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Abstract

Both, photosynthetic organisms such as diatoms as well as heterotrophic bacteria, play important roles in the oceanic carbon cycle and the biological pump. A model system for studying diatom-bacteria interactions consisting of the marine Gammaproteobacterium M. adhaerens HP15 and the diatom T. weissflogii had been established in order to better understand their interactions and the role in the mentioned processes. The initial work of this study focused on the growth behavior of both organisms, the optimization of the protein extraction and the proteomics analysis of M. adhaerens HP15 during the interaction with T. weissflogii. To achieve these aims, growth curve experiments and attachment assay experiments with subsequent proteomics analysis via 2D gel electrophoresis and MALDI-ToF –MS were conducted. Growth curve experiments showed that M. adhaerens HP15 is capable to grow both in the complex MB medium as well as in minimal f/2 medium supplemented with glutamate. Microscopic observations during the attachment assay experiment revealed that several bacterial cells attach to a single diatom cell. In the three replicates of this experiment 36.2, 24.3, and 48.3 % of bacterial cells attached to diatoms, respectively. In the proteomics analysis differences in the protein patterns of samples of free-living and attached bacteria could be observed on 2D gels. However, only proteins excised from gels with samples of free- living bacteria could be identified by MALDI-ToF –MS. These included for example proteins involved in chemotaxis and proteins typical for fast growing bacteria. These findings suggest that chemotaxis is an important process during diatom-bacteria interactions and that free- living bacteria might grow faster than attached bacteria. Why proteins from the attached fraction could not be identified with MALDI-ToF –MS so far remained unclear. The future PhD thesis work will first focus the trouble-shooting and technical optimization of the protein extraction and proteomics analysis in order to get highly reliable results and to be able to apply MALDI-ToF –MS on proteins from attached bacteria. Subsequently, the proteomics of M. adhaerens HP15 during the direct and indirect interaction as well as during the aggregation process with T. weissflogii will be analyzed with attachment assays and rolling tank experiments under defined environmental conditions. Both types of experiments will also be conducted in a comparative mutant analysis with already constructed M. adhaerens HP15 mutants. Additionally new gene–specific mutants of M. adhaerens HP15 will be generated and phenotypically characterized in order to identify new genes involved in diatom-bacteria interactions.