Diversität und Ökologie von limnischen Actinobakterien

Abstract

Actinobacteria are highly abundant in freshwater bacterioplankton. However, little is known about their diversity, distribution, physiology and ecological role in the aquatic environment. In this thesis the rRNA approach was applied to elucidate the diversity and distribution of freshwater Actinobacteria (limnic Actinobacteria = LAB). The selected cloning strategy and a specifically designed screening method yielded many nearly complete 16S rRNA sequences originating from diverse freshwater habitats. Thereby, the available sequence dataset and the awareness of diversity of LAB was comprehensively expanded.Comparative Sequence analysis revealed that most sequence motifs along with published sequences from mainly freshwater habitats formed four monophyletic clades. The newly defined clusters acI to acIV were distinct from those from marine habitats and soils. Based on phylogenetic reconstructions specific probes for fluorescence-in situ-hybridization (FISH) were designed to analyze the population structure of LAB. For five probes stringent hybridization conditions were determined by custom-developed testing strategies. The application of these probes demonstrated the abundance and importance of the actinobacterial acI-cluster in freshwater bacterioplankton. A subset of three probes targeting subgroups of the acI-cluster revealed differences in the community composition between different lakes. Clones of the acII-cluster appeared equally abundant in 16S rRNA gene libraries, but the respective organisms were of minor numerical importance in the investigated habitats.In a field study ten high mountain lakes in the Tyrolean Alps (Austria) were sampled. These lakes differed in e.g. their altitude and the water visibility leading to distinct exposures of aquatic organisms to UV-B radiation. The UV-B radiation damages DNA mainly by dimerisation of adjacent thymin bases. The genomic DNA of Actinobacteria contains a high molar G C content and thus a higher number of CC-dimers than TT-dimers. CC-dimers are thermodynamically more stable. Thus, bacteria with genomes rich in G C likely are more resistant to UV-B radiation. Therefore the hypothesis was tested, if there is a link between the relative abundance of LAB and the exposure to UV-B in surface layers of high mountain lakes. A positive correlation (r2=0,73) between the abundances of Actinobacteria and UV-B was shown for high mountain lakes above the timberline, whereas no correlation was found for two lakes at lower altitudes. Previous studies have speculated that LAB are relatively inactive organisms surviving mainly due to their resistance to grazing by protists. To test this hypothesis DNA-synthesizing populations in high mountain lakes were identified. The results indicate that the proportion of active cells of all Actinobacteria was similar to the average proportion of active bacterial cells of total bacteria. A clear preference of LAB for glucose-uptake was found in lake Fuchskuhle as demonstrated by combined micro-autoradiography and FISH (MAR-FISH). In cultivation attempts in the laboratory, growth of representatives of the environmentally important acI cluster was successfully stimulated. Future studies will focus on isolation of these bacteria into pure culture. This thesis gives clear evidence that Actinobacteria of the acI cluster are an abundant and metabolically active group of freshwater bacterioplankton. Hence, they constitute an important autochthonous part of such microbial communities.