Molecular ecology of key organisms in sulfur and carbon cycling in marine sediments

Abstract

The World s oceans host a variety of sulfidic habitats. Yet, microorganisms oxidizing reduced inorganic sulfur compounds have mostly been studied at hydrothermal vents, in anoxic basins, conspicuous microbial mats and symbioses but rarely in coastal sediments. In this thesis sulfur-oxidizing prokaryotes (SOP) of a eutrophic intertidal sand flat in the German Wadden Sea were investigated by molecular techniques. The diversity, abundance and activity of SOP were analyzed in particular among the Gammaproteobacteria. Comparative sequence analysis of the 16S rRNA and three genes involved in sulfur oxidation revealed a high diversity of mainly gammaproteobacterial SOP. Most of them were closely related to thiotrophic symbionts, including those of the tubeworm genus Oligobrachia. A group of free-living relatives accounted for up to 4% of all cells (~1.3 × 108 cells ml-1). Consistent with a presumed chemolithoautotrophic utilization of inorganic sulfur compounds, these and numerous other members of the Gammaproteobacteria incorporated 14CO2 as revealed by microautoradiography (MAR). The findings demonstrate that non-filamentous Gammaproteobacteria are important catalysts of sedimentary sulfur oxidation and contribute to CO2-fixation in coastal surface sediments. Similarly, Roseobacter clade bacteria (RCB) accounted for unexpectedly high abundances of up to 10% of all cells in surface sediments (~2.5 × 108 cells ml-1). A RCB-related genome fragment of 35 kb was recovered from a metagenomic fosmid library. It encoded genes of the SOX multienzyme system including the sulfur dehydogenase SoxCD, but also the complete rDSR pathway, a gene arrangement that is unique among SOP. Gene-targeted FISH confirmed the presence of the gene dsrA in sedimentary RCB enriched in anaerobic sulfidic medium. In addition, a novel gene, which encodes a putative dioxygenase, designated as dsrU, was identified in the rDSR pathway. Protocols were developed for application of MAR and nano-scale secondary ion mass spectroscopy (nanoSIMS) to marine sediment samples to follow assimilation of acetate in single cells. Members of the Gammaproteobacteria appeared to assimilate slightly more acetate than RCB, whereas sulfate-reducing bacteria showed no significant incorporation. Particularly the combination of flow cytometry and nanoSIMS proved to be powerful for up-scaling of the analysis of substrate uptake by sediment bacteria enabling an efficient, high-resolution profiling of single cells from complex microbial communities.