Centimeter-long electron transport in marine sediments via conductive minerals

© 2015 International Society for Microbial Ecology All rights reserved. Centimeter-long electron conduction through marine sediments, in which electrons derived from sulfide in anoxic sediments are transported to oxygen in surficial sediments, may have an important influence on sediment geochemistry. Filamentous bacteria have been proposed to mediate the electron transport, but the filament conductivity could not be verified and other mechanisms are possible. Surprisingly, previous investigations have never actually measured the sediment conductivity or its basic physical properties. Here we report direct measurements that demonstrate centimeter-long electron flow through marine sediments, with conductivities sufficient to account for previously estimated electron fluxes. Conductivity was lost for oxidized sediments, which contrasts with the previously described increase in the conductivity of microbial biofilms upon oxidation. Adding pyrite to the sediments significantly enhanced the conductivity. These results suggest that the role of conductive minerals, which are more commonly found in sediments than centimeter-long microbial filaments, need to be considered when modeling marine sediment biogeochemistry

Effect of methanogenic substrates on anaerobic oxidation of methane and sulfate reduction by an anaerobic methanotrophic enrichment

Anaerobic oxidation of methane (AOM) coupled to sulfate reduction (SR) is assumed to be a syntrophic process, in which methanotrophic archaea produce an interspecies electron carrier (IEC), which is subsequently utilized by sulfate-reducing bacteria. In this paper, six methanogenic substrates are tested as candidate-IECs by assessing their effect on AOM and SR by an anaerobic methanotrophic enrichment. The presence of acetate, formate or hydrogen enhanced SR, but did not inhibit AOM, nor did these substrates trigger methanogenesis. Carbon monoxide also enhanced SR but slightly inhibited AOM. Methanol did not enhance SR nor did it inhibit AOM, and methanethiol inhibited both SR and AOM completely. Subsequently, it was calculated at which candidate-IEC concentrations no more Gibbs free energy can be conserved from their production from methane at the applied conditions. These concentrations were at least 1,000 times lower can the final candidate-IEC concentration in the bulk liquid. Therefore, the tested candidate-IECs could not have been produced from methane during the incubations. Hence, acetate, formate, methanol, carbon monoxide, and hydrogen can be excluded as sole IEC in AOM coupled to SR. Methanethiol did inhibit AOM and can therefore not be excluded as IEC by this study

Application of a Colorimetric Assay to Identify Putative Ribofuranosylaminobenzene 5'-Phosphate Synthase Genes Expressed with Activity in Escherichia coli

Tetrahydromethanopterin (H(4)MPT) is a tetrahydrofolate analog originally discovered in methanogenic archaea, but later found in other archaea and bacteria. The extent to which H(4)MPT occurs among living organisms is unknown. The key enzyme which distinguishes the biosynthetic pathways of H(4)MPT and tetrahydrofolate is ribofuranosylaminobenzene 5'-phosphate synthase (RFAP synthase). Given the importance of RFAP synthase in H(4)MPT biosynthesis, the identification of putative RFAP synthase genes and measurement of RFAP synthase activity would provide an indication of the presence of H(4)MPT in untested microorganisms. Investigation of putative archaeal RFAP synthase genes has been hampered by the tendency of the resulting proteins to form inactive inclusion bodies in Escherichia coli. The current work describes a colorimetric assay for measuring RFAP synthase activity, and two modified procedures for expressing recombinant RFAP synthase genes to produce soluble, active enzyme. By lowering the incubation temperature during expression, RFAP synthase from Archaeoglobus fulgidus was produced in E. coli and purified to homogeneity. The production of active RFAP synthase from Methanothermobacter thermautotrophicus was achieved by coexpression of the gene MTH0830 with a molecular chaperone. This is the first direct biochemical identification of a methanogen gene that codes for an active RFAP synthase

Microbial diversity arising from thermodynamic constraints

The microbial world displays an immense taxonomic diversity. This diversity is manifested also in a multitude of metabolic pathways that can utilize different substrates and produce different products. Here, we propose that these observations directly link to thermodynamic constraints that inherently arise from the metabolic basis of microbial growth. We show that thermodynamic constraints can enable coexistence of microbes that utilise the same substrate but produce different end products. We find that this thermodynamics-driven emergence of diversity is most relevant for metabolic conversions with low free energy as seen for example under anaerobic conditions, where population dynamics is governed by thermodynamic effects rather than kinetic factors such as substrate uptake rates. These findings provide a general understanding of the microbial diversity based on the first-principles of thermodynamics. As such they provide a thermodynamics-based framework for explaining the observed microbial diversity in different natural and synthetic environments

Massive Peatland Carbon Banks Vulnerable to Rising Temperatures

Peatlands contain one-third of the world’s soil carbon (C). If destabilized, decomposition of this vast C bank could accelerate climate warming; however, the likelihood of this outcome remains unknown. Here, we examine peatland C stability through five years of whole-ecosystem warming and two years of elevated atmospheric carbon dioxide concentrations (eCO2). Warming exponentially increased methane (CH4) emissions and enhanced CH4 production rates throughout the entire soil profile; although surface CH4 production rates remain much greater than those at depth. Additionally, older deeper C sources played a larger role in decomposition following prolonged warming. Most troubling, decreases in CO2:CH4 ratios in gas production, porewater concentrations, and emissions, indicate that the peatland is becoming more methanogenic with warming. We observed limited evidence of eCO2 effects. Our results suggest that ecosystem responses are largely driven by surface peat, but that the vast C bank at depth in peatlands is responsive to prolonged warming

Genome Sequence of a Mesophilic Hydrogenotrophic Methanogen Methanocella paludicola, the First Cultivated Representative of the Order Methanocellales

We report complete genome sequence of a mesophilic hydrogenotrophic methanogen Methanocella paludicola, the first cultured representative of the order Methanocellales once recognized as an uncultured key archaeal group for methane emission in rice fields. The genome sequence of M. paludicola consists of a single circular chromosome of 2,957,635 bp containing 3004 protein-coding sequences (CDS). Genes for most of the functions known in the methanogenic archaea were identified, e.g. a full complement of hydrogenases and methanogenesis enzymes. The mixotrophic growth of M. paludicola was clarified by the genomic characterization and re-examined by the subsequent growth experiments. Comparative genome analysis with the previously reported genome sequence of RC-IMRE50, which was metagenomically reconstructed, demonstrated that about 70% of M. paludicola CDSs were genetically related with RC-IMRE50 CDSs. These CDSs included the genes involved in hydrogenotrophic methane production, incomplete TCA cycle, assimilatory sulfate reduction and so on. However, the genetic components for the carbon and nitrogen fixation and antioxidant system were different between the two Methanocellales genomes. The difference is likely associated with the physiological variability between M. paludicola and RC-IMRE50, further suggesting the genomic and physiological diversity of the Methanocellales methanogens. Comparative genome analysis among the previously determined methanogen genomes points to the genome-wide relatedness of the Methanocellales methanogens to the orders Methanosarcinales and Methanomicrobiales methanogens in terms of the genetic repertoire. Meanwhile, the unique evolutionary history of the Methanocellales methanogens is also traced in an aspect by the comparative genome analysis among the methanogens

Evaluation and optimization of PCR primers for selective and quantitative detection of marine ANME subclusters involved in sulfate-dependent anaerobic methane oxidation

Since the discovery that anaerobic methanotrophic archaea (ANME) are involved in the anaerobic oxidation of methane coupled to sulfate reduction in marine sediments, different primers and probes specifically targeting the 16S rRNA gene of these archaea have been developed. Microbial investigation of the different ANME subtypes (ANME-1; ANME-2a, b, and c; and ANME-3) was mainly done in sediments where specific subtypes of ANME were highly enriched and methanogenic cell numbers were low. In different sediments with higher archaeal diversity and abundance, it is important that primers and probes targeting different ANME subtypes are very specific and do not detect other ANME subtypes or methanogens that are also present. In this study, primers and probes that were regularly used in AOM studies were tested in silico on coverage and specificity. Most of the previously developed primers and probes were not specific for the ANME subtypes, thereby not reflecting the actual ANME population in complex samples. Selected primers that showed good coverage and high specificity for the subclades ANME-1, ANME-2a/b, and ANME-2c were thoroughly validated using quantitative polymerase chain reaction (qPCR). From these qPCR tests, only certain combinations seemed suitable for selective amplification. After optimization of these primer sets, we obtained valid primer combinations for the selective detection and quantification of ANME-1, ANME-2a/b, and ANME-2c in samples where different ANME subtypes and possibly methanogens could be present. As a result of this work, we propose a standard workflow to facilitate selection of suitable primers for qPCR experiments on novel environmental samples.This research is supported by the Dutch Technology Foundation STW (project 10711), which is part of the Netherlands Organization for Scientific Research (NWO), and which is partly funded by the Ministry of Economic Affairs. Research of AJMS is supported by ERC grant (project 323009). Research of PHATand AJMS is supported by the SIAM Gravitation grant (project 024.002.002) of the Netherlands Ministry of Education, Culture and Science and the Netherlands Science Foundation (NWO).info:eu-repo/semantics/publishedVersio

Role of nickel in high rate methanol degradation in anaerobic granular sludge bioreactors

The effect of nickel deprivation from the influent of a mesophilic (30°C) methanol fed upflow anaerobic sludge bed (UASB) reactor was investigated by coupling the reactor performance to the evolution of the Methanosarcina population of the bioreactor sludge. The reactor was operated at pH 7.0 and an organic loading rate (OLR) of 5–15 g COD l−1 day−1 for 191 days. A clear limitation of the specific methanogenic activity (SMA) on methanol due to the absence of nickel was observed after 129 days of bioreactor operation: the SMA of the sludge in medium with the complete trace metal solution except nickel amounted to 1.164 (±0.167) g CH4-COD g VSS−1 day−1 compared to 2.027 (±0.111) g CH4-COD g VSS−1 day−1 in a medium with the complete (including nickel) trace metal solution. The methanol removal efficiency during these 129 days was 99%, no volatile fatty acid (VFA) accumulation was observed and the size of the Methanosarcina population increased compared to the seed sludge. Continuation of the UASB reactor operation with the nickel limited sludge lead to incomplete methanol removal, and thus methanol accumulation in the reactor effluent from day 142 onwards. This methanol accumulation subsequently induced an increase of the acetogenic activity in the UASB reactor on day 160. On day 165, 77% of the methanol fed to the system was converted to acetate and the Methanosarcina population size had substantially decreased. Inclusion of 0.5 μM Ni (dosed as NiCl2) to the influent from day 165 onwards lead to the recovery of the methanol removal efficiency to 99% without VFA accumulation within 2 days of bioreactor operation

Satellite remote sensing data can be used to model marine microbial metabolite turnover

Sampling ecosystems, even at a local scale, at the temporal and spatial resolution necessary to capture natural variability in microbial communities are prohibitively expensive. We extrapolated marine surface microbial community structure and metabolic potential from 72 16S rRNA amplicon and 8 metagenomic observations using remotely sensed environmental parameters to create a system-scale model of marine microbial metabolism for 5904 grid cells (49 km2) in the Western English Chanel, across 3 years of weekly averages. Thirteen environmental variables predicted the relative abundance of 24 bacterial Orders and 1715 unique enzyme-encoding genes that encode turnover of 2893 metabolites. The genes’ predicted relative abundance was highly correlated (Pearson Correlation 0.72, P-value <10−6) with their observed relative abundance in sequenced metagenomes. Predictions of the relative turnover (synthesis or consumption) of CO2 were significantly correlated with observed surface CO2 fugacity. The spatial and temporal variation in the predicted relative abundances of genes coding for cyanase, carbon monoxide and malate dehydrogenase were investigated along with the predicted inter-annual variation in relative consumption or production of ~3000 metabolites forming six significant temporal clusters. These spatiotemporal distributions could possibly be explained by the co-occurrence of anaerobic and aerobic metabolisms associated with localized plankton blooms or sediment resuspension, which facilitate the presence of anaerobic micro-niches. This predictive model provides a general framework for focusing future sampling and experimental design to relate biogeochemical turnover to microbial ecology

The exchange activities of [Fe] hydrogenase (iron–sulfur-cluster-free hydrogenase) from methanogenic archaea in comparison with the exchange activities of [FeFe] and [NiFe] hydrogenases

[Fe] hydrogenase (iron–sulfur-cluster-free hydrogenase) catalyzes the reversible reduction of methenyltetrahydromethanopterin (methenyl-H4MPT+) with H2 to methylene-H4MPT, a reaction involved in methanogenesis from H2 and CO2 in many methanogenic archaea. The enzyme harbors an iron-containing cofactor, in which a low-spin iron is complexed by a pyridone, two CO and a cysteine sulfur. [Fe] hydrogenase is thus similar to [NiFe] and [FeFe] hydrogenases, in which a low-spin iron carbonyl complex, albeit in a dinuclear metal center, is also involved in H2 activation. Like the [NiFe] and [FeFe] hydrogenases, [Fe] hydrogenase catalyzes an active exchange of H2 with protons of water; however, this activity is dependent on the presence of the hydride-accepting methenyl-H4MPT+. In its absence the exchange activity is only 0.01% of that in its presence. The residual activity has been attributed to the presence of traces of methenyl-H4MPT+ in the enzyme preparations, but it could also reflect a weak binding of H2 to the iron in the absence of methenyl-H4MPT+. To test this we reinvestigated the exchange activity with [Fe] hydrogenase reconstituted from apoprotein heterologously produced in Escherichia coli and highly purified iron-containing cofactor and found that in the absence of added methenyl-H4MPT+ the exchange activity was below the detection limit of the tritium method employed (0.1 nmol min−1 mg−1). The finding reiterates that for H2 activation by [Fe] hydrogenase the presence of the hydride-accepting methenyl-H4MPT+ is essentially required. This differentiates [Fe] hydrogenase from [FeFe] and [NiFe] hydrogenases, which actively catalyze H2/H2O exchange in the absence of exogenous electron acceptors