Flow sorting of marine bacteria for targeted metabolic studies
Half of Earth’s primary production takes place in the marine environment, where CO2 from the atmosphere is transformed by marine phytoplankton into organic biomass. This newly produced biomass is then rapidly consumed by heterotrophic microorganisms such as bacteria and archaea. Heterotrophic microo...
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| Format: | Thesis |
| Language: | English |
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Universität Bremen
2020
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| Online Access: | https://dx.doi.org/10.26092/elib/725 https://media.suub.uni-bremen.de/handle/elib/4928 |
| Summary: | Half of Earth’s primary production takes place in the marine environment, where CO2 from the atmosphere is transformed by marine phytoplankton into organic biomass. This newly produced biomass is then rapidly consumed by heterotrophic microorganisms such as bacteria and archaea. Heterotrophic microorganisms thus contribute to a large part of the turnover of carbon in the ocean, affecting the global carbon cycle. Among the complex communities of marine microorganisms, the contribution of individual organisms to the utilization of organic matter can vary greatly, depending on the type of organic matter and the organism’s specific capabilities for substrate processing. The focus of this work was on the targeted identification of metabolically active bacteria. The first two chapters deal with the bacterial utilization of polysaccharides, a type of substrate that can only be utilized by some members of microbial communities, even though it is one of the main constituents of freshly produced photosynthetic biomass. The first chapter provides methodological guidelines for incubation experiments with fluorescently labeled polysaccharides (FLA-PS) that can be used to simultaneously investigate extracellular hydrolysis and selfish uptake – two distinct modes of bacterial polysaccharide utilization. Using FLA-PS incubation experiments, we could show in chapter 2 the balance of both polysaccharide utilization mechanisms at Helgoland over the course of a year, with increasing and more rapid selfish uptake for laminarin and xylan towards summer and a shift to high rates of extracellular hydrolysis for xylan and chondroitin sulfate in autumn. Flow cytometric sorting of intensely FLA-laminarin stained cells with subsequent taxonomic identification through 16S rRNA sequencing and fluorescence in situ hybridization revealed a contribution of the verrucomicrobial Pedosphaeraceae to fast uptake of laminarin in summer. Bulk community analysis from that time did not hint at the contribution of these bacteria to polysaccharide turnover, implying that bulk analysis alone can lead to a misestimation of polysaccharide turnover. In the third chapter, we used sensitive radiotracer incubation experiments with amino acids at ambient picomolar concentrations to measure the growth and metabolic activity of a deep-ocean bacterioplankton community in the subtropical North Atlantic. Flow cytometry was proven to be a central tool in the combination of different techniques for a targeted linkage of bacterial identity to metabolic activity for selected bacteria from a complex environmental community. The investigation of polysaccharide and amino acid utilization reveals a new perspective on varying roles played by specific members of complex communities, advances our understanding of heterotrophic organic matter utilization, and provides further insight to their contributions to global biogeochemical cycles. |
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