Reconstruction of molecular interactions between microalgae and associated bacteria using simplified models

The relationships between microalgae and bacteria represent one of the most important cross-kingdom associations in aquatic environments. These microscopic interactions have fundamental ecological importance since they mediate nutrient cycling and recycling, regulate the productivity and stability of aquatic food webs, and shape the ecosystem diversity. The interactions between microalgae and bacteria take place within the region immediately surrounding individual microalgal cells. This microenvironment is known as the phycosphere. Microalgal-bacterial relationships are expansive and can span mutualism, commensalism, parasitism, and competition. It has been demonstrated that these complex and dynamic interactions are based on the exchange of nutrients and signalling molecules and can be either species-specific or non-specific. In this thesis, the molecular interactions between microalgae and associated bacteria inside the phycophere were analyzed, by using simplified systems composed of microalga-bacterium co-cultures, under controlled laboratory conditions. The model diatom Phaeodactylum tricornutum was used to set-up all the thesis experiments, while the key bacterial players were changed through the diverse studied co-culture systems. Different levels of interactions were examined: growth dynamics, chemical modification of the phycosphere and trascriptome analysis. Firstly, it was set up a two members co-culture system composed of the diatom Phaeodactylum tricornutum and the bacterium Pseudoalteromonas haloplanktis TAC125 and used to study the growth dynamics existing between the two microorganisms. It was demonstrated that the bacterium can use both diatom dead cells and diatom-derived compounds to grow. On the contrary, the diatom growth was not influenced by the presence of the bacterium. Then, the obtained growth data was used to validate the mathematical model describing these dynamics. It was revealed that the interactions existing between P. tricornutum and P. haloplanktis modify the chemical composition of the synthetic phycosphere reconstructed in laboratory conditions. Both the monosaccharide composition of exopolysaccharides (EPS) released by the two microorganisms, and the extracellular metabolites (exo-metabolites) detected with NMR analysis were evaluated. Different EPS sugar profiles in co-culture respect to the bacterial and diatom controls were observed; a changing monosaccharidic composition of EPS was revealed also during the different growth phases. The NMR analysis was not informative enough to reconstruct the metabolic interactions between the diatom and the bacterium due to low metabolites concentration. The interactions between P. tricornutum and its associated bacteria, isolated from its own phycosphere, were analyzed with a transcriptomic approach. Four isolates (Marinobacter adhaerens, Labrenzia alexandrii, Roseovarius tolerans and Alteromonas naphthalenivorans), commonly found to be associated with diatoms, were chosen to set-up four different co-cultures; the effect of the isolates on the growth and transcriptome of the diatom was evaluated. Marinobacter, Labrenzia, and Roseovarius showed a positive effect on the growth of P. tricornutum, while Alteromonas exhibited a negative effect on diatom growth. Transcriptomic data revealed a differentially pathways regulation of P. tricornutum in the presence of the different isolates, suggesting the existence of species-specific interactions between the diatom and the associated bacteria. The present thesis brings a step forward in the current knowledge on microalgal-bacterial interactions, though an in-depth understanding of the interaction mechanisms still requires further research. This research field can provide useful insights into globally significant biogeochemical processes and the basal function of aquatic ecosystems.