii to rescue populations of nitrate-reductase deficient P. tricornutum from nitrogen starvation with and without the addition of carbon. This study provides key insights into the roles of carbon and nitrogen in phytoplankton-bacteria dynamics and lays the foundation for developing a mechanistic understanding of these interactions using co-culturing and genetic manipulation.