Methane production controls in a young thermokarst lake formed by abrupt permafrost thaw.

Methane (CH4 ) release to the atmosphere from thawing permafrost contributes significantly to global CH4 emissions. However, constraining the effects of thaw that control the production and emission of CH4 is needed to anticipate future Arctic emissions. Here are presented robust rate measurements of CH4 production and cycling in a region of rapidly degrading permafrost. Big Trail Lake, located in central Alaska, is a young, actively expanding thermokarst lake. The lake was investigated by taking two 1 m cores of sediment from different regions. Two independent methods of measuring microbial CH4  production, long term (CH4 accumulation) and short term (14 C tracer), produced similar average rates of 11 ± 3.5 and 9 ± 3.6 nmol cm-3  d-1 , respectively. The rates had small variations between the different lithological units, indicating homogeneous CH4 production despite heterogeneous lithology in the surface ~1 m of sediment. To estimate the total CH4 production, the CH4 production rates were multiplied through the 10-15 m deep talik (thaw bulb). This estimate suggests that CH4  production is higher than emission by a maximum factor of ~2, which is less than previous estimates. Stable and radioactive carbon isotope measurements showed that 50% of dissolved CH4 in the first meter was produced further below. Interestingly, labeled 14 C incubations with 2-14 C acetate and 14 C CO2 indicate that variations in the pathway used by microbes to produce CH4 depends on the age and type of organic matter in the sediment, but did not appear to influence the rates at which CH4  was produced. This study demonstrates that at least half of the CH4 produced by microbial breakdown of organic matter in actively expanding thermokarst is emitted to the atmosphere, and that the majority of this CH4 is produced in the deep sediment.

Thiomicrorhabdus streamers and sulfur cycling in perennial hypersaline cold springs in the Canadian high Arctic.

The Gypsum Hill (GH) springs on Axel Heiberg Island in the Canadian high Arctic are host to chemolithoautotrophic, sulfur-oxidizing streamers that flourish in the high Arctic winter in water temperatures from -1.3 to 7°C with ~8% salinity in a high Arctic winter environment with air temperatures commonly less than -40°C and an average annual air temperature of -15°C. Metagenome sequencing and binning of streamer samples produced a 96% complete Thiomicrorhabdus sp. metagenome-assembled genome representing a possible new species or subspecies. This is the most cold- and salt-extreme source environment for a Thiomicrorhabdus genome yet described. Metaproteomic and metatranscriptomic analysis attributed nearly all gene expression in the streamers to the Thiomicrorhabdus sp. and suggested that it is active in CO2 fixation and oxidation of sulfide to elemental sulfur. In situ geochemical and isotopic analyses of the fractionation of multiple sulfur isotopes determined the biogeochemical transformation of sulfur from its source in Carboniferous evaporites to biotic processes occurring in the sediment and streamers. These complementary molecular tools provided a functional link between the geochemical substrates and the collective traits and activity that define the microbial community's interactions within a unique polar saline habitat where Thiomicrorhabdus-dominated streamers form and flourish.

The effect of temperature on sulfur and oxygen isotope fractionation by sulfate reducing bacteria (Desulfococcus multivorans).

Temperature influences microbiological growth and catabolic rates. Between 15 and 35 °C the growth rate and cell specific sulfate reduction rate of the sulfate reducing bacterium Desulfococcus multivorans increased with temperature. Sulfur isotope fractionation during sulfate reduction decreased with increasing temperature from 27.2 ‰ at 15 °C to 18.8 ‰ at 35 °C which is consistent with a decreasing reversibility of the metabolic pathway as the catabolic rate increases. Oxygen isotope fractionation, in contrast, decreased between 15 and 25 °C and then increased again between 25 and 35 °C, suggesting increasing reversibility in the first steps of the sulfate reducing pathway at higher temperatures. This points to a decoupling in the reversibility of sulfate reduction between the steps from the uptake of sulfate into the cell to the formation of sulfite, relative to the whole pathway from sulfate to sulfide. This observation is consistent with observations of increasing sulfur isotope fractionation when sulfate reducing bacteria are living near their upper temperature limit. The oxygen isotope decoupling may be a first signal of changing physiology as the bacteria cope with higher temperatures.

Large sulfur isotope fractionation by bacterial sulfide oxidation.

A sulfide-oxidizing microorganism, Desulfurivibrio alkaliphilus (DA), generates a consistent enrichment of sulfur-34 (34 S) in the produced sulfate of +12.5 per mil or greater. This observation challenges the general consensus that the microbial oxidation of sulfide does not result in large 34 S enrichments and suggests that sedimentary sulfides and sulfates may be influenced by metabolic activity associated with sulfide oxidation. Since the DA-type sulfide oxidation pathway is ubiquitous in sediments, in the modern environment, and throughout Earth history, the enrichments and depletions in 34 S in sediments may be the combined result of three microbial metabolisms: microbial sulfate reduction, the disproportionation of external sulfur intermediates, and microbial sulfide oxidation.

The Biogeochemical Sulfur Cycle of Marine Sediments.

Microbial dissimilatory sulfate reduction to sulfide is a predominant terminal pathway of organic matter mineralization in the anoxic seabed. Chemical or microbial oxidation of the produced sulfide establishes a complex network of pathways in the sulfur cycle, leading to intermediate sulfur species and partly back to sulfate. The intermediates include elemental sulfur, polysulfides, thiosulfate, and sulfite, which are all substrates for further microbial oxidation, reduction or disproportionation. New microbiological discoveries, such as long-distance electron transfer through sulfide oxidizing cable bacteria, add to the complexity. Isotope exchange reactions play an important role for the stable isotope geochemistry and for the experimental study of sulfur transformations using radiotracers. Microbially catalyzed processes are partly reversible whereby the back-reaction affects our interpretation of radiotracer experiments and provides a mechanism for isotope fractionation. We here review the progress and current status in our understanding of the sulfur cycle in the seabed with respect to its microbial ecology, biogeochemistry, and isotope geochemistry.

A Critical Look at the Combined Use of Sulfur and Oxygen Isotopes to Study Microbial Metabolisms in Methane-Rich Environments.

Separating the contributions of anaerobic oxidation of methane and organoclastic sulfate reduction in the overall sedimentary sulfur cycle of marine sediments has benefited from advances in isotope biogeochemistry. Particularly, the coupling of sulfur and oxygen isotopes measured in the residual sulfate pool (δ18OSO4 vs. δ34SSO4). Yet, some important questions remain. Recent works have observed patterns that are inconsistent with previous interpretations. We differentiate the contributions of oxygen and sulfur isotopes to separating the anaerobic oxidation of methane and organoclastic sulfate reduction into three phases; first evidence from conventional high methane vs. low methane sites suggests a clear relationship between oxygen and sulfur isotopes in porewater and the metabolic process taking place. Second, evidence from pure cultures and organic matter rich sites with low levels of methane suggest the signatures of both processes overlap and cannot be differentiated. Third, we take a critical look at the use of oxygen and sulfur isotopes to differentiate metabolic processes (anaerobic oxidation of methane vs. organoclastic sulfate reduction). We identify that it is essential to develop a better understanding of the oxygen kinetic isotope effect, the degree of isotope exchange with sulfur intermediates as well as establishing their relationships with the cell-specific metabolic rates if we are to develop this proxy into a reliable tool to study the sulfur cycle in marine sediments and the geological record.

Sulfur Isotope Fractionation by Sulfate-Reducing Microbes Can Reflect Past Physiology.

Sulfur (S) isotope fractionation by sulfate-reducing microorganisms is a direct manifestation of their respiratory metabolism. This fractionation is apparent in the substrate (sulfate) and waste (sulfide) produced. The sulfate-reducing metabolism responds to variability in the local environment, with the response determined by the underlying genotype, resulting in the expression of an "isotope phenotype". Sulfur isotope phenotypes have been used as a diagnostic tool for the metabolic activity of sulfate-reducing microorganisms in the environment. Our experiments with Desulfovibrio vulgaris Hildenborough (DvH) grown in batch culture suggest that the S isotope phenotype of sulfate respiring microbes may lag environmental changes on time scales that are longer than generational. When inocula from different phases of growth are assayed under the same environmental conditions, we observed that DvH exhibited different net apparent fractionations of up to -9‰. The magnitude of fractionation was weakly correlated with physiological parameters but was strongly correlated to the age of the initial inoculum. The S isotope fractionation observed between sulfate and sulfide showed a positive correlation with respiration rate, contradicting the well-described negative dependence of fractionation on respiration rate. Quantitative modeling of S isotope fractionation shows that either a large increase (≈50×) in the abundance of sulfate adenylyl transferase (Sat) or a smaller increase in sulfate transport proteins (≈2×) is sufficient to account for the change in fractionation associated with past physiology. Temporal transcriptomic studies with DvH imply that expression of sulfate permeases doubles over the transition from early exponential to early stationary phase, lending support to the transport hypothesis proposed here. As it is apparently maintained for multiple generations (≈1-6) of subsequent growth in the assay environment, we suggest that this fractionation effect acts as a sort of isotopic "memory" of a previous physiological and environmental state. Whatever its root cause, this physiological hysteresis effect can explain variations in fractionations observed in many environments. It may also enable new insights into life at energetic limits, especially if its historical footprint extends deeper than generational.

Deconstructing the Dissimilatory Sulfate Reduction Pathway: Isotope Fractionation of a Mutant Unable of Growth on Sulfate.

The sulfur isotope record provides key insight into the history of Earth's redox conditions. A detailed understanding of the metabolisms driving this cycle, and specifically microbial sulfate reduction (MSR), is crucial for accurate paleoenvironmental reconstructions. This includes a precise knowledge of the step-specific sulfur isotope effects during MSR. In this study, we aim at resolving the cellular-level fractionation factor during dissimilatory sulfite reduction to sulfide within MSR, and use this measured isotope effect as a calibration to enhance our understanding of the biochemistry of sulfite reduction. For this, we merge measured isotope effects associated with dissimilatory sulfite reduction with a quantitative model that explicitly links net fractionation, reaction reversibility, and intracellular metabolite levels. The highly targeted experimental aspect of this study was possible by virtue of the availability of a deletion mutant strain of the model sulfate reducer Desulfovibrio vulgaris (strain Hildenborough), in which the sulfite reduction step is isolated from the rest of the metabolic pathway owing to the absence of its QmoABC complex (ΔQmo). This deletion disrupts electron flux and prevents the reduction of adenosine phosphosulfate (APS) to sulfite. When grown in open-system steady-state conditions at 10% maximum growth rate in the presence of sulfite and lactate as electron donor, sulfur isotope fractionation factors averaged -15.9‰ (1 σ = 0.4), which appeared to be statistically indistinguishable from a pure enzyme study with dissimilatory sulfite reductase. We coupled these measurements with an understanding of step-specific equilibrium and kinetic isotope effects, and furthered our mechanistic understanding of the biochemistry of sulfite uptake and ensuing reduction. Our metabolically informed isotope model identifies flavodoxin as the most likely electron carrier performing the transfer of electrons to dissimilatory sulfite reductase. This is in line with previous work on metabolic strategies adopted by sulfate reducers under different energy regimes, and has implications for our understanding of the plasticity of this metabolic pathway at the center of our interpretation of modern and palaeo-environmental records.

The seasonal fluctuations and accumulation of iodine-129 in relation to the hydrogeochemistry of the Wolf Creek Research Basin, a discontinuous permafrost watershed.

The long lived radioisotope (129)I is a uranium fission product, and an environmental contaminant of the nuclear age. Consequently, it can trace anthropogenic releases of (129)I in watersheds, and has been identified as a potential means to distinguish water sources in discharge (Nimz, 1998). The purpose of this work was to identify the sources and mass input of (129)I and trace the transport, partitioning and mass balance of (129)I over time in a remote watershed. We monitored (129)I and other geochemical and isotope tracers (e.g. δ(14)CDIC, δ(13)CDIC, δ(2)H, δ(18)O, etc.) in precipitation and discharge from the Wolf Creek Research Basin (WCRB), a discontinuous permafrost watershed in the Yukon Territory, Canada, and evaluated the use of (129)I as a water end-member tracer. Radiocarbon and geochemical tracers of weathering show that discharge is comprised of (i) groundwater baseflow that has recharged under open system conditions, (ii) spring freshet meltwater that has derived solutes through closed-system interaction with saturated soils, and (iii) active layer drainage. The abundance of (129)I and the (129)I/(127)I ratio correlated with geochemical tracers suggests varying contributions of these three water end-members to discharge. The (129)I concentration was highest at the onset of freshet, reaching 17.4×10(6) atoms/L, and likely reflects the lack of interaction between meltwater and organic matter at that time. This peak in (129)I was followed by a decline over the summer to its lowest value. Mass balance calculations of the (129)I budget show that the input to the watershed via precipitation is nearly one order of magnitude higher than the output suggesting that such arctic watersheds accumulate nearly 90% of the annual input, primarily in soil organic matter. Temporal variations in discharge (129)I concentrations correlated with changes in discharge water sources suggesting that (129)I is a promising hydrologic tracer, particularly when used in concert with other stable and radioisotopes.

Sulfur isotope fractionation during the evolutionary adaptation of a sulfate-reducing bacterium.

Dissimilatory sulfate reduction is a microbial catabolic pathway that preferentially processes less massive sulfur isotopes relative to their heavier counterparts. This sulfur isotope fractionation is recorded in ancient sedimentary rocks and generally is considered to reflect a phenotypic response to environmental variations rather than to evolutionary adaptation. Modern sulfate-reducing microorganisms isolated from similar environments can exhibit a wide range of sulfur isotope fractionations, suggesting that adaptive processes influence the sulfur isotope phenotype. To date, the relationship between evolutionary adaptation and isotopic phenotypes has not been explored. We addressed this by studying the covariation of fitness, sulfur isotope fractionation, and growth characteristics in Desulfovibrio vulgaris Hildenborough in a microbial evolution experiment. After 560 generations, the mean fitness of the evolved lineages relative to the starting isogenic population had increased by ∼ 17%. After 927 generations, the mean fitness relative to the initial ancestral population had increased by ∼ 20%. Growth rate in exponential phase increased during the course of the experiment, suggesting that this was a primary influence behind the fitness increases. Consistent changes were observed within different selection intervals between fractionation and fitness. Fitness changes were associated with changes in exponential growth rate but changes in fractionation were not. Instead, they appeared to be a response to changes in the parameters that govern growth rate: yield and cell-specific sulfate respiration rate. We hypothesize that cell-specific sulfate respiration rate, in particular, provides a bridge that allows physiological controls on fractionation to cross over to the adaptive realm.

Microbial diversity in endostromatolites (cf. Fissure Calcretes) and in the surrounding permafrost landscape, Haughton impact structure region, Devon Island, Canada.

In recent years, endostromatolites, which consist of finely laminated calcite columns that grow orthogonally within millimeter- to centimeter-thick fissures in limestone bedrock outcrops, have been discovered in dolomitic outcrops in the Haughton impact structure region, Devon Island, Canada. The growth mechanism of the endostromatolites is believed to be very slow and possibly intertwined with biotic and abiotic processes. Therefore, to discern how endostromatolites form in this polar desert environment, the composition of the microbial community of endostromatolites was determined by means of molecular phylogenetic analysis and compared to the microbial communities found in the surrounding soils. The microbial community present within endostromatolites can be inferred to be (given the predominant metabolic traits of related organisms) mostly aerobic and chemoheterotrophic, and belongs in large part to the phylum Actinobacteria and the subphylum Alphaproteobacteria. The identification of these bacteria suggests that the conditions within the fissure were mostly oxidizing during the growth of endostromatolite. The DNA sequences also indicate that a number of bacteria that closely resemble Rubrobacter radiotolerans are abundant in the endostromatolites as well as other Actinobacteria and Alphaproteobacteria. Some of these taxa have been associated with calcite precipitation, which suggests that the endostromatolites might in fact be microbially mediated. Bacterial communities from nearby permanently frozen soils were more diverse and harbored all the phyla found in the endostromatolites with additional taxa. This study on the microbial communities preserved in potentially microbially mediated secondary minerals in the Arctic could help in the search for evidence of life-forms near the edge of habitability on other planetary bodies.