Ubiquitous Presence and Novel Diversity of Anaerobic Alkane Degraders in Cold Marine Sediments

Alkanes are major constituents of crude oil and are released to the marine environment by natural seepage and from anthropogenic sources. Due to their chemical inertness, their removal from anoxic marine sediments is primarily controlled by the activity of anaerobic alkane-degrading microorganisms. To facilitate comprehensive cultivation-independent surveys of the diversity and distribution of anaerobic alkane degraders, we designed novel PCR primers that cover all known diversity of the 1-methylalkyl succinate synthase gene (masD/assA), which catalyzes the initial activation of alkanes. We studied masD/assA gene diversity in pristine and seepage-impacted Danish coastal sediments, as well as in sediments and alkane-degrading enrichment cultures from the Middle Valley (MV) hydrothermal vent system in the Pacific Northwest. MasD/assA genes were ubiquitously present, and the primers captured the diversity of both known and previously undiscovered masD/assA gene diversity. Seepage sediments were dominated by a single masD/assA gene cluster, which is presumably indicative of a substrate-adapted community, while pristine sediments harbored a diverse range of masD/assA phylotypes including those present in seepage sediments. This rare biosphere of anaerobic alkane degraders will likely increase in abundance in the event of seepage or accidental oil spillage. Nanomolar concentrations of short-chain alkanes (SCA) were detected in pristine and seepage sediments. Interestingly, anaerobic alkane degraders closely related to strain BuS5, the only SCA degrader in pure culture, were found in mesophilic MV enrichments, but not in cold sediments from Danish waters. We propose that the new masD/assA gene lineages in these sediments represent novel phylotypes that are either fueled by naturally occurring low levels of SCA or that metabolize medium- to long-chain alkanes. Our study highlights that masD/assA genes are a relevant diagnostic marker to identify seepage and microseepage, e.g., during prospecting for oil and gas, and may act as an indicator of anthropogenic oil spills in marine sediments

Lignin Preservation and Microbial Carbohydrate Metabolism in Permafrost Soils

Permafrost-affected soils in the northern circumpolar region store more than 1,000 Pg soil organic carbon (OC), and are strongly vulnerable to climatic warming. However, the extent to which changing soil environmental conditions with permafrost thaw affects different compounds of soil organic matter (OM) is poorly understood. Here, we assessed the fate of lignin and non-cellulosic carbohydrates in density fractionated soils (light fraction, LF vs. heavy fraction, HF) from three permafrost regions with decreasing continentality, expanding from east to west of northern Siberia (Cherskiy, Logata, Tazovskiy, respectively). In soils at the Tazovskiy site with thicker active layers, the LF showed smaller OC-normalized contents of lignin-derived phenols and plant-derived sugars and a decrease of these compounds with soil depth, while a constant or even increasing trend was observed in soils with shallower active layers (Cherskiy and Logata). Also in the HF, soils at the Tazovskiy site had smaller contents of OC-normalized lignin-derived phenols and plant-derived sugars along with more pronounced indicators of oxidative lignin decomposition and production of microbial-derived sugars. Active layer deepening, thus, likely favors the decomposition of lignin and plant-derived sugars, that is, lignocelluloses, by increasing water drainage and aeration. Our study suggests that climate-induced degradation of permafrost soils may promote carbon losses from lignin and associated polysaccharides by abolishing context-specific preservation mechanisms. However, relations of OC-based lignin-derived phenols and sugars in the HF with mineralogical properties suggest that future OM transformation and carbon losses will be modulated in addition by reactive soil minerals.publishedVersio

Storage and transformation of organic matter fractions in cryoturbated permafrost soils across the Siberian Arctic

In permafrost soils, the temperature regime and the resulting cryogenic processes are important determinants of the storage of organic carbon (OC) and its small-scale spatial variability. For cryoturbated soils, there is a lack of research assessing pedon-scale heterogeneity in OC stocks and the transformation of functionally different organic matter (OM) fractions, such as particulate and mineral-associated OM. Therefore, pedons of 28 Turbels were sampled in 5 m wide soil trenches across the Siberian Arctic to calculate OC and total nitrogen (TN) stocks based on digital profile mapping. Density fractionation of soil samples was performed to distinguish between particulate OM (light fraction, LF, 1.6 g cm−3), and a mobilizable dissolved pool (mobilizable fraction, MoF). Across all investigated soil profiles, the total OC storage was 20.2 ± 8.0 kg m−2 (mean ± SD) to 100 cm soil depth. Fifty-four percent of this OC was located in the horizons of the active layer (annual summer thawing layer), showing evidence of cryoturbation, and another 35 % was present in the upper permafrost. The HF-OC dominated the overall OC stocks (55 %), followed by LF-OC (19 % in mineral and 13 % in organic horizons). During fractionation, approximately 13 % of the OC was released as MoF, which likely represents a readily bioavailable OM pool. Cryogenic activity in combination with cold and wet conditions was the principle mechanism through which large OC stocks were sequestered in the subsoil (16.4 ± 8.1 kg m−2; all mineral B, C, and permafrost horizons). Approximately 22 % of the subsoil OC stock can be attributed to LF material subducted by cryoturbation, whereas migration of soluble OM along freezing gradients appeared to be the principle source of the dominant HF (63 %) in the subsoil. Despite the unfavourable abiotic conditions, low C / N ratios and high δ13C values indicated substantial microbial OM transformation in the subsoil, but this was not reflected in altered LF and HF pool sizes. Partial least-squares regression analyses suggest that OC accumulates in the HF fraction due to co-precipitation with multivalent cations (Al, Fe) and association with poorly crystalline iron oxides and clay minerals. Our data show that, across all permafrost pedons, the mineral-associated OM represents the dominant OM fraction, suggesting that the HF-OC is the OM pool in permafrost soils on which changing soil conditions will have the largest impact.Russian Ministry of Education and Science/14.B25.31.0031German Federal Ministry of Education and Research/03F0616AEvangelisches Studienwerk VilligstDF

Input of easily available organic C and N stimulates microbial decomposition of soil organic matter in arctic permafrost soil

Rising temperatures in the Arctic can affect soil organic matter (SOM) decomposition directly and indirectly, by increasing plant primary production and thus the allocation of plant-derived organic compounds into the soil. Such compounds, for example root exudates or decaying fine roots, are easily available for microorganisms, and can alter the decomposition of older SOM ("priming effect"). We here report on a SOM priming experiment in the active layer of a permafrost soil from the central Siberian Arctic, comparing responses of organic topsoil, mineral subsoil, and cryoturbated subsoil material (i.e., poorly decomposed topsoil material subducted into the subsoil by freeze-thaw processes) to additions of 13C-labeled glucose, cellulose, a mixture of amino acids, and protein (added at levels corresponding to approximately 1% of soil organic carbon). SOM decomposition in the topsoil was barely affected by higher availability of organic compounds, whereas SOM decomposition in both subsoil horizons responded strongly. In the mineral subsoil, SOM decomposition increased by a factor of two to three after any substrate addition (glucose, cellulose, amino acids, protein), suggesting that the microbial decomposer community was limited in energy to break down more complex components of SOM. In the cryoturbated horizon, SOM decomposition increased by a factor of two after addition of amino acids or protein, but was not significantly affected by glucose or cellulose, indicating nitrogen rather than energy limitation. Since the stimulation of SOM decomposition in cryoturbated material was not connected to microbial growth or to a change in microbial community composition, the additional nitrogen was likely invested in the production of extracellular enzymes required for SOM decomposition. Our findings provide a first mechanistic understanding of priming in permafrost soils and suggest that an increase in the availability of organic carbon or nitrogen, e.g., by increased plant productivity, can change the decomposition of SOM stored in deeper layers of permafrost soils, with possible repercussions on the global climate.Austrian Science Fund (FWF)/CryoCAR

Effects of soil organic matter properties and microbial community composition on enzyme activities in cryoturbated arctic soils

Enzyme-mediated decomposition of soil organic matter (SOM) is controlled, amongst other factors, by organic matter properties and by the microbial decomposer community present. Since microbial community composition and SOM properties are often interrelated and both change with soil depth, the drivers of enzymatic decomposition are hard to dissect. We investigated soils from three regions in the Siberian Arctic, where carbon rich topsoil material has been incorporated into the subsoil (cryoturbation). We took advantage of this subduction to test if SOM properties shape microbial community composition, and to identify controls of both on enzyme activities. We found that microbial community composition (estimated by phospholipid fatty acid analysis), was similar in cryoturbated material and in surrounding subsoil, although carbon and nitrogen contents were similar in cryoturbated material and topsoils. This suggests that the microbial community in cryoturbated material was not well adapted to SOM properties. We also measured three potential enzyme activities (cellobiohydrolase, leucine-amino-peptidase and phenoloxidase) and used structural equation models (SEMs) to identify direct and indirect drivers of the three enzyme activities. The models included microbial community composition, carbon and nitrogen contents, clay content, water content, and pH. Models for regular horizons, excluding cryoturbated material, showed that all enzyme activities were mainly controlled by carbon or nitrogen. Microbial community composition had no effect. In contrast, models for cryoturbated material showed that enzyme activities were also related to microbial community composition. The additional control of microbial community composition could have restrained enzyme activities and furthermore decomposition in general. The functional decoupling of SOM properties and microbial community composition might thus be one of the reasons for low decomposition rates and the persistence of 400 Gt carbon stored in cryoturbated material

Soil organic matter quality exerts a stronger control than stoichiometry on microbial substrate use efficiency along a latitudinal transect

A substantial portion of soil organic matter (SOM) is of microbial origin. The efficiency with which soil mi-croorganisms can convert their substrate carbon (C) into biomass, compared to how much is lost as respiration, thus co-determines the carbon storage potential of soils. Despite increasing insight into soil microbial C cycling, empirical measurements of microbial C processing across biomes and across soil horizons remain sparse. The theory of ecological stoichiometry predicts that microbial carbon use efficiency (CUE), i.e. growth over uptake of organic C, strongly depends on the relative availability of C and nutrients, particularly N, as microorganisms will either respire excess C or conserve C while mineralising excess nutrients. Microbial CUE is thus expected to increase from high to low latitudes and from topsoil to subsoil as the soil C:N and the stoichiometric imbalance between SOM and the microbial biomass decrease. To test these hypotheses, we collected soil samples from the organic topsoil, mineral topsoil, and mineral subsoil of seven sites along a 1500-km latitudinal transect in Western Siberia. As a proxy for CUE, we measured the microbial substrate use efficiency (SUE) of added sub-strates by incubating soil samples with a mixture of 13 C labelled sugars, amino sugars, amino acids, and organic acids and tracing 13 C into microbial biomass and released CO 2 . In addition to soil and microbial C:N stoichio-metry, we also determined the potential extracellular enzyme activities of cellobiohydrolase (CBH) and phenol oxidase (POX) and used the CBH:POX ratio as an indicator of SOM substrate quality. We found an overall decrease of SUE with latitude, corresponding to a decrease in mean annual temperature, in mineral soil horizons. SUE decreased with decreasing stoichiometric imbalance in the organic and mineral topsoil, while a relationship of SUE with soil C:N was only found in the mineral topsoil. However, contrary to our hypothesis, SUE did not increase with soil depth and mineral subsoils displayed lower average SUE than mineral topsoils. Both within individual horizons and across all horizons SUE was strongly correlated with CBH:POX ratio as well as with climate variables. Since enzyme activities likely reflect the chemical properties of SOM, our results indicate that SOM quality exerts a stronger control on SUE than SOM stoichiometry, particularly in subsoils were SOM has been turned over repeatedly and there is little variation in SOM elemental ratios

A communal catalogue reveals Earth's multiscale microbial diversity

Our growing awareness of the microbial world's importance and diversity contrasts starkly with our limited understanding of its fundamental structure. Despite recent advances in DNA sequencing, a lack of standardized protocols and common analytical frameworks impedes comparisons among studies, hindering the development of global inferences about microbial life on Earth. Here we present a meta-analysis of microbial community samples collected by hundreds of researchers for the Earth Microbiome Project. Coordinated protocols and new analytical methods, particularly the use of exact sequences instead of clustered operational taxonomic units, enable bacterial and archaeal ribosomal RNA gene sequences to be followed across multiple studies and allow us to explore patterns of diversity at an unprecedented scale. The result is both a reference database giving global context to DNA sequence data and a framework for incorporating data from future studies, fostering increasingly complete characterization of Earth's microbial diversity.Peer reviewe

A communal catalogue reveals Earth’s multiscale microbial diversity

Our growing awareness of the microbial world’s importance and diversity contrasts starkly with our limited understanding of its fundamental structure. Despite recent advances in DNA sequencing, a lack of standardized protocols and common analytical frameworks impedes comparisons among studies, hindering the development of global inferences about microbial life on Earth. Here we present a meta-analysis of microbial community samples collected by hundreds of researchers for the Earth Microbiome Project. Coordinated protocols and new analytical methods, particularly the use of exact sequences instead of clustered operational taxonomic units, enable bacterial and archaeal ribosomal RNA gene sequences to be followed across multiple studies and allow us to explore patterns of diversity at an unprecedented scale. The result is both a reference database giving global context to DNA sequence data and a framework for incorporating data from future studies, fostering increasingly complete characterization of Earth’s microbial diversity

Identity and abundance of active sulfate-reducing bacteria in deep tidal flat sediments determined by directed cultivation and CARD-FISH analysis

The identity and abundance of potentially active sulfate-reducing bacteria (SRB) in several metre deep sediments of a tidal sand flat in the German Wadden Sea were assessed by directed cultivation and cultivation-independent CARD-FISH analysis (catalysed reporter deposition fluorescence in situ hybridization). Presumably abundant SRB from different sediment layers between 0.5 and 4 m depth were selectively enriched in up to million-fold diluted cultures supplemented with lactate, acetate or hydrogen. Partial 16S rRNA gene sequences obtained from highest dilution steps showing sulfide formation indicated growth of deltaproteobacterial SRB belonging to the Desulfobulbaceae and the Desulfobacteraceae as well as of members of the Firmicutes. Subsequent isolation resulted in 10 novel phylotypes of both litho- and organotrophic sulfate-reducing Deltaproteobacteria. Molecular pre-screening identified six isolates as members of the Desulfobulbaceae, sharing highest identities with either candidatus ‘Desulfobacterium corrodens’ (95–97%) or Desulfobacterium catecholicum (98%), and four isolates as members of Desulfobacteraceae, being related to either Desulfobacter psychrotolerans (98%) or Desulfobacula phenolica (95–97%). Relatives of D. phenolica were exlusively isolated from 50 and 100 cm deep sediments with 10 and 2 mM of pore water sulfate respectively. In contrast, relatives of D. corrodens, D. psychrotolerans and D. catecholicum were also obtained from layers deeper than 100 cm and with less than 2 mM sulfate. The high in situ abundance of members of both families in sediment layers beneath 50 cm could be confirmed via CARD-FISH analysis performed with a set of six SRB-specific oligonucleotide probes. Moreover, SRB represented a numerically significant fraction of the microbial community throughout the sediment (up to 7%) and reached even higher cell numbers in deep, sulfate-poor layers than in the sulfate-rich surface sediment. This relatively large community size of potentially active SRB in deep sandy sediments might on the one hand be a result of their syntrophic association with other anaerobes. Our results furthermore support the hypothesis that enhanced advective pore water transport might supply nutrients to microbial communities in deep sandy sediments and point to their so far unrecognized contribution to biogeochemical processes in Wadden Sea sediments

Prokaryotic Community Structure and Sulfate Reducer Activity in Water from High-Temperature Oil Reservoirs with and without Nitrate Treatment▿ †

Sulfate-reducing prokaryotes (SRP) cause severe problems like microbial corrosion and reservoir souring in seawater-injected oil production systems. One strategy to control SRP activity is the addition of nitrate to the injection water. Production waters from two adjacent, hot (80°C) oil reservoirs, one with and one without nitrate treatment, were compared for prokaryotic community structure and activity of SRP. Bacterial and archaeal 16S rRNA gene analyses revealed higher prokaryotic abundance but lower diversity for the nitrate-treated field. The 16S rRNA gene clone libraries from both fields were dominated by sequences affiliated with Firmicutes (Bacteria) and Thermococcales (Archaea). Potential heterotrophic nitrate reducers (Deferribacterales) were exclusively found at the nitrate-treated field, possibly stimulated by nitrate addition. Quantitative PCR of dsrAB genes revealed that archaeal SRP (Archaeoglobus) dominated the SRP communities, but with lower relative abundance at the nitrate-treated site. Bacterial SRP were found in only low abundance at both sites and were nearly exclusively affiliated with thermophilic genera (Desulfacinum and Desulfotomaculum). Despite the high abundance of archaeal SRP, no archaeal SRP activity was detected in [35S]sulfate incubations at 80°C. Sulfate reduction was found at 60°C in samples from the untreated field and accompanied by the growth of thermophilic bacterial SRP in batch cultures. Samples from the nitrate-treated field generally lacked SRP activity. These results indicate that (i) Archaeoglobus can be a major player in hot oil reservoirs, and (ii) nitrate may act in souring control—not only by inhibiting SRP, but also by changing the overall community structure, including the stimulation of competitive nitrate reducers