Isolation and Characterization of a Rhizobium Bacterium Associated with the Toxic Dinoflagellate Gambierdiscus balechii

Algae-bacteria associations are increasingly being recognized to be important in shaping the growth of both algae and bacteria. Bacteria belonging to order Rhizobiales are important symbionts of legumes often developing as nodules on plant roots, but have not been widely documented in association with algae. Here, we detected, isolated, and characterized a Rhizobium species from the toxic benthic dinoflagellate Gambierdiscus culture. The sequence of 16S rDNA showed 99% identity with that of Rhizobium rosettiformans. To further characterize the bacterium, we amplified and sequenced a cell wall hydrolase (CWH)-encoding gene; phylogenetic analysis indicated that this sequence was similar to the homologs of Martellela sp. and Hoeflea sp, of order Rhizobiales. We performed PCR using nifH primers to determine whether this bacterium can fix N2; however, the results of sequencing analysis showed that it was closer to chlorophyllide a reductase-encoding gene (bchX), which is similar to nifH. Results of 16S rDNA qPCR showed that compared to that in the early exponential phase, the abundance of this bacterium increased during the late exponential growth phase of Gambierdiscus. When the dinoflagellate culture was subjected to N limitation, the abundance of the bacterium represented by both 16S rDNA and CWH increased. Based on these results and published literature, it is apparent that this Rhizobium bacterium benefits from the association with Gambierdiscus by hydrolyzing and utilizing the extracellular organic matter exudates released by the dinoflagellate. This is the first report of Rhizobium species being associated with dinoflagellates, which will shed light on the algae-bacteria relationships.\n\nIMPORTANCEPhytoplankton are the undisputed primary producers in the aquatic ecosystems and contribute approximately half of the global net primary productivity.\n\nDinoflagellates are one of the most important phytoplankton in the marine ecosystems. Commonly, they do not exist autonomously in the marine environment but rather co-live with many bacteria that interact with dinoflagellates, producing a dynamic microbial ecosystem. Their interactions play a major role in important processes such as carbon fluxes and nutrient regeneration in the ocean, ultimately influencing the global carbon cycle and the climate. Hence, there is a need to understand the association and relationships between dinoflagellates and bacteria. Here, we tried to elucidate these interactions through isolating and characterizing a bacterium from a benthic toxic dinoflagellate culture. Our study is the first report of such bacterium being recorded to be associated with a dinoflagellate in this genus, providing new insights into the dinoflagellate-bacteria association for future research.