Identifizierung aktiver mikrobieller Populationen mit Hilfe stabiler Isotope

Prokaryoten spielen eine entscheidende Rolle am anaeroben Abbau und der Remineralisierung von sedimentiertem organischem Material in den anoxischen Sedimentschichten des Profundals von Süßwasserseen. Am Beispiel des Profundalsediments des See Genezareth wurde hier erstmalig durch die Kombination bio...

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Bibliographic Details
Main Author: Schwarz, Julia Ingeborg Klara
Contributors: Conrad, Ralf (Prof. Dr.) (Thesis advisor)
Format: Dissertation
Language:German
Published: Philipps-Universität Marburg 2006
Biologie
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Table of Contents: The microbial community of freshwater sediments plays a crucial role in the degradation and transformation of organic matter. In this study, microorganisms involved in the anaerobic degradation of algal biomass and acetate in the profundal sediment of Lake Kinneret were identified for the first time by applying a combination of biogeochemical and molecular methods as well as by using 13C-labelled substrates. First, the microbial community structure of three independent sediment samples was analysed by culture-independent molecular methods. Species composition and relative abundance of the overall microbial community turned out to be very stable. The numbers of Archaea and Bacteria, quantified by real-time PCR targeting 16S rRNA genes, showed that Bacteria account for the larger fraction of the total prokaryotic community. Species composition of the bacterial and archaeal community was analysed by terminal restriction fragment length polymorphism and comparative sequence analysis. The bacterial community showed a high phylogenetic diversity, comprising a lot of yet uncultivated species. It was dominated by members of the Deltaproteobacteria and the Bacteroidetes phylum. In contrast to Bacteria, Archaea were phylogenetically less divers und were dominated by hydrogenotrophic Methanomicrobiales and acetoclastic Methanosaeta spp.. Until now it was unknown, how microbial communities of profundal lake sediments react to the input of organic material. In Lake Kinneret the highest input of organic material occurs after the annual bloom and sedimentation of the algae Peridinium gatunense. This sedimentation event was simulated by addition of P. gatunense cells (unlabelled and 13C-labelled) to anaerobic sediment slurries. Acetate, Propionate and H2 were formed as key intermediates during the anaerobic degradation of the algal biomass. Immediate accumulation of acetate, propionate, H2, CO2 and CH4 with the onset of the incubation indicated a fast reaction of the microbial community to the input of algal biomass. This response correlated with temporal shifts in the composition of ribosomal RNA (16S rRNA) of active bacterial and archaeal populations. After one day of incubation active Bacteria were dominated by Deltaproteobacteria. Additionally, members of Clostridia Cluster I were detected. This Cluster I contains saccharolytic and proteolytic as well as fermentative bacteria. However, after six days of incubation active bacterial populations were characterized by the occurrence of members of the Bacteriodetes phylum, which are known as degraders of biopolymers. This was continued by analysis of sediment slurries incubated after addition of 13C-labelled algal biomass by 16S rRNA-based stable-isotope probing (RNA-SIP) that indicated an enhanced activation of Bacteriodetes-related bacteria at that time. Furthermore, addition of algal biomass resulted in a slight 13C-enrichment of archaeal 16S rRNA. More specifically, a decrease in the concentration of acetate, an increased methane production rate and an increase in the relative abundance of Methanosaeta spp. suggested an activation of acetoclastic methanogens. To get insights into the conversion of acetate as the most important intermediate in anaerobic degradation of the algal biomass the respiratory index (RI) for the conversion of [2-14C]acetate was determined. RI values showed that more than 80% of the acetate added was converted to methane by acetoclastic cleavage but only a minor fraction was oxidized to CO2. To identify microorganisms involved in these processes sediment slurries were incubated with [U-13C]acetate (RNA-SIP). Subsequent analysis of the 13C-labelled RNA showed a strong activation of archaeal populations and that Methanosaeta spp. was responsible for the degradation of acetate by acetoclastic cleavage. Although only a minor part of the Bacteria incorporated the 13C-label into their RNA, RNA-SIP could identify those bacteria. Most of the active Bacteria were affiliated with members of the Thermodesulfovibrio-„Magnetobacterium“-group of the Nitrospira phylum and also with Nitrosomonadales, Burkholderiales and Rhodocyclales within the Betaproteobacteria. In summary, our study provides a comprehensive insight into the function of microbial communities in this habitat. Besides the characterization of the total microbial community of a profundal lake sediment, we identified Bacteria and Archaea actively involved in the anaerobic degradation of algal biomass and acetate.