Ulrike Jaekel

Anaerobic degradation of propane and butane by sulfate-reducing bacteria enriched from marine hydrocarbon cold seeps

The short-chain, non-methane hydrocarbons propane and butane can contribute significantly to the carbon and sulfur cycles in marine environments affected by oil or natural gas seepage. In the present study, we enriched and identified novel propane and butane-degrading sulfate reducers from marine oil and gas cold seeps in the Gulf of Mexico and Hydrate Ridge. The enrichment cultures obtained were able to degrade simultaneously propane and butane, but not other gaseous alkanes. They were cold-adapted, showing highest sulfate-reduction rates between 16 and 20 °C. Analysis of 16S rRNA gene libraries, followed by whole-cell hybridizations with sequence-specific oligonucleotide probes showed that each enrichment culture was dominated by a unique phylotype affiliated with the Desulfosarcina-Desulfococcus cluster within the Deltaproteobacteria. These phylotypes formed a distinct phylogenetic cluster of propane and butane degraders, including sequences from environments associated with hydrocarbon seeps. Incubations with 13C-labeled substrates, hybridizations with sequence-specific probes and nanoSIMS analyses showed that cells of the dominant phylotypes were the first to become enriched in 13C, demonstrating that they were directly involved in hydrocarbon degradation. Furthermore, using the nanoSIMS data, carbon assimilation rates were calculated for the dominant cells in each enrichment culture.

Determination of the Diversity of Rhodopirellula Isolates from European Seas by Multilocus Sequence Analysis

ABSTRACT In the biogeography of microorganisms, the habitat size of an attached-living bacterium has never been investigated. We approached this theme with a multilocus sequence analysis (MLSA) study of new strains of Rhodopirellula sp., an attached-living planctomycete. The development of an MLSA for Rhodopirellula baltica enabled the characterization of the genetic diversity at the species level, beyond the resolution of the 16S rRNA gene. The alleles of the nine housekeeping genes acsA, guaA, trpE, purH, glpF, fumC, icd, glyA, and mdh indicated the presence of 13 genetically defined operational taxonomic units (OTUs) in our culture collection. The MLSA-based OTUs coincided with the taxonomic units defined by DNA-DNA hybridization experiments. BOX-PCR supported the MLSA-based differentiation of two closely related OTUs. This study established a taxon-area relationship of cultivable Rhodopirellula species. In European seas, three closely related species covered the Baltic Sea and the eastern North Sea, the North Atlantic region, and the southern North Sea to the Mediterranean. The last had regional genotypes, as revealed by BOX-PCR. This suggests a limited habitat size of attached-living Rhodopirellula species.

Carbon and hydrogen stable isotope fractionation associated with the anaerobic degradation of propane and butane by marine sulfate-reducing bacteria

The anaerobic degradation of propane and butane is typically initiated by activation via addition to fumarate. Here we investigated the mechanism of activation under sulfate-reducing conditions by one pure culture (strain BuS5) and three enrichment cultures employing stable isotope analysis. Stable isotope fractionation was compared for cultures incubated with or without substrate diffusion limitation. Bulk enrichment factors were significantly higher in mixed vs. static incubations. Two dimensional factors, given by the correlation of stable isotope fractionation of both carbon and hydrogen at their reactive positions (Lambda reactive position, Λrp), were compared to analyse the activation mechanisms. A characteristic reactive position isotope fractionation pattern was observed, distinct from aerobic degradation. Λrp values ranged from 10.5 to 11.8 for propane and from 7.8 to 9.4 for butane. Incubations of strain BuS5 with deuterium-labelled n-alkanes indicated that butane was activated solely at the subterminal C atom. In contrast, propane was activated mainly at the subterminal C atom but also significantly at the terminal C atoms. A conservative estimate suggests that about 70% of the propane activation events occurred at the subterminal C atom and about 30% at the terminal C atoms.

Anaerobic degradation of cyclohexane by sulfate-reducing bacteria from hydrocarbon-contaminated marine sediments

The fate of cyclohexane, often used as a model compound for the biodegradation of cyclic alkanes due to its abundance in crude oils, in anoxic marine sediments has been poorly investigated. In the present study, we obtained an enrichment culture of cyclohexane-degrading sulfate-reducing bacteria from hydrocarbon-contaminated intertidal marine sediments. Microscopic analyses showed an apparent dominance by oval cells of 1.5 × 0.8 μm. Analysis of a 16S rRNA gene library, followed by whole-cell hybridization with group- and sequence-specific oligonucleotide probes showed that these cells belonged to a single phylotype, and were accounting for more than 80% of the total cell number. The dominant phylotype, affiliated with the Desulfosarcina-Desulfococcus cluster of the Deltaproteobacteria, is proposed to be responsible for the degradation of cyclohexane. Quantitative growth experiments showed that cyclohexane degradation was coupled with the stoichiometric reduction of sulfate to sulfide. Substrate response tests corroborated with hybridization with a sequence-specific oligonucleotide probe suggested that the dominant phylotype apparently was able to degrade other cyclic and n-alkanes, including the gaseous alkane n-butane. Based on GC-MS analyses of culture extracts cyclohexylsuccinate was identified as a metabolite, indicating an activation of cyclohexane by addition to fumarate. Other metabolites detected were 3-cyclohexylpropionate and cyclohexanecarboxylate providing evidence that the overall degradation pathway of cyclohexane under anoxic conditions is analogous to that of n-alkanes.

A distinct and active bacterial community in cold oxygenated fluids circulating beneath the western flank of the Mid-Atlantic ridge.

The rock-hosted, oceanic crustal aquifer is one of the largest ecosystems on Earth, yet little is known about its indigenous microorganisms. Here we provide the first phylogenetic and functional description of an active microbial community residing in the cold oxic crustal aquifer. Using subseafloor observatories, we recovered crustal fluids and found that the geochemical composition is similar to bottom seawater, as are cell abundances. However, based on relative abundances and functional potential of key bacterial groups, the crustal fluid microbial community is heterogeneous and markedly distinct from seawater. Potential rates of autotrophy and heterotrophy in the crust exceeded those of seawater, especially at elevated temperatures (25 °C) and deeper in the crust. Together, these results reveal an active, distinct, and diverse bacterial community engaged in both heterotrophy and autotrophy in the oxygenated crustal aquifer, providing key insight into the role of microbial communities in the ubiquitous cold dark subseafloor biosphere.

Nitrogen Cycling of Active Bacteria within Oligotrophic Sediment of the Mid-Atlantic Ridge Flank

ABSTRACT Microbial ecology within oligotrophic marine sediment is poorly understood, yet is critical for understanding geochemical cycles. Here, 16S rRNA sequences from RNA and DNA inform the structure of active and total microbial communities in oligotrophic sediment on the western flank of the Mid-Atlantic Ridge. Sequences identified as Bacillariophyta chloroplast were detected within DNA, but undetectable within RNA, suggesting preservation in 5.6-million-year-old sediment. Statistical analysis revealed that RNA-based microbial populations correlated significantly with nitrogen concentrations, whereas DNA-based populations did not correspond to measured geochemical analytes. Bioenergetic calculations determined which metabolisms could yield energy in situ, and found that denitrification, nitrification, and nitrogen fixation were all favorable. A metagenome was produced from one sample, and included genes mediating nitrogen redox processes. Nitrogen respiration by active bacteria is an important metabolic strategy in North Pond sediments, and could be widespread in the oligotrophic sedimentary biosphere.

Microbial decomposition of marine dissolved organic matter in cool oceanic crust

Marine dissolved organic carbon (DOC) is one of the largest active reservoirs of reduced carbon on Earth. In the deep ocean, DOC has been described as biologically recalcitrant and has a radiocarbon age of 4,000 to 6,000 years, which far exceeds the timescale of ocean overturning. However, abiotic removal mechanisms cannot account for the full magnitude of deep-ocean DOC loss. Deep-ocean water circulates at low temperatures through volcanic crust on ridge flanks, but little is known about the associated biogeochemical processes and carbon cycling. Here we present analyses of DOC in fluids from two borehole observatories installed in crustal rocks west of the Mid-Atlantic Ridge, and show that deep-ocean DOC is removed from these cool circulating fluids. The removal mechanism is isotopically selective and causes a shift in specific features of molecular composition, consistent with microbe-mediated oxidation. We suggest organic molecules with an average radiocarbon age of 3,200 years are bioavailable to crustal microbes, and that this removal mechanism may account for at least 5% of the global loss of DOC in the deep ocean. Cool crustal circulation probably contributes to maintaining the deep ocean as a reservoir of ‘aged’ and refractory DOC by discharging the surviving organic carbon constituents that are molecularly degraded and depleted in 14C and 13C into the deep ocean.Microbe-mediated oxidation may account for at least 5% of the global dissolved organic carbon loss from the deep ocean, according to carbon isotope analyses on cool crustal fluids circulating through the Mid-Atlantic Ridge.

The Green Berry Consortia of the Sippewissett Salt Marsh: Millimeter-Sized Aggregates of Diazotrophic Unicellular Cyanobacteria

Microbial interactions driving key biogeochemical fluxes often occur within multispecies consortia that form spatially heterogeneous microenvironments. Here, we describe the “green berry” consortia of the Sippewissett salt marsh (Falmouth, MA, United States): millimeter-sized aggregates dominated by an uncultured, diazotrophic unicellular cyanobacterium of the order Chroococcales (termed GB-CYN1). We show that GB-CYN1 is closely related to Crocosphaera watsonii (UCYN-B) and “Candidatus Atelocyanobacterium thalassa” (UCYN-A), two groups of unicellular diazotrophic cyanobacteria that play an important role in marine primary production. Other green berry consortium members include pennate diatoms and putative heterotrophic bacteria from the Alphaproteobacteria and Bacteroidetes. Tight coupling was observed between photosynthetic oxygen production and heterotrophic respiration. When illuminated, the green berries became supersaturated with oxygen. From the metagenome, we observed that GB-CYN1 encodes photosystem II genes and thus has the metabolic potential for oxygen production unlike UCYN-A. In darkness, respiratory activity rapidly depleted oxygen creating anoxia within the aggregates. Metagenomic data revealed a suite of nitrogen fixation genes encoded by GB-CYN1, and nitrogenase activity was confirmed at the whole-aggregate level by acetylene reduction assays. Metagenome reads homologous to marker genes for denitrification were observed and suggest that heterotrophic denitrifiers might co-occur in the green berries, although the physiology and activity of facultative anaerobes in these aggregates remains uncharacterized. Nitrogen fixation in the surface ocean was long thought to be driven by filamentous cyanobacterial aggregates, though recent work has demonstrated the importance of unicellular diazotrophic cyanobacteria (UCYN) from the order Chroococcales. The green berries serve as a useful contrast to studies of open ocean UCYN and may provide a tractable model system to investigate microbial dynamics within phytoplankton aggregates, a phenomenon of global importance to the flux of particulate organic carbon and nitrogen in surface waters.