Epibiotic bacterial community composition in red-tide dinoflagellate Akashiwo sanguinea culture under various growth conditions

The phycosphere of phytoplankton harbors a diversity of microbes that can potentially interact with the host phytoplankton cells. In spite of increasing reports regarding the interactions between some model phytoplankton and bacteria, there remain many unknowns regarding interactions between the dinoflagellate Akashiwo sanguinea and its phycosphere bacteria. Here we used both cultivation and metagenomic sequencing methods to investigate the microbial community composition in A. sanguinea batch cultures under various growth conditions. The microbial community dynamics under different growth stages and nutrient (nitrogen and iron) depletion scenarios were determined by Illumina MiSeq sequencing of 16S rRNA gene amplicons. Rhodobacteraceae, Alteromonadaceae and Flavobacteriaceae were the main bacterial families and varied significantly under different states of the host A. sanguinea. Selective fluorescence in situ hybridization was also performed to label Rhodobacterales and Alteromonas clade bacteria to confirm their attachment to A. sanguinea cells under confocal laser scanning microscopy. Plate streaking protocol isolated 19 bacterial strains from A. sanguinea cultures, identified through 16S rRNA analysis. Most culturable strains (11 of 19) belonged to Rhodobacteraceae consistent with Illumina MiSeq sequencing data. The possible interaction pathway between these epibiotic bacteria and A. sanguinea in the phycosphere is also discussed.