Combined in vivo and in situ genome-resolved metagenomics reveals novel symbiotic nitrogen fixing interactions between non-cyanobacterial diazotrophs and microalgae

BackgroundNon-cyanobacteria diazotrophs (NCDs) were shown to dominate in surface waters shifting the long-held paradigm of cyanobacteria dominance and raising fundamental questions on how these putative heterotrophic bacteria thrive in sunlit oceans. The absence of laboratory cultures of these bacteria significantly limits our ability to understand their behavior in natural environments and, consequently, their contribution to the marine nitrogen cycle. ResultsHere, we used a multidisciplinary approach and report an unprecedented finding in the diatom Phaeodactylum tricornutum (Pt) of NCDs in the phycosphere or the pelagic community sustaining its survival in the absence of bioavailable nitrogen. We sequenced the bacterial metacommunity associated with Pt and assembled several bacterial genomes, identifying multiple NCDs from the Rhizobiales order, including Bradyrhizobium, Mesorhizobium, Georhizobium and Methylobacterium. We demonstrated the nitrogen-fixing ability of PtNCDs through in silico identification of nitrogen fixation genes, or by using PCR, acetylene reduction, or 15N incorporation. We showed the wide occurrence of this type of interactions with the isolation of NCDs from other microalgae, their identification in the environment, and their predicted associations with photosynthetic microalgae. ConclusionsOur study underscores the importance of microalgae interactions with NCDs to permit and support nitrogen fixation. This work provides a unique model Pt-NCDs to study the ecology of this interaction advancing our understanding of the key drivers of global marine nitrogen fixation.