Polysaccharide utilization by bacteria associated with micro- and macroalgae

Abstract

Algae play a crucial role in marine carbon cycling. This involves various polysaccharides of different functions to which algae convert a significant proportion of their photoassimilated organic carbon. Suitable conditions provided, microalgae (phytoplankton) can proliferate very fast. During such blooms, substantial quantities of dissolved and particulate polysaccharide-rich organic matter are released to the environment, both by exudation of living and by decomposition of dead algal cells. While research on the community and functional dynamics of free-living bacteria during phytoplankton blooms is extensive, knowledge about particle-attached bacteria, especially regarding taxonomic composition and associated gene functions, is limited. Like planktonic microalgae, sessile macroalgae are also rich in polysaccharides and harbor bacteria with significant potential for polysaccharide degradation, particularly for complex polysaccharides. However, research on macroalgal phycosphere microbes so far primarily focused on community composition studies via 16S rRNA gene amplicon sequencing and (meta-)genomic analysis of single algal species or a limited number of samples. This thesis provides a comprehensive exploration of microalgae-associated microbial communities during phytoplankton blooms and of macroalgae-associated phycophere communities, both with a focus on polysaccharide utilization functions. Chapter I establishes the theoretical foundation of this thesis. It starts out with a review of the critical roles that micro- and macroalgae play within the marine carbon budgets. It then narrows down to explore phytoplankton blooms, key events in marine carbon cycling marked by rapid carbon fixation, remineralization, and release. The studies of marine particles are summarized. Afterwards, common polysaccharides of micro- and macroalgae are introduced, as well as the mechanisms of bacterial polysaccharide utilization. Then research methodologies in microbial ecology are outlined with a focus on the analysis of omics data, which form the core methodologies in the two subsequent chapters, and finally the chapter closes with a summary of the research questions that this thesis aims to address. In Chapter II, I present a study in which I explored dynamic shifts in community composition and polysaccharide degradation functions of particle-attached bacteria throughout an extensive phytoplankton bloom in contrast to those of free-living bacteria. Bloom progression was captured using microscopic, chlorophyll a and 18S rRNA gene amplicon data at high temporal resolution. I identified abundant bacterial clades using corresponding high-resolution 16S rRNA gene amplicon sequencing across three filter size fractions during the bloom. Through metagenome data analysis using both short- and long-read sequencing, I then identified and explored abundant polysaccharide degrading bacteria. Additionally, metaproteome data were analyzed to correlate bacterial proteins with the breakdown of algal glycans. The combined results indicated that, similar to their free-living counterparts, particle-attached polysaccharide-degrading bacteria targeted soluble and structurally simple polysaccharides such as laminarin. However, they also possessed abundant genes dedicated to the degradation of insoluble and structurally complex polysaccharides, setting them apart in their functional capabilities. As detailed in Chapter III, I had a major part in analyzing the composition and polysaccharide utilization functions of macroalgal phycosphere bacteria on four types of macroalgae across the four seasons. Comparisons were made with microbial communities in surrounding seawater and sediment, revealing fourteen core genera consistently present on all algae. Metagenome analysis focused on polysaccharide degradation and secondary metabolite production, while cultivation techniques yielded pure isolates for draft genome sequencing. A high cultivability of macroalgal phycosphere bacteria enabled sequencing of numerous isolates, offering insights into a manifold of polysaccharide utilization loci and their possible polysaccharide substrates. Chapter IV presents a comparative analysis of particle-attached bacteria originating from microalgae versus those from macroalgal phycospheres, a pioneering endeavor not previously undertaken. This way, I identified dominant bacteria shared between both studied communities, alongside unique clades specific to each community. Furthermore, the advantages of utilizing PacBio metagenome sequencing, drafting genome sequences of cultivable strains, and the application of multiple databases for functional annotation are discussed, including insights to enhance the study of particle-attached bacteria and isolated strains. The thesis then closes with an outlook with a proposal for future projects.