Single-Cell Genome and Group-Specific dsrAB Sequencing Implicate Marine Members of the Class Dehalococcoidia (Phylum Chloroflexi) in Sulfur Cycling.

UNLABELLED: The marine subsurface sediment biosphere is widely inhabited by bacteria affiliated with the class Dehalococcoidia (DEH), phylum Chloroflexi, and yet little is known regarding their metabolisms. In this report, genomic content from a single DEH cell (DEH-C11) with a 16S rRNA gene that was affiliated with a diverse cluster of 16S rRNA gene sequences prevalent in marine sediments was obtained from sediments of Aarhus Bay, Denmark. The distinctive gene content of this cell suggests metabolic characteristics that differ from those of known DEH and Chloroflexi The presence of genes encoding dissimilatory sulfite reductase (Dsr) suggests that DEH could respire oxidized sulfur compounds, although Chloroflexi have never been implicated in this mode of sulfur cycling. Using long-range PCR assays targeting DEH dsr loci, dsrAB genes were amplified and sequenced from various marine sediments. Many of the amplified dsrAB sequences were affiliated with the DEH Dsr clade, which we propose equates to a family-level clade. This provides supporting evidence for the potential for sulfite reduction by diverse DEH species. DEH-C11 also harbored genes encoding reductases for arsenate, dimethyl sulfoxide, and halogenated organics. The reductive dehalogenase homolog (RdhA) forms a monophyletic clade along with RdhA sequences from various DEH-derived contigs retrieved from available metagenomes. Multiple facts indicate that this RdhA may not be a terminal reductase. The presence of other genes indicated that nutrients and energy may be derived from the oxidation of substituted homocyclic and heterocyclic aromatic compounds. Together, these results suggest that marine DEH play a previously unrecognized role in sulfur cycling and reveal the potential for expanded catabolic and respiratory functions among subsurface DEH. IMPORTANCE: Sediments underlying our oceans are inhabited by microorganisms in cell numbers similar to those estimated to inhabit the oceans. Microorganisms in sediments consist of various diverse and uncharacterized groups that contribute substantially to global biogeochemical cycles. Since most subsurface microorganisms continue to evade cultivation, possibly due to very slow growth, we obtained and analyzed genomic information from a representative of one of the most widespread and abundant, yet uncharacterized bacterial groups of the marine subsurface. We describe several key features that may contribute to their widespread distribution, such as respiratory flexibility and the potential to use oxidized sulfur compounds, which are abundant in marine environments, as electron acceptors. Together, these data provide important information that can be used to assist in designing enrichment strategies or other postgenomic studies, while also improving our understanding of the diversity and distribution of dsrAB genes, which are widely used functional marker genes for sulfur-cycling microbes.

Ammonia-oxidizing Bacteria of the Nitrosospira cluster 1 dominate over ammonia-oxidizing Archaea in oligotrophic surface sediments near the South Atlantic Gyre.

Sediments across the Namibian continental margin feature a strong microbial activity gradient at their surface. This is reflected in ammonium concentrations of < 10 µM in oligotrophic abyssal plain sediments near the South Atlantic Gyre compared with ammonium concentrations of > 700 µM in upwelling areas near the coast. Here we address changes in apparent abundance and structure of ammonia-oxidizing archaeal and bacterial communities (AOA and AOB) along a transect of seven sediment stations across the Namibian shelf by analysing their respective ammonia monooxygenase genes (amoA). The relative abundance of archaeal and bacterial amoA (g(-1) DNA) decreased with increasing ammonium concentrations, and bacterial amoA frequently outnumbered archaeal amoA at the sediment-water interface [0-1 cm below seafloor (cmbsf)]. In contrast, AOA were apparently as abundant as AOB or dominated in several deeper (> 10 cmbsf), anoxic sediment layers. Phylogenetic analyses showed a change within the AOA community along the transect, from two clusters without cultured representatives at the gyre to Nitrososphaera and Nitrosopumilus clusters in the upwelling region. AOB almost exclusively belonged to the Nitrosospira cluster 1. Our results suggest that this predominantly marine AOB lineage without cultured representatives can thrive at low ammonium concentrations and is active in the marine nitrogen cycle.

Genome sequencing of a single cell of the widely distributed marine subsurface Dehalococcoidia, phylum Chloroflexi.

Bacteria of the class Dehalococcoidia (DEH), phylum Chloroflexi, are widely distributed in the marine subsurface, yet metabolic properties of the many uncultivated lineages are completely unknown. This study therefore analysed genomic content from a single DEH cell designated 'DEH-J10' obtained from the sediments of Aarhus Bay, Denmark. Real-time PCR showed the DEH-J10 phylotype was abundant in upper sediments but was absent below 160 cm below sea floor. A 1.44 Mbp assembly was obtained and was estimated to represent up to 60.8% of the full genome. The predicted genome is much larger than genomes of cultivated DEH and appears to confer metabolic versatility. Numerous genes encoding enzymes of core and auxiliary beta-oxidation pathways were identified, suggesting that this organism is capable of oxidising various fatty acids and/or structurally related substrates. Additional substrate versatility was indicated by genes, which may enable the bacterium to oxidise aromatic compounds. Genes encoding enzymes of the reductive acetyl-CoA pathway were identified, which may also enable the fixation of CO2 or oxidation of organics completely to CO2. Genes encoding a putative dimethylsulphoxide reductase were the only evidence for a respiratory terminal reductase. No evidence for reductive dehalogenase genes was found. Genetic evidence also suggests that the organism could synthesise ATP by converting acetyl-CoA to acetate by substrate-level phosphorylation. Other encoded enzymes putatively conferring marine adaptations such as salt tolerance and organo-sulphate sulfohydrolysis were identified. Together, these analyses provide the first insights into the potential metabolic traits that may enable members of the DEH to occupy an ecological niche in marine sediments.

Influence of oxic/anoxic fluctuations on ammonia oxidizers and nitrification potential in a wet tropical soil.

Ammonia oxidation is a key process in the global nitrogen cycle. However, in tropical soils, little is known about ammonia-oxidizing microorganisms and how characteristically variable oxygen regimes affect their activity. We investigated the influence of brief anaerobic periods on ammonia oxidation along an elevation, moisture, and oxygen availability gradient in wet tropical soils. Soils from three forest types were incubated for up to 36 weeks in lab microcosms under three regimes: (1) static aerobic; (2) static anaerobic; and (3) fluctuating (aerobic/anaerobic). Nitrification potential was measured in field-fresh soils and incubated soils. The native ammonia-oxidizing community was also characterized, based on diversity assessments (clone libraries) and quantification of the ammonia monooxygenase α-subunit (amoA) gene. These relatively low pH soils appear to be dominated by ammonia-oxidizing archaea (AOA), and AOA communities in the three soil types differed significantly in their ability to oxidize ammonia. Soils from an intermediate elevation, and those incubated with fluctuating redox conditions, tended to have the highest nitrification potential following an influx of oxygen, although all soils retained the capacity to nitrify even after long anoxic periods. Together, these results suggest that wet tropical soil AOA are tolerant of extended periods of anoxia.

Predominant archaea in marine sediments degrade detrital proteins.

Half of the microbial cells in the Earth's oceans are found in sediments. Many of these cells are members of the Archaea, single-celled prokaryotes in a domain of life separate from Bacteria and Eukaryota. However, most of these archaea lack cultured representatives, leaving their physiologies and placement on the tree of life uncertain. Here we show that the uncultured miscellaneous crenarchaeotal group (MCG) and marine benthic group-D (MBG-D) are among the most numerous archaea in the marine sub-sea floor. Single-cell genomic sequencing of one cell of MCG and three cells of MBG-D indicated that they form new branches basal to the archaeal phyla Thaumarchaeota and Aigarchaeota, for MCG, and the order Thermoplasmatales, for MBG-D. All four cells encoded extracellular protein-degrading enzymes such as gingipain and clostripain that are known to be effective in environments chemically similar to marine sediments. Furthermore, we found these two types of peptidase to be abundant and active in marine sediments, indicating that uncultured archaea may have a previously undiscovered role in protein remineralization in anoxic marine sediments.

Pyrene toxicity is affected by the nutrient status of a marine sediment community: implications for risk assessment.

Risk assessment of toxicants often disregards that environmental conditions, like changing nutrient status, may affect ecosystem response to a toxicant even within an ecosystem. We investigated if effects of pyrene on shallow-water sediments depended on nutrient status of the sediment during 58 days of incubation. Natural undisturbed sediment cores were pre-exposed to two concentrations of inorganic nutrients (nitrogen and phosphorous) for 14 days. After terminating nutrient additions, pyrene was applied once to half the Nu(high) and half the Nu(low) cores in a concentration of 2 mg/kg DW, normalized to 1% TOC. Pyrene affected the sediment systems in both Nu(high) and Nu(low), but effects of pyrene differed between nutrient regimes. In the Nu(low) system, effects of pyrene were mainly seen on the sediment community structure, such as meiofauna community structure. On the contrary, effects of pyrene in Nu(high) were mainly seen on community functions, such as changes in inorganic nutrient fluxes. Direct and indirect effects of pyrene were observed in both nutrient regimes, but they operated on different timescales depending on the variable in focus. This study shows the need to include environmental factors such as nutrient status in risk assessment of toxicants.

Phototoxicity of pyrene affects benthic algae and bacteria from the Arctic.

Phototoxicity of polycyclic aromatic hydrocarbons (PAHs) in the Arctic is important to study since the future PAH load is likely to increase. In combination with the increased UV-light penetration due to ozone layer thinning, phototoxicity may be a potential problem for arctic areas. The aim of this study was to evaluate effects of pyrene and phototoxicity of pyrene on natural algae and bacteria from arctic sediments. Sediments from a shallow-water marine baywere spiked with different pyrene concentrations. Microcosms containing the sediment were incubated under three light regimes, natural sunlight with UV-light, natural sunlight without UV-light, and dark. Significant effects were evident at low pyrene concentrations, particularly in presence of UV-light, indicating phototoxicity. The microalgae were especially sensitive to the phototoxicity of pyrene. Already atthe lowest pyrene concentration (Cfree: 4 nM) algal 14C-incorporation and chlorophyll a content were reduced. The toxic effects of pyrene on the microalgae probably led to the release of organic matter. In agreement with this, bacterial activity increased at high pyrene concentrations indicated by increased oxygen consumption and increased release of inorganic N and P from the sediment. This study indicates that phototoxicity of PAHs may be relevant for sediment communities from shallow marine arctic areas at environmentally relevant pyrene concentrations.