Unicellular Cyanobacterium Symbiotic with a Single-Celled Eukaryotic Alga

Abstract

Symbioses between nitrogen (N)(2)-fixing prokaryotes and photosynthetic eukaryotes are important for nitrogen acquisition in N-limited environments. Recently, a widely distributed planktonic uncultured nitrogen-fixing cyanobacterium (UCYN-A) was found to have unprecedented genome reduction, including the lack of oxygen-evolving photosystem II and the tricarboxylic acid cycle, which suggested partnership in a symbiosis. We showed that UCYN-A has a symbiotic association with a unicellular prymnesiophyte, closely related to calcifying taxa present in the fossil record. The partnership is mutualistic, because the prymnesiophyte receives fixed N in exchange for transferring fixed carbon to UCYN-A. This unusual partnership between a cyanobacterium and a unicellular alga is a model for symbiosis and is analogous to plastid and organismal evolution, and if calcifying, may have important implications for past and present oceanic N(2) fixation.

High levels of heterogeneity in diazotroph diversity and activity within a putative hotspot for marine nitrogen fixation

“…Despite high silicate concentrations and pigment indications of diatom-dominated phytoplankton communities in the ATS, only one OTU associated with the heterocystous cyanobacterial symbiont of diatoms, Richelia, was observed in our sequence data, and this represented a total of only six nifH sequences (data not shown). The significant negative correlation observed between silicate and UCYN-A1 and γ-24774A11 transcripts, and UCYN-A1 and fucoxanthin, could be indicative of shifting phytoplankton communities between the ATS and Coral Sea, given that UCYN-A is known to live in association with a prymnesiophyte host (Thompson et al, 2012;Hagino et al, 2013;Krupke et al, 2013). Currently, the lifestyle (for example, free-living, particle attached or symbiont) of γ-24774A11 remains unknown (Langlois et al, 2015); however, it has been speculated that it may depend upon phytoplankton-produced dissolved organic carbon (Moisander et al, 2012(Moisander et al, , 2014.…”

Coupling FACS and Genomic Methods for the Characterization of Uncultivated Symbionts

“…In these cases, FACS can be coupled to nested PCR, which is applied to sorted single-events (host-symbiont complexes), to amplify phylogenetically relevant genes for sequencing. This is a relatively low cost method of identifying symbiotic partners before further sequencing efforts (Thompson, et al, 2012) (Figure 1). …”

“…The freeze-thaw cycle should disrupt any delicate intercellular connections and leave the one symbiont intact as in a study on symbiotic cyanobacteria Thompson, et al, 2012). Once thawed, the concentrated sample is used as soon as possible, and certainly within a few hours.…”

“…Preventing disruption of intercellular connections by turbulence, freezing, or by chemical treatment is essential to exploring intact associations. For open ocean studies, the Influx TM can be equipped for the field (Thompson, et al, 2012;Worden, et al, 2012), which is required for analyzing untreated open ocean samples. The target symbiont can be located among cell populations from the untreated samples with the same sort strategy described in Section 3.…”

“…Phylogenetic analysis and alignment of the sequences can be conducted through BLASTn to GenBank (Benson, et al, 2013) or through other tools such as the Ribosomal Database Project (Wang, et al, 2007), SILVA (Quast, et al, 2013), or SINA (Pruesse, et al, 2012). Negative controls often contain amplification products after the qPCR and nested PCR procedures, so it is important to work under as clean conditions as possible for highest efficiency of sequencing nested PCR products (Thompson, et al, 2012). The sorts from each row and column are screened with a qPCR assay speci c to one symbiont.…”