Identity And Activity of Marine Microbial Populations As Revealed By Single Cell Techniques

In most aquatic habitats the mere quantification of bacterial taxa in situ does not appear to provide sufficient information about their ecological role. Consequently, there is a need for in situ approaches that allow simultaneous microbial identification and an estimate of microbial activity. These approaches should optimally provide a resolution at the level of single populations or even cells as bulk activity measurements seldom correlate with total abundances of bacteria and specific microbial populations may mediate central biogeochemical processes. At the end of the 1990's, a methodological approach was developed to track substrate uptake by specific prokaryotic groups. This was achieved by the combination of microautoradiography and fluorescence in situ hybridization (MARFISH). However, the original MARFISH method had several drawbacks for its application in marine samples. The first aim of this study was to overcome these limitations by introducing three major modifications that rendered the method more sensitive, accurate, and suitable for high-throughput sample processing. In the second half of this work this improved protocol was employed for two studies on the ecology of particular picoplankton populations in the coastal North Sea. In the first application the potential for anaerobic metabolism of pelagic bacteria was investigated. It has been suggested that in coastal environments the potential for anaerobic metabolism might be a common feature of bacterioplankton, but no direct evidence had been provided to support this hypothesis. Incorporation of glucose under anoxic conditions was found in Alphaproteobacteria, Gammaproteobacteria and the Cytophaga-Flavobacteria. Moreover, specific populations of copiotrophic bacteria (Alteromonas, Pseudoalteromonas) showed preferential glucose incorporation under anoxic conditions. In a second application, concentration-dependent uptake of glucose and leucine was assayed before and during a spring phytoplankton bloom. Coastal pelagic environments are characterized by concentration gradients of dissolved organic carbon, and by pronounced seasonal differences in substrate availability for the picoplankton. Microbial taxa that co-exist in such habitats might thus differ in their ability to incorporate substrates at various concentrations. Our results supported this hypothesis. Three patterns were observed for monomer uptake: high numbers of active cells regardless the substrate concentration (Roseobacter), preference for a specific concentration (SAR11 bacteria), and increasing numbers of active cells with increasing substrate concentration (SAR86, DE2 cluster of Bacteroidetes, and Euryarchaeota).