Background/Question/Methods The marine cyanobacterium, Prochlorococcus, is the most abundant oxygenic phototroph in the world's oceans, with an estimated global population size of 10²⁷ cells.  It forms the base of the marine food web in the oligotrophic gyres of the tropical and subtropical oceans, among the largest contiguous ecosystems on Earth.  Since it's discovery in 1988, dozens of strains have been isolated.  Surprisingly, none of these cultures could use nitrate (NO₃^-), a major source of nitrogen in marine environments.  Genome sequences from 12 Prochlorococcus isolates confirmed this conclusion, revealing the absence of genes encoding nitrate assimilation proteins.  The lack of these genes in Prochlorococcus was even more puzzling given that closely related, and co-occurring, Synechococcus cells have them.  However, most Prochlorococcus cultures were isolated in ammonium-based medium, which may have selected for cells incapable of growth using nitrate.  We hypothesized that some environments provide selective pressures that would favor the retention of nitrate assimilation capabilities in wild Prochlorococcus populations, and looked for evidence of this trait in metagenomic data from the Global Ocean Survey, an extensive DNA sequence dataset containing over 10 million sequence reads.  We first did a broad search for DNA fragments carrying the gene for assimilatory nitrate reductase, and then within this pool looked for fragments that also contained genes with close homology to known Prochlorococcus genes.

Results/Conclusions Our results confirmed that uncultured Prochlorococcus do possess the genetic potential for nitrate assimilation, and metatranscriptomics data from the field confirmed that these genes are expressed (A.C. Martiny, S. Kathuria, and P.M. Berube. 2009. Proc. Natl. Acad. Sci. USA 106:10787–10792).  Our data suggest that the ability to assimilate nitrate is associated with microdiverse lineages within several known Prochlorococcus ecotypes.  Recently, we have been successful in cultivating both high-light and low-light adapted ecotypes capable of growing solely on nitrate as a nitrogen source, and will soon sequence these novel strains.  We are also studying the relative frequency of nitrate utilizing Prochlorococcus in different ocean habitats and using single-cell genomics to further explore the distribution, abundance, and diversity of uncultivated Prochlorococcus.  This work has significantly changed our understanding of nitrogen metabolism in Prochlorococcus and has implications for understanding its biogeography and role in the oceanic carbon and nitrogen cycles.