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Sample records for co-grown methanosarcina acetivorans

  1. Reductive nitrosylation of Methanosarcina acetivorans protoglobin: A comparative study

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    Ascenzi, Paolo, E-mail: ascenzi@uniroma3.it [Laboratorio Interdipartimentale di Microscopia Elettronica, Università Roma Tre, Via della Vasca Navale 79, I-00146 Roma (Italy); Istituto di Biochimica delle Proteine, CNR, Via Pietro Castellino 111, I-80131 Napoli (Italy); Pesce, Alessandra [Dipartimento di Fisica, Università di Genova, I-16146 Genova (Italy); Nardini, Marco; Bolognesi, Martino [Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133 Milano (Italy); Ciaccio, Chiara; Coletta, Massimo [Dipartimento di Scienze Cliniche e Medicina Traslazionale, Università di Roma Tor Vergata, Via Montpellier 1, I-00133 Roma (Italy); Consorzio Interuniversitario di Ricerca in Chimica dei Metalli nei Sistemi Biologici, Piazza Umberto I 1, I-70121 Bari (Italy); Dewilde, Sylvia [Department of Biomedical Sciences, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp (Belgium)

    2013-01-25

    Highlights: ► Methanosarcina acetivorans is a strictly anaerobic non-motile methane-producing Archaea. ► M. acetivorans protoglobin binds preferentially O{sub 2} rather than CO. ► Reductive nitrosylation of ferric M. acetivorans protoglobin. ► Nitrosylation of ferrious M. acetivorans protoglobin. ► M. acetivorans protoglobin is a scavenger of RNS and ROS. -- Abstract: Methanosarcina acetivorans is a strictly anaerobic non-motile methane-producing Archaea expressing protoglobin (Pgb) which might either facilitate O{sub 2} detoxification or act as a CO sensor/supplier in methanogenesis. Unusually, M. acetivorans Pgb (MaPgb) binds preferentially O{sub 2} rather than CO and displays anticooperativity in ligand binding. Here, kinetics and/or thermodynamics of ferric and ferrous MaPgb (MaPgb(III) and MaPgb(II), respectively) nitrosylation are reported. Data were obtained between pH 7.2 and 9.5, at 22.0 °C. Addition of NO to MaPgb(III) leads to the transient formation of MaPgb(III)–NO in equilibrium with MaPgb(II)–NO{sup +}. In turn, MaPgb(II)–NO{sup +} is converted to MaPgb(II) by OH{sup −}-based catalysis. Then, MaPgb(II) binds NO very rapidly leading to MaPgb(II)–NO. The rate-limiting step for reductive nitrosylation of MaPgb(III) is represented by the OH{sup −}-mediated reduction of MaPgb(II)–NO{sup +} to MaPgb(II). Present results highlight the potential role of MaPgb in scavenging of reactive nitrogen and oxygen species.

  2. The Methanosarcina barkeri genome: comparative analysis withMethanosarcina acetivorans and Methanosarcina mazei reveals extensiverearrangement within methanosarcinal genomes

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    Maeder, Dennis L.; Anderson, Iain; Brettin, Thomas S.; Bruce,David C.; Gilna, Paul; Han, Cliff S.; Lapidus, Alla; Metcalf, William W.; Saunders, Elizabeth; Tapia, Roxanne; Sowers, Kevin R.

    2006-05-19

    We report here a comparative analysis of the genome sequence of Methanosarcina barkeri with those of Methanosarcina acetivorans and Methanosarcina mazei. All three genomes share a conserved double origin of replication and many gene clusters. M. barkeri is distinguished by having an organization that is well conserved with respect to the other Methanosarcinae in the region proximal to the origin of replication with interspecies gene similarities as high as 95%. However it is disordered and marked by increased transposase frequency and decreased gene synteny and gene density in the proximal semi-genome. Of the 3680 open reading frames in M. barkeri, 678 had paralogs with better than 80% similarity to both M. acetivorans and M. mazei while 128 nonhypothetical orfs were unique (non-paralogous) amongst these species including a complete formate dehydrogenase operon, two genes required for N-acetylmuramic acid synthesis, a 14 gene gas vesicle cluster and a bacterial P450-specific ferredoxin reductase cluster not previously observed or characterized in this genus. A cryptic 36 kbp plasmid sequence was detected in M. barkeri that contains an orc1 gene flanked by a presumptive origin of replication consisting of 38 tandem repeats of a 143 nt motif. Three-way comparison of these genomes reveals differing mechanisms for the accrual of changes. Elongation of the large M. acetivorans is the result of multiple gene-scale insertions and duplications uniformly distributed in that genome, while M. barkeri is characterized by localized inversions associated with the loss of gene content. In contrast, the relatively short M. mazei most closely approximates the ancestral organizational state.

  3. Structure of the surface layer of the methanogenic archaean Methanosarcina acetivorans

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    Arbing, Mark A.; Chan, Sum; Shin, Annie; Phan, Tung; Ahn, Christine J.; Rohlin, Lars; Gunsalus, Robert P.

    2012-01-01

    Archaea have a self-assembling proteinaceous surface (S-) layer as the primary and outermost boundary of their cell envelopes. The S-layer maintains structural rigidity, protects the organism from adverse environmental elements, and yet provides access to all essential nutrients. We have determined the crystal structure of one of the two “homologous” tandem polypeptide repeats that comprise the Methanosarcina acetivorans S-layer protein and propose a high-resolution model for a microbial S-la...

  4. Air-adapted Methanosarcina acetivorans shows high methane production and develops resistance against oxygen stress.

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    Ricardo Jasso-Chávez

    Full Text Available Methanosarcina acetivorans, considered a strict anaerobic archaeon, was cultured in the presence of 0.4-1% O2 (atmospheric for at least 6 months to generate air-adapted cells; further, the biochemical mechanisms developed to deal with O2 were characterized. Methane production and protein content, as indicators of cell growth, did not change in air-adapted cells respect to cells cultured under anoxia (control cells. In contrast, growth and methane production significantly decreased in control cells exposed for the first time to O2. Production of reactive oxygen species was 50 times lower in air-adapted cells versus control cells, suggesting enhanced anti-oxidant mechanisms that attenuated the O2 toxicity. In this regard, (i the transcripts and activities of superoxide dismutase, catalase and peroxidase significantly increased; and (ii the thiol-molecules (cysteine + coenzyme M-SH + sulfide and polyphosphate contents were respectively 2 and 5 times higher in air-adapted cells versus anaerobic-control cells. Long-term cultures (18 days of air-adapted cells exposed to 2% O2 exhibited the ability to form biofilms. These data indicate that M. acetivorans develops multiple mechanisms to contend with O2 and the associated oxidative stress, as also suggested by genome analyses for some methanogens.

  5. A Heme-based Redox Sensor in the Methanogenic Archaeon Methanosarcina acetivorans*

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    Molitor, Bastian; Stassen, Marc; Modi, Anuja; El-Mashtoly, Samir F.; Laurich, Christoph; Lubitz, Wolfgang; Dawson, John H.; Rother, Michael; Frankenberg-Dinkel, Nicole

    2013-01-01

    Based on a bioinformatics study, the protein MA4561 from the methanogenic archaeon Methanosarcina acetivorans was originally predicted to be a multidomain phytochrome-like photosensory kinase possibly binding open-chain tetrapyrroles. Although we were able to show that recombinantly produced and purified protein does not bind any known phytochrome chromophores, UV-visible spectroscopy revealed the presence of a heme tetrapyrrole cofactor. In contrast to many other known cytoplasmic heme-containing proteins, the heme was covalently attached via one vinyl side chain to cysteine 656 in the second GAF domain. This GAF domain by itself is sufficient for covalent attachment. Resonance Raman and magnetic circular dichroism data support a model of a six-coordinate heme species with additional features of a five-coordination structure. The heme cofactor is redox-active and able to coordinate various ligands like imidazole, dimethyl sulfide, and carbon monoxide depending on the redox state. Interestingly, the redox state of the heme cofactor has a substantial influence on autophosphorylation activity. Although reduced protein does not autophosphorylate, oxidized protein gives a strong autophosphorylation signal independent from bound external ligands. Based on its genomic localization, MA4561 is most likely a sensor kinase of a two-component system effecting regulation of the Mts system, a set of three homologous corrinoid/methyltransferase fusion protein isoforms involved in methyl sulfide metabolism. Consistent with this prediction, an M. acetivorans mutant devoid of MA4561 constitutively synthesized MtsF. On the basis of our results, we postulate a heme-based redox/dimethyl sulfide sensory function of MA4561 and propose to designate it MsmS (methyl sulfide methyltransferase-associated sensor). PMID:23661702

  6. Structure of the surface layer of the methanogenic archaean Methanosarcina acetivorans.

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    Arbing, Mark A; Chan, Sum; Shin, Annie; Phan, Tung; Ahn, Christine J; Rohlin, Lars; Gunsalus, Robert P

    2012-07-17

    Archaea have a self-assembling proteinaceous surface (S-) layer as the primary and outermost boundary of their cell envelopes. The S-layer maintains structural rigidity, protects the organism from adverse environmental elements, and yet provides access to all essential nutrients. We have determined the crystal structure of one of the two "homologous" tandem polypeptide repeats that comprise the Methanosarcina acetivorans S-layer protein and propose a high-resolution model for a microbial S-layer. The molecular features of our hexameric S-layer model recapitulate those visualized by medium resolution electron microscopy studies of microbial S-layers and greatly expand our molecular view of S-layer dimensions, porosity, and symmetry. The S-layer model reveals a negatively charged molecular sieve that presents both a charge and size barrier to restrict access to the cell periplasmic-like space. The β-sandwich folds of the S-layer protein are structurally homologous to eukaryotic virus envelope proteins, suggesting that Archaea and viruses have arrived at a common solution for protective envelope structures. These results provide insight into the evolutionary origins of primitive cell envelope structures, of which the S-layer is considered to be among the most primitive: it also provides a platform for the development of self-assembling nanomaterials with diverse functional and structural properties.

  7. Structure of the surface layer of the methanogenic archaean Methanosarcina acetivorans

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    Arbing, Mark A.; Chan, Sum; Shin, Annie; Phan, Tung; Ahn, Christine J.; Rohlin, Lars; Gunsalus, Robert P. (UCLA)

    2012-09-05

    Archaea have a self-assembling proteinaceous surface (S-) layer as the primary and outermost boundary of their cell envelopes. The S-layer maintains structural rigidity, protects the organism from adverse environmental elements, and yet provides access to all essential nutrients. We have determined the crystal structure of one of the two 'homologous' tandem polypeptide repeats that comprise the Methanosarcina acetivorans S-layer protein and propose a high-resolution model for a microbial S-layer. The molecular features of our hexameric S-layer model recapitulate those visualized by medium resolution electron microscopy studies of microbial S-layers and greatly expand our molecular view of S-layer dimensions, porosity, and symmetry. The S-layer model reveals a negatively charged molecular sieve that presents both a charge and size barrier to restrict access to the cell periplasmic-like space. The {beta}-sandwich folds of the S-layer protein are structurally homologous to eukaryotic virus envelope proteins, suggesting that Archaea and viruses have arrived at a common solution for protective envelope structures. These results provide insight into the evolutionary origins of primitive cell envelope structures, of which the S-layer is considered to be among the most primitive: it also provides a platform for the development of self-assembling nanomaterials with diverse functional and structural properties.

  8. Structural Bases for the Regulation of CO Binding in the Archaeal Protoglobin from Methanosarcina acetivorans.

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    Lesley Tilleman

    Full Text Available Studies of CO ligand binding revealed that two protein states with different ligand affinities exist in the protoglobin from Methanosarcina acetivorans (in MaPgb*, residue Cys(E20101 was mutated to Ser. The switch between the two states occurs upon the ligation of MaPgb*. In this work, site-directed mutagenesis was used to explore the role of selected amino acids in ligand sensing and stabilization and in affecting the equilibrium between the "more reactive" and "less reactive" conformational states of MaPgb*. A combination of experimental data obtained from electronic and resonance Raman absorption spectra, CO ligand-binding kinetics, and X-ray crystallography was employed. Three amino acids were assigned a critical role: Trp(60B9, Tyr(61B10, and Phe(93E11. Trp(60B9 and Tyr(61B10 are involved in ligand stabilization in the distal heme pocket; the strength of their interaction was reflected by the spectra of the CO-ligated MaPgb* and by the CO dissociation rate constants. In contrast, Phe(93E11 is a key player in sensing the heme-bound ligand and promotes the rotation of the Trp(60B9 side chain, thus favoring ligand stabilization. Although the structural bases of the fast CO binding rate constant of MaPgb* are still unclear, Trp(60B9, Tyr(61B10, and Phe(93E11 play a role in regulating heme/ligand affinity.

  9. Activation of methanogenesis by cadmium in the marine archaeon Methanosarcina acetivorans.

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    Elizabeth Lira-Silva

    Full Text Available Methanosarcina acetivorans was cultured in the presence of CdCl(2 to determine the metal effect on cell growth and biogas production. With methanol as substrate, cell growth and methane synthesis were not altered by cadmium, whereas with acetate, cadmium slightly increased both, growth and methane rate synthesis. In cultures metabolically active, incubations for short-term (minutes with 10 µM total cadmium increased the methanogenesis rate by 6 and 9 folds in methanol- and acetate-grown cells, respectively. Cobalt and zinc but not copper or iron also activated the methane production rate. Methanogenic carbonic anhydrase and acetate kinase were directly activated by cadmium. Indeed, cells cultured in 100 µM total cadmium removed 41-69% of the heavy metal from the culture and accumulated 231-539 nmol Cd/mg cell protein. This is the first report showing that (i Cd(2+ has an activating effect on methanogenesis, a biotechnological relevant process in the bio-fuels field; and (ii a methanogenic archaea is able to remove a heavy metal from aquatic environments.

  10. Apo and ligand-bound structures of ModA from the archaeon Methanosarcina acetivorans.

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    Chan, Sum; Giuroiu, Iulia; Chernishof, Irina; Sawaya, Michael R; Chiang, Janet; Gunsalus, Robert P; Arbing, Mark A; Perry, L Jeanne

    2010-03-01

    The trace-element oxyanion molybdate, which is required for the growth of many bacterial and archaeal species, is transported into the cell by an ATP-binding cassette (ABC) transporter superfamily uptake system called ModABC. ModABC consists of the ModA periplasmic solute-binding protein, the integral membrane-transport protein ModB and the ATP-binding and hydrolysis cassette protein ModC. In this study, X-ray crystal structures of ModA from the archaeon Methanosarcina acetivorans (MaModA) have been determined in the apoprotein conformation at 1.95 and 1.69 A resolution and in the molybdate-bound conformation at 2.25 and 2.45 A resolution. The overall domain structure of MaModA is similar to other ModA proteins in that it has a bilobal structure in which two mixed alpha/beta domains are linked by a hinge region. The apo MaModA is the first unliganded archaeal ModA structure to be determined: it exhibits a deep cleft between the two domains and confirms that upon binding ligand one domain is rotated towards the other by a hinge-bending motion, which is consistent with the 'Venus flytrap' model seen for bacterial-type periplasmic binding proteins. In contrast to the bacterial ModA structures, which have tetrahedral coordination of their metal substrates, molybdate-bound MaModA employs octahedral coordination of its substrate like other archaeal ModA proteins.

  11. Identification of the major expressed S-layer and cell surface-layer-related proteins in the model methanogenic archaea: Methanosarcina barkeri Fusaro and Methanosarcina acetivorans C2A.

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    Rohlin, Lars; Leon, Deborah R; Kim, Unmi; Loo, Joseph A; Ogorzalek Loo, Rachel R; Gunsalus, Robert P

    2012-01-01

    Many archaeal cell envelopes contain a protein coat or sheath composed of one or more surface exposed proteins. These surface layer (S-layer) proteins contribute structural integrity and protect the lipid membrane from environmental challenges. To explore the species diversity of these layers in the Methanosarcinaceae, the major S-layer protein in Methanosarcina barkeri strain Fusaro was identified using proteomics. The Mbar_A1758 gene product was present in multiple forms with apparent sizes of 130, 120, and 100 kDa, consistent with post-translational modifications including signal peptide excision and protein glycosylation. A protein with features related to the surface layer proteins found in Methanosarcina acetivorans C2A and Methanosarcina mazei Goel was identified in the M. barkeri genome. These data reveal a distinct conserved protein signature with features and implied cell surface architecture in the Methanosarcinaceae that is absent in other archaea. Paralogous gene expression patterns in two Methanosarcina species revealed abundant expression of a single S-layer paralog in each strain. Respective promoter elements were identified and shown to be conserved in mRNA coding and upstream untranslated regions. Prior M. acetivorans genome annotations assigned S-layer or surface layer associated roles of eighty genes: however, of 68 examined none was significantly expressed relative to the experimentally determined S-layer gene.

  12. Carbon-dependent control of electron transfer and central carbon pathway genes for methane biosynthesis in the Archaean, Methanosarcina acetivorans strain C2A

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    Gunsalus Robert P

    2010-02-01

    Full Text Available Abstract Background The archaeon, Methanosarcina acetivorans strain C2A forms methane, a potent greenhouse gas, from a variety of one-carbon substrates and acetate. Whereas the biochemical pathways leading to methane formation are well understood, little is known about the expression of the many of the genes that encode proteins needed for carbon flow, electron transfer and/or energy conservation. Quantitative transcript analysis was performed on twenty gene clusters encompassing over one hundred genes in M. acetivorans that encode enzymes/proteins with known or potential roles in substrate conversion to methane. Results The expression of many seemingly "redundant" genes/gene clusters establish substrate dependent control of approximately seventy genes for methane production by the pathways for methanol and acetate utilization. These include genes for soluble-type and membrane-type heterodisulfide reductases (hdr, hydrogenases including genes for a vht-type F420 non-reducing hydrogenase, molybdenum-type (fmd as well as tungsten-type (fwd formylmethanofuran dehydrogenases, genes for rnf and mrp-type electron transfer complexes, for acetate uptake, plus multiple genes for aha- and atp-type ATP synthesis complexes. Analysis of promoters for seven gene clusters reveal UTR leaders of 51-137 nucleotides in length, raising the possibility of both transcriptional and translational levels of control. Conclusions The above findings establish the differential and coordinated expression of two major gene families in M. acetivorans in response to carbon/energy supply. Furthermore, the quantitative mRNA measurements demonstrate the dynamic range for modulating transcript abundance. Since many of these gene clusters in M. acetivorans are also present in other Methanosarcina species including M. mazei, and in M. barkeri, these findings provide a basis for predicting related control in these environmentally significant methanogens.

  13. Development of β-Lactamase as a Tool for Monitoring Conditional Gene Expression by a Tetracycline-Riboswitch in Methanosarcina acetivorans

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    Shemsi Demolli

    2014-01-01

    Full Text Available The use of reporter gene fusions to assess cellular processes such as protein targeting and regulation of transcription or translation is established technology in archaeal, bacterial, and eukaryal genetics. Fluorescent proteins or enzymes resulting in chromogenic substrate turnover, like β-galactosidase, have been particularly useful for microscopic and screening purposes. However, application of such methodology is of limited use for strictly anaerobic organisms due to the requirement of molecular oxygen for chromophore formation or color development. We have developed β-lactamase from Escherichia coli (encoded by bla in conjunction with the chromogenic substrate nitrocefin into a reporter system usable under anaerobic conditions for the methanogenic archaeon Methanosarcina acetivorans. By using a signal peptide of a putative flagellin from M. acetivorans and different catabolic promoters, we could demonstrate growth substrate-dependent secretion of β-lactamase, facilitating its use in colony screening on agar plates. Furthermore, a series of fusions comprised of a constitutive promoter and sequences encoding variants of the synthetic tetracycline-responsive riboswitch (tc-RS was created to characterize its influence on translation initiation in M. acetivorans. One tc-RS variant resulted in more than 11-fold tetracycline-dependent regulation of bla expression, which is in the range of regulation by naturally occurring riboswitches. Thus, tc-RS fusions represent the first solely cis-active, that is, factor-independent system for controlled gene expression in Archaea.

  14. Analysis of the MT1/MT2 Systems Involved in the Metabolism of One-Carbon Compounds in "Methanosarcina acetivorans" C2A

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    Opulencia, Rina Bagsic

    2009-01-01

    Methanogens are strictly anaerobic Archaea that derive their energy for growth by reducing a limited number of substrates to methane. "Methanosarcina" spp. utilize the methylotrophic pathway to grow on methanol, methylamines and methylsulfides. These compounds enter the methylotrophic pathway as methyl-coenzyme M, the synthesis of which is…

  15. Transfer of Methanolobus siciliae to the genus Methanosarcina, naming it Methanosarcina siciliae, and emendation of the genus Methanosarcina

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    Ni, S.; Woese, C. R.; Aldrich, H. C.; Boone, D. R.

    1994-01-01

    A sequence analysis of the 16S rRNA of Methanolobus siciliae T4/M(T) (T = type strain) showed that this strain is closely related to members of the genus Methanosarcina, especially Methanosarcina acetivorans C2A(T). Methanolobus siciliae T4/M(T) and HI350 were morphologically more similar to members of the genus Methanosarcina than to members of the genus Methanolobus in that they both formed massive cell aggregates with pseudosarcinae. Thus, we propose that Methanolobus siciliae should be transferred to the genus Methanosarcina as Methanosarcina siciliae.

  16. Mining Proteomic Data to Expose Protein Modifications in Methanosarcina mazei strain Gö1

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    Deborah eLeon

    2015-03-01

    Full Text Available Proteomic tools identify constituents of complex mixtures, often delivering long lists of identified proteins. The high-throughput methods excel at matching tandem mass spectrometry data to spectra predicted from sequence databases. Unassigned mass spectra are ignored, but could, in principle, provide valuable information on unanticipated modifications and improve protein annotations while consuming limited quantities of material. Strategies to mine information from these discards are presented, along with discussion of features that, when present, provide strong support for modifications. In this study we mined LC-MS/MS datasets of proteolytically-digested concanavalin A pull down fractions from Methanosarcina mazei Gö1 cell lysates. Analyses identified 154 proteins. Many of the observed proteins displayed post-translationally modified forms, including O-formylated and methyl-esterified segments that appear biologically relevant (i.e., not artifacts of sample handling. Interesting cleavages and modifications (e.g., S-cyanylation and trimethylation were observed near catalytic sites of methanogenesis enzymes. Of 31 Methanosarcina protein N-termini recovered by concanavalin A binding or from a previous study, only M. mazei S-layer protein MM1976 and its M. acetivorans C2A orthologue, MA0829, underwent signal peptide excision. Experimental results contrast with predictions from algorithms SignalP 3.0 and Exprot, which were found to over-predict the presence of signal peptides. Proteins MM0002, MM0716, MM1364, and MM1976 were found to be glycosylated, and employing chromatography tailored specifically for glycopeptides will likely reveal more.This study supplements limited, existing experimental datasets of mature archaeal N-termini, including presence or absence of signal peptides, translation initiation sites, and other processing. Methanosarcina surface and membrane proteins are richly modified.

  17. Genetic manipulation of Methanosarcina spp.

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    Petra Regine Adelheid Kohler

    2012-07-01

    Full Text Available The discovery of the third domain of life, the Archaea, is one of the most exciting findings of the last century. These remarkable prokaryotes are well known for their adaptations to extreme environments; however, Archaea have also conquered moderate environments. Many of the archaeal biochemical processes, such as methane production, are unique in nature and therefore of great scientific interest. Although formerly restricted to biochemical and physiological studies, sophisticated systems for genetic manipulation have been developed during the last two decades for methanogenic archaea, halophilic archaea and thermophilic, sulfur-metabolizing archaea. The availability of these tools has allowed for more complete studies of archaeal physiology and metabolism and most importantly provides the basis for the investigation of gene expression, regulation and function. In this review we provide an overview of methods for genetic manipulation of Methanosarcina spp., a group of methanogenic archaea that are key players in the global carbon cycle and which can be found in a variety of anaerobic environments.

  18. A multienzyme complex channels substrates and electrons through acetyl-CoA and methane biosynthesis pathways in Methanosarcina.

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    Dillon J Lieber

    Full Text Available Multienzyme complexes catalyze important metabolic reactions in many organisms, but little is known about the complexes involved in biological methane production (methanogenesis. A crosslinking-mass spectrometry (XL-MS strategy was employed to identify proteins associated with coenzyme M-coenzyme B heterodisulfide reductase (Hdr, an essential enzyme in all methane-producing archaea (methanogens. In Methanosarcina acetivorans, Hdr forms a multienzyme complex with acetyl-CoA decarbonylase synthase (ACDS, and F420-dependent methylene-H4MPT reductase (Mer. ACDS is essential for production of acetyl-CoA during growth on methanol, or for methanogenesis from acetate, whereas Mer is essential for methanogenesis from all substrates. Existence of a Hdr:ACDS:Mer complex is consistent with growth phenotypes of ACDS and Mer mutant strains in which the complex samples the redox status of electron carriers and directs carbon flux to acetyl-CoA or methanogenesis. We propose the Hdr:ACDS:Mer complex comprises a special class of multienzyme redox complex which functions as a "biological router" that physically links methanogenesis and acetyl-CoA biosynthesis pathways.

  19. Mixotrophic growth of two thermophilic Methanosarcina strains, Methanosarcina thermophila TM-1 and Methanosarcina sp. SO-2P, on methanol and hydrogen/carbon dioxide

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    Mladenovska, Zuzana; Ahring, Birgitte Kiær

    1997-01-01

    Two thermophilic strains, Methanosarcina thermophila TM-1 and Methanosarcina sp. SO-2P, were capable of mixotrophic growth on methanol and H-2/CO2. Activated carbon was, however, found to be necessary to support good growth. Both strains used hydrogen and methanol simultaneously. When methanol wa...... was depleted, hydrogen utilization continued and methane was further produced with concurrent cell growth. UV epifluorescence microscopy revealed that aggregates of both strains exhibited a bright red fluorescence besides the usual blue-green fluorescence....

  20. Mixotrophic growth of two thermophilic Methanosarcina strains, Methanosarcina thermophila TM-1 and Methanosarcina sp. SO-2P, on methanol and hydrogen/carbon dioxide

    DEFF Research Database (Denmark)

    Mladenovska, Zuzana; Ahring, Birgitte Kiær

    1997-01-01

    Two thermophilic strains, Methanosarcina thermophila TM-1 and Methanosarcina sp. SO-2P, were capable of mixotrophic growth on methanol and H-2/CO2. Activated carbon was, however, found to be necessary to support good growth. Both strains used hydrogen and methanol simultaneously. When methanol wa...... was depleted, hydrogen utilization continued and methane was further produced with concurrent cell growth. UV epifluorescence microscopy revealed that aggregates of both strains exhibited a bright red fluorescence besides the usual blue-green fluorescence....

  1. Direct interspecies electron transfer between Geobacter metallireducens and Methanosarcina barkeri

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    Rotaru, Amelia-Elena; Shrestha, Pravin Malla; Liu, Fanghua;

    2014-01-01

    Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri, to participate in DIET was evaluated in co-culture with Geobacter...

  2. Microbial succession during thermophilic digestion: the potential of Methanosarcina sp.

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    Paul Illmer

    Full Text Available A distinct succession from a hydrolytic to a hydrogeno- and acetotrophic community was well documented by DGGE (denaturing gradient gel electrophoresis and dHPLC (denaturing high performance liquid chromatography, and confirmed by qPCR (quantitative PCR measurements and DNA sequence analyses. We could prove that Methanosarcina thermophila has been the most important key player during the investigated anaerobic digestion process. This organism was able to terminate a stagnation phase, most probable caused by a decreased pH and accumulated acetic acid following an initial hydrolytic stage. The lack in Methanosarcina sp. could not be compensated by high numbers of Methanothermobacter sp. or Methanoculleus sp., which were predominant during the initial or during the stagnation phase of the fermentation, respectively.

  3. Direct interspecies electron transfer between Geobacter metallireducens and Methanosarcina barkeri

    DEFF Research Database (Denmark)

    Rotaru, Amelia-Elena; Shrestha, Pravin Malla; Liu, Fanghua

    2014-01-01

    Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri, to participate in DIET was evaluated in co-culture with Geobacter...... metallireducens. Co-cultures formed aggregates that shared electrons via DIET during the stoichiometric conversion of ethanol to methane. Co-cultures could not be initiated with a pilin-deficient G. metallireducens, suggesting that long-range electron transfer along pili was important for DIET. Amendments...... physical contact was not necessary for interspecies H2 transfer. M. barkeri is the second methanogen found to accept electrons via DIET and the first methanogen known to be capable of using either H2 or electrons derived from DIET for CO2 reduction. Furthermore, M. barkeri is genetically tractable, making...

  4. Growth kinetics and competition between Methanosarcina and Methanosaeta in mesophilic anaerobic digestion.

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    Conklin, Anne; Stensel, H David; Ferguson, John

    2006-05-01

    Methanosarcina species with a high maximum specific growth rate (mumax) and high half-saturation coefficient (KS) and Methanosaeta species with a low mumax and low KS are the only known aceticlastic methanogens. Because of Methanosaeta's low KS, the low acetate concentrations in conventional, mesophilic anaerobic digestion yield Methanosaeta dominance. However, Methanosarcina absorbs increases in acetate more efficiently and thus promotes more stable digestion. This paper tests the hypothesis that decreasing digester feeding frequencies can increase Methanosarcina predominance. Two acetate-fed reactors were established at a 17-day solids retention time. One reactor was fed hourly, and one was fed once daily. Microscopic and molecular methods were used to verify that the hourly fed reactor enriched for Methanosaeta, while the daily fed reactor enriched for Methanosarcina. Growth and substrate-use kinetics were measured for each reactor. A digester overload condition was simulated, and the Methanosarcina-enriched reactor was found to perform better than the Methanosaeta-enriched reactor. These findings indicate that Methanosarcina dominance can be achieved with infrequent feedings, leading to more stable digestion.

  5. Mineral transformations associated with goethite reduction by Methanosarcina barkeri

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    Liu, D.; Wang, Hongfang; Dong, H.; Qiu, X.; Dong, X.; Cravotta, C.A.

    2011-01-01

    To investigate the interaction between methanogens and iron-containing minerals in anoxic environments, we conducted batch culture experiments with Methanosarcina barkeri in a phosphate-buffered basal medium (PBBM) to bioreduce structural Fe(III) in goethite with hydrogen as the sole substrate. Fe(II) and methane concentrations were monitored over the course of the bioreduction experiments with wet chemistry and gas chromatography, respectively. Subsequent mineralogical changes were characterized with X-ray diffraction (XRD) and scanning electron microscopy (SEM). In the presence of an electron shuttle anthraquinone-2,6-disulfonate (AQDS), 30% Fe(III) in goethite (weight basis) was reduced to Fe(II). In contrast, only 2% Fe(III) (weight basis) was bioreduced in the absence of AQDS. Most of the bioproduced Fe(II) was incorporated into secondary minerals including dufr??nite and vivianite. Our data implied a dufr??nite-vivianite transformation mechanism where a metastable dufr??nite transformed to a more stable vivianite over extended time in anaerobic conditions. Methanogenesis was greatly inhibited by bioreduction of goethite Fe(III). These results have important implications for the methane flux associated with Fe(III) bioreduction and ferrous iron mineral precipitation in anaerobic soils and sediments. ?? 2011 Elsevier B.V.

  6. Direct interspecies electron transfer between Geobacter metallireducens and Methanosarcina barkeri.

    Science.gov (United States)

    Rotaru, Amelia-Elena; Shrestha, Pravin Malla; Liu, Fanghua; Markovaite, Beatrice; Chen, Shanshan; Nevin, Kelly P; Lovley, Derek R

    2014-08-01

    Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri to participate in DIET was evaluated in coculture with Geobacter metallireducens. Cocultures formed aggregates that shared electrons via DIET during the stoichiometric conversion of ethanol to methane. Cocultures could not be initiated with a pilin-deficient G. metallireducens strain, suggesting that long-range electron transfer along pili was important for DIET. Amendments of granular activated carbon permitted the pilin-deficient G. metallireducens isolates to share electrons with M. barkeri, demonstrating that this conductive material could substitute for pili in promoting DIET. When M. barkeri was grown in coculture with the H2-producing Pelobacter carbinolicus, incapable of DIET, M. barkeri utilized H2 as an electron donor but metabolized little of the acetate that P.carbinolicus produced. This suggested that H2, but not electrons derived from DIET, inhibited acetate metabolism. P. carbinolicus-M. barkeri cocultures did not aggregate, demonstrating that, unlike DIET, close physical contact was not necessary for interspecies H2 transfer. M. barkeri is the second methanogen found to accept electrons via DIET and the first methanogen known to be capable of using either H2 or electrons derived from DIET for CO2 reduction. Furthermore, M. barkeri is genetically tractable,making it a model organism for elucidating mechanisms by which methanogens make biological electrical connections with other cells.

  7. Methanosarcina subterranea sp. nov., a methanogenic archaeon isolated from a deep subsurface diatomaceous shale formation.

    Science.gov (United States)

    Shimizu, Satoru; Ueno, Akio; Naganuma, Takeshi; Kaneko, Katsuhiko

    2015-04-01

    A methanogenic archaeon, strain HC-2(T), was isolated from a deep diatomaceous shale formation. The strain grew on methanol, monomethylamine, dimethylamine, trimethylamine and dimethylsulphide, but not on acetate, H2/CO2, formate, 2-propanol, 2-butanol or cyclopentanol. Cells were Gram-stain-negative, non-motile, and coccus-like, 0.9-1.4 µm in diameter, and occurred singly, in pairs, or as aggregates. The strain grew at 10-40 °C (optimum 35 °C), pH 5.9-7.4 (optimum pH 6.6-6.8) and in 0-0.6 M NaCl (optimum 0.1-0.2 M). The genomic DNA G+C content was 41.5 mol% and the 16S rRNA gene sequence was closely related to those of Methanosarcina lacustris DSM 13486(T) (99.1%) and Methanosarcina siciliae DSM 3028(T) (98.3%). Values for DNA-DNA hybridization with these strains were less than 30%. The phenotypic and phylogenetic features of HC-2(T) indicate that it represents a novel species of the genus Methanosarcina , for which the name Methanosarcina subterranea sp. nov. is proposed. The type strain is HC-2(T) ( = DSM 22503(T) = JCM 15540(T) = NBRC 102578(T)). © 2015 IUMS.

  8. Optimization of RNA Isolation from the Archaebacterium Methanosarcina Barkeri and Validation for Oligonucleotide Microarray Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Culley, David E.; Kovacik, William P.; Brockman, Fred J.; Zhang, Weiwen

    2006-10-01

    ABSTRACT-The recent completion of a draft genome sequence for Methanosarcina barkeri has allowed the application of various high throughput post-genomics technologies, such as nucleic acid microarrays and mass spectrometry of proteins to detect global changes in transcription and translation that occur in response to experimental treatments...

  9. Methanosarcina spelaei sp. nov., a methanogenic archaeon isolated from a floating biofilm of a subsurface sulphurous lake.

    Science.gov (United States)

    Ganzert, Lars; Schirmack, Janosch; Alawi, Mashal; Mangelsdorf, Kai; Sand, Wolfgang; Hillebrand-Voiculescu, Alexandra; Wagner, Dirk

    2014-10-01

    A novel methanogenic archaeon, strain MC-15(T), was isolated from a floating biofilm on a sulphurous subsurface lake in Movile Cave (Mangalia, Romania). Cells were non-motile sarcina-like cocci with a diameter of 2-4 µm, occurring in aggregates. The strain was able to grow autotrophically on H2/CO2. Additionally, acetate, methanol, monomethylamine, dimethylamine and trimethylamine were utilized, but not formate or dimethyl sulfide. Trypticase peptone and yeast extract were not required for growth. Optimal growth was observed at 33 °C, pH 6.5 and a salt concentration of 0.05 M NaCl. The predominant membrane lipids of MC-15(T) were archaeol and hydroxyarchaeol phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylinositol as well as hydroxyarchaeol phosphatidylserine and archaeol glycosaminyl phosphatidylinositol. The closely related species, Methanosarcina vacuolata and Methanosarcina horonobensis, had a similar composition of major membrane lipids to strain MC-15(T). The 16S rRNA gene sequence of strain MC-15(T) was similar to those of Methanosarcina vacuolata DSM 1232(T) (sequence similarity 99.3%), Methanosarcina horonobensis HB-1(T) (98.8%), Methanosarcina barkeri DSM 800(T) (98.7%) and Methanosarcina siciliae T4/M(T) (98.4%). DNA-DNA hybridization revealed 43.3% relatedness between strain MC-15(T) and Methanosarcina vacuolata DSM 1232(T). The G+C content of the genomic DNA was 39.0 mol%. Based on physiological, phenotypic and genotypic differences, strain MC-15(T) represents a novel species of the genus Methanosarcina, for which the name Methanosarcina spelaei sp. nov. is proposed. The type strain is MC-15(T) ( = DSM 26047(T) = JCM 18469(T)). © 2014 IUMS.

  10. The Geoglobus acetivorans genome: Fe(III) reduction, acetate utilization, autotrophic growth, and degradation of aromatic compounds in a hyperthermophilic archaeon.

    Science.gov (United States)

    Mardanov, Andrey V; Slododkina, Galina B; Slobodkin, Alexander I; Beletsky, Alexey V; Gavrilov, Sergey N; Kublanov, Ilya V; Bonch-Osmolovskaya, Elizaveta A; Skryabin, Konstantin G; Ravin, Nikolai V

    2015-02-01

    Geoglobus acetivorans is a hyperthermophilic anaerobic euryarchaeon of the order Archaeoglobales isolated from deep-sea hydrothermal vents. A unique physiological feature of the members of the genus Geoglobus is their obligate dependence on Fe(III) reduction, which plays an important role in the geochemistry of hydrothermal systems. The features of this organism and its complete 1,860,815-bp genome sequence are described in this report. Genome analysis revealed pathways enabling oxidation of molecular hydrogen, proteinaceous substrates, fatty acids, aromatic compounds, n-alkanes, and organic acids, including acetate, through anaerobic respiration linked to Fe(III) reduction. Consistent with the inability of G. acetivorans to grow on carbohydrates, the modified Embden-Meyerhof pathway encoded by the genome is incomplete. Autotrophic CO2 fixation is enabled by the Wood-Ljungdahl pathway. Reduction of insoluble poorly crystalline Fe(III) oxide depends on the transfer of electrons from the quinone pool to multiheme c-type cytochromes exposed on the cell surface. Direct contact of the cells and Fe(III) oxide particles could be facilitated by pilus-like appendages. Genome analysis indicated the presence of metabolic pathways for anaerobic degradation of aromatic compounds and n-alkanes, although an ability of G. acetivorans to grow on these substrates was not observed in laboratory experiments. Overall, our results suggest that Geoglobus species could play an important role in microbial communities of deep-sea hydrothermal vents as lithoautotrophic producers. An additional role as decomposers would close the biogeochemical cycle of carbon through complete mineralization of various organic compounds via Fe(III) respiration.

  11. Methanosarcina soligelidi sp. nov., a desiccation- and freeze-thaw-resistant methanogenic archaeon from a Siberian permafrost-affected soil.

    Science.gov (United States)

    Wagner, Dirk; Schirmack, Janosch; Ganzert, Lars; Morozova, Daria; Mangelsdorf, Kai

    2013-08-01

    A methanogenic archaeon, strain SMA-21(T), was isolated from a permafrost-affected soil by serial dilution in liquid medium. The cells were non-motile, stained Gram-negative and grew as irregular cocci with a diameter of 1.3-2.5 µm. Optimal growth was observed at 28 °C, pH 7.8 and 0.02 M NaCl. The strain grew on H2/CO2, methanol and acetate, but not on formate, ethanol, 2-butanol, 2-propanol, monomethylamine, dimethylamine, trimethylamine or dimethyl sulfide. Major membrane lipids of strain SMA-21(T) were archaeol phosphatidylglycerol, archaeol phosphatidylethanolamine and the corresponding hydroxyarchaeol compounds. The G+C content of the genomic DNA was 40.9 mol%. The 16S rRNA gene sequence was closely related to those of Methanosarcina mazei DSM 2053(T) (similarity 99.9 %) and Methanosarcina horonobensis HB-1(T) (similarity 98.7 %). On basis of the level of DNA-DNA hybridization (22.1 %) between strain SMA-21(T) and Methanosarcina mazei DSM 2053(T) as well as of phenotypic and genotypic differences, strain SMA-21(T) was assigned to a novel species of the genus Methanosarcina, for which the name Methanosarcina soligelidi sp. nov. is proposed. The type strain is SMA-21(T) (=DSM 26065(T) [corrected] = JCM 18468).

  12. Co-occurrence of Methanosarcina mazei and Geobacteraceae in an iron(III-reducing enrichment culture

    Directory of Open Access Journals (Sweden)

    Shiling eZheng

    2015-09-01

    Full Text Available Methanosaeta harundinacea and Methanosarcina barkeri, known as classic acetoclastic methanogens, are capable of directly accepting electrons from Geobacter metallireducens for the reduction of carbon dioxide to methane, having been revealed as direct interspecies electron transfer (DIET in the laboratory co-cultures. However, whether their co-occurrences are ubiquitous in the iron (III-reducing environments and the other species of acetoclastic methanogens such as Methanosarcina mazei are capable of DIET are still unknown. Instead of initiating the co-cultures with pure cultures, two-step cultivation was employed to selectively enrich iron (III-reducing microorganisms in a coastal gold mining river, Jiehe River, with rich iron content in the sediments. First, iron (III reducers including Geobacteraceae were successfully enriched by 3-months successive culture on amorphous Fe(III oxides as electron acceptor and acetate as electron donor. High-throughput Illumina sequencing, terminal restriction fragment length polymorphism (T-RFLP and clone library analysis based on 16S rRNA genes revealed that the enrichment cultures actively contained the bacteria belong to Geobacteraceae and Bacilli, exclusively dominated by the archaea belong to Methanosarcinaceae. Second, the enrichment cultures including methanogens and Geobacteraceae were transferred with ethanol as alternative electron donor. Remarkably, aggregates were successively formed in the enrichments after three transfers. The results revealed by RNA-based analysis demonstrate that the co-occurrence of Methanosarcina mazei and Geobacteraceae in an iron (III-reducing enrichment culture. Furthermore, the aggregates, as close physical contact, formed in the enrichment culture, indicate that DIET could be a possible option for interspecies electron transfer in the aggregates.

  13. Synthesis of acetyl coenzyme A by carbon monoxide dehydrogenase complex from acetate-grown Methanosarcina thermophila.

    OpenAIRE

    Abbanat, D R; Ferry, J G

    1990-01-01

    The carbon monoxide dehydrogenase (CODH) complex from Methanosarcina thermophila catalyzed the synthesis of acetyl coenzyme A (acetyl-CoA) from CH3I, CO, and coenzyme A (CoA) at a rate of 65 nmol/min/mg at 55 degrees C. The reaction ended after 5 min with the synthesis of 52 nmol of acetyl-CoA per nmol of CODH complex. The optimum temperature for acetyl-CoA synthesis in the assay was between 55 and 60 degrees C; the rate of synthesis at 55 degrees C was not significantly different between pHs...

  14. Transmethylation reactions during methanogenesis from acetate in Methanosarcina barkeri. [Annual report], August 1, 1991--April 1, 1993

    Energy Technology Data Exchange (ETDEWEB)

    Krzycki, J.A.

    1993-07-01

    Methanosarcina possesses an abundance corrinoid, which acts as enzyme prosthetic group and functions in methyl transfer reactions. When this work was initiated only two corrinoid proteins had been described from Methanosarcina: methyltransferase I, which catalyses the first step m methanogenesis from methanol; and the corrinoid/iron sulfur protein, which is the primary methyl accepting group in the cleavage of acetyl-CoA during methanogenesis from acetate. Using our technique of in vitro methylation of corrinoid proteins by radioactive methanogenic substrates we have found several novel corrinoid proteins are present in this organism which can function as methytransferases, which are methylated by growth substrate, and are apparently regulated by growth substrate.

  15. Functional organization of a single nif cluster in the mesophilic archaeon Methanosarcina mazei strain Gö1

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    Claudia Ehlers

    2002-01-01

    Full Text Available The mesophilic methanogenic archaeon Methanosarcina mazei strain Gö1 is able to utilize molecular nitrogen (N2 as its sole nitrogen source. We have identified and characterized a single nitrogen fixation (nif gene cluster in M. mazei Gö1 with an approximate length of 9 kbp. Sequence analysis revealed seven genes with sequence similarities to nifH, nifI1, nifI2, nifD, nifK, nifE and nifN, similar to other diazotrophic methanogens and certain bacteria such as Clostridium acetobutylicum, with the two glnB-like genes (nifI1 and nifI2 located between nifH and nifD. Phylogenetic analysis of deduced amino acid sequences for the nitrogenase structural genes of M. mazei Gö1 showed that they are most closely related to Methanosarcina barkeri nif2 genes, and also closely resemble those for the corresponding nif products of the gram-positive bacterium C. acetobutylicum. Northern blot analysis and reverse transcription PCR analysis demonstrated that the M. mazei nif genes constitute an operon transcribed only under nitrogen starvation as a single 8 kb transcript. Sequence analysis revealed a palindromic sequence at the transcriptional start site in front of the M. mazei nifH gene, which may have a function in transcriptional regulation of the nif operon.

  16. Functional organization of a single nif cluster in the mesophilic archaeon Methanosarcina mazei strain Gö1.

    Science.gov (United States)

    Ehlers, Claudia; Veit, Katharina; Gottschalk, Gerhard; Schmitz, Ruth A

    2002-09-01

    The mesophilic methanogenic archaeon Methanosarcina mazei strain Gö1 is able to utilize molecular nitrogen (N2) as its sole nitrogen source. We have identified and characterized a single nitrogen fixation (nif) gene cluster in M. mazei Gö1 with an approximate length of 9 kbp. Sequence analysis revealed seven genes with sequence similarities to nifH, nifI1, nifI2, nifD, nifK, nifE and nifN, similar to other diazotrophic methanogens and certain bacteria such as Clostridium acetobutylicum, with the two glnB-like genes (nifI1 and nifI2) located between nifH and nifD. Phylogenetic analysis of deduced amino acid sequences for the nitrogenase structural genes of M. mazei Gö1 showed that they are most closely related to Methanosarcina barkeri nif2 genes, and also closely resemble those for the corresponding nif products of the gram-positive bacterium C. acetobutylicum. Northern blot analysis and reverse transcription PCR analysis demonstrated that the M. mazei nif genes constitute an operon transcribed only under nitrogen starvation as a single 8 kb transcript. Sequence analysis revealed a palindromic sequence at the transcriptional start site in front of the M. mazei nifH gene, which may have a function in transcriptional regulation of the nif operon.

  17. 2,6,10,15,19-Pentamethylicosenes in Methanolobus bombayensis, a marine methanogenic archaeon, and in Methanosarcina mazei

    NARCIS (Netherlands)

    VanderMaarel, MJEC; Huber, R; Damste, JSS; Sinninghe Damsté, Jaap S.

    1997-01-01

    2,6,10,15,19-Pentamethylicosenes (PMEs) containing three to five double bonds have been found in the methanogenic archaea Methanosarcina mazei (DSM 3338), a strain isolated from sewage sludge, and in Methanolobus bombayensis (OCM 438), a non-extremophilic archaeon isolated from a marine sediment.

  18. Isolation and characterization of a new Methanosarcina mazei strain GFJ07 from a mountain forest pond

    Directory of Open Access Journals (Sweden)

    Yi, Y.

    2012-01-01

    Full Text Available Aims: A new methanogenic strain, named GFJ07, was isolated from a pond of mountain forests in Guangxi, China. Cells grown in liquid culture tended to form aggregates with pseudosarcina-like or irregular shape.Methodology and Results: The optimum temperature, pH and NaCl concentration were 35 ℃, 7.0 and 0.5%, respectively. The isolate used methanol, trimethylamine, acetate and H2-CO2 as substrates. Analysis of the 16S rDNA sequences revealed strain GFJ07 showed the highest sequence similarity of 99.9% to Methanosarcina mazei.Conclusion, significance and impact of study: The cells were Gram positive and nonmotile. Most of single cell grew as a sausage-like clinder about 0.5 μm in diameter and 1.0 μm in length.

  19. Effects of Nitrogen and Carbon Sources on Transcription of Soluble Methyltransferases in Methanosarcina mazei Strain Gö1†

    OpenAIRE

    Veit, Katharina; Ehlers, Claudia; Schmitz, Ruth A.

    2005-01-01

    The methanogenic archaeon Methanosarcina mazei strain Gö1 uses versatile carbon sources and is able to fix molecular nitrogen with methanol as carbon and energy sources. Here, we demonstrate that when growing on trimethylamine (TMA), nitrogen fixation does not occur, indicating that ammonium released during TMA degradation is sufficient to serve as a nitrogen source and represses nif gene induction. We further report on the transcriptional regulation of soluble methyltransferases, which catal...

  20. Purification and characterization of dimethylamine:5-hydroxybenzimidazolyl-cobamide methyltransferase from Methanosarcina barkeri Fusaro.

    Science.gov (United States)

    Wassenaar, R W; Keltjens, J T; van der Drift, C; Vogels, G D

    1998-05-01

    Dimethylamine:5-hydroxybenzimidazolylcobamide methyltransferase (DMA-MT) was purified from cells of Methanosarcina barkeri Fusaro grown on trimethylamine. In the presence of methylcobalamine:coenzyme M methyltransferase isoenzyme II [MT2(II)] the enzyme quite specifically catalyzed the stoichiometric conversion of dimethylamine (apparent Km = 0.45 mM) and 2-mercaptoethane-sulfonate (coenzyme M) to monomethylamine and methyl-coenzyme M. Monomethylamine was a competitive inhibitor of the reaction (Ki = 4.5 mM). The apparent molecular mass of DMA-MT was 100 kDa and the enzyme was found to be a dimer, composed of identical 50-kDa subunits. A corrinoid content of 0.9 +/- 0.1 mol B12/mol holoenzyme was calculated from HPLC analysis. The as-isolated methyltransferase was inactive, but it could be reductively reactivated. Activation required the presence of methyltransferase-activating protein, ATP and dimethylamine. Incubation with these compounds resulted in the methylation of the corrinoid prosthetic group.

  1. The Alternative Route to Heme in the Methanogenic Archaeon Methanosarcina barkeri

    Directory of Open Access Journals (Sweden)

    Melanie Kühner

    2014-01-01

    Full Text Available In living organisms heme is formed from the common precursor uroporphyrinogen III by either one of two substantially different pathways. In contrast to eukaryotes and most bacteria which employ the so-called “classical” heme biosynthesis pathway, the archaea use an alternative route. In this pathway, heme is formed from uroporphyrinogen III via the intermediates precorrin-2, sirohydrochlorin, siroheme, 12,18-didecarboxysiroheme, and iron-coproporphyrin III. In this study the heme biosynthesis proteins AhbAB, AhbC, and AhbD from Methanosarcina barkeri were functionally characterized. Using an in vivo enzyme activity assay it was shown that AhbA and AhbB (Mbar_A1459 and Mbar_A1460 together catalyze the conversion of siroheme into 12,18-didecarboxysiroheme. The two proteins form a heterodimeric complex which might be subject to feedback regulation by the pathway end-product heme. Further, AhbC (Mbar_A1793 was shown to catalyze the formation of iron-coproporphyrin III in vivo. Finally, recombinant AhbD (Mbar_A1458 was produced in E. coli and purified indicating that this protein most likely contains two [4Fe-4S] clusters. Using an in vitro enzyme activity assay it was demonstrated that AhbD catalyzes the conversion of iron-coproporphyrin III into heme.

  2. Biochar alleviates combined stress of ammonium and acids by firstly enriching Methanosaeta and then Methanosarcina.

    Science.gov (United States)

    Lü, Fan; Luo, Chenghao; Shao, Liming; He, Pinjing

    2016-03-01

    This investigation evaluated the effectiveness of biochar of different particle sizes in alleviating ammonium (NH4(+)) inhibition (up to 7 g-N/L) during anaerobic digestion of 6 g/L glucose. Compared to the control treatment without biochar addition, treatments that included biochar particles 2-5 mm, 0.5-1 mm and 75-150 μm in size reduced the methanization lag phase by 23.9%, 23.8% and 5.9%, respectively, and increased the maximum methane production rate by 47.1%, 23.5% and 44.1%, respectively. These results confirmed that biochar accelerated the initiation of methanization during anaerobic digestion under double inhibition risk from both ammonium and acids. Furthermore, fine biochar significantly promoted the production of volatile fatty acids (VFAs). Comparative analysis on the archaeal and bacterial diversity at the early and later stages of digestion, and in the suspended, biochar loosely bound, and biochar tightly bound fractions suggested that, in suspended fractions, hydrogenotrophic Methanobacterium was actively resistant to ammonium. However, acetoclastic Methanosaeta can survive at VFAs concentrations up to 60-80 mmol-C/L by improved affinity to conductive biochar, resulting in the accelerated initiation of acetate degradation. Improved methanogenesis was followed by the colonization of the biochar tightly bound fractions by Methanosarcina. The selection of appropriate biochar particles sizes was important in facilitating the initial colonization of microbial cells.

  3. A novel inducible protein production system and neomycin resistance as selection marker for Methanosarcina mazei.

    Science.gov (United States)

    Mondorf, Sebastian; Deppenmeier, Uwe; Welte, Cornelia

    2012-01-01

    Methanosarcina mazei is one of the model organisms for the methanogenic order Methanosarcinales whose metabolism has been studied in detail. However, the genetic toolbox is still limited. This study was aimed at widening the scope of utilizable methods in this group of organisms. (i) Proteins specific to methanogens are oftentimes difficult to produce in E. coli. However, a protein production system is not available for methanogens. Here we present an inducible system to produce Strep-tagged proteins in Ms. mazei. The promoter p1687, which directs the transcription of methyl transferases that demethylate methylamines, was cloned into plasmid pWM321 and its activity was determined by monitoring β-glucuronidase production. The promoter was inactive during growth on methanol but was rapidly activated when trimethylamine was added to the medium. The gene encoding the β-glucuronidase from E. coli was fused to a Strep-tag and was cloned downstream of the p1687 promoter. The protein was overproduced in Ms. mazei and was purified in an active form by affinity chromatography. (ii) Puromycin is currently the only antibiotic used as a selectable marker in Ms. mazei and its relatives. We established neomycin resistance as a second selectable marker by designing a plasmid that confers neomycin resistance in Ms. mazei.

  4. A Novel Inducible Protein Production System and Neomycin Resistance as Selection Marker for Methanosarcina mazei

    Directory of Open Access Journals (Sweden)

    Sebastian Mondorf

    2012-01-01

    Full Text Available Methanosarcina mazei is one of the model organisms for the methanogenic order Methanosarcinales whose metabolism has been studied in detail. However, the genetic toolbox is still limited. This study was aimed at widening the scope of utilizable methods in this group of organisms. (i Proteins specific to methanogens are oftentimes difficult to produce in E. coli. However, a protein production system is not available for methanogens. Here we present an inducible system to produce Strep-tagged proteins in Ms. mazei. The promoter p1687, which directs the transcription of methyl transferases that demethylate methylamines, was cloned into plasmid pWM321 and its activity was determined by monitoring β-glucuronidase production. The promoter was inactive during growth on methanol but was rapidly activated when trimethylamine was added to the medium. The gene encoding the β-glucuronidase from E. coli was fused to a Strep-tag and was cloned downstream of the p1687 promoter. The protein was overproduced in Ms. mazei and was purified in an active form by affinity chromatography. (ii Puromycin is currently the only antibiotic used as a selectable marker in Ms. mazei and its relatives. We established neomycin resistance as a second selectable marker by designing a plasmid that confers neomycin resistance in Ms. mazei.

  5. Production and biophysical characterization of the CorA transporter from Methanosarcina mazei.

    Science.gov (United States)

    Veesler, David; Blangy, Stéphanie; Siponen, Marina; Vincentelli, Renaud; Cambillau, Christian; Sciara, Giuliano

    2009-05-01

    We report here a general strategy to overproduce and characterize membrane transporters. To illustrate our approach, we selected one member of the CorA transporter family among four tested that belonged to different species. This approach is transposable to other membrane proteins and involves the following steps: (i) cloning by homologous recombination, (ii) high-throughput expression screening, (iii) fermenter-based large-scale production, (iv) high-throughput detergent solubilization screening, (v) protein purification, (vi) multiangle static light scattering/refractometry characterization of purified proteins, (vii) circular dichroism spectroscopy, and (viii) detergent concentration measurements by Fourier transform infrared (FT-IR) spectroscopy. Methanosarcina mazei CorA was expressed in milligram quantities and purified (> 95% pure). n-Dodecyl-beta-D-maltopyranoside (DDM) retained the pentameric native structure of this transporter; thus, we selected it as working detergent. Furthermore, we measured the detergent concentration in our purified and concentrated protein sample by FT-IR to maintain it as low as possible. Our strategy can be adapted to many structural biology approaches as well as for study of single membrane proteins in a variety of conditions.

  6. Effects of various environmental conditions on the transformation of chlorinated solvents by Methanosarcina thermophila cell exudates.

    Science.gov (United States)

    Baeseman, J L; Novak, P J

    2001-12-20

    Several microbiologically produced biomolecules have been shown to degrade chlorinated contaminants found in groundwater systems. It was discovered that the cell-free exudates of the methanogen Methanosarcina thermophila were capable of carbon tetrachloride (CT) and chloroform (CF) degradation. Characterization of the exudates suggested that the active agents were porphorinogen-type molecules, possibly containing zinc. This research was performed to determine if the exudates from M. thermophila could be used for remediation purposes. The cell exudates were found to be capable of degrading CT, CF, tetrachloroethene, trichloroethene, and 1,1,1-trichloroethane. CT degradation was used to gauge exudate activity under a variety of conditions that would be encountered in the environment. The cell exudates were active when incubated in two types of soil matrices and at temperatures ranging from 4 to 23 degrees C. Over a 35-day period approximately 10.2 micromoles of CT were degraded by M. thermophila exudates. To test the hypothesis that the exudates contained either a zinc porphorinogen or a quinone, experiments were performed with zinc 5,10,15,20-tetra (4-pyridyl)-21 H, 23 H-porphine tetrakis, protoporphyrin IX zinc, and juglone. The two zinc porphyrins were capable of mediating CT degradation at rates comparable to those observed with the M. thermophila exudates; however, juglone was only capable of very slow CT transformation. The electron-transfer activity of the M. thermophila cell exudates was therefore more consistent with the activity of porphorinogens rather than quinones. Finally, in two enrichment cultures established from aquifer material and marine sediment, the possibility of excreted agents capable of degrading CT was evident.

  7. Characterization of cytochromes from Methanosarcina strain Göl and their involvement in electron transport during growth on methanol.

    Science.gov (United States)

    Kamlage, B; Blaut, M

    1992-01-01

    Methanosarcina strain Gö1 was tested for the presence of cytochromes. Low-temperature spectroscopy, hemochrome derivative spectroscopy, and redox titration revealed the presence of two b-type (b559 and b564) and two c-type (c547 and c552) cytochromes in membranes from Methanosarcina strain Gö1. The midpoint potentials determined were Em,7 = -135 +/- 5 and -240 +/- 11 mV (b-type cytochromes) and Em,7 = -140 +/- 10 and -230 +/- 10 mV (c-type cytochromes). The protoheme IX and the heme c contents were 0.21 to 0.24 and 0.09 to 0.28 mumol/g of membrane protein, respectively. No cytochromes were detectable in the cytoplasmic fraction. Of various electron donors and acceptors tested, only the reduced form of coenzyme F420 (coenzyme F420H2) and the heterodisulfide of coenzyme M and 7-mercaptoheptanoylthreonine phosphate (CoM-S-S-HTP) were capable of reducing and oxidizing the cytochromes at a high rate, respectively. Addition of CoM-S-S-HTP to reduced cytochromes and subsequent low-temperature spectroscopy revealed the oxidation of cytochrome b564. On the basis of these results, we suggest that one or several cytochromes participate in the coenzyme F420H2-dependent reduction of the heterodisulfide. PMID:1597414

  8. Genome sequencing of methanogenic Archaea Methanosarcina mazei TUC01 strain isolated from an Amazonian Flooded Area

    Science.gov (United States)

    Baraúna, R. A.; Graças, D. A.; Ramos, R. T.; Carneiro, A. R.; Lopes, T. S.; Lima, A. R.; Zahlouth, R. L.; Pellizari, V. H.; Silva, A.

    2013-05-01

    Methanosarcina mazei is a strictly anaerobic methanogen from the Methanosarcinales order. This species is known for its broad catabolic range among methanogens and is widespread throughout diverse environments. The draft genome of a strain cultivated from the sediment of the Tucuruí hydroelectric power station, the fourth largest hydroelectric dam in the world, is described here. Approximately 80% of methane is produced by biogenic sources, such as methanogenic archaea from M. mazei species. Although the methanogenesis pathway is well known, some aspects of the core genome, genome evolution and shared genes are still unclear. A sediment sample from the Tucuruí hydropower station reservoir was inoculated in mineral media supplemented with acetate and methanol. This media was maintained in an H2:CO2 (80:20) atmosphere to enrich and cultivate M. mazei. The enrichment was conducted at 30°C under standard anaerobic conditions. After several molecular and cellular analyses, total DNA was extracted from a non-pure culture of M. mazei, amplified using phi29 DNA polymerase (BioLabs) and finally used as a source template for genome sequencing. The draft genome was obtained after two rounds of sequencing. First, the genome was sequenced using a SOLiD System V3 with a mate-paired library, which yielded 24,405,103 and 24,399,268 reads (50 bp) for the R3 and F3 tags, respectively. The second round of sequencing was performed using the SOLiD 5500 XL platform with a mate-paired library, resulting in a total of 113,588,848 reads (60 bp) for each tag (F3 and R3). All reads obtained by this procedure were filtered using Quality Assessment software, whereby reads with an average quality score below Phred 20 were removed. Velvet and Edena were used to assemble the reads, and Simplifier was used to remove the redundant sequences. After this, a total of 16,811 contigs were obtained. M. mazei GO1 (AE008384) genome was used to map the contigs and generate the scaffolds. We used the

  9. Enhanced methane production in an anaerobic digestion and microbial electrolysis cell coupled system with co-cultivation of Geobacter and Methanosarcina.

    Science.gov (United States)

    Yin, Qi; Zhu, Xiaoyu; Zhan, Guoqiang; Bo, Tao; Yang, Yanfei; Tao, Yong; He, Xiaohong; Li, Daping; Yan, Zhiying

    2016-04-01

    The anaerobic digestion (AD) and microbial electrolysis cell (MEC) coupled system has been proved to be a promising process for biomethane production. In this paper, it was found that by co-cultivating Geobacter with Methanosarcina in an AD-MEC coupled system, methane yield was further increased by 24.1%, achieving to 360.2 mL/g-COD, which was comparable to the theoretical methane yield of an anaerobic digester. With the presence of Geobacter, the maximum chemical oxygen demand (COD) removal rate (216.8 mg COD/(L·hr)) and current density (304.3A/m(3)) were both increased by 1.3 and 1.8 fold compared to the previous study without Geobacter, resulting in overall energy efficiency reaching up to 74.6%. Community analysis demonstrated that Geobacter and Methanosarcina could coexist together in the biofilm, and the electrochemical activities of both were confirmed by cyclic voltammetry. Our study observed that the carbon dioxide content in total gas generated from the AD reactor with Geobacter was only half of that generated from the same reactor without Geobacter, suggesting that Methanosarcina may obtain the electron transferred from Geobacter for the reduction of carbon dioxide to methane. Taken together, Geobacter not only can improve the performance of the MEC system, but also can enhance methane production.

  10. sRNA154 a newly identified regulator of nitrogen fixation in Methanosarcina mazei strain Gö1.

    Science.gov (United States)

    Prasse, Daniela; Förstner, Konrad U; Jäger, Dominik; Backofen, Rolf; Schmitz, Ruth A

    2017-03-15

    Trans-encoded sRNA154 is exclusively expressed under nitrogen (N)-deficiency in Methanosarcina mazei strain Gö1. The sRNA154 deletion strain showed a significant decrease in growth under N-limitation, pointing toward a regulatory role of sRNA154 in N-metabolism. Aiming to elucidate its regulatory function we characterized sRNA154 by means of biochemical and genetic approaches. 24 homologs of sRNA154 were identified in recently reported draft genomes of Methanosarcina strains, demonstrating high conservation in sequence and predicted secondary structure with two highly conserved single stranded loops. Transcriptome studies of sRNA154 deletion mutants by an RNA-seq approach uncovered nifH- and nrpA-mRNA, encoding the α-subunit of nitrogenase and the transcriptional activator of the nitrogen fixation (nif)-operon, as potential targets besides other components of the N-metabolism. Furthermore, results obtained from stability, complementation and western blot analysis, as well as in silico target predictions combined with electrophoretic mobility shift-assays, argue for a stabilizing effect of sRNA154 on the polycistronic nif-mRNA and nrpA-mRNA by binding with both loops. Further identified N-related targets were studied, which demonstrates that translation initiation of glnA2-mRNA, encoding glutamine synthetase2, appears to be affected by sRNA154 masking the ribosome binding site, whereas glnA1-mRNA appears to be stabilized by sRNA154. Overall, we propose that sRNA154 has a crucial regulatory role in N-metabolism in M. mazei by stabilizing the polycistronic mRNA encoding nitrogenase and glnA1-mRNA, as well as allowing a feed forward regulation of nif-gene expression by stabilizing nrpA-mRNA. Consequently, sRNA154 represents the first archaeal sRNA, for which a positive posttranscriptional regulation is demonstrated as well as inhibition of translation initiation.

  11. Single-cell analysis reveals gene-expression heterogeneity in syntrophic dual-culture of Desulfovibrio vulgaris with Methanosarcina barkeri

    Science.gov (United States)

    Qi, Zhenhua; Pei, Guangsheng; Chen, Lei; Zhang, Weiwen

    2014-12-01

    Microbial syntrophic metabolism has been well accepted as the heart of how methanogenic and other anaerobic microbial communities function. In this work, we applied a single-cell RT-qPCR approach to reveal gene-expression heterogeneity in a model syntrophic system of Desulfovibrio vulgaris and Methanosarcina barkeri, as compared with the D. vulgaris monoculture. Using the optimized primers and single-cell analytical protocol, we quantitatively determine gene-expression levels of 6 selected target genes in each of the 120 single cells of D. vulgaris isolated from its monoculture and dual-culture with M. barkeri. The results demonstrated very significant cell-to-cell gene-expression heterogeneity for the selected D. vulgaris genes in both the monoculture and the syntrophic dual-culture. Interestingly, no obvious increase in gene-expression heterogeneity for the selected genes was observed for the syntrophic dual-culture when compared with its monoculture, although the community structure and cell-cell interactions have become more complicated in the syntrophic dual-culture. In addition, the single-cell RT-qPCR analysis also provided further evidence that the gene cluster (DVU0148-DVU0150) may be involved syntrophic metabolism between D. vulgaris and M. barkeri. Finally, the study validated that single-cell RT-qPCR analysis could be a valuable tool in deciphering gene functions and metabolism in mixed-cultured microbial communities.

  12. Establishing a Markerless Genetic Exchange System for Methanosarcina mazei Strain Gö1 for Constructing Chromosomal Mutants of Small RNA Genes

    Directory of Open Access Journals (Sweden)

    Claudia Ehlers

    2011-01-01

    Full Text Available A markerless genetic exchange system was successfully established in Methanosarcina mazei strain Gö1 using the hpt gene coding for hypoxanthine phosphoribosyltransferase. First, a chromosomal deletion mutant of the hpt gene was generated conferring resistance to the purine analog 8-aza-2,6-diaminopurine (8-ADP. The nonreplicating allelic exchange vector (pRS345 carrying the pac-resistance cassette for direct selection of chromosomal integration, and the hpt gene for counterselection was introduced into this strain. By a pop-in and ultimately pop-out event of the plasmid from the chromosome, allelic exchange is enabled. Using this system, we successfully generated a M. mazei deletion mutant of the gene encoding the regulatory non-coding RNA sRNA154. Characterizing M. mazei Δ154 under nitrogen limiting conditions demonstrated differential expression of at least three cytoplasmic proteins and reduced growth strongly arguing for a prominent role of sRNA154 in regulation of nitrogen fixation by posttranscriptional regulation.

  13. Relationship of Intracellular Coenzyme F(420) Content to Growth and Metabolic Activity of Methanobacterium bryantii and Methanosarcina barkeri.

    Science.gov (United States)

    Heine-Dobbernack, E; Schoberth, S M; Sahm, H

    1988-02-01

    The use of F(420) as a parameter for growth or metabolic activity of methanogenic bacteria was investigated. Two representative species of methanogens were grown in batch culture: Methanobacterium bryantii (strain M.o.H.G.) on H(2) and CO(2), and Methanosarcina barkeri (strain Fusaro) on methanol or acetate. The total intracellular content of coenzyme F(420) was followed by high-resolution fluorescence spectroscopy. F(420) concentration in M. bryantii ranged from 1.84 to 3.65 mumol . g of protein; and in M. barkeri grown with methanol it ranged from 0.84 to 1.54 mumol . g depending on growth conditions. The content of F(420) in M. barkeri was influenced by a factor of 2 depending on the composition of the medium (minimal or complex) and by a factor of 3 to 4 depending on whether methanol or acetate was used as the carbon source. A comparison of F(420) content with protein, cell dry weight, optical density, and specific methane production rate showed that the intracellular content of F(420) approximately followed the increase in biomass in both strains. In contrast, no correlation was found between specific methane production rate and intracellular F(420) content. However, qCH(4)(F(420)), calculated by dividing the methane production rate by the coenzyme F(420) concentration, almost paralleled qCH(4)(protein). These results suggest that F(420) may be used as a specific parameter for estimating the biomass, but not the metabolic activity, of methanogens; hence qCH(4)(F(420)) determined in mixed populations with complex carbon substrates must be considered as measure of the actual methanogenic activity and not as a measure of potential activity.

  14. The genes coding for the hsp70(dnaK) molecular chaperone machine occur in the moderate thermophilic archaeon Methanosarcina thermophila TM-1

    DEFF Research Database (Denmark)

    Hofman-Bang, H Jacob Peider; Lange, Marianne; Ahring, Birgitte Kiær

    1999-01-01

    The hsp70 (dnaK) locus of the moderate thermophilic archaeon Methanosarcina thermophila TM-1 was cloned, sequenced, and tested in vitro to measure gene induction by heat and ammonia, i.e., stressors pertinent to the biotechnological ecosystem of this methanogen that plays a key role in anaerobic...... thermoautotrophicum Delta H, from another genus, in which trkA is not part of the locus. The proteins encoded in the TM-1 genes are very similar to the S-6 homologs, but considerably less similar to the Delta H proteins. The TM-1 Hsp70(DnaK) protein has the 23-amino acid deletion-by comparison with homologs from Gram...

  15. Electron paramagnetic resonance spectroscopic and electrochemical characterization of the partially purified N5-methyltetrahydromethanopterin:coenzyme M methyltransferase from Methanosarcina mazei Gö1.

    Science.gov (United States)

    Lu, W P; Becher, B; Gottschalk, G; Ragsdale, S W

    1995-01-01

    The N5-methyltetrahydromethanopterin:coenzyme M methyltransferase is a membrane-bound cobalamin-containing protein of Methanosarcina mazei Gö1 that couples the methylation of coenzyme M by methyltetra-hydrosarcinopterin to the translocation of Na+ across the cell membrane (B. Becher, V. Müller, and G. Gottschalk, J. Bacteriol. 174:7656-7660, 1992). We have partially purified this enzyme and shown that, in addition to the cobamide, at least one iron-sulfur cluster is essential for the transmethylation reaction. The membrane fraction or the partly purified protein contains a "base-on" cobamide with a standard reduction potential (Eo') for the Co2+/1+ couple of -426 mV. The iron-sulfur cluster appears to be a [4Fe-4S]2+/1+ type with an Eo' value of -215 mV. We have determined the methyltransferase activity at various controlled redox potentials and demonstrated that the enzyme activity is activated by a one-electron reduction with half-maximum activity occurring at -235 mV in the presence of ATP and -450 mV in its absence. No activation was observed when ATP was replaced by other nucleoside triphosphates or nonhydrolyzable ATP analogs. PMID:7730249

  16. The stimulating role of subunit F in ATPase activity inside the A1-complex of the Methanosarcina mazei Gö1 A1AO ATP synthase.

    Science.gov (United States)

    Singh, Dhirendra; Sielaff, Hendrik; Sundararaman, Lavanya; Bhushan, Shashi; Grüber, Gerhard

    2016-02-01

    A1AO ATP synthases couple ion-transport of the AO sector and ATP synthesis/hydrolysis of the A3B3-headpiece via their stalk subunits D and F. Here, we produced and purified stable A3B3D- and A3B3DF-complexes of the Methanosarcina mazei Gö1 A-ATP synthase as confirmed by electron microscopy. Enzymatic studies with these complexes showed that the M. mazei Gö1 A-ATP synthase subunit F is an ATPase activating subunit. The maximum ATP hydrolysis rates (Vmax) of A3B3D and A3B3DF were determined by substrate-dependent ATP hydrolysis experiments resulting in a Vmax of 7.9 s(-1) and 30.4 s(-1), respectively, while the KM is the same for both. Deletions of the N- or C-termini of subunit F abolished the effect of ATP hydrolysis activation. We generated subunit F mutant proteins with single amino acid substitutions and demonstrated that the subunit F residues S84 and R88 are important in stimulating ATP hydrolysis. Hybrid formation of the A3B3D-complex with subunit F of the related eukaryotic V-ATPase of Saccharomyces cerevisiae or subunit ε of the F-ATP synthase from Mycobacterium tuberculosis showed that subunit F of the archaea and eukaryotic enzymes are important in ATP hydrolysis.

  17. Dicty_cDB: Contig-U15070-1 [Dicty_cDB

    Lifescience Database Archive (English)

    Full Text Available _2742( AE010299 |pid:none) Methanosarcina acetivorans str.... 37 1.9 AJ006017_1( AJ006017 |pid:none) Hamster...in Kilh... 37 2.5 AJ006016_1( AJ006016 |pid:none) Hamster polyomavirus VP2 gene f

  18. Tetrahydrofolate-specific enzymes in Methanosarcina barkeri and growth dependence of this methanogenic archaeon on folic acid or p-aminobenzoic acid.

    Science.gov (United States)

    Buchenau, Bärbel; Thauer, Rudolf K

    2004-10-01

    Methanogenic archaea are generally thought to use tetrahydromethanopterin or tetrahydrosarcinapterin (H4SPT) rather than tetrahydrofolate (H4F) as a pterin C1 carrier. However, the genome sequence of Methanosarcina species recently revealed a cluster of genes, purN, folD, glyA and metF, that are predicted to encode for H4F-specific enzymes. We show here for folD and glyA from M. barkeri that this prediction is correct: FolD (bifunctional N5,N10-methylene-H4F dehydrogenase/N5,N10-methenyl-H4F cyclohydrolase) and GlyA (serine:H4F hydroxymethyltransferase) were heterologously overproduced in Escherichia coli, purified and found to be specific for methylene-H4F and H4F, respectively (apparent Km below 5 microM). Western blot analyses and enzyme activity measurements revealed that both enzymes were synthesized in M. barkeri. The results thus indicate that M. barkeri should contain H4F, which was supported by the finding that growth of M. barkeri was dependent on folic acid and that the vitamin could be substituted by p-aminobenzoic acid, a biosynthetic precursor of H4F. From the p-aminobenzoic acid requirement, an intracellular H4F concentration of approximately 5 M was estimated. Evidence is presented that the p-aminobenzoic acid taken up by the growing cells was not required for the biosynthesis of H4SPT, which was found to be present in the cells at a concentration above 3 mM. The presence of both H4SPT and H4F in M. barkeri is in agreement with earlier isotope labeling studies indicating that there are two separate C1 pools in these methanogens.

  19. Single-cell analysis of the methanogenic archaeon Methanosarcina soligelidi from Siberian permafrost by means of confocal Raman microspectrocopy for astrobiological research

    Science.gov (United States)

    Serrano, Paloma; Wagner, Dirk; Böttger, Ute; de Vera, Jean-Pierre; Lasch, Peter; Hermelink, Antje

    2014-08-01

    Methanogenic archaea from Siberian permafrost are suitable model organisms that meet many of the preconditions for survival on the martian subsurface. These microorganisms have proven to be highly resistant when exposed to diverse stress factors such as desiccation, radiation and other thermo-physical martian conditions. In addition, the metabolic requirements of methanogenic archaea are in principle compatible with the environmental conditions of the Red Planet. The ExoMars mission will deploy a rover carrying a Raman spectrometer among the analytical instruments in order to search for signatures of life and to investigate the martian geochemistry. Raman spectroscopy is known as a powerful nondestructive optical technique for biosignature detection that requires only little sample preparation. In this study, we describe the use of confocal Raman microspectroscopy (CRM) as a rapid and sensitive technique for characterization of the methanogenic archaeon Methanosarcina soligelidi SMA-21 at the single cell level. These studies involved acquisition of Raman spectra from individual cells isolated from microbial cultures at different stages of growth. Spectral analyses indicated a high degree of heterogeneity between cells of individual cultures and also demonstrated the existence of growth-phase specific Raman patterns. For example, besides common Raman patterns of microbial cells, CRM additionally revealed the presence of lipid vesicles and CaCO3 particles in microbial preparations of M. soligelidi SMA-21, a finding that could be confirmed by electron microscopy. The results of this study suggest that heterogeneity and diversity of microorganisms have to be considered when using Raman-based technologies in future space exploration missions.

  20. Methanosarcina Play an Important Role in Anaerobic Co-Digestion of the Seaweed Ulva lactuca: Taxonomy and Predicted Metabolism of Functional Microbial Communities

    Science.gov (United States)

    FitzGerald, Jamie A.; Allen, Eoin; Wall, David M.; Jackson, Stephen A.; Murphy, Jerry D.; Dobson, Alan D. W.

    2015-01-01

    Macro-algae represent an ideal resource of third generation biofuels, but their use necessitates a refinement of commonly used anaerobic digestion processes. In a previous study, contrasting mixes of dairy slurry and the macro-alga Ulva lactuca were anaerobically digested in mesophilic continuously stirred tank reactors for 40 weeks. Higher proportions of U. lactuca in the feedstock led to inhibited digestion and rapid accumulation of volatile fatty acids, requiring a reduced organic loading rate. In this study, 16S pyrosequencing was employed to characterise the microbial communities of both the weakest (R1) and strongest (R6) performing reactors from the previous work as they developed over a 39 and 27-week period respectively. Comparing the reactor communities revealed clear differences in taxonomy, predicted metabolic orientation and mechanisms of inhibition, while constrained canonical analysis (CCA) showed ammonia and biogas yield to be the strongest factors differentiating the two reactor communities. Significant biomarker taxa and predicted metabolic activities were identified for viable and failing anaerobic digestion of U. lactuca. Acetoclastic methanogens were inhibited early in R1 operation, followed by a gradual decline of hydrogenotrophic methanogens. Near-total loss of methanogens led to an accumulation of acetic acid that reduced performance of R1, while a slow decline in biogas yield in R6 could be attributed to inhibition of acetogenic rather than methanogenic activity. The improved performance of R6 is likely to have been as a result of the large Methanosarcina population, which enabled rapid removal of acetic acid, providing favourable conditions for substrate degradation. PMID:26555136

  1. Methanosarcina Play an Important Role in Anaerobic Co-Digestion of the Seaweed Ulva lactuca: Taxonomy and Predicted Metabolism of Functional Microbial Communities.

    Directory of Open Access Journals (Sweden)

    Jamie A FitzGerald

    Full Text Available Macro-algae represent an ideal resource of third generation biofuels, but their use necessitates a refinement of commonly used anaerobic digestion processes. In a previous study, contrasting mixes of dairy slurry and the macro-alga Ulva lactuca were anaerobically digested in mesophilic continuously stirred tank reactors for 40 weeks. Higher proportions of U. lactuca in the feedstock led to inhibited digestion and rapid accumulation of volatile fatty acids, requiring a reduced organic loading rate. In this study, 16S pyrosequencing was employed to characterise the microbial communities of both the weakest (R1 and strongest (R6 performing reactors from the previous work as they developed over a 39 and 27-week period respectively. Comparing the reactor communities revealed clear differences in taxonomy, predicted metabolic orientation and mechanisms of inhibition, while constrained canonical analysis (CCA showed ammonia and biogas yield to be the strongest factors differentiating the two reactor communities. Significant biomarker taxa and predicted metabolic activities were identified for viable and failing anaerobic digestion of U. lactuca. Acetoclastic methanogens were inhibited early in R1 operation, followed by a gradual decline of hydrogenotrophic methanogens. Near-total loss of methanogens led to an accumulation of acetic acid that reduced performance of R1, while a slow decline in biogas yield in R6 could be attributed to inhibition of acetogenic rather than methanogenic activity. The improved performance of R6 is likely to have been as a result of the large Methanosarcina population, which enabled rapid removal of acetic acid, providing favourable conditions for substrate degradation.

  2. The transcriptional activator NrpA is crucial for inducing nitrogen fixation in Methanosarcina mazei Gö1 under nitrogen-limited conditions.

    Science.gov (United States)

    Weidenbach, Katrin; Ehlers, Claudia; Schmitz, Ruth A

    2014-08-01

    With the aim of unraveling their potential involvement in the regulation of nitrogen metabolism in Methanosarcina mazei strain Gö1, we characterized five genes that are differentially transcribed in response to changing nitrogen availability and encoding putative transcriptional regulators. Study of the respective mutant strains under nitrogen-limited conditions revealed a growth delay for M. mazei MM0444::pac and MM1708::pac, and strongly reduced diazotrophic growth for MM0872::pac, whereas the absence of MM2441 or MM2525 did not affect growth behaviour. Transcriptome analyses further demonstrated that only MM1708 - encoding a CxxCG zinc finger protein - plays a regulatory role in nitrogen metabolism, most likely by specifically enhancing transcription of the N2 fixation (nif) operon under nitrogen-limited conditions. In agreement with this, a palindromic binding motif was predicted in silico in the nifH promoter region, nine nucleotides upstream of the BRE box, and confirmed to bind purified maltose-binding protein-MM1708 by electromobility shift assays. As MM1708 itself is under the control of the global nitrogen repressor NrpR, this adds a secondary level to the transcriptional regulation of the nif genes, and is most likely crucial for maximal nif induction under nitrogen-limited conditions. This is in accordance with the finding that protein expression of NifH is highly reduced in the absence of MM1708 under nitrogen-limited conditions. On the basis of our findings, we hypothesize that, in M. mazei, nitrogen fixation is controlled by a hierarchical network of two transcriptional regulators, the global nitrogen repressor NrpR, and the newly identified activator NrpA (MM1708), thereby providing tight control of N2 fixation.

  3. Nonsense and sense suppression abilities of original and derivative Methanosarcina mazei pyrrolysyl-tRNA synthetase-tRNA(Pyl pairs in the Escherichia coli BL21(DE3 cell strain.

    Directory of Open Access Journals (Sweden)

    Keturah A Odoi

    Full Text Available Systematic studies of nonsense and sense suppression of the original and three derivative Methanosarcina mazei PylRS-tRNA(Pyl pairs and cross recognition between nonsense codons and various tRNA(Pyl anticodons in the Escherichia coli BL21(DE3 cell strain are reported. tRNA(CUA(Pyl is orthogonal in E. coli and able to induce strong amber suppression when it is co-expressed with pyrrolysyl-tRNA synthetase (PylRS and charged with a PylRS substrate, N(ε-tert-butoxycarbonyl-L-lysine (BocK. Similar to tRNA(CUA(Pyl, tRNA(UUA(Pyl is also orthogonal in E. coli and can be coupled with PylRS to genetically incorporate BocK at an ochre mutation site. Although tRNA(UUA(Pyl is expected to recognize a UAG codon based on the wobble hypothesis, the PylRS-tRNA(UUA(Pyl pair does not give rise to amber suppression that surpasses the basal amber suppression level in E. coli. E. coli itself displays a relatively high opal suppression level and tryptophan (Trp is incorporated at an opal mutation site. Although the PylRS-tRNA(UCA(Pyl pair can be used to encode BocK at an opal codon, the pair fails to suppress the incorporation of Trp at the same site. tRNA(CCU(Pyl fails to deliver BocK at an AGG codon when co-expressed with PylRS in E. coli.

  4. The archetype gamma-class carbonic anhydrase (Cam) contains iron when synthesized in vivo.

    Science.gov (United States)

    Macauley, Sheridan R; Zimmerman, Sabrina A; Apolinario, Ethel E; Evilia, Caryn; Hou, Ya-Ming; Ferry, James G; Sowers, Kevin R

    2009-02-10

    A recombinant protein overproduction system was developed in Methanosarcina acetivorans to facilitate biochemical characterization of oxygen-sensitive metalloenzymes from strictly anaerobic species in the Archaea domain. The system was used to overproduce the archetype of the independently evolved gamma-class carbonic anhydrase. The overproduced enzyme was oxygen sensitive and had full incorporation of iron instead of zinc observed when overproduced in Escherichia coli. This, the first report of in vivo iron incorporation for any carbonic anhydrase, supports the need to reevaluate the role of iron in all classes of carbonic anhydrases derived from anaerobic environments.

  5. Trace methane oxidation studied in several Euryarchaeota under diverse conditions

    Directory of Open Access Journals (Sweden)

    James J. Moran

    2005-01-01

    Full Text Available We used 13C-labeled methane to document the extent of trace methane oxidation by Archaeoglobus fulgidus, Archaeoglobus lithotrophicus, Archaeoglobus profundus, Methanobacterium thermoautotrophicum, Methanosarcina barkeri and Methanosarcina acetivorans. The results indicate trace methane oxidation during growth varied among different species and among methanogen cultures grown on different substrates. The extent of trace methane oxidation by Mb. thermoautotrophicum (0.05 ± 0.04%, ± 2 standard deviations of the methane produced during growth was less than that by M. barkeri (0.15 ± 0.04%, grown under similar conditions with H2 and CO2. Methanosarcina acetivorans oxidized more methane during growth on trimethylamine (0.36 ± 0.05% than during growth on methanol (0.07 ± 0.03%. This may indicate that, in M. acetivorans, either a methyltransferase related to growth on trimethylamine plays a role in methane oxidation, or that methanol is an intermediate of methane oxidation. Addition of possible electron acceptors (O2, NO3–, SO22–, SO32– or H2 to the headspace did not substantially enhance or diminish methane oxidation in M. acetivorans cultures. Separate growth experiments with FAD and NAD+ showed that inclusion of these electron carriers also did not enhance methane oxidation. Our results suggest trace methane oxidized during methanogenesis cannot be coupled to the reduction of these electron acceptors in pure cultures, and that the mechanism by which methane is oxidized in methanogens is independent of H2 concentration. In contrast to the methanogens, species of the sulfate-reducing genus Archaeoglobus did not significantly oxidize methane during growth (oxidizing 0.003 ± 0.01% of the methane provided to A. fulgidus, 0.002 ± 0.009% to A. lithotrophicus and 0.003 ± 0.02% to A. profundus. Lack of observable methane oxidation in the three Archaeoglobus species examined may indicate that methyl-coenzyme M reductase, which is not present in

  6. A Method for Identification of Selenoprotein Genes in Archaeal Genomes

    Institute of Scientific and Technical Information of China (English)

    Mingfeng Li; Yanzhao Huang; Yi Xiao

    2009-01-01

    The genetic codon UGA has a dual function: serving as a terminator and encoding selenocysteine. However, most popular gene annotation programs only take it as a stop signal, resulting in misannotation or completely missing selenoprotein genes. We developed a computational method named Asec-Prediction that is specific for the prediction of archaeal selenoprotein genes. To evaluate its effectiveness, we first applied it to 14 archaeal genomes with previously known selenoprotein genes, and Asec-Prediction identified all reported selenoprotein genes without redundant results. When we applied it to 12 archaeal genomes that had not been researched for selenoprotein genes, Asec-Prediction detected a novel selenoprotein gene in Methanosarcina acetivorans. Further evidence was also collected to support that the predicted gene should be a real selenoprotein gene. The result shows that Asec-Prediction is effective for the prediction of archaeal selenoprotein genes.

  7. Dicty_cDB: Contig-U10802-1 [Dicty_cDB

    Lifescience Database Archive (English)

    Full Text Available :none) Burkholderia vietnamiensis G4 ch... 52 5e-11 CP000352_1110( CP000352 |pid:none) Ralstonia metallid...:none) Clostridium botulinum A2 str. K... 67 1e-09 AL939123_131( AL939123 |pid:none) Streptomyces coelicolor A3(2) co...:none) Streptomyces coelicolor A3(2) com... 194 7e-70 CP000083_2628( CP000083 |pid:none) Colwellia psychrerythrae...:none) Methanosarcina acetivorans str. ... 60 8e-14 CP000036_2527( CP000036 |pid:none) Shigella boydii Sb227, co...:none) Clostridium difficile 630 compl... 60 7e-15 CP001230_873( CP001230 |pid:none) Persephonella marina

  8. Genome-scale Metabolic Reaction Modeling: a New Approach to Geomicrobial Kinetics

    Science.gov (United States)

    McKernan, S. E.; Shapiro, B.; Jin, Q.

    2014-12-01

    Geomicrobial rates, rates of microbial metabolism in natural environments, are a key parameter of theoretical and practical problems in geobiology and biogeochemistry. Both laboratory- and field-based approaches have been applied to study rates of geomicrobial processes. Laboratory-based approaches analyze geomicrobial kinetics by incubating environmental samples under controlled laboratory conditions. Field methods quantify geomicrobial rates by observing the progress of geomicrobial processes. To take advantage of recent development in biogeochemical modeling and genome-scale metabolic modeling, we suggest that geomicrobial rates can also be predicted by simulating metabolic reaction networks of microbes. To predict geomicrobial rates, we developed a genome-scale metabolic model that describes enzyme reaction networks of microbial metabolism, and simulated the network model by accounting for the kinetics and thermodynamics of enzyme reactions. The model is simulated numerically to solve cellular enzyme abundance and hence metabolic rates under the constraints of cellular physiology. The new modeling approach differs from flux balance analysis of system biology in that it accounts for the thermodynamics and kinetics of enzymatic reactions. It builds on subcellular metabolic reaction networks, and hence also differs from classical biogeochemical reaction modeling. We applied the new approach to Methanosarcina acetivorans, an anaerobic, marine methanogen capable of disproportionating acetate to carbon dioxide and methane. The input of the new model includes (1) enzyme reaction network of acetoclastic methanogenesis, and (2) representative geochemical conditions of freshwater sedimentary environments. The output of the simulation includes the proteomics, metabolomics, and energy and matter fluxes of M. acetivorans. Our simulation results demonstrate the predictive power of the new modeling approach. Specifically, the results illustrate how methanogenesis rates vary

  9. Effects of the elemental layer thickness on the properties of Fe/Co grown at 200 °C

    Energy Technology Data Exchange (ETDEWEB)

    Carbucicchio, M., E-mail: massimo.carbucicchio@fis.unipr.it [Physics Department, University of Parma, Parco Area delle Scienze 7/A, 43124 Parma (Italy); Ciprian, R. [Physics Department, University of Parma, Parco Area delle Scienze 7/A, 43124 Parma (Italy); Nasi, L. [CNR-IMEM Institute, Parco Area delle Scienze 37/A, 43124 Parma (Italy)

    2013-09-30

    Thin Fe/Co multilayers were grown at 200 °C onto glass and naturally oxidized Si substrates, changing the elemental layer thickness. Onto glass substrates, the multilayers show a large in-plane uniaxial magnetocrystalline anisotropy, which strengthens by increasing the Fe layer thickness. Onto naturally oxidized Si substrates, an appreciable out-of-plane contribution to the magnetization vector is present. This can be due to the absence in the multilayer stack of a pure-Co layer as a consequence of a large intermixing occurring at the Fe/Co interfaces, that gives rise to a structure only constituted by intercalated Fe and FeCo layers. However, by increasing the Co and Fe layer thickness, the intermixing lowers because of a change in the sample morphology and microstructure, which determines the disappearance of the out-of-plane tilting of the magnetization vector while promoting the establishing of an in-plane anisotropy. - Highlights: ► Fe/Co multilayers were e-beam evaporated onto glass and Si substrates at 200 °C. ► The elemental layer thickness was changed. On glass, large in-plane anisotropy ► Onto Si, perpendicular anisotropy because of a large intermixing ► The perpendicular anisotropy is due to the absence of a pure Co layer. ► The type of in-plane anisotropy is controlled by the Fe/Co thickness ratio.

  10. Stable carbon isotope fractionation of six strongly fractionating microorganisms is not affected by growth temperature under laboratory conditions

    Science.gov (United States)

    Penger, Jörn; Conrad, Ralf; Blaser, Martin

    2014-09-01

    Temperature is the major driving force for many biological as well as chemical reactions and may impact the fractionation of stable carbon isotopes. Thus, a good correlation between temperature and fractionation is observed in many chemical systems that are controlled by an equilibrium isotope effect. In contrast, biological systems that are usually controlled by a kinetic isotope effect are less well studied with respect to temperature effects and have shown contrasting results. We studied three different biological pathways (methylotrophic methanogenesis, hydrogenotrophic methanogenesis, acetogenesis by the acetyl-CoA pathway) which are characterized by very strong carbon isotope enrichment factors (-50‰ to -83‰). The microorganisms (Methanosarcina barkeri, Methanosarcina acetivorans, Methanolobus zinderi, Methanothermobacter marburgensis, Methanothermobacter thermoautotrophicus, Thermoanaerobacter kivui) exhibiting these pathways were grown at different temperatures ranging between 25 and 68 °C, and the fractionation factors were determined from 13C/12C isotope discrimination during substrate depletion and product formation. Our experiments showed that the fractionation factors were different for the different metabolic pathways but were not much affected by the different growth temperatures. Slight variations were well within the standard errors of replication and regression analysis. Our results showed that temperature had no significant effect on the fractionation of stable carbon isotopes during anaerobic microbial metabolism with relatively strong isotope fractionation.

  11. New Perspectives on Acetate and One-Carbon Metabolism in the Methanoarchaea

    Energy Technology Data Exchange (ETDEWEB)

    Ferry, James [Pennsylvania State Univ., University Park, PA (United States)

    2017-03-20

    Carbonic anhydrases catalyze the reversible hydration of carbon dioxide to bicarbonate. Although widespread in prokaryotes of the domains Bacteria and Archaea, few have been investigated and the physiological functions are largely unknown. Carbonic anhydrases are of biotechnological interest for carbon dioxide capture and sequestration at point sources. Prokaryotes encode three independently evolved classes. The alpha-class is restricted to a few pathogens and the other two are uniformly distributed in phylogenetically and physiologically diverse species. Although wide-spread in prokaryotes, only three gamma-class enzymes have been biochemically characterized and the physiological functions have not been investigated. The gamma-class is prominent in anaerobic acetate-utilizing methane-producing species of the genus Methanosarcina that encode three subclasses. Enzymes from two of the subclasses, Cam and CamH from Methanosarcina thermophila, have been characterized and found to utilize iron in the active site which is the first example of an iron-containing carbonic anhydrase. No representative of the third subclass has been isolated, although this subclass constitutes the great majority of the β-class. This grant application proposed to characterize gamma-class carbonic anhydrases from diverse anaerobic prokaryotes from the domains Bacteria and Archaea to broaden the understanding of this enzyme. In particular, the three subclasses present the genetically tractable acetate-utilizing methanogen Methanosarcina acetivorans will be investigated to extend studies of acetate and one-carbon metabolism in this species. A genetic approach will be taken to ascertain the physiological functions. It is also proposed to delve deeper into the mechanism of Cam from M. thermophila, the archetype of the gamma-class, via a high resolution neutron structure and kinetic analysis of site-specific amino acid replacement variants. In the course of the investigation, goals were added to

  12. Reducing the genetic code induces massive rearrangement of the proteome.

    Science.gov (United States)

    O'Donoghue, Patrick; Prat, Laure; Kucklick, Martin; Schäfer, Johannes G; Riedel, Katharina; Rinehart, Jesse; Söll, Dieter; Heinemann, Ilka U

    2014-12-02

    Expanding the genetic code is an important aim of synthetic biology, but some organisms developed naturally expanded genetic codes long ago over the course of evolution. Less than 1% of all sequenced genomes encode an operon that reassigns the stop codon UAG to pyrrolysine (Pyl), a genetic code variant that results from the biosynthesis of Pyl-tRNA(Pyl). To understand the selective advantage of genetically encoding more than 20 amino acids, we constructed a markerless tRNA(Pyl) deletion strain of Methanosarcina acetivorans (ΔpylT) that cannot decode UAG as Pyl or grow on trimethylamine. Phenotypic defects in the ΔpylT strain were evident in minimal medium containing methanol. Proteomic analyses of wild type (WT) M. acetivorans and ΔpylT cells identified 841 proteins from >7,000 significant peptides detected by MS/MS. Protein production from UAG-containing mRNAs was verified for 19 proteins. Translation of UAG codons was verified by MS/MS for eight proteins, including identification of a Pyl residue in PylB, which catalyzes the first step of Pyl biosynthesis. Deletion of tRNA(Pyl) globally altered the proteome, leading to >300 differentially abundant proteins. Reduction of the genetic code from 21 to 20 amino acids led to significant down-regulation in translation initiation factors, amino acid metabolism, and methanogenesis from methanol, which was offset by a compensatory (100-fold) up-regulation in dimethyl sulfide metabolic enzymes. The data show how a natural proteome adapts to genetic code reduction and indicate that the selective value of an expanded genetic code is related to carbon source range and metabolic efficiency.

  13. 产甲烷的常温古细菌和嗜热古细菌的代谢网络比对研究%Studies on the Metabolic Network Alignment of Mesophilic and Thermophilic Methanogenic Archaea

    Institute of Scientific and Technical Information of China (English)

    陈璟; 须文波

    2015-01-01

    生物网络比对是生物体的结构、功能和进化分析的重要研究手段.以从KFGG数据库获得的产甲烷的常温古细菌Methanosarcina acetivorans(M.acetivorans)和嗜热古细菌Methanopyrus kandleri(M.kandleri)的代谢网络为对象,采用了网络比对算法Matching-based Integrative GRAph Aligner(MI-GRAAL)对它们的全局代谢网络以及hub模块网络进行了比对.比对结果表明采用度、聚集系数以及离心率三个度量参数相结合的网络比对结果明显优于其它度量参数的计算结果,且结果更稳定.同时发现常温产甲烷菌M.acetivorans的hub模块与嗜热产甲烷菌M.kandleri的hub模块相似代谢途径的拓扑基本一致,不相似的代谢网络中有81.8%以上的节点都在嗜热产甲烷菌M.kandleri的最紧密的7-核中,推测嗜热菌的耐热性可能与受到胞内酪氨酸的影响.

  14. A Ferredoxin- and F420H2-Dependent, Electron-Bifurcating, Heterodisulfide Reductase with Homologs in the Domains Bacteria and Archaea

    Science.gov (United States)

    Yan, Zhen

    2017-01-01

    ABSTRACT Heterodisulfide reductases (Hdr) of the HdrABC class are ancient enzymes and a component of the anaerobic core belonging to the prokaryotic common ancestor. The ancient origin is consistent with the widespread occurrence of genes encoding putative HdrABC homologs in metabolically diverse prokaryotes predicting diverse physiological functions; however, only one HdrABC has been characterized and that was from a narrow metabolic group of obligate CO2-reducing methanogenic anaerobes (methanogens) from the domain Archaea. Here we report the biochemical characterization of an HdrABC homolog (HdrA2B2C2) from the acetate-utilizing methanogen Methanosarcina acetivorans with unusual properties structurally and functionally distinct from the only other HdrABC characterized. Homologs of the HdrA2B2C2 archetype are present in phylogenetically and metabolically diverse species from the domains Bacteria and Archaea. The expression of the individual HdrA2, HdrB2, and HdrB2C2 enzymes in Escherichia coli, and reconstitution of an active HdrA2B2C2 complex, revealed an intersubunit electron transport pathway dependent on ferredoxin or coenzyme F420 (F420H2) as an electron donor. Remarkably, HdrA2B2C2 couples the previously unknown endergonic oxidation of F420H2 and reduction of ferredoxin with the exergonic oxidation of F420H2 and reduction of the heterodisulfide of coenzyme M and coenzyme B (CoMS-SCoB). The unique electron bifurcation predicts a role for HdrA2B2C2 in Fe(III)-dependent anaerobic methane oxidation (ANME) by M. acetivorans and uncultured species from ANME environments. HdrA2B2C2, ubiquitous in acetotrophic methanogens, was shown to participate in electron transfer during acetotrophic growth of M. acetivorans and proposed to be essential for growth in the environment when acetate is limiting. PMID:28174314

  15. Enzymology of the Pathway for Acetate Conversion to Methane in Methanosarcina thermophilia

    Energy Technology Data Exchange (ETDEWEB)

    Ferry, James G.

    1999-05-04

    These topics are covered: Regulation of enzyme synthesis; Activation of acetate to acetyl-CoA; Biochemistry of acetyl-CoA cleavage; Electron transport; Other enzymes implicated in the pathway of acetate conversion to methane; and publications resulting from this work.

  16. Relationships between anaerobic consortia and removal efficiencies in an UASB reactor degrading 2,4 dichlorophenol (DCP).

    Science.gov (United States)

    Sponza, Delia Teresa; Cigal, Canan

    2008-04-01

    To gain more insight into the interactions between anaerobic bacteria and reactor performances (chemical oxygen demand-COD, 2,4 dichlorophenol-2,4 DCP removals, volatile fatty acid-VFA, and methane gas productions) and how they depended on operational conditions the microbial variations in the anaerobic granular sludge from an upflow anaerobic sludge blanket (UASB) reactor treating 2,4 DCP was studied. The study was composed of two parts. In the first part, the numbers of methanogens and acedogens in the anaerobic granular sludge were counted at different COD removal efficiencies. The relationships between the numbers of methanogens, the methane gas production and VFA production were investigated. The COD removal efficiencies increased to 74% from 30% while the number of total acedogens decreased to 10 from 30 cfu ml(-1). The number of total methanogens and acedogens varied between 11 x 10(3) and 10 x 10(9)MPN g(-1) and 10 and 30 cfu ml(-1) as the 2,4 DCP removal efficiencies were obtained between 60% and 99%, respectively. It was seen that, as the number of total acedogens decreased, the COD removal efficiencies increased. However, the number of total methanogens increased as the COD removal efficiencies increased. Correlations between the bacterial number and with the removal efficiencies obtained in different operational conditions were investigated. From the results presented in this paper a high correlation between the number of bacteria, COD removals, methane gas percentage, 2,4 DCP removals and VFA was observed. In the second part, methanogen bacteria in the anaerobic granular sludge were identified. Microbial observations and biochemical tests were applied to identify the anaerobic microorganisms from the anaerobic granular sludge. In the reactor treating 2,4 DCP, Methanobacterium bryantii, Methanobacterium formicicum, Methanobrevibacter smithii, Methanococcus voltae, Methanosarcina mazei, Methanosarcina acetivorans, Methanogenium bourgense and

  17. The stepwise evolution of early life driven by energy conservation.

    Science.gov (United States)

    Ferry, James G; House, Christopher H

    2006-06-01

    Two main theories have emerged for the origin and early evolution of life based on heterotrophic versus chemoautotrophic metabolisms. With the exception of a role for CO, the theories have little common ground. Here we propose an alternative theory for the early evolution of the cell which combines principal features of the widely disparate theories. The theory is based on the extant pathway for conversion of CO to methane and acetate, largely deduced from the genomic analysis of the archaeon Methanosarcina acetivorans. In contrast to current paradigms, we propose that an energy-conservation pathway was the major force which powered and directed the early evolution of the cell. We envision the proposed primitive energy-conservation pathway to have developed sometime after a period of chemical evolution but prior to the establishment of diverse protein-based anaerobic metabolisms. We further propose that energy conservation played the predominant role in the later evolution of anaerobic metabolisms which explains the origin and evolution of extant methanogenic pathways.

  18. Isotope fractionation during the anaerobic consumption of acetate by methanogenic and sulfate-reducing microorganisms

    Science.gov (United States)

    Gövert, D.; Conrad, R.

    2009-04-01

    During the anaerobic degradation of organic matter in anoxic sediments and soils acetate is the most important substrate for the final step in production of CO2 and/or CH4. Sulfate-reducing bacteria (SRB) and methane-producing archaea both compete for the available acetate. Knowledge about the fractionation of 13C/12C of acetate carbon by these microbial groups is still limited. Therefore, we determined carbon isotope fractionation in different cultures of acetate-utilizing SRB (Desulfobacter postgatei, D. hydrogenophilus, Desulfobacca acetoxidans) and methanogens (Methanosarcina barkeri, M. acetivorans). Including literature values (e.g., Methanosaeta concilii), isotopic enrichment factors (epsilon) ranged between -35 and +2 permil, possibly involving equilibrium isotope effects besides kinetic isotope effects. The values of epsilon were dependent on the acetate-catabolic pathway of the particular microorganism, the methyl or carboxyl position of acetate, and the relative availability or limitation of the substrate acetate. Patterns of isotope fractionation in anoxic lake sediments and rice field soil seem to reflect the characteristics of the microorganisms actively involved in acetate catabolism. Hence, it might be possible using environmental isotopic information to determine the type of microbial metabolism converting acetate to CO2 and/or CH4.

  19. Towards a Computational Model of a Methane Producing Archaeum

    Directory of Open Access Journals (Sweden)

    Joseph R. Peterson

    2014-01-01

    Full Text Available Progress towards a complete model of the methanogenic archaeum Methanosarcina acetivorans is reported. We characterized size distribution of the cells using differential interference contrast microscopy, finding them to be ellipsoidal with mean length and width of 2.9 μm and 2.3 μm, respectively, when grown on methanol and 30% smaller when grown on acetate. We used the single molecule pull down (SiMPull technique to measure average copy number of the Mcr complex and ribosomes. A kinetic model for the methanogenesis pathways based on biochemical studies and recent metabolic reconstructions for several related methanogens is presented. In this model, 26 reactions in the methanogenesis pathways are coupled to a cell mass production reaction that updates enzyme concentrations. RNA expression data (RNA-seq measured for cell cultures grown on acetate and methanol is used to estimate relative protein production per mole of ATP consumed. The model captures the experimentally observed methane production rates for cells growing on methanol and is most sensitive to the number of methyl-coenzyme-M reductase (Mcr and methyl-tetrahydromethanopterin:coenzyme-M methyltransferase (Mtr proteins. A draft transcriptional regulation network based on known interactions is proposed which we intend to integrate with the kinetic model to allow dynamic regulation.

  20. Enhancement of bioenergy production from organic wastes by two-stage anaerobic hydrogen and methane production process.

    Science.gov (United States)

    Luo, Gang; Xie, Li; Zhou, Qi; Angelidaki, Irini

    2011-09-01

    The present study investigated a two-stage anaerobic hydrogen and methane process for increasing bioenergy production from organic wastes. A two-stage process with hydraulic retention time (HRT) 3d for hydrogen reactor and 12d for methane reactor, obtained 11% higher energy compared to a single-stage methanogenic process (HRT 15 d) under organic loading rate (OLR) 3 gVS/(L d). The two-stage process was still stable when the OLR was increased to 4.5 gVS/(Ld), while the single-stage process failed. The study further revealed that by changing the HRT(hydrogen):HRT(methane) ratio of the two-stage process from 3:12 to 1:14, 6.7%, more energy could be obtained. Microbial community analysis indicated that the dominant bacterial species were different in the hydrogen reactors (Thermoanaerobacterium thermosaccharolyticum-like species) and methane reactors (Clostridium thermocellum-like species). The changes of substrates and HRT did not change the dominant species. The archaeal community structures in methane reactors were similar both in single- and two- stage reactors, with acetoclastic methanogens Methanosarcina acetivorans-like organisms as the dominant species.

  1. Methanosarcina soligelidi sp. nov., a desiccation- and freeze-thaw-resistant methanogenic archaeon from a Siberian permafrost-affected soil

    National Research Council Canada - National Science Library

    Wagner, Dirk; Schirmack, Janosch; Ganzert, Lars; Morozova, Daria; Mangelsdorf, Kai

    2013-01-01

    .... Optimal growth was observed at 28 °C, pH 7.8 and 0.02 M NaCl. The strain grew on H2/CO2, methanol and acetate, but not on formate, ethanol, 2-butanol, 2-propanol, monomethylamine, dimethylamine, trimethylamine or dimethyl sulfide...

  2. Design of carrier tRNAs and selection of four-base codons for efficient incorporation of various nonnatural amino acids into proteins in Spodoptera frugiperda 21 (Sf21) insect cell-free translation system.

    Science.gov (United States)

    Taki, Masumi; Tokuda, Yasunori; Ohtsuki, Takashi; Sisido, Masahiko

    2006-12-01

    Spodoptera frugiperda 21 (Sf21) insect cell-free protein synthesizing system was expanded to include nonnatural amino acids. Orthogonal tRNAs that work as carriers of nonnatural amino acids in the insect system were explored. Four-base codons for assigning the positions of nonnatural amino acids were also selected. Mutated streptavidin mRNAs that contained different four-base codons were prepared and added to the insect cell-free system in the presence of various tRNAs possessing the corresponding four-base anticodons. The tRNAs were chemically aminoacylated with various types of nonnatural amino acids to examine their incorporation efficiencies. Using p-nitrophenylalanine as the nonnatural amino acid and streptavidin as the target protein, tRNA sequences and the types of four-base codons were optimized to maximize the yield of the nonnatural mutant and to minimize production of full-length proteins that do not contain the nonnatural amino acid. Among the tRNA sequences taken from a variety of tRNAs of nonstandard structures, the tRNA derived from Methanosarcina acetivorans tRNA(Pyl) was the most efficient and orthogonal tRNA. Of the CGGN-type four-base codons, CGGA and CGGG were the most efficient ones for assigning the positions of nonnatural amino acids. p-Nitrophenylalanine and 2-naphthylalanine were efficiently incorporated as in the case of Escherichia coli and rabbit reticulocyte cell-free systems. Much less efficient incorporation was observed, however, for other nonnatural amino acids, indicating that the insect system is less tolerant to the structural diversity of amino acids than the E. coli cell-free system.

  3. Betaine: New Oxidant in the Stickland Reaction and Methanogenesis from Betaine and l-Alanine by a Clostridium sporogenes-Methanosarcina barkeri Coculture

    OpenAIRE

    Naumann, Evelyn; Hippe, Hans; Gottschalk, Gerhard

    1983-01-01

    Growing and nongrowing cells of Clostridium sporogenes fermented betaine with l-alanine, l-valine, l-leucine, and l-isoleucine as electron donors in a coupled oxidation-reduction reaction (Stickland reaction). For the substrate combinations betaine and l-alanine and betaine and l-valine balance studies were performed; the results were in agreement with the following fermentation equation: 1 R- CH(NH2)-COOH + 2 betaine + 2 H2O → 1 R-COOH + 1 CO2 + 1 NH3 + 2 trimethylamine + 2 acetate. Growth a...

  4. Biophysical Characterisation of Globins and Multi-Heme Cytochromes Using Electron Paramagnetic Resonance and Optical Spectroscopy

    Science.gov (United States)

    Desmet, Filip

    Heme proteins of different families were investigated in this work, using a combination of pulsed and continuous-wave electron paramagnetic resonance (EPR) spectroscopy, optical absorption spectroscopy, resonance Raman spectroscopy and laser flash photolysis. The first class of proteins that were investigated, were the globins. The globin-domain of the globin-coupled sensor of the bacterium Geobacter sulfurreducens was studied in detail using different pulsed EPR techniques (HYSCORE and Mims ENDOR). The results of this pulsed EPR study are compared with the results of the optical investigation and the crystal structure of the protein. The second globin, which was studied, is the Protoglobin of Methanosarcina acetivorans, various mutants of this protein were studied using laser flash photolysis and Raman spectroscopy to unravel the link between this protein's unusual structure and its ligand-binding kinetics. In addition to this, the CN -bound form of this protein was investigated using EPR and the influence of the strong deformation of the heme on the unusual low gz values is discussed. Finally, the neuroglobins of three species of fishes, Danio rerio, Dissostichus mawsoni and Chaenocephalus aceratus are studied. The influence of the presence or absence of two cysteine residues in the C-D and D-region of the protein on the EPR spectrum, and the possible formation of a disulfide bond is studied. The second group of proteins that were studied in this thesis belong to the family of the cytochromes. First the Mouse tumor suppressor cytochrome b561 was studied, the results of a Raman and EPR investigation are compared to the Human orthologue of the protein. Secondly, the tonoplast cytochrome b561 of Arabidopsis was investigated in its natural form and in two double-mutant forms, in which the heme at the extravesicular side was removed. The results of this investigation are then compared with two models in literature that predict the localisation of the hemes in this

  5. Computational Modeling of Fluctuations in Energy and Metabolic Pathways of Methanogenic Archaea

    Energy Technology Data Exchange (ETDEWEB)

    Luthey-Schulten, Zaida [Univ. of Illinois, Urbana-Champaign, IL (United States). Dept. of Chemistry; Carl R. Woese Inst. for Genomic Biology

    2017-01-04

    investigations of the methanogen Methanosarcina acetivorans. By integrating an unprecedented transcriptomics dataset for growth of the methanogen on many substrates with an in silico model, heterogeneity in metabolic pathway usage and methane production were examined. This lent insight into the physiological requirements of the organism under different environmental conditions and uncovered the unique regulatory role that mRNA half-life has in shaping metabolic flux distributions in this organism.

  6. NCBI nr-aa BLAST: CBRC-ACAR-01-0779 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-ACAR-01-0779 ref|NP_632766.1| Arsenical-resistance protein [Methanosarcina maz...ei Go1] gb|AAM30438.1| Arsenical-resistance protein [Methanosarcina mazei Go1] NP_632766.1 0.39 30% ...

  7. NCBI nr-aa BLAST: CBRC-AGAM-02-0012 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-AGAM-02-0012 ref|YP_304864.1| hypothetical protein Mbar_A1321 [Methanosarcina barker...i str. Fusaro] gb|AAZ70284.1| hypothetical protein Mbar_A1321 [Methanosarcina barkeri str. Fusaro] YP_304864.1 0.39 34% ...

  8. NCBI nr-aa BLAST: CBRC-DMEL-02-0058 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DMEL-02-0058 ref|YP_305810.1| leucine-rich-repeat protein [Methanosarcina barker...i str. Fusaro] gb|AAZ71230.1| leucine-rich-repeat protein [Methanosarcina barkeri str. Fusaro] YP_305810.1 6e-19 33% ...

  9. NCBI nr-aa BLAST: CBRC-TTRU-01-0874 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-TTRU-01-0874 ref|YP_305025.1| hypothetical protein Mbar_A1490 [Methanosarcina barker...i str. Fusaro] gb|AAZ70445.1| hypothetical protein Mbar_A1490 [Methanosarcina barkeri str. Fusaro] YP_305025.1 0.001 21% ...

  10. NCBI nr-aa BLAST: CBRC-DNOV-01-2526 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DNOV-01-2526 ref|YP_306419.1| hypothetical protein Mbar_A2944 [Methanosarcina barker...i str. Fusaro] gb|AAZ71839.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_306419.1 0.072 19% ...

  11. NCBI nr-aa BLAST: CBRC-DYAK-02-0045 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DYAK-02-0045 ref|YP_305810.1| leucine-rich-repeat protein [Methanosarcina barker...i str. Fusaro] gb|AAZ71230.1| leucine-rich-repeat protein [Methanosarcina barkeri str. Fusaro] YP_305810.1 2e-18 32% ...

  12. NCBI nr-aa BLAST: CBRC-DSIM-02-0062 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DSIM-02-0062 ref|YP_305810.1| leucine-rich-repeat protein [Methanosarcina barker...i str. Fusaro] gb|AAZ71230.1| leucine-rich-repeat protein [Methanosarcina barkeri str. Fusaro] YP_305810.1 4e-21 34% ...

  13. NCBI nr-aa BLAST: CBRC-AGAM-04-0028 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-AGAM-04-0028 ref|YP_305972.1| hypothetical protein Mbar_A2479 [Methanosarcina barker...i str. Fusaro] gb|AAZ71392.1| hypothetical protein Mbar_A2479 [Methanosarcina barkeri str. Fusaro] YP_305972.1 9e-19 35% ...

  14. NCBI nr-aa BLAST: CBRC-CFAM-29-0004 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-CFAM-29-0004 ref|YP_305025.1| hypothetical protein Mbar_A1490 [Methanosarcina barker...i str. Fusaro] gb|AAZ70445.1| hypothetical protein Mbar_A1490 [Methanosarcina barkeri str. Fusaro] YP_305025.1 1.4 26% ...

  15. NCBI nr-aa BLAST: CBRC-DDIS-04-0035 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DDIS-04-0035 ref|YP_303708.1| hypothetical protein Mbar_A0143 [Methanosarcina barker...i str. Fusaro] gb|AAZ69128.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_303708.1 1e-10 42% ...

  16. NCBI nr-aa BLAST: CBRC-DMEL-01-0076 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DMEL-01-0076 ref|YP_306433.1| hypothetical protein Mbar_A2958 [Methanosarcina barker...i str. Fusaro] gb|AAZ71853.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_306433.1 3.6 30% ...

  17. NCBI nr-aa BLAST: CBRC-ACAR-01-0286 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-ACAR-01-0286 ref|YP_304217.1| hypothetical protein Mbar_A0658 [Methanosarcina barker...i str. Fusaro] gb|AAZ69637.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_304217.1 0.19 25% ...

  18. NCBI nr-aa BLAST: CBRC-DYAK-02-0000 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-DYAK-02-0000 ref|YP_306433.1| hypothetical protein Mbar_A2958 [Methanosarcina barker...i str. Fusaro] gb|AAZ71853.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_306433.1 0.029 34% ...

  19. NCBI nr-aa BLAST: CBRC-OPRI-01-1140 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-OPRI-01-1140 ref|YP_303598.1| hypothetical protein Mbar_A0027 [Methanosarcina barker...i str. Fusaro] gb|AAZ69018.1| hypothetical protein Mbar_A0027 [Methanosarcina barkeri str. Fusaro] YP_303598.1 0.078 24% ...

  20. NCBI nr-aa BLAST: CBRC-CJAC-01-1179 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-CJAC-01-1179 ref|YP_305972.1| hypothetical protein Mbar_A2479 [Methanosarcina barker...i str. Fusaro] gb|AAZ71392.1| hypothetical protein Mbar_A2479 [Methanosarcina barkeri str. Fusaro] YP_305972.1 1e-09 31% ...

  1. NCBI nr-aa BLAST: CBRC-XTRO-01-2145 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-XTRO-01-2145 ref|YP_305913.1| hypothetical protein Mbar_A2413 [Methanosarcina barker...i str. Fusaro] gb|AAZ71333.1| conserved hypothetical protein [Methanosarcina barkeri str. Fusaro] YP_305913.1 0.12 24% ...

  2. NCBI nr-aa BLAST: CBRC-XTRO-01-1006 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-XTRO-01-1006 ref|YP_306263.1| cell surface protein [Methanosarcina barkeri str.... Fusaro] gb|AAZ71683.1| cell surface protein [Methanosarcina barkeri str. Fusaro] YP_306263.1 0.032 22% ...

  3. NCBI nr-aa BLAST: CBRC-ACAR-01-0339 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available CBRC-ACAR-01-0339 ref|YP_300087.1| hypothetical protein Mbar_B3757 [Methanosarcina barker...i str. Fusaro] gb|AAZ68991.1| hypothetical protein Mbar_B3757 [Methanosarcina barkeri str. Fusaro] YP_300087.1 1.3 24% ...

  4. Biotic and abiotic dynamics of a high solid-state anaerobic digestion box-type container system.

    Science.gov (United States)

    Walter, Andreas; Probst, Maraike; Hinterberger, Stephan; Müller, Horst; Insam, Heribert

    2016-03-01

    A solid-state anaerobic digestion box-type container system for biomethane production was observed in 12 three-week batch fermentations. Reactor performance was monitored using physico-chemical analysis and the methanogenic community was identified using ANAEROCHIP-microarrays and quantitative PCR. A resilient community was found in all batches, despite variations in inoculum to substrate ratio, feedstock quality, and fluctuating reactor conditions. The consortia were dominated by mixotrophic Methanosarcina that were accompanied by hydrogenotrophic Methanobacterium, Methanoculleus, and Methanocorpusculum. The relationship between biotic and abiotic variables was investigated using bivariate correlation analysis and univariate analysis of variance. High amounts of biogas were produced in batches with high copy numbers of Methanosarcina. High copy numbers of Methanocorpusculum and extensive percolation, however, were found to negatively correlate with biogas production. Supporting these findings, a negative correlation was detected between Methanocorpusculum and Methanosarcina. Based on these results, this study suggests Methanosarcina as an indicator for well-functioning reactor performance.

  5. Green house gas emissions from termite ecosystem

    African Journals Online (AJOL)

    USER

    Key words: Macrotermes, methane, carbondioxide, GHG, methanobacteria, methanosarcina. INTRODUCTION ... from its natural role of conversion of woody and cellulosic ..... responses to the presence of oxygen and their sensitivity to it vary ...

  6. Threshold Acetate Concentrations for Acetate Catabolism by Aceticlastic Methanogenic Bacteria

    OpenAIRE

    Westermann, Peter; Ahring, Birgitte K.; Mah, Robert A.

    1989-01-01

    Marked differences were found for minimum threshold concentrations of acetate catabolism by Methanosarcina barkeri 227 (1.180 mM), Methanosarcina mazei S-6 (0.396 mM), and a Methanothrix sp. (0.069 mM). This indicates that the aceticlastic methanogens responsible for the conversion of acetate to methane in various ecosystems might be different, depending on the prevailing in situ acetate concentrations.

  7. Microbial ecology of thermophilic anaerobic digestion. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Stephen H. Zinder

    2000-04-15

    This grant supported research on methanogenic archaea. The two major areas that were supported were conversion of acetic acid to methane and nitrogen fixation by Methanosarcina. Among the achievements of this research were the isolation of novel methanogenic cultures, elucidation of the pathways from acetate to methane, description of a specific DNA-binding complex in nitrogen fixing methanogens, and demonstration of an alternative nitrogenase in Methanosarcina.

  8. Gene Regulation of Methanogenesis from Acetate in the Acetotrophic Methane Producing Archaebacteria.

    Science.gov (United States)

    1988-06-27

    planned to use CO dehydrogenase from 6 M. thermophila provided by J.G. Ferry to produce probes for genomic libraries of M. acetivorans on the assumption...regions of the proteins are conserved. 4) Antibody production and screening. M. thermophila genomic libraries were screened with antisera directed...be used in construction of gene shuttle vectors with the evencual goal of developing an gene transformation system, and iii) generation of genomic

  9. Final Technical Report for Award # ER64999

    Energy Technology Data Exchange (ETDEWEB)

    Metcalf, William W. [University of Illinois

    2014-10-08

    This report provides a summary of activities for Award # ER64999, a Genomes to Life Project funded by the Office of Science, Basic Energy Research. The project was entitled "Methanogenic archaea and the global carbon cycle: a systems biology approach to the study of Methanosarcina species". The long-term goal of this multi-investigator project was the creation of integrated, multiscale models that accurately and quantitatively predict the role of Methanosarcina species in the global carbon cycle under dynamic environmental conditions. To achieve these goals we pursed four specific aims: (1) genome sequencing of numerous members of the Order Methanosarcinales, (2) identification of genomic sources of phenotypic variation through in silico comparative genomics, (3) elucidation of the transcriptional networks of two Methanosarcina species, and (4) development of comprehensive metabolic network models for characterized strains to address the question of how metabolic models scale with genetic distance.

  10. Thermophilic anaerobic acetate-utilizing methanogens and their metabolism

    DEFF Research Database (Denmark)

    Mladenovska, Zuzana

    Methanosarcina strains required vitamins as growth factor. The isolates differed in cardinal growth pH and temperatures, the kinetic parameters - maximum specific growth rate and half saturation constant - and threshold values during growth on acetate. Analysis of total cell proteins by SDS-page gel...

  11. Promoting interspecies electron transfer with biochar

    DEFF Research Database (Denmark)

    Chen, Shanshan; Rotaru, Amelia-Elena; Shrestha, Pravin Malla;

    2014-01-01

    to that previously reported for granular activated carbon (GAC). Although the biochars investigated were 1000 times less conductive than GAC, they stimulated DIET in co-cultures of Geobacter metallireducens with Geobacter sulfurreducens or Methanosarcina barkeri in which ethanol was the electron donor. Cells were...

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

    NARCIS (Netherlands)

    Fermoso, F.G.; Collins, G.; Bartacek, J.; O'Flaherty, V.; Lens, P.N.L.

    2008-01-01

    The effect of nickel deprivation from the influent of a mesophilic (30 degrees 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.

  13. The impact of Co and Ni speciation on methanogenesis in sulfidic media - Biouptake versus Metal dissolution

    NARCIS (Netherlands)

    Jansen, S.; Gonzalez-Gil, G.; Leeuwen, van H.P.

    2007-01-01

    The speciation of the trace nutrients Co(II) and Ni(II) in sulfide containing media can control the methanogenic activity of Methanosarcina sp., which is of importance for the optimisation of anaerobic treatment of wastewater containing methanol. To obtain more insight in the mechanistic backgrounds

  14. The impact of Co and Ni speciation on methanogenesis in sulfidic media - Biouptake versus Metal dissolution

    NARCIS (Netherlands)

    Jansen, S.; Gonzalez-Gil, G.; Leeuwen, van H.P.

    2007-01-01

    The speciation of the trace nutrients Co(II) and Ni(II) in sulfide containing media can control the methanogenic activity of Methanosarcina sp., which is of importance for the optimisation of anaerobic treatment of wastewater containing methanol. To obtain more insight in the mechanistic backgrounds

  15. The impact of Co and Ni speciation on methanogenesis in sulfidic media - Biouptake versus Metal dissolution

    NARCIS (Netherlands)

    Jansen, S.; Gonzalez-Gil, G.; Leeuwen, van H.P.

    2007-01-01

    The speciation of the trace nutrients Co(II) and Ni(II) in sulfide containing media can control the methanogenic activity of Methanosarcina sp., which is of importance for the optimisation of anaerobic treatment of wastewater containing methanol. To obtain more insight in the mechanistic

  16. Effect of shallow donors on Curie–Weiss temperature of Co-doped ZnO

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Shuxia, E-mail: gsx0391@sina.com [Department of Physics, Jiaozuo Teachers College, Jiaozuo 454001 (China); Key Laboratory for Special Functional Materials of Ministry of Education, Henan University, Kaifeng 475004 (China); Li, Jiwu [Department of Physics, Jiaozuo Teachers College, Jiaozuo 454001 (China); Du, Zuliang [Key Laboratory for Special Functional Materials of Ministry of Education, Henan University, Kaifeng 475004 (China)

    2014-12-15

    Co-doped ZnO and Al, Co co-doped ZnO polycrystalline powders were synthesized by co-precipitation method. The magnetization curves measured at 2 K show no hysteresis neither remanence for all samples. ZnO:Co grown at low temperature has a positive Curie–Weiss temperature Θ, and ZnO:Co grown at high temperature has a negative Θ. But Al-doped ZnO:Co grown at high temperature has a positive Θ. Positive Curie–Weiss temperature Θ was considered to have relation to the presence of shallow donors in the samples. - Highlights: • Co-doped ZnO and Al, Co co-doped ZnO polycrystalline powders were synthesized. • No hysteresis is observed for all samples. • The Curie–Weiss temperature Θ changes its sign by Al doping. • Positive Θ should be related to shallow donors.

  17. Enzymology of acetoclastic methanogenesis. Final technical report, August 1994--July 1997

    Energy Technology Data Exchange (ETDEWEB)

    Ragsdale, S.W.

    1997-12-31

    The overall aims of the proposal were to identify intermediates in the process of methane formation from acetate. The specific aims were: (1) to characterize the mechanism of acetyl-CoA conversion to CH{sub 3}-H{sub 4}MPT, CO, and CoA and elucidate the mechanism of CO oxidation by the methanosarcina thermophila CO dehydrogenase/acetyl-CoA synthase (CODH/ACS), and (2) to study the mechanism of methyl-CoM reduction to methane by developing an activation protocol for methyl-CoM reductase (MCR) and identifying intermediates in the MCR mechanism. The authors also made significant progress toward accomplishing two other specific aims that were not included in the original proposal. These were: (1) to characterize the M. thermophila heterodisulfide reductase (HDR), and (2) to characterize the methyltetrahydromethanopterin: Coenzyme M methyltransferase from Methanosarcina mazei strain Goe1 in collaboration with Gerhard Gottschalk`s group. Results from these four studies are summarized.

  18. Population dynamics of biofilm development during start-up of a butyrate-degrading fluidized-bed reactor

    Energy Technology Data Exchange (ETDEWEB)

    Zellner, G.; Geveke, M.; Diekmann, H. (Hannover Univ. (Germany). Inst. fuer Mikrobiologie); Conway de Macario, E. (New York State Dept. of Health, Albany, NY (United States). Wadsworth Center for Laboratories and Research)

    1991-12-01

    Population dynamics during start-up of a fluidized-bed reactor with butyrate or butyrate plus acetate as sole substrates as well as biofilm development on the sand substratum were studied microbiologically, immunologically and by scanning electron microscopy. An adapted syntrophic consortium consisting of Syntrophospora sp., Methanothrix soehngenii, Methanosarcina mazei and Methanobrevibacter arboriphilus or Methanogenium sp. achieved high-rate butyrate degradation to methane and carbon dioxide. Desulfovibrio sp., Methanocorpusculum sp., and Methanobacterium sp. were also present in lower numbers. Immunological analysis demonstrated methanogens antigenically related to Methanobrevibacter ruminantium M1, Methanosarcina mazei S6, M. thermophila TM1, Methanobrevibacter arboriphilus AZ and Methanothrix soehngenii Opfikon in the biofilm. Immunological analysis also showed that the organisms isolated from the butyrate-degrading culture used as a source of inoculum were related to M. soehngenii Opfikon, Methanobacterium formicium MF and Methanospirillum hungatei JF1. (orig.).

  19. [Community Structure and Succession of Methanogens in Beishenshu Landfill, Beijing].

    Science.gov (United States)

    Song, Li-na; Wang, Lei; Xia, Meng-jing; Su, Yue; Li, Zhen-shan

    2015-09-01

    Methanogens are the key microorganisms for landfill stabilization. RT-PCR and qPCR detecting system were employed to determine the types and abundance of methanogens in 2-15 year-old solid wastes that sampled from Beishenshu Landfill, Beijing. The organic components were almost stable and the pH values were in alkaline range, which indicated that the landfill was in the methanogenic process. Methanobacterials, Methanosaeta, and Methanosarcina were detected, among which Methanosaeta and Methanosarcina are acetoclastic, and Methanobacterials are hydrogenotrophic. As landfill processing, within this time range, although the bacterial abundance was significantly decreased, the amount of methanogens was first increased and then decreased, and finally became stable after being landfilled for 9 years. Methanosarcina was the dominate taxa. Significant correlations were found between the methanogens and the volatile fatty acids, but the correlations between methanogens and larger molecular organic matters were relatively weak or even absent. Taken together, our study revealed that the amount of methanogens were affected by substrates, but hardly influenced by the conversion of large molecules in these wastes landfilled for more than 2 years.

  20. Squalenes, phytanes and other isoprenoids as major neutral lipids of methanogenic and thermoacidophilic archaebacteria

    Energy Technology Data Exchange (ETDEWEB)

    Tornabene, T.G. (Colorado State Univ., Fort Collins); Langworthy, T.A.; Holzer, G.; Oro, J.

    1979-01-01

    The neutral lipids of nine species of methanogenic bacteria including five methanobacilli, two methanococci, a methanospirillum, one methanosarcina as well as two thermoacidophilic bacteria, Thermoplasma and Sulfolobus, were analyzed. The major components were C/sub 30/, C/sub 25/ and/or C/sub 20/ acyclic isoprenoid hydrocarbons with a continuous range of hydroisoprenoid homologues. The range of acyclic isoprenoids detected were from C/sub 14/ to C/sub 30/. Apart from Methanosarcina barkeri, squalene and/or hydrosqualene derivatives were the predominant components in all species studied. The components of Methanosarcina barkeri were a family of C/sub 25/ homologues. The distribution of the neutral lipid components and their specific variations in relative intensities emphasized the differences between the test organisms while the generic nature of the isoprenoid hydrocarbons demonstrated similarities between the diverse bacteria. The neutral lipid compositions from these bacteria, many of which exist in environmental conditions like those described for the various evolutionary stages of the archean ecology, resemble the isoprenoid distribution isolated from ancient sediments and petroleum. Therefore, these findings may have major implications to biological and biogeochemical evolution.

  1. [Conversion of acetic acid to methane by thermophiles: Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Zinder, S.

    1991-12-31

    The objective of this project is to provide an understanding of thermophilic anaerobic microorganisms capable of breaking down acetic acid, the precursor of two-thirds of the methane produced by anaerobic bioreactors. Recent results include: (1) the isolation of Methanothrix strain CALLS-1, which grows much more rapidly than mesophilic strains; (2) the demonstration that thermophilic cultures of Methanosarcina and Methanothrix show minimum thresholds for acetate utilization of 1--2.5 mM and 10--20{mu}m respectively, in agreement with ecological data indicating that Methanothrix is favored by low acetate concentration; (3) the demonstration of high levels of thermostable acetyl-coA synthetase and carbon monoxide dehydrogenase in cell-free extracts of Methanothrix strains CALS-1; (4) the demonstration of methanogenesis from acetate and ATP in cell free extracts of strain CALS-1. (5) the demonstration that methanogenesis from acetate required 2 ATP/methane, and, in contrast to Methanosarcina, was independent of hydrogen and other electron donors; (6) the finding that entropy effects must be considered when predicting the level of hydrogen in thermophilic syntrophic cultures. (7) the isolation and characterization of the Desulfotomaculum thermoacetoxidans. Current research is centered on factors which allow thermophilic Methanothrix to compete with Methanosarcina.

  2. (Conversion of acetic acid to methane by thermophiles: Progress report)

    Energy Technology Data Exchange (ETDEWEB)

    Zinder, S.

    1991-01-01

    The objective of this project is to provide an understanding of thermophilic anaerobic microorganisms capable of breaking down acetic acid, the precursor of two-thirds of the methane produced by anaerobic bioreactors. Recent results include: (1) the isolation of Methanothrix strain CALLS-1, which grows much more rapidly than mesophilic strains; (2) the demonstration that thermophilic cultures of Methanosarcina and Methanothrix show minimum thresholds for acetate utilization of 1--2.5 mM and 10--20{mu}m respectively, in agreement with ecological data indicating that Methanothrix is favored by low acetate concentration; (3) the demonstration of high levels of thermostable acetyl-coA synthetase and carbon monoxide dehydrogenase in cell-free extracts of Methanothrix strains CALS-1; (4) the demonstration of methanogenesis from acetate and ATP in cell free extracts of strain CALS-1. (5) the demonstration that methanogenesis from acetate required 2 ATP/methane, and, in contrast to Methanosarcina, was independent of hydrogen and other electron donors; (6) the finding that entropy effects must be considered when predicting the level of hydrogen in thermophilic syntrophic cultures. (7) the isolation and characterization of the Desulfotomaculum thermoacetoxidans. Current research is centered on factors which allow thermophilic Methanothrix to compete with Methanosarcina.

  3. Searching for links in the biotic characteristics and abiotic parameters of nine different biogas plants

    Science.gov (United States)

    Walter, Andreas; Knapp, Brigitte A.; Farbmacher, Theresa; Ebner, Christian; Insam, Heribert; Franke‐Whittle, Ingrid H.

    2012-01-01

    Summary To find links between the biotic characteristics and abiotic process parameters in anaerobic digestion systems, the microbial communities of nine full‐scale biogas plants in South Tyrol (Italy) and Vorarlberg (Austria) were investigated using molecular techniques and the physical and chemical properties were monitored. DNA from sludge samples was subjected to microarray hybridization with the ANAEROCHIP microarray and results indicated that sludge samples grouped into two main clusters, dominated either by Methanosarcina or by Methanosaeta, both aceticlastic methanogens. Hydrogenotrophic methanogens were hardly detected or if detected, gave low hybridization signals. Results obtained using denaturing gradient gel electrophoresis (DGGE) supported the findings of microarray hybridization. Real‐time PCR targeting Methanosarcina and Methanosaeta was conducted to provide quantitative data on the dominating methanogens. Correlation analysis to determine any links between the microbial communities found by microarray analysis, and the physicochemical parameters investigated was conducted. It was shown that the sludge samples dominated by the genus Methanosarcina were positively correlated with higher concentrations of acetate, whereas sludge samples dominated by representatives of the genus Methanosaeta had lower acetate concentrations. No other correlations between biotic characteristics and abiotic parameters were found. Methanogenic communities in each reactor were highly stable and resilient over the whole year. PMID:22950603

  4. AcEST: DK960495 [AcEST

    Lifescience Database Archive (English)

    Full Text Available t : Swiss-Prot sp_hit_id Q5F464 Definition sp|Q5F464|LPP_CHICK Lipoma-preferred partner homolog OS=Gallus ga...producing significant alignments: (bits) Value sp|Q5F464|LPP_CHICK Lipoma-preferred...... 30 9.0 sp|Q8TSH7|MDH_METAC Malate dehydrogenase OS=Methanosarcina aceti... 30 9.0 >sp|Q5F464|LPP_CHICK Lipoma-preferred

  5. Diphytanyl and dibiphytanyl glycerol ether lipids of methanogenic archaebacteria

    Energy Technology Data Exchange (ETDEWEB)

    Tornabene, T.G. (Colorado State Univ., Fort Collins); Langworthy, T.A.

    1979-01-05

    The lipids of nine different methanogenic bacterial strains are comprised of diphytanyl glycerol diethers, previously known only in extremely halophilic bacteria, as well as dibiphytanyl diglycerol tetraethers, known formerly only in the extremely thermoacidophilic bacteria Thermoplasma and Sulfolobus. Of the methanogens examined from four representative taxonomic groups, Methanobacterium and Methanospirillum contained both types of isopranyl ethers in nearly equal proportions, whereas the coccal forms, Methanosarcina and Methanococcus, possessed diphytanyl glycerol diethers, but with only a trace of or no dibiphytanyl diglycerol tetraethers. The occurrence of both types of isopranyl glycerol ethers in methanogenic bacteria supports the proposal that they have a close genealogical relationship to the extremely halophilic and thermoacidophilic bacteria.

  6. There is a baby in the bath water: AcrB contamination is a major problem in membrane-protein crystallization.

    Science.gov (United States)

    Veesler, David; Blangy, Stéphanie; Cambillau, Christian; Sciara, Giuliano

    2008-10-01

    In the course of a crystallographic study of the Methanosarcina mazei CorA transporter, the membrane protein was obtained with at least 95% purity and was submitted to crystallization trials. Small crystals (contaminating protein, acriflavine resistance protein B (AcrB), that was present at less than 5% in the protein preparations. AcrB contamination is a major problem when expressing membrane proteins in E. coli since it binds naturally to immobilized metal-ion affinity chromatography (IMAC) resins. Here, the structure is compared with previously deposited AcrB structures and strategies are proposed to avoid this contamination.

  7. A detailed phylogeny for the Methanomicrobiales

    Science.gov (United States)

    Rouviere, P.; Mandelco, L.; Winker, S.; Woese, C. R.

    1992-01-01

    The small subunit rRNA sequence of twenty archaea, members of the Methanomicrobiales, permits a detailed phylogenetic tree to be inferred for the group. The tree confirms earlier studies, based on far fewer sequences, in showing the group to be divided into two major clusters, temporarily designated the "methanosarcina" group and the "methanogenium" group. The tree also defines phylogenetic relationships within these two groups, which in some cases do not agree with the phylogenetic relationships implied by current taxonomic names--a problem most acute for the genus Methanogenium and its relatives. The present phylogenetic characterization provides the basis for a consistent taxonomic restructuring of this major methanogenic taxon.

  8. Development of methanogenic consortia in fluidized-bed batches using sepiolite of different particle size.

    Science.gov (United States)

    Sánchez, J M; Rodríguez, F; Valle, L; Muñoz, M A; Moriñigo, M A; Borrego, J J

    1996-09-01

    The addition of support materials, such as sepiolite, to fluidized-bed anaerobic digesters enhances the methane production by increasing the colonization by syntrophic microbiota. However, the efficiency in the methanogenesis depends on the particle size of the support material, the highest level of methane production being obtained by the smaller particle size sepiolite. Because of the porosity and physico-chemical characteristics of these support materials, the anaerobic microbial consortia formed quickly (after one week of incubation). The predominant methanogenic bacteria present in the active granules, detected both by immunofluorescence using specific antibodies and by scanning electron microscopy, were acetoclastic methanogens, mainly Methanosarcina and Methanosaeta.

  9. A detailed phylogeny for the Methanomicrobiales

    Science.gov (United States)

    Rouviere, P.; Mandelco, L.; Winker, S.; Woese, C. R.

    1992-01-01

    The small subunit rRNA sequence of twenty archaea, members of the Methanomicrobiales, permits a detailed phylogenetic tree to be inferred for the group. The tree confirms earlier studies, based on far fewer sequences, in showing the group to be divided into two major clusters, temporarily designated the "methanosarcina" group and the "methanogenium" group. The tree also defines phylogenetic relationships within these two groups, which in some cases do not agree with the phylogenetic relationships implied by current taxonomic names--a problem most acute for the genus Methanogenium and its relatives. The present phylogenetic characterization provides the basis for a consistent taxonomic restructuring of this major methanogenic taxon.

  10. Dicty_cDB: Contig-U10274-1 [Dicty_cDB

    Lifescience Database Archive (English)

    Full Text Available :none) Burkholderia vietnamiensis G4 c... 75 2e-12 CR378672_161( CR378672 |pid...:none) Methanosarcina mazei strain Goe1... 89 1e-16 CP000053_198( CP000053 |pid:none) Rickettsia felis URRWXCal2, co...romyxobacter sp. K, complete... 87 7e-16 AL939131_102( AL939131 |pid:none) Streptomyces coelicolor A3(2) co... 3500... 79 1e-13 AL939112_114( AL939112 |pid:none) Streptomyces coelicolor A3(2) co....la str. Sg (Schizaphis graminum)... 44 7.1 1 >( C94358 ) Dictyostelium discoideum slug

  11. Trace elements induce predominance among methanogenic activity in anaerobic digestion

    Directory of Open Access Journals (Sweden)

    Babett Wintsche

    2016-12-01

    Full Text Available Trace elements play an essential role in all organisms due to their functions in enzyme complexes. In anaerobic digesters, control and supplementation of trace elements lead to stable and more efficient methane production processes while trace element deficits cause process imbalances. However, the underlying metabolic mechanisms and the adaptation of the affected microbial communities to such deficits are not yet fully understood. Here, we investigated the microbial community dynamics and resulting process changes induced by trace element deprivation. Two identical lab-scale continuous stirred tank reactors fed with distiller’s grains and supplemented with trace elements (cobalt, molybdenum, nickel, tungsten and a commercial iron additive were operated in parallel. After 72 weeks of identical operation, the feeding regime of one reactor was changed by omitting trace element supplements and reducing the amount of iron additive. Both reactors were operated for further 21 weeks. Various process parameters (biogas production and composition, total solids and volatile solids, trace element concentration, volatile fatty acids, total ammonium nitrogen, total organic acids/alkalinity ratio, and pH and the composition and activity of the microbial communities were monitored over the total experimental time. While the methane yield remained stable, the concentrations of hydrogen sulfide, total ammonia nitrogen, and acetate increased in the trace element-depleted reactor compared to the well-supplied control reactor. Methanosarcina and Methanoculleus dominated the methanogenic communities in both reactors. However, the activity ratio of these two genera was shown to depend on trace element supplementation explainable by different trace element requirements of their energy conservation systems. Methanosarcina dominated the well-supplied anaerobic digester, pointing to acetoclastic methanogenesis as the dominant methanogenic pathway. Under trace element

  12. Biodegradability potential of two experimental landfills in Brazil

    Directory of Open Access Journals (Sweden)

    Vazoller Rosana Filomena

    2001-01-01

    Full Text Available Solid wastes anaerobic biodegradability, methane production potential and microbiological composition of two experimental sanitary landfills in Brazil, running for one year, were evaluated. The two landfills showed a similar organic matter stabilization during the methane production phase, despite the high heterogeneity of the solid wastes. Both landfills presented the same level of methane (around 91.5 L CH4 / kg Volatile Total Solids and organic acids, mainly acetic and butyric acids, in the leachate. Bacterial isolates belonged to genera Megasphaera, Selenomonas, Methanobacterium, Methanobrevibacter and Methanosarcina.

  13. Genome Sequence of Desulfurella amilsii Strain TR1 and Comparative Genomics of Desulfurellaceae Family.

    Science.gov (United States)

    Florentino, Anna P; Stams, Alfons J M; Sánchez-Andrea, Irene

    2017-01-01

    The acidotolerant sulfur reducer Desulfurella amilsii was isolated from sediments of Tinto River, an extremely acidic environment. Its ability to grow in a broad range of pH and to tolerate certain heavy metals offers potential for metal recovery processes. Here we report its high-quality draft genome sequence and compare it to the available genome sequences of other members of Desulfurellaceae family: D. acetivorans. D. multipotens, Hippea maritima. H. alviniae, H. medeae, and H. jasoniae. For most species, pairwise comparisons for average nucleotide identity (ANI) and in silico DNA-DNA hybridization (DDH) revealed ANI values from 67.5 to 80% and DDH values from 12.9 to 24.2%. D. acetivorans and D. multipotens, however, surpassed the estimated thresholds of species definition for both DDH (98.6%) and ANI (88.1%). Therefore, they should be merged to a single species. Comparative analysis of Desulfurellaceae genomes revealed different gene content for sulfur respiration between Desulfurella and Hippea species. Sulfur reductase is only encoded in D. amilsii, in which it is suggested to play a role in sulfur respiration, especially at low pH. Polysulfide reductase is only encoded in Hippea species; it is likely that this genus uses polysulfide as electron acceptor. Genes encoding thiosulfate reductase are present in all the genomes, but dissimilatory sulfite reductase is only present in Desulfurella species. Thus, thiosulfate respiration via sulfite is only likely in this genus. Although sulfur disproportionation occurs in Desulfurella species, the molecular mechanism behind this process is not yet understood, hampering a genome prediction. The metabolism of acetate in Desulfurella species can occur via the acetyl-CoA synthetase or via acetate kinase in combination with phosphate acetyltransferase, while in Hippea species, it might occur via the acetate kinase. Large differences in gene sets involved in resistance to acidic conditions were not detected among the

  14. Unexpected competitiveness of Methanosaeta populations at elevated acetate concentrations in methanogenic treatment of animal wastewater.

    Science.gov (United States)

    Chen, Si; Cheng, Huicai; Liu, Jiang; Hazen, Terry C; Huang, Vicki; He, Qiang

    2017-02-01

    Acetoclastic methanogenesis is a key metabolic process in anaerobic digestion, a technology with broad applications in biogas production and waste treatment. Acetoclastic methanogenesis is known to be performed by two archaeal genera, Methanosaeta and Methanosarcina. The conventional model posits that Methanosaeta populations are more competitive at low acetate levels (competitiveness of Methanosaeta at elevated acetate was further supported by the enrichment of Methanosaeta with high concentrations of acetate (20 mM). The dominance of Methanosaeta in the methanogen community could be reproduced in anaerobic digesters with the direct addition of acetate to above 20 mM, again supporting the competitiveness of Methanosaeta over Methanosarcina at elevated acetate levels. This study for the first time systematically demonstrated that the dominance of Methanosaeta populations in anaerobic digestion could be linked to the competitiveness of Methanosaeta at elevated acetate concentrations. Given the importance of acetoclastic methanogenesis in biological methane production, findings from this study could have major implications for developing strategies for more effective control of methanogenic treatment processes.

  15. The role of different methanogen groups evaluated by Real-Time qPCR as high-efficiency bioindicators of wet anaerobic co-digestion of organic waste

    Science.gov (United States)

    2011-01-01

    Methanogen populations and their domains are poorly understood; however, in recent years, research on this topic has emerged. The relevance of this field has also been enhanced by the growing economic interest in methanogen skills, particularly the production of methane from organic substrates. Management attention turned to anaerobic wastes digestion because the volume and environmental impact reductions. Methanogenesis is the biochemically limiting step of the process and the industrially interesting phase because it connects to the amount of biogas production. For this reason, several studies have evaluated the structure of methanogen communities during this process. Currently, it is clear that the methanogen load and diversity depend on the feeding characteristics and the process conditions, but not much data is available. In this study, we apply a Real-Time Polymerase Chain Reaction (RT-PCR) method based on mcrA target to evaluate, by specific probes, some subgroups of methanogens during the mesophilic anaerobic digestion process fed wastewater sludge and organic fraction of the municipal solid waste with two different pre-treatments. The obtained data showed the prevalence of Methanomicrobiales and significantly positive correlation between Methanosarcina and Methanosaetae and the biogas production rate (0.744 p < 0.01 and 0.641 p < 0.05). Methanosarcina detected levels are different during the process after the two pre-treatment of the input materials (T-test p < 0.05). Moreover, a role as diagnostic tool could be suggested in digestion optimisation. PMID:21982396

  16. Presence of an unusual methanogenic bacterium in coal gasification waste.

    Science.gov (United States)

    Tomei, F A; Rouse, D; Maki, J S; Mitchell, R

    1988-12-01

    Methanogenic bacteria growing on a pilot-scale, anaerobic filter processing coal gasification waste were enriched in a mineral salts medium containing hydrogen and acetate as potential energy sources. Transfer of the enrichments to methanol medium resulted in the initial growth of a strain of Methanosarcina barkeri, but eventually small cocci became dominant. The cocci growing on methanol produced methane and exhibited the typical fluorescence of methanogenic bacteria. They grew in the presence of the cell wall synthesis-inhibiting antibiotics d-cycloserine, fosfomycin, penicillin G, and vancomycin as well as in the presence of kanamycin, an inhibitor of protein synthesis in eubacteria. The optimal growth temperature was 37 degrees C, and the doubling time was 7.5 h. The strain lysed after reaching stationary phase. The bacterium grew poorly with hydrogen as the energy source and failed to grow on acetate. Morphologically, the coccus shared similarities with Methanosarcina sp. Cells were 1 mum wide, exhibited the typical thick cell wall and cross-wall formation, and formed tetrads. Packets and cysts were not formed.

  17. Molecular Biology and Genetics of the Acetate-Utilizing Methanogenic Bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Robert P. Gunsalus

    2003-07-21

    Methane biosynthesis by the Methanosarcina species, in contrast to other methanogens, occurs from the full range of methanogenic substrates that include acetate, methanol, tri-methyl, di-methyl, and methyl-amine, methyl-sulfides, and in limited instances, H2/CO2. The Methanosarcina are also versatile in their ability to adapt and grow in habitats of varying osmolarity ranging from fresh water environments, marine environments, and to hyper saline environments (ca to 1.2 M NaCl). To facilitate studies that address the biochemistry, molecular biology and physiology of these organisms, we have constructed a whole-genome microarray to identify classes of differentially expressed genes in M. mazei strain Goe1. We propose to further identify and examine how genes and their proteins involved in the synthesis and transport of osmolytes in the cell are regulated. These compounds include N-epsilon-acetyl-beta-lysine, alpha-glutamate, betaine, and potassium whose levels within the cell are modulated in order to provide appropriate osmotic balance. We will identify differentially expressed genes involved in hydrogen and carbon dioxide sequestration since M. mazei strain Goe1 is currently the only practical model for such study. Finally, we will explore the essential roles of two metals, molybdate and tungstate, in methanogen regulation and metabolism of these environmentally essential organsims. The above studies will advance our general understanding of how methanogens respond to their environmental signals, and adapt by adjusting their physiology to thrive in changing anaerobic habitats whether natural or man-made.

  18. Effect of ferrihydrite biomineralization on methanogenesis in an anaerobic incubation from paddy soil

    Science.gov (United States)

    Zhuang, Li; Xu, Jielong; Tang, Jia; Zhou, Shungui

    2015-05-01

    Microbial reduction of Fe(III) can be one of the major factors controlling methane production from anaerobic sedimentary environments, such as paddy soils and wetlands. Although secondary iron mineralization following Fe(III) reduction is a process that occurs naturally over time, it has not yet been considered in methanogenic systems. This study performed a long-term anaerobic incubation of a paddy soil and ferrihydrite-supplemented soil cultures to investigate methanogenesis during ferrihydrite biomineralization. The results revealed that the long-term effect of ferrihydrite on methanogenesis may be enhancement rather than suppression documented in previous studies. During initial microbial ferrihydrite reduction, methanogenesis was suppressed; however, the secondary minerals of magnetite formation was simultaneous with facilitated methanogenesis in terms of average methane production rate and acetate utilization rate. In the phase of magnetite formation, microbial community analysis revealed a strong stimulation of the bacterial Geobacter, Bacillus, and Sedimentibacter and the archaeal Methanosarcina in the ferrihydrite-supplemented cultures. Direct electric syntrophy between Geobacter and Methanosarcina via conductive magnetite is the plausible mechanism for methanogenesis acceleration along with magnetite formation. Our data suggested that a change in iron mineralogy might affect the conversion of anaerobic organic matter to methane and might provide a fresh perspective on the mitigation of methane emissions from paddy soils by ferric iron fertilization.

  19. Similar evolution in delta 13CH4 and model-predicted relative rate of aceticlastic methanogenesis during mesophilic methanization of municipal solid wastes.

    Science.gov (United States)

    Vavilin, V A; Qu, X; Qu, X; Mazéas, L; Lemunier, M; Duquennoi, C; Mouchel, J M; He, P; Bouchez, T

    2009-01-01

    Similar evolution was obtained for the stable carbon isotope signatures delta (13)CH(4) and the model-predicted relative rate of aceticlastic methanogenesis during mesophilic methanization of municipal solid wastes. In batch incubations, the importance of aceticlastic and hydrogenotrophic methanogenesis changes in time. Initially, hydrogenotrophic methanogenesis dominated, but increasing population of Methanosarcina sp. enhances aceticlastic methanogenesis. Later, hydrogenotrophic methanogenesis intensified again. A mathematical model was developed to evaluate the relative contribution of hydrogenotrophic and aceticlastic pathways of methane generation during mesophilic batch anaerobic biodegradation of the French and the Chinese Municipal Solid Wastes (FMSW and CMSW). Taking into account molecular biology analysis reported earlier three groups of methanogens including strictly hydrogenotrophic methanogens, strictly aceticlastic methanogens (Methanosaeta sp.) and Methanosarcina sp., consuming both acetate and H(2)/H(2)CO(3) were considered in the model. The total organic and inorganic carbon concentrations, methane production volume, methane and carbon dioxide partial pressures values were used for the model calibration and validation. Methane isotopic composition (delta (13)CH(4)) evolution during the incubations was used to independently validate the model results. The model demonstrated that only the putrescible solid waste was totally converted to methane.

  20. Effect of Nickel Levels on Hydrogen Partial Pressure and Methane Production in Methanogens

    Science.gov (United States)

    2016-01-01

    Hydrogen (H2) consumption and methane (CH4) production in pure cultures of three different methanogens were investigated during cultivation with 0, 0.2 and 4.21 μM added nickel (Ni). The results showed that the level of dissolved Ni in the anaerobic growth medium did not notably affect CH4 production in the cytochrome-free methanogenic species Methanobacterium bryantii and Methanoculleus bourgensis MAB1, but affected CH4 formation rate in the cytochrome-containing Methanosarcina barkeri grown on H2 and CO2. Methanosarcina barkeri also had the highest amounts of Ni in its cells, indicating that more Ni is needed by cytochrome-containing than by cytochrome-free methanogenic species. The concentration of Ni affected threshold values of H2 partial pressure (pH2) for all three methanogen species studied, with M. bourgensis MAB1 reaching pH2 values as low as 0.1 Pa when Ni was available in amounts used in normal anaerobic growth medium. To our knowledge, this is the lowest pH2 threshold recorded to date in pure methanogen culture, which suggests that M.bourgensis MAB1 have a competitive advantage over other species through its ability to grow at low H2 concentrations. Our study has implications for research on the H2-driven deep subsurface biosphere and biogas reactor performance. PMID:27992585

  1. Biomethanation and microbial community changes in a digester treating sludge from a brackish aquaculture recirculation system.

    Science.gov (United States)

    Zhang, Xuedong; Tao, Yu; Hu, Jianmei; Liu, Gang; Spanjers, Henri; van Lier, Jules B

    2016-08-01

    Using a high-salinity-adapted inoculum and a moderate stepwise-increased organic loading rate (OLR), a stable digester performance was achieved in treating sludge from a brackish aquaculture recirculation system. The specific methane yield was distinctly enhanced, reaching 0.203LCH4/gCODadded, compared to literature values (0.140-0.154LCH4/gCODadded) from the salty sludges. OLR adjustment and the fecal substrate substantially influenced population changes in the digester. Within the bacterial subpopulations, the relative abundance of Bacillus and Bacteroides declined, accompanied by the increase of Clostridium and Trigonala over time. The results show Trigonala was derived from the substrate and accumulated inside the digester. The most abundant methanogen was Methanosarcina in the inoculum and the digestates. The Methanosarcina proliferation can be ascribed to its metabolic versatility, probably a feature of crucial importance for high-salinity environments. Other frequently observed methanogens were outcompeted. The population similarity at the genus level between inoculum and digestates declined during the initial stage and afterwards increased.

  2. [Conversion of acetic acid to methane by thermophiles]. Annual progress report

    Energy Technology Data Exchange (ETDEWEB)

    Zinder, S.H.

    1994-02-01

    Acetate is the precursor of approximately two-thirds of the methane produced by anaerobic bioreactors and many other methanogenic habitats. Besides their intrinsic interest, thermophilic acetotrophic methanogenic cultures usually grow at least twice as fast as their mesophilic counterparts, making them more amenable to study. In recent years, attention has been mainly focused on the thermophilic acetate utilizing methanogen Methanothrix strain CALS-1. Methanothrix, also called Methanosaeta, is one of only two methanogenic genera known to convert acetate to methane, the other being Methanosarcina. The faster-growing more versatile Methanosarcina has been better studied. However, when one examines anaerobic digestor contents, Methanothrix is often the dominant acetate-utilizing methanogen. As described in previous progress reports, the authors have achieved methanogenesis from acetate in cell-free extracts of Methanothrix strain CALS-1 grown in a pH auxostat. Using these cell extracts, specific activities for methanogenesis from acetate and ATP of 100--300 nmol/min were routinely obtained, levels comparable to the rate in whole cells, which is not usually the case in methanogenic extracts. Recently obtained results are given and discussed for the following: Methanogenesis in crude extracts; Role of the cell membrane in methanogenesis from acetate; Carbon monoxide dehydrogenase; Novel thermophilic cultures converting acetate to methane; and Methanol-utilizing methanogen.

  3. High-rate, High Temperature Acetotrophic Methanogenesis Governed by a Three Population Consortium in Anaerobic Bioreactors

    Science.gov (United States)

    Ho, Dang; Jensen, Paul; Gutierrez-Zamora, Maria-Luisa; Beckmann, Sabrina; Manefield, Mike; Batstone, Damien

    2016-01-01

    A combination of acetate oxidation and acetoclastic methanogenesis has been previously identified to enable high-rate methanogenesis at high temperatures (55 to 65°C), but this capability had not been linked to any key organisms. This study combined RNA–stable isotope probing on 13C-labelled acetate and 16S amplicon sequencing to identify the active micro-organisms involved in high-rate methanogenesis. Active biomass was harvested from three bench-scale thermophilic bioreactors treating waste activated sludge at 55, 60 and 65°C, and fed with 13-C labelled and 12C-unlabelled acetate. Acetate uptake and cumulative methane production were determined and kinetic parameters were estimated using model-based analysis. Pyrosequencing performed on 13C- enriched samples indicated that organisms accumulating labelled carbon were Coprothermobacter (all temperatures between 55 and 65°C), acetoclastic Methanosarcina (55 to 60°C) and hydrogenotrophic Methanothermobacter (60 to 65°C). The increased relative abundance of Coprothermobacter with increased temperature corresponding with a shift to syntrophic acetate oxidation identified this as a potentially key oxidiser. Methanosarcina likely acts as both a hydrogen utilising and acetoclastic methanogen at 55°C, and is replaced by Methanothermobacter as a hydrogen utiliser at higher temperatures. PMID:27490246

  4. Anaerobic treatment performance and microbial population of thermophilic upflow anaerobic filter reactor treating awamori distillery wastewater.

    Science.gov (United States)

    Tang, Yue-Qin; Fujimura, Yutaka; Shigematsu, Toru; Morimura, Shigeru; Kida, Kenji

    2007-10-01

    Distillery wastewater from awamori making was anaerobically treated for one year using thermophilic upflow anaerobic filter (UAF) reactors packed with pyridinium group-containing nonwoven fabric material. The microbial structure and spatial distribution of microorganisms on the support material were characterized using molecular biological methods. The reactor steadily achieved a high TOC loading rate of 18 g/l/d with approximately 80% TOC removal efficiency when non-diluted wastewater was fed. The maximum TOC loading rate increased to 36 g/l/d when treating thrice-diluted wastewater. However, the TOC removal efficiency and gas evolution rate decreased compared with that when non-diluted wastewater was used. Methanogens closely related to Methanosarcina thermophila and Methanoculleus bourgensis and bacteria in the phyla Firmicutes and Bacteroidetes were predominant methanogens and bacteria in the thermophilic UFA reactor, as indicated by 16S rRNA gene clone analysis. Fluorescence in situ hybridization (FISH) results showed that a large quantity of bacterial cells adhered throughout the whole support, and Methanosarcina-like methanogens existed mainly in the relative outside region while Methanoculleus cells were located in the relative inner part of the support. The support material used proved to be an excellent carrier for microorganisms, and a UAF reactor using this kind of support can be used for high-rate treatment of awamori/shochu distillery wastewater.

  5. Influence of the addition of sulphate and ferric ions in a methanogenic anaerobic packed-bed reactor treating gasoline-contaminated water.

    Science.gov (United States)

    Fernandes, B S; Chinalia, F A; Sarti, A; Silva, A J; Foresti, E; Zaiat, M

    2006-01-01

    Benzene, toluene and xylene (BTX) are relatively soluble aromatic compounds of gasoline. Gasoline storage tank leakages generally lead to an extensive contamination of groundwater. In the natural environment for instance, these compounds might be biodegraded under a variety of reducing potentials. The objective of this work was to examine the influence of the addition of sulphate and Fe(OH)3 in a methanogenic horizontal-flow anaerobic immobilized-biomass reactor treating gasoline-contaminated water. Three different conditions were evaluated: methanogenic, sulphidogenic and sulphidogenic with the addition of ferric ions. Methanogenic condition showed the higher BTX degradation rates and the addition of sulphate negatively affected BTX removal rates with the production of H2S. However, the addition of ferric ions resulted in the precipitation of sulphur, improving BTX degradation by the consortium. Metanosphaera sp., Methanosarcina barkeri and Methanosaeta concilii were identified in the consortium by means of 16S and directly related to the addition of ferric ions.

  6. Changes of the microbial population structure in an overloaded fed-batch biogas reactor digesting maize silage.

    Science.gov (United States)

    Kampmann, Kristina; Ratering, Stefan; Geißler-Plaum, Rita; Schmidt, Michael; Zerr, Walter; Schnell, Sylvia

    2014-12-01

    Two parallel, stable operating biogas reactors were fed with increasing amounts of maize silage to monitor microbial community changes caused by overloading. Changes of microorganisms diversity revealed by SSCP (single strand conformation polymorphism) indicating an acidification before and during the pH-value decrease. The earliest indicator was the appearance of a Methanosarcina thermophila-related species. Diversity of dominant fermenting bacteria within Bacteroidetes, Firmicutes and other Bacteria decreased upon overloading. Some species became dominant directly before and during acidification and thus could be suitable as possible indicator organisms for detection of futurity acidification. Those bacteria were related to Prolixibacter bellariivorans and Streptococcus infantarius subsp. infantarius. An early detection of community shifts will allow better feeding management for optimal biogas production.

  7. Evaluation of marine sediments as microbial sources for methane production from brown algae under high salinity.

    Science.gov (United States)

    Miura, Toyokazu; Kita, Akihisa; Okamura, Yoshiko; Aki, Tsunehiro; Matsumura, Yukihiko; Tajima, Takahisa; Kato, Junichi; Nakashimada, Yutaka

    2014-10-01

    Various marine sediments were evaluated as promising microbial sources for methane fermentation of Saccharina japonica, a brown alga, at seawater salinity. All marine sediments tested produced mainly acetate among volatile fatty acids. One marine sediment completely converted the produced volatile fatty acids to methane in a short period. Archaeal community analysis revealed that acetoclastic methanogens belonging to the Methanosarcina genus dominated after cultivation. Measurement of the specific conversion rate at each step of methane production under saline conditions demonstrated that the marine sediments had higher conversion rates of butyrate and acetate than mesophilic methanogenic granules. These results clearly show that marine sediments can be used as microbial sources for methane production from algae under high-salt conditions without dilution. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Methanogenic burst in the end-Permian carbon cycle.

    Science.gov (United States)

    Rothman, Daniel H; Fournier, Gregory P; French, Katherine L; Alm, Eric J; Boyle, Edward A; Cao, Changqun; Summons, Roger E

    2014-04-15

    The end-Permian extinction is associated with a mysterious disruption to Earth's carbon cycle. Here we identify causal mechanisms via three observations. First, we show that geochemical signals indicate superexponential growth of the marine inorganic carbon reservoir, coincident with the extinction and consistent with the expansion of a new microbial metabolic pathway. Second, we show that the efficient acetoclastic pathway in Methanosarcina emerged at a time statistically indistinguishable from the extinction. Finally, we show that nickel concentrations in South China sediments increased sharply at the extinction, probably as a consequence of massive Siberian volcanism, enabling a methanogenic expansion by removal of nickel limitation. Collectively, these results are consistent with the instigation of Earth's greatest mass extinction by a specific microbial innovation.

  9. Biological treatment of TMAH (tetra-methyl ammonium hydroxide) in a full-scale TFT-LCD wastewater treatment plant.

    Science.gov (United States)

    Hu, Tai-Ho; Whang, Liang-Ming; Liu, Pao-Wen Grace; Hung, Yu-Ching; Chen, Hung-Wei; Lin, Li-Bin; Chen, Chia-Fu; Chen, Sheng-Kun; Hsu, Shu Fu; Shen, Wason; Fu, Ryan; Hsu, Romel

    2012-06-01

    This study evaluated biological treatment of TMAH in a full-scale methanogenic up-flow anaerobic sludge blanket (UASB) followed by an aerobic bioreactor. In general, the UASB was able to perform a satisfactory TMAH degradation efficiency, but the effluent COD of the aerobic bioreactor seemed to increase with an increased TMAH in the influent wastewater. The batch test results confirmed that the UASB sludge under methanogenic conditions would be favored over the aerobic ones for TMAH treatment due to its superb ability of handling high strength of TMAH-containing wastewaters. Based on batch experiments, inhibitory chemicals present in TFT-LCD wastewater like surfactants and sulfate should be avoided to secure a stable methanogenic TMAH degradation. Finally, molecular monitoring of Methanomethylovorans hollandica and Methanosarcina mazei in the full-scale plant, the dominant methanogens in the UASB responsible for TMAH degradation, may be beneficial for a stable TMAH treatment performance.

  10. Link between capacity for current production and syntrophic growth in Geobacter species

    DEFF Research Database (Denmark)

    Rotaru, Amelia-Elena; Woodard, Trevor; Nevin, Kelly;

    2015-01-01

    -culture with Methanosarcina barkeri, which is capable of direct interspecies electron transfer (DIET), but not with Methanospirillium hungatei capable only of H2 or formate transfer. Conductive granular activated carbon (GAC) stimulated metabolism of the G. hydrogenophilus - M. barkeri co-culture, consistent with electron......Electrodes are unnatural electron acceptors, and it is yet unknown how some Geobacter species evolved to use electrodes as terminal electron acceptors. Analysis of different Geobacter species revealed that they varied in their capacity for current production. G. metallireducens and G....... hydrogenophilus generated high current densities (ca. 0.05 mA/cm2), comparable to G. sulfurreducens. G. bremensis, G. chapellei, G. humireducens, and G. uranireducens, produced much lower currents (ca. 0.05 mA/cm2) and G. bemidjiensis was previously found to not produce current. There was no correspondence...

  11. Promoting Interspecies Electron Transfer with Biochar

    Science.gov (United States)

    Chen, Shanshan; Rotaru, Amelia-Elena; Shrestha, Pravin Malla; Malvankar, Nikhil S.; Liu, Fanghua; Fan, Wei; Nevin, Kelly P.; Lovley, Derek R.

    2014-01-01

    Biochar, a charcoal-like product of the incomplete combustion of organic materials, is an increasingly popular soil amendment designed to improve soil fertility. We investigated the possibility that biochar could promote direct interspecies electron transfer (DIET) in a manner similar to that previously reported for granular activated carbon (GAC). Although the biochars investigated were 1000 times less conductive than GAC, they stimulated DIET in co-cultures of Geobacter metallireducens with Geobacter sulfurreducens or Methanosarcina barkeri in which ethanol was the electron donor. Cells were attached to the biochar, yet not in close contact, suggesting that electrons were likely conducted through the biochar, rather than biological electrical connections. The finding that biochar can stimulate DIET may be an important consideration when amending soils with biochar and can help explain why biochar may enhance methane production from organic wastes under anaerobic conditions. PMID:24846283

  12. Promoting interspecies electron transfer with biochar

    DEFF Research Database (Denmark)

    Chen, Shanshan; Rotaru, Amelia-Elena; Shrestha, Pravin Malla

    2014-01-01

    Biochar, a charcoal-like product of the incomplete combustion of organic materials, is an increasingly popular soil amendment designed to improve soil fertility. We investigated the possibility that biochar could promote direct interspecies electron transfer (DIET) in a manner similar...... to that previously reported for granular activated carbon (GAC). Although the biochars investigated were 1000 times less conductive than GAC, they stimulated DIET in co-cultures of Geobacter metallireducens with Geobacter sulfurreducens or Methanosarcina barkeri in which ethanol was the electron donor. Cells were...... attached to the biochar, yet not in close contact, suggesting that electrons were likely conducted through the biochar, rather than biological electrical connections. The finding that biochar can stimulate DIET may be an important consideration when amending soils with biochar and can help explain why...

  13. Methane production potentials, pathways, and communities of methanogens in vertical sediment profiles of river Sitka.

    Science.gov (United States)

    Mach, Václav; Blaser, Martin B; Claus, Peter; Chaudhary, Prem P; Rulík, Martin

    2015-01-01

    Biological methanogenesis is linked to permanent water logged systems, e.g., rice field soils or lake sediments. In these systems the methanogenic community as well as the pathway of methane formation are well-described. By contrast, the methanogenic potential of river sediments is so far not well-investigated. Therefore, we analyzed (a) the methanogenic potential (incubation experiments), (b) the pathway of methane production (stable carbon isotopes and inhibitor studies), and (c) the methanogenic community composition (terminal restriction length polymorphism of mcrA) in depth profiles of sediment cores of River Sitka, Czech Republic. We found two depth-related distinct maxima for the methanogenic potentials (a) The pathway of methane production was dominated by hydrogenotrophic methanogenesis (b) The methanogenic community composition was similar in all depth layers (c) The main TRFs were representative for Methanosarcina, Methanosaeta, Methanobacterium, and Methanomicrobium species. The isotopic signals of acetate indicated a relative high contribution of chemolithotrophic acetogenesis to the acetate pool.

  14. Reduction of the hydraulic retention time at constant high organic loading rate to reach the microbial limits of anaerobic digestion in various reactor systems.

    Science.gov (United States)

    Ziganshin, Ayrat M; Schmidt, Thomas; Lv, Zuopeng; Liebetrau, Jan; Richnow, Hans Hermann; Kleinsteuber, Sabine; Nikolausz, Marcell

    2016-10-01

    The effects of hydraulic retention time (HRT) reduction at constant high organic loading rate on the activity of hydrogen-producing bacteria and methanogens were investigated in reactors digesting thin stillage. Stable isotope fingerprinting was additionally applied to assess methanogenic pathways. Based on hydA gene transcripts, Clostridiales was the most active hydrogen-producing order in continuous stirred tank reactor (CSTR), fixed-bed reactor (FBR) and anaerobic sequencing batch reactor (ASBR), but shorter HRT stimulated the activity of Spirochaetales. Further decreasing HRT diminished Spirochaetales activity in systems with biomass retention. Based on mcrA gene transcripts, Methanoculleus and Methanosarcina were the predominantly active in CSTR and ASBR, whereas Methanosaeta and Methanospirillum activity was more significant in stably performing FBR. Isotope values indicated the predominance of aceticlastic pathway in FBR. Interestingly, an increased activity of Methanosaeta was observed during shortening HRT in CSTR and ASBR despite high organic acids concentrations, what was supported by stable isotope data.

  15. Molecular analysis of methanogens involved in methanogenic degradation of tetramethylammonium hydroxide in full-scale bioreactors.

    Science.gov (United States)

    Whang, Liang-Ming; Hu, Tai-Ho; Liu, Pao-Wen Grace; Hung, Yu-Ching; Fukushima, Toshikazu; Wu, Yi-Ju; Chang, Shao-Hsiung

    2015-02-01

    This study investigated methanogenic communities involved in degradation of tetramethylammonium hydroxide (TMAH) in three full-scale bioreactors treating TMAH-containing wastewater. Based on the results of terminal-restriction fragment-length polymorphism (T-RFLP) and quantitative PCR analyses targeting the methyl-coenzyme M reductase alpha subunit (mcrA) genes retrieved from three bioreactors, Methanomethylovorans and Methanosarcina were the dominant methanogens involved in the methanogenic degradation of TMAH in the bioreactors. Furthermore, batch experiments were conducted to evaluate mcrA messenger RNA (mRNA) expression during methanogenic TMAH degradation, and the results indicated that a higher level of TMAH favored mcrA mRNA expression by Methansarcina, while Methanomethylovorans could only express considerable amount of mcrA mRNA at a lower level of TMAH. These results suggest that Methansarcina is responsible for methanogenic TMAH degradation at higher TMAH concentrations, while Methanomethylovorans may be important at a lower TMAH condition.

  16. Reducing gas content of coal deposits by means of bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Godlewska-Lipowa, A.A.; Kozlowski, B.

    1981-07-01

    This paper discusses the results of experiments carried out in Poland under laboratory conditions on efficiency of methane control using bacteria from Methanosarcina and Methanomonas groups. Malashenko and Whittenburry culture mediums were used. Bacteria growth in an atmosphere of air and methane (48.2%, 8.6% and 5.21%) was observed. Temperature ranged from 19 to 20 C. Investigations show that the bacteria are characterized by high oxidation activity. Depending on methane concentration in the air the bacteria consume from 75% to 100% of methane during biosynthesis. The bacteria reduce methane and oxygen content and increase carbon dioxide content in the air. Using bacteria methane concentration in the air was reduced from 48.2% to 12.3%, from 8.6% to 0.0% and from 5.21% to 0.01%. (7 refs.) (In Polish)

  17. Application of a novel microtitre plate-based assay for the discovery of new inhibitors of DNA gyrase and DNA topoisomerase VI.

    Science.gov (United States)

    Taylor, James A; Mitchenall, Lesley A; Rejzek, Martin; Field, Robert A; Maxwell, Anthony

    2013-01-01

    DNA topoisomerases are highly exploited targets for antimicrobial drugs. The spread of antibiotic resistance represents a significant threat to public health and necessitates the discovery of inhibitors that target topoisomerases in novel ways. However, the traditional assays for topoisomerase activity are not suitable for the high-throughput approaches necessary for drug discovery. In this study we validate a novel assay for screening topoisomerase inhibitors. A library of 960 compounds was screened against Escherichia coli DNA gyrase and archaeal Methanosarcina mazei DNA topoisomerase VI. Several novel inhibitors were identified for both enzymes, and subsequently characterised in vitro and in vivo. Inhibitors from the M. mazei topoisomerase VI screen were tested for their ability to inhibit Arabidopsis topoisomerase VI in planta. The data from this work present new options for antibiotic drug discovery and provide insight into the mechanism of topoisomerase VI.

  18. Hydrogen and methane production from desugared molasses using a two‐stage thermophilic anaerobic process

    DEFF Research Database (Denmark)

    Kongjan, Prawit; O-Thong, Sompong; Angelidaki, Irini

    2013-01-01

    Hydrogen and methane production from desugared molasses by a two‐stage thermophilic anaerobic process was investigated in a series of two up‐flow anaerobic sludge blanket (UASB) reactors. The first reactor that was dominated with hydrogen‐producing bacteria of Thermoanaerobacterium...... thermosaccharolyticum and Thermoanaerobacterium aciditolerans could generate a high hydrogen production rate of 5600 mL H2/day/L, corresponding to a yield of 132 mL H2/g volatile solid (VS). The effluent from the hydrogen reactor was further converted to methane in the second reactor with the optimal production rate...... of 3380 mL CH4/day/L, corresponding to a yield of 239 mL CH4/g VS. Aceticlastic Methanosarcina mazei was the dominant methanogen in the methanogenesis stage. This work demonstrates that biohydrogen production can be very efficiently coupled with a subsequent step of methane production using desugared...

  19. iTRAQ quantitative proteomic analysis reveals the pathways for methanation of propionate facilitated by magnetite

    DEFF Research Database (Denmark)

    Jing, Yuhang; Wan, Jingjing; Angelidaki, Irini

    2017-01-01

    by around 44% in batch experiments, and both direct interspecies electron transfer and interspecies H2 transfer were thermodynamically feasible with the addition of magnetite. The methanation of propionate facilitated by magnetite was also demonstrated in a long-term operated continuous reactor. The methane...... enriched with the addition of magnetite. iTRAQ quantitative proteomic analysis, which was used in mixed culture for the first time, showed that magnetite induced the changes of protein expression levels involved in various pathways during the methanation of propionate. The up-regulation of proteins...... electron transfer considering its up-regulation with the addition of magnetite and origination from Thauera. Most of the up-regulated proteins in methane metabolism were originated from Methanosaeta, while most of the enzymes with down-regulated proteins were originated from Methanosarcina. However, the up-regulated...

  20. Localization of Methyl-Coenzyme M Reductase as Metabolic Marker for Diverse Methanogenic Archaea

    Directory of Open Access Journals (Sweden)

    Christoph Wrede

    2013-01-01

    Full Text Available Methyl-Coenzyme M reductase (MCR as key enzyme for methanogenesis as well as for anaerobic oxidation of methane represents an important metabolic marker for both processes in microbial biofilms. Here, the potential of MCR-specific polyclonal antibodies as metabolic marker in various methanogenic Archaea is shown. For standard growth conditions in laboratory culture, the cytoplasmic localization of the enzyme in Methanothermobacter marburgensis, Methanothermobacter wolfei, Methanococcus maripaludis, Methanosarcina mazei, and in anaerobically methane-oxidizing biofilms is demonstrated. Under growth limiting conditions on nickel-depleted media, at low linear growth of cultures, a fraction of 50–70% of the enzyme was localized close to the cytoplasmic membrane, which implies “facultative” membrane association of the enzyme. This feature may be also useful for assessment of growth-limiting conditions in microbial biofilms.

  1. Effects of temperature and hydraulic retention time on acetotrophic pathways and performance in high-rate sludge digestion.

    Science.gov (United States)

    Ho, Dang; Jensen, Paul; Batstone, Damien

    2014-06-01

    High-rate anaerobic digestion of organic solids requires rapid hydrolysis and enhanced methanogenic growth rates, which can be achieved through elevated temperature (>55 °C) at short hydraulic retention times (HRT). This study assesses the effect of temperatures between 55 °C and 65 °C and HRTs between 2 and 4 days on process performance, microbial community structure, microbial capability, and acetotrophic pathways in thermophilic anaerobic reactors. Increasing the temperature did not enhance volatile solids (VS) destruction above the base value of 37% achieved at 55 °C and 4 days HRT. Stable isotopic signatures (δ13C) revealed that elevated temperature promoted syntrophic acetate oxidation, which accounted for 60% of the methane formation at 55 °C, and increasing substantially to 100% at 65 °C. The acetate consumption capacity dropped with increasing temperature (from 0.69-0.81 gCOD gVS(-1) d(-1) at 55 °C to 0.21-0.35 gCOD gVS(-1) d(-1) at 65 °C), based on specific activity testing of reactor contents. Community analysis using 16S rRNA pyrosequencing revealed the dominance of Methanosarcina at 55-60 °C. However, a further increase to 65 °C resulted in loss of Methanosarcina, with an accumulation of organic acids and reduced methane production. Similar issues were observed when reducing the HRT to 2 days, indicating that temperature3 days are critical to operate these systems stably.

  2. Laser spectroscopic real time measurements of methanogenic activity under simulated Martian subsurface analog conditions

    Science.gov (United States)

    Schirmack, Janosch; Böhm, Michael; Brauer, Chris; Löhmannsröben, Hans-Gerd; de Vera, Jean-Pierre; Möhlmann, Diedrich; Wagner, Dirk

    2014-08-01

    On Earth, chemolithoautothrophic and anaerobic microorganisms such as methanogenic archaea are regarded as model organisms for possible subsurface life on Mars. For this reason, the methanogenic strain Methanosarcina soligelidi (formerly called Methanosarcina spec. SMA-21), isolated from permafrost-affected soil in northeast Siberia, has been tested under Martian thermo-physical conditions. In previous studies under simulated Martian conditions, high survival rates of these microorganisms were observed. In our study we present a method to measure methane production as a first attempt to study metabolic activity of methanogenic archaea during simulated conditions approaching conditions of Mars-like environments. To determine methanogenic activity, a measurement technique which is capable to measure the produced methane concentration with high precision and with high temporal resolution is needed. Although there are several methods to detect methane, only a few fulfill all the needed requirements to work within simulated extraterrestrial environments. We have chosen laser spectroscopy, which is a non-destructive technique that measures the methane concentration without sample taking and also can be run continuously. In our simulation, we detected methane production at temperatures down to -5 °C, which would be found on Mars either temporarily in the shallow subsurface or continually in the deep subsurface. The pressure of 50 kPa which we used in our experiments, corresponds to the expected pressure in the Martian near subsurface. Our new device proved to be fully functional and the results indicate that the possible existence of methanogenic archaea in Martian subsurface habitats cannot be ruled out.

  3. Methanogens at the top of the world: occurrence and potential activity of methanogens in newly deglaciated soils in high-altitude cold deserts in the Western Himalayas.

    Science.gov (United States)

    Aschenbach, Katrin; Conrad, Ralf; Reháková, Klára; Doležal, Jiří; Janatková, Kateřina; Angel, Roey

    2013-01-01

    Methanogens typically occur in reduced anoxic environments. However, in recent studies it has been shown that many aerated upland soils, including desert soils also host active methanogens. Here we show that soil samples from high-altitude cold deserts in the western Himalayas (Ladakh, India) produce CH4 after incubation as slurry under anoxic conditions at rates comparable to those of hot desert soils. Samples of matured soil from three different vegetation belts (arid, steppe, and subnival) were compared with younger soils originating from frontal and lateral moraines of receding glaciers. While methanogenic rates were higher in the samples from matured soils, CH4 was also produced in the samples from the recently deglaciated moraines. In both young and matured soils, those covered by a biological soil crust (biocrust) were more active than their bare counterparts. Isotopic analysis showed that in both cases CH4 was initially produced from H2/CO2 but later mostly from acetate. Analysis of the archaeal community in the in situ soil samples revealed a clear dominance of sequences related to Thaumarchaeota, while the methanogenic community comprised only a minor fraction of the archaeal community. Similar to other aerated soils, the methanogenic community was comprised almost solely of the genera Methanosarcina and Methanocella, and possibly also Methanobacterium in some cases. Nevertheless, ~10(3) gdw(-1) soil methanogens were already present in the young moraine soil together with cyanobacteria. Our results demonstrate that Methanosarcina and Methanocella not only tolerate atmospheric oxygen but are also able to survive in these harsh cold environments. Their occurrence in newly deglaciated soils shows that they are early colonizers of desert soils, similar to cyanobacteria, and may play a role in the development of desert biocrusts.

  4. Hexavalent Chromium Reduction under Fermentative Conditions with Lactate Stimulated Native Microbial Communities

    Energy Technology Data Exchange (ETDEWEB)

    Somenahally, Anil C [ORNL; Mosher, Jennifer J [ORNL; Yuan, Tong [University of Oklahoma; Phelps, Tommy Joe [ORNL; Brown, Steven D [ORNL; Yang, Zamin Koo [ORNL; Hazen, Terry C [ORNL; Arkin, Adam [Lawrence Berkeley National Laboratory (LBNL); Palumbo, Anthony Vito [ORNL; Van Nostrand, Dr. Joy D. [Oklahoma University; Zhou, Jizhong [University of Oklahoma; Elias, Dwayne A [ORNL

    2013-01-01

    Microbial reduction of toxic hexavalent chromium (Cr(VI)) in-situ is a plausible bioremediation strategy in electron-acceptor limited environments. However, higher [Cr(VI)] may impose stress on syntrophic communities and impact community structure and function. The study objectives were to understand the impacts of Cr(VI) concentrations on community structure and on the Cr(VI)-reduction potential of groundwater communities at Hanford, WA. Steady state continuous flow bioreactors were used to grow native communities enriched with lactate (30 mM) and continuously amended with Cr(VI) at 0.0 (No-Cr), 0.1 (Low-Cr) and 3.0 (High-Cr) mg/L. Microbial growth, metabolites, Cr(VI), 16S rRNA gene sequences and GeoChip based functional gene composition were monitored for 15 weeks. Temporal trends and differences in growth, metabolite profiles, and community composition were observed, largely between Low-Cr and High-Cr bioreactors. In both High-Cr and Low-Cr bioreactors, Cr(VI) levels were below detection from week 1 until week 15. With lactate enrichment, native bacterial diversity substantially decreased as Pelosinus spp., and Sporotalea spp., became the dominant groups, but did not significantly differ between Cr concentrations. The Archaea diversity also substantially decreased after lactate enrichment from Methanosaeta (35%), Methanosarcina (17%) and others, to mostly Methanosarcina spp. (95%). Methane production was lower in High-Cr reactors suggesting some inhibition of methanogens. Several key functional genes were distinct in Low-Cr bioreactors compared to High-Cr. Among the Cr resistant microbes, Burkholderia vietnamiensis, Comamonas testosterone and Ralstonia pickettii proliferated in Cr amended bioreactors. In-situ fermentative conditions facilitated Cr(VI) reduction, and as a result 3.0 mg/L Cr(VI) did not impact the overall bacterial community structure.

  5. Hexavalent chromium reduction under fermentative conditions with lactate stimulated native microbial communities.

    Science.gov (United States)

    Somenahally, Anil C; Mosher, Jennifer J; Yuan, Tong; Podar, Mircea; Phelps, Tommy J; Brown, Steven D; Yang, Zamin K; Hazen, Terry C; Arkin, Adam P; Palumbo, Anthony V; Van Nostrand, Joy D; Zhou, Jizhong; Elias, Dwayne A

    2013-01-01

    Microbial reduction of toxic hexavalent chromium (Cr(VI)) in-situ is a plausible bioremediation strategy in electron-acceptor limited environments. However, higher [Cr(VI)] may impose stress on syntrophic communities and impact community structure and function. The study objectives were to understand the impacts of Cr(VI) concentrations on community structure and on the Cr(VI)-reduction potential of groundwater communities at Hanford, WA. Steady state continuous flow bioreactors were used to grow native communities enriched with lactate (30 mM) and continuously amended with Cr(VI) at 0.0 (No-Cr), 0.1 (Low-Cr) and 3.0 (High-Cr) mg/L. Microbial growth, metabolites, Cr(VI), 16S rRNA gene sequences and GeoChip based functional gene composition were monitored for 15 weeks. Temporal trends and differences in growth, metabolite profiles, and community composition were observed, largely between Low-Cr and High-Cr bioreactors. In both High-Cr and Low-Cr bioreactors, Cr(VI) levels were below detection from week 1 until week 15. With lactate enrichment, native bacterial diversity substantially decreased as Pelosinus spp., and Sporotalea spp., became the dominant groups, but did not significantly differ between Cr concentrations. The Archaea diversity also substantially decreased after lactate enrichment from Methanosaeta (35%), Methanosarcina (17%) and others, to mostly Methanosarcina spp. (95%). Methane production was lower in High-Cr reactors suggesting some inhibition of methanogens. Several key functional genes were distinct in Low-Cr bioreactors compared to High-Cr. Among the Cr resistant microbes, Burkholderia vietnamiensis, Comamonas testosterone and Ralstonia pickettii proliferated in Cr amended bioreactors. In-situ fermentative conditions facilitated Cr(VI) reduction, and as a result 3.0 mg/L Cr(VI) did not impact the overall bacterial community structure.

  6. Influence of hexavalent chromium on lactate-enriched Hanford groundwater microbial communities.

    Energy Technology Data Exchange (ETDEWEB)

    Somenahally, Anil C [ORNL; Mosher, Jennifer J [ORNL; Yuan, Tong [University of Oklahoma; Podar, Mircea [ORNL; Phelps, Tommy Joe [ORNL; Brown, Steven D [ORNL; Yang, Zamin Koo [ORNL; Hazen, Terry C [ORNL; Arkin, Adam [Lawrence Berkeley National Laboratory (LBNL); Palumbo, Anthony Vito [ORNL; Zhou, Jizhong [University of Oklahoma; Elias, Dwayne A [ORNL

    2013-01-01

    Microbial reduction and immobilization of chromate (Cr(VI)) is a plausible bioremediation strategy. However, higher Cr(VI) concentrations may impose stress on native Cr-reducing communities. We sought to determine if Cr(VI) would influence the lactate enriched native microbial community structure and function in groundwater from the Cr contaminated site at Hanford, WA. Steady state continuous flow bioreactors were amended with lactate and Cr(VI) (0.0, 0.1 and 3.0 mg/L). Microbial growth, metabolites, Cr(VI) concentrations, 16S rRNA gene sequences and GeoChip based functional gene composition in bioreactors were monitored for 15 weeks. Temporal trends and some differences in growth, metabolite profiles, and community composition were observed, largely between Low-Cr and High-Cr bioreactors. In both High-Cr and Low-Cr bioreactors, Cr(VI) was reduced in the bioreactors. With lactate enrichment, the native communities did not significantly differ between Cr concentrations. Native bacterial communities were diverse, whereas after lactate enrichment, Pelosinus spp., and Sporotalea spp., were the most predominant groups in all bioreactors. Similarly, the Archaea diversity significantly decreased from Methanosaeta (35%), Methanosarcina (17%), Halobacteriales (12%), Methanoregula (8%) and others, to mostly Methanosarcina spp. (95%) after lactate enrichment. Composition of several key functional genes was distinct in Low-Cr bioreactors compared to High-Cr. Among the Cr resistant probes (chrA), Burkholderia vietnamiensis, Comamonas testosterone and Ralstonia pickettii proliferated in Cr amended bioreactors. In-situ fermentative conditions facilitated Cr(VI) reduction, and as a result the 3.0 mg/L Cr(VI) did not appear to give chromate reducing strains a competitive advantage for proliferation or for increasing Cr-reduction.

  7. Hexavalent chromium reduction under fermentative conditions with lactate stimulated native microbial communities.

    Directory of Open Access Journals (Sweden)

    Anil C Somenahally

    Full Text Available Microbial reduction of toxic hexavalent chromium (Cr(VI in-situ is a plausible bioremediation strategy in electron-acceptor limited environments. However, higher [Cr(VI] may impose stress on syntrophic communities and impact community structure and function. The study objectives were to understand the impacts of Cr(VI concentrations on community structure and on the Cr(VI-reduction potential of groundwater communities at Hanford, WA. Steady state continuous flow bioreactors were used to grow native communities enriched with lactate (30 mM and continuously amended with Cr(VI at 0.0 (No-Cr, 0.1 (Low-Cr and 3.0 (High-Cr mg/L. Microbial growth, metabolites, Cr(VI, 16S rRNA gene sequences and GeoChip based functional gene composition were monitored for 15 weeks. Temporal trends and differences in growth, metabolite profiles, and community composition were observed, largely between Low-Cr and High-Cr bioreactors. In both High-Cr and Low-Cr bioreactors, Cr(VI levels were below detection from week 1 until week 15. With lactate enrichment, native bacterial diversity substantially decreased as Pelosinus spp., and Sporotalea spp., became the dominant groups, but did not significantly differ between Cr concentrations. The Archaea diversity also substantially decreased after lactate enrichment from Methanosaeta (35%, Methanosarcina (17% and others, to mostly Methanosarcina spp. (95%. Methane production was lower in High-Cr reactors suggesting some inhibition of methanogens. Several key functional genes were distinct in Low-Cr bioreactors compared to High-Cr. Among the Cr resistant microbes, Burkholderia vietnamiensis, Comamonas testosterone and Ralstonia pickettii proliferated in Cr amended bioreactors. In-situ fermentative conditions facilitated Cr(VI reduction, and as a result 3.0 mg/L Cr(VI did not impact the overall bacterial community structure.

  8. Methanogens at the top of the world: occurrence and potential activity of methanogens in newly deglaciated soils in high-altitude cold deserts in the Western Himalayas

    Directory of Open Access Journals (Sweden)

    Katrin eAschenbach

    2013-12-01

    Full Text Available Methanogens typically occur in reduced anoxic environments. However, in recent studies it has been shown that many aerated upland soils, including desert soils also host active methanogens. Here we show that soil samples from high–altitude cold deserts in the western Himalayas (Ladakh, India produce CH4 after incubation as slurry under anoxic conditions at rates comparable to those of hot desert soils. Samples of matured soil from three different vegetation belts (arid, steppe, and subnival were compared with younger soils originating from frontal and lateral moraines of receding glaciers. While methanogenic rates were higher in the samples from matured soils, CH4 was also produced in the samples from the recently deglaciated moraines. In both young and matured soils, those covered by a biological soil crust (biocrust were more active than their bare counterparts. Isotopic analysis showed that in both cases CH4 was initially produced from H2/CO2 but later mostly from acetate. Analysis of the archaeal community in the in situ soil samples revealed a clear dominance of sequences related to Thaumarchaeota, while the methanogenic community comprised only a minor fraction of the archaeal community. Similar to other aerated soils, the methanogenic community was comprised almost solely of the genera Methanosarcina and Methanocella, and possibly also Methanobacterium in some cases. Nevertheless, approximately 103 gdw-1 soil methanogens were already present in the young moraine soil together with cyanobacteria. Our results demonstrate that Methanosarcina and Methanocella not only tolerate atmospheric oxygen but are also able to survive in these harsh cold environments. Their occurrence in newly deglaciated soils shows that they are early colonisers of desert soils, similar to cyanobacteria, and may play a role in the development of desert biocrusts.

  9. One carbon metabolism in anaerobic bacteria: Regulation of carbon and electron flow during organic acid production

    Energy Technology Data Exchange (ETDEWEB)

    Zeikus, J.G.; Jain, M.

    1993-12-31

    The project deals with understanding the fundamental biochemical mechanisms that physiologically control and regulate carbon and electron flow in anaerobic chemosynthetic bacteria that couple metabolism of single carbon compounds and hydrogen to the production of organic acids (formic, acetic, butyric, and succinic) or methane. The authors compare the regulation of carbon dioxide and hydrogen metabolism by fermentation, enzyme, and electron carrier analysis using Butyribacterium methylotrophicum, Anaeroblospirillum succiniciproducens, Methanosarcina barkeri, and a newly isolated tri-culture composed of a syntrophic butyrate degrader strain IB, Methanosarcina mazei and Methanobacterium formicicum as model systems. To understand the regulation of hydrogen metabolism during butyrate production or acetate degradation, hydrogenase activity in B. methylotrophicum or M. barkeri is measured in relation to growth substrate and pH; hydrogenase is purified and characterized to investigate number of hydrogenases; their localization and functions; and, their sequences are determined. To understand the mechanism for catabolic CO{sub 2} fixation to succinate the PEP carboxykinase enzyme and gene of A. succiniciproducens are purified and characterized. Genetically engineered strains of Escherichia coli containing the phosphoenolpyruvate (PEP) carboxykinase gene are examined for their ability to produce succinate in high yield. To understand the mechanism of fatty acid degradation by syntrophic acetogens during mixed culture methanogenesis formate and hydrogen production are characterized by radio tracer studies. It is intended that these studies provide strategies to improve anaerobic fermentations used for the production of organic acids or methane and, new basic understanding on catabolic CO{sub 2} fixation mechanisms and on the function of hydrogenase in anaerobic bacteria.

  10. Magnetic properties and interlayer coupling of epitaxial Co/Cu films on Si

    Energy Technology Data Exchange (ETDEWEB)

    Mansell, R.; Petit, D. C. M. C.; Fernández-Pacheco, A.; Lavrijsen, R.; Lee, J. H.; Cowburn, R. P. [Cavendish Laboratory, University of Cambridge, JJ Thomson Avenue, Cambridge CB3 0HE (United Kingdom)

    2014-08-14

    Thin films of Co and Co/Cu/Co trilayers with wedged Cu interlayers were grown epitaxially on Cu buffer layers on hydrogen passivated Si(001) wafers. We find that single Co layers have a well-defined four-fold anisotropy but with smaller in-plane anisotropies than observed in Co grown on Cu crystals. Ruderman–Kittel–Kasuya–Yosida (RKKY) interlayer coupling is observed in one Co/Cu/Co sample which is the smoothest of the films as measured by atomic force microscopy. Some of the films also form a dot-like structure on the surface. Intermixing at elevated temperatures between the Cu buffer and Si limits the ability to form flat surfaces to promote RKKY coupling.

  11. Spin-Orbit Effects in Spin-Resolved L2,3 Core Level Photoemission of 3d Ferromagnetic Thin Films

    Energy Technology Data Exchange (ETDEWEB)

    Komesu, T; Waddill, G D; Yu, S W; Butterfield, M; Tobin, J G

    2007-10-02

    We present spin-resolved 2p core level photoemission for the 3d transition metal films of Fe and Co grown on Cu(100). We observe clear spin asymmetry in the main 2p core level photoemission peaks of Fe and Co films consistent with trends in the bulk magnetic moments. The spin polarization can be strongly enhanced, by variation of the experimental geometry, when the photoemission is undertaken with circularly polarized light, indicating that spin-orbit interaction can have a profound in spin polarized photoemission. Further spin polarized photoemission studies using variable circularly polarized light at high photon energies, high flux are indicated, underscoring the value of synchrotron measurements at facilities with increased beam stability.

  12. Performance of a three-stage membrane bioprocess treating the organic fraction of municipal solid waste and evolution of its archaeal and bacterial ecology.

    Science.gov (United States)

    Trzcinski, Antoine P; Ray, Michael J; Stuckey, David C

    2010-03-01

    A novel three-stage bioprocess achieved 75% volatile solids (VS) removal at an organic loading rate (OLR) of 4g VSL(-1)day, a solids retention time (SRT) of 66days, a hydraulic retention time (HRT) of 20days, at a temperature of 35 degrees C. The bioprocess consisted of an anaerobic hydrolytic reactor (HR) where the solids and liquid fractions of the Organic Fraction of the Municipal Solid Waste (OFMSW) were separated with a mesh. The leachate was pumped to a Submerged Anaerobic Membrane Bioreactor (SAMBR) and the treated permeate was polished in an Aerobic Membrane Bioreactor (AMBR). Denaturing Gradient Gel Electrophoresis (DGGE) and DNA sequencing analyses indicated that the increase in methane content in the HR caused by the excess sludge recycle from the SAMBR was associated with an increase in the number of hydrogenotrophic species, mainly Methanobrevibacter sp., Methanobacterium formicicum and Methanosarcina sp. At 20 degrees C VS removal dropped to 50% in the HR and some DGGE bands disappeared when compared to 35 degrees C samples, while some bands such as the one corresponding to Ruminococcus flavefaciens were reduced in intensity. The species associated with the COD-polishing properties of the AMBR correspond to the genera Pseudomonas, Hyphomonas and Hyphomicrobiaceae. These results highlight the positive effect of recycling the excess sludge from the SAMBR to re-inoculate the HR with hydrogenotrophic species.

  13. Chemical shifts assignments of the archaeal MC1 protein and a strongly bent 15 base pairs DNA duplex in complex.

    Science.gov (United States)

    Loth, Karine; Landon, Céline; Paquet, Françoise

    2015-04-01

    MC1 is the most abundant architectural protein present in Methanosarcina thermophila CHTI55 in laboratory growth conditions and is structurally unrelated to other DNA-binding proteins. MC1 functions are to shape and to protect DNA against thermal denaturation by binding to it. Therefore, MC1 has a strong affinity for any double-stranded DNA. However, it recognizes and preferentially binds to bent DNA, such as four-way junctions and negatively supercoiled DNA minicircles. Combining NMR data, electron microscopy data, biochemistry, molecular modelisation and docking approaches, we proposed recently a new type of DNA/protein complex, in which the monomeric protein MC1 binds on the concave side of a strongly bent 15 base pairs DNA. We present here the NMR chemical shifts assignments of each partner in the complex, (1)H (15)N MC1 protein and (1)H (13)C (15)N bent duplex DNA, as first step towards the first experimental 3D structure of this new type of DNA/protein complex.

  14. Methanogenic community dynamics in anaerobic co-digestion of fruit and vegetable waste and food waste

    Institute of Scientific and Technical Information of China (English)

    Jia Lin; Jiane Zuo; Ruofan Ji; Xiaojie Chen; Fenglin Liu; Kaijun Wang; Yunfeng Yang

    2012-01-01

    A lab-scale continuously-stirred tank reactor (CSTR),used for anaerobic co-digestion of fruit and vegetable waste (FVW) and food waste (FW) at different mixture ratios,was operated for 178 days at the organic loading rate of 3 kg VS (volatile solids)/(m3.day).The dynamics of the Archaeal community and the correlations between environmental variables and methanogenic community structure were analyzed by polymerase chain reactions - denaturing gradient gel electrophoresis (PCR-DGGE) and redundancy analysis (RDA),respectively.PCR-DGGE results demonstrated that the mixture ratio of FVW to FW altered the community composition of Aachaea.As the FVW/FW ratio increased,Methanoculleus,Methanosaeta and Methanosarcina became the predominant methanogens in the community.Redundancy analysis results indicated that the shift of the methanogenic community was significantly correlated with the composition of acidogenic products and methane production yield.Different mixture ratios of substrates led to different compositions of intermediate metabolites,which may affect the methanogenic community.These results suggested that the analysis of microbial communities could be used to diagnose anaerobic processes.

  15. Stereochemical studies of acyclic isoprenoids-XII. Lipids of methanogenic bacteria and possible contributions to sediments

    Science.gov (United States)

    Risatti, J.B.; Rowland, S.J.; Yon, D.A.; Maxwell, J.R.

    1984-01-01

    Abundant volatile lipids of Methanobacterium thermoautotrophicum and Methanosarcina barkeri include isoprenoid hydrocarbons (??? C30), and C15, C20 and C25 isoprenoid alcohols. M. barkeri contains 2,6,10,15,19-pentamethyleicosane, whose relative stereochemistry is the same as found in marine sediments, indicating that it is a marker of methanogenic activity. The C20, C30 and C25 alkenes in M. thermoautotrophicum also have a preferred sterochemistry; the latter have the 2,6,10,14,18-pentamethyleicosanyl skeleton, suggesting that the alkane in marine sediments may derive from methanogens. The stereochemistry of squalane in a marine sediment is also compatible with an origin in methanogens; in contrast, the stereochemistry of pristane in M. thermoautotrophicum indicates a fossil fuel contaminant origin, suggesting that this and certain other alkanes reported in archaebacteria might also be of contaminant origin. There is, therefore, little evidence at present that the pristane in immature marine sediments originates in methanogens. The C15 and C20 saturated alcohols in M. thermoautotrophicum have mainly the all-R configuration. If this is generally true for methanogens, the C20 alcohol in the Messel shale may originate mainly from methanogens, whereas that in the Green River shale may originate mainly from photosynthetic organisms. ?? 1984.

  16. Ribosomal intergenic spacer analysis as a tool for monitoring methanogenic Archaea changes in an anaerobic digester.

    Science.gov (United States)

    Ciesielski, Slawomir; Bułkowska, Katarzyna; Dabrowska, Dorota; Kaczmarczyk, Dariusz; Kowal, Przemyslaw; Możejko, Justyna

    2013-08-01

    The applicability of a newly-designed PCR primer pair in examination of methanogenic Archaea in a digester treating plant biomass was evaluated by Ribosmal Intergenic Spacer Analysis (RISA). To find a suitable approach, three variants of RISA were tested: (1) standard, polyacrylamide gel-based, (2) automated, utilized capillary electrophoresis (GA-ARISA), and (3) automated microfluidics-based (MF-ARISA). All three techniques yielded a consistent picture of archaeal community structure changes during anaerobic digestion monitored for more than 6 weeks. While automated variants were more practical for handling and rapid analysis of methanogenic Archaea, the gel-based technique was advantageous when micro-organism identification was required. A DNA-sequence analysis of dominant bands extracted from the gel revealed that the main role in methane synthesis was played by micro-organisms affiliated with Methanosarcina barkeri. The obtained results revealed that RISA is a robust method allowing for detailed analysis of archaeal community structure during organic biomass conversion into biogas. In addition, our results showed that GA-ARISA has a higher resolution and reproducibility than other variants of RISA and could be used as a technique for tracking changes in methanogenic Archaea in an anaerobic digester.

  17. Dynamic variation of the microbial community structure during the long-time mono-fermentation of maize and sugar beet silage

    Science.gov (United States)

    Klang, Johanna; Theuerl, Susanne; Szewzyk, Ulrich; Huth, Markus; Tölle, Rainer; Klocke, Michael

    2015-01-01

    This study investigated the development of the microbial community during a long-term (337 days) anaerobic digestion of maize and sugar beet silage, two feedstocks that significantly differ in their chemical composition. For the characterization of the microbial dynamics, the community profiling method terminal restriction fragment length polymorphism (TRFLP) in combination with a cloning-sequencing approach was applied. Our results revealed a specific adaptation of the microbial community to the supplied feedstocks. Based on the high amount of complex compounds, the anaerobic conversion rate of maize silage was slightly lower compared with the sugar beet silage. It was demonstrated that members from the phylum Bacteroidetes are mainly involved in the degradation of low molecular weight substances such as sugar, ethanol and acetate, the main compounds of the sugar beet silage. It was further shown that species of the genus Methanosaeta are highly sensitive against sudden stress situations such as a strong decrease in the ammonium nitrogen (NH4+-N) concentration or a drop of the pH value. In both cases, a functional compensation by members of the genera Methanoculleus and/or Methanosarcina was detected. However, the overall biomass conversion of both feedstocks proceeded efficiently as a steady state between acid production and consumption was recorded, which further resulted in an equal biogas yield. PMID:25712194

  18. Functionally redundant but dissimilar microbial communities within biogas reactors treating maize silage in co-fermentation with sugar beet silage

    Science.gov (United States)

    Langer, Susanne G; Ahmed, Sharif; Einfalt, Daniel; Bengelsdorf, Frank R; Kazda, Marian

    2015-01-01

    Numerous observations indicate a high flexibility of microbial communities in different biogas reactors during anaerobic digestion. Here, we describe the functional redundancy and structural changes of involved microbial communities in four lab-scale continuously stirred tank reactors (CSTRs, 39°C, 12 L volume) supplied with different mixtures of maize silage (MS) and sugar beet silage (SBS) over 80 days. Continuously stirred tank reactors were fed with mixtures of MS and SBS in volatile solid ratios of 1:0 (Continuous Fermenter (CF) 1), 6:1 (CF2), 3:1 (CF3), 1:3 (CF4) with equal organic loading rates (OLR 1.25 kgVS m−3 d−1) and showed similar biogas production rates in all reactors. The compositions of bacterial and archaeal communities were analysed by 454 amplicon sequencing approach based on 16S rRNA genes. Both bacterial and archaeal communities shifted with increasing amounts of SBS. Especially pronounced were changes in the archaeal composition towards Methanosarcina with increasing proportion of SBS, while Methanosaeta declined simultaneously. Compositional shifts within the microbial communities did not influence the respective biogas production rates indicating that these communities adapted to environmental conditions induced by different feedstock mixtures. The diverse microbial communities optimized their metabolism in a way that ensured efficient biogas production. PMID:26200922

  19. Microbial population dynamics during startup of a full-scale anaerobic digester treating industrial food waste in Kyoto eco-energy project.

    Science.gov (United States)

    Ike, Michihiko; Inoue, Daisuke; Miyano, Tomoki; Liu, Tong Tong; Sei, Kazunari; Soda, Satoshi; Kadoshin, Shiro

    2010-06-01

    The microbial community in a full-scale anaerobic digester (2300m3) treating industrial food waste in the Kyoto Eco-Energy Project was analyzed using terminal restriction fragment length polymorphism for eubacterial and archaeal 16S rRNA genes. Both thermophilic and mesophilic sludge of treated swine waste were seeded to the digestion tank. During the 150-day startup period, coffee grounds as a main food waste, along with potato, kelp and boiled beans, tofu, bean curd lees, and deep-fried bean curd were fed to the digestion process step-by-step (max. 40t/d). Finally, the methane yield reached 360m3/t-feed with 40days' retention time, although temporary accumulation of propionate was observed. Eubacterial communities that formed in the thermophilic digestion tank differed greatly from both thermophilic and mesophilic types of seed sludge. Results suggest that the Actinomyces/Thermomonospora and Ralstonia/Shewanella were contributors for hydrolyzation and degradation of food waste into volatile fatty acids. Acetate-utilizing methanogens, Methanosaeta, were dominant in seed sludges of both types, but they decreased drastically during processing in the digestion tank. Methanosarcina and Methanobrevibacter/Methanobacterium were, respectively, possible main contributors for methane production from acetate and H2 plus CO2. Copyright 2010 Elsevier Ltd. All rights reserved.

  20. Stimulation of methane generation from nonproductive coal by addition of nutrients or a microbial consortium.

    Science.gov (United States)

    Jones, Elizabeth J P; Voytek, Mary A; Corum, Margo D; Orem, William H

    2010-11-01

    Biogenic formation of methane from coal is of great interest as an underexploited source of clean energy. The goal of some coal bed producers is to extend coal bed methane productivity and to utilize hydrocarbon wastes such as coal slurry to generate new methane. However, the process and factors controlling the process, and thus ways to stimulate it, are poorly understood. Subbituminous coal from a nonproductive well in south Texas was stimulated to produce methane in microcosms when the native population was supplemented with nutrients (biostimulation) or when nutrients and a consortium of bacteria and methanogens enriched from wetland sediment were added (bioaugmentation). The native population enriched by nutrient addition included Pseudomonas spp., Veillonellaceae, and Methanosarcina barkeri. The bioaugmented microcosm generated methane more rapidly and to a higher concentration than the biostimulated microcosm. Dissolved organics, including long-chain fatty acids, single-ring aromatics, and long-chain alkanes accumulated in the first 39 days of the bioaugmented microcosm and were then degraded, accompanied by generation of methane. The bioaugmented microcosm was dominated by Geobacter sp., and most of the methane generation was associated with growth of Methanosaeta concilii. The ability of the bioaugmentation culture to produce methane from coal intermediates was confirmed in incubations of culture with representative organic compounds. This study indicates that methane production could be stimulated at the nonproductive field site and that low microbial biomass may be limiting in situ methane generation. In addition, the microcosm study suggests that the pathway for generating methane from coal involves complex microbial partnerships.

  1. Biogas production from coumarin-rich plants--inhibition by coumarin and recovery by adaptation of the bacterial community.

    Science.gov (United States)

    Popp, Denny; Schrader, Steffi; Kleinsteuber, Sabine; Harms, Hauke; Sträuber, Heike

    2015-09-01

    Plants like sweet clover (Melilotus spp.) are not suitable as fodder for cattle because of harmful effects of the plant secondary metabolite coumarin. As an alternative usage, the applicability of coumarin-rich plants as substrates for biogas production was investigated. When coumarin was added to continuous fermentation processes codigesting grass silage and cow manure, it caused a strong inhibition noticeable as decrease of biogas production by 19% and increase of metabolite concentrations to an organic acids/alkalinity ratio higher than 0.3(gorganic acids) gCaCO3 (-1). Microbial communities of methanogenic archaea were dominated by the genera Methanosarcina (77%) and Methanoculleus (11%). This community composition was not influenced by coumarin addition. The bacterial community analysis unraveled a divergence caused by coumarin addition correlating with the anaerobic degradation of coumarin and the recovery of the biogas process. As a consequence, biogas production resumed similar to the coumarin-free control with a biogas yield of 0.34 LN g(volatile solids) (-1) and at initial metabolite concentrations (∼ 0.2 g(organic acids) gCaCO3 (-1)). Coumarin acts as inhibitor and as substrate during anaerobic digestion. Hence, coumarin-rich plants might be suitable for biogas production, but should only be used after adaptation of the microbial community to coumarin.

  2. Potential Role of Acetyl-CoA Synthetase (acs) and Malate Dehydrogenase (mae) in the Evolution of the Acetate Switch in Bacteria and Archaea.

    Science.gov (United States)

    Barnhart, Elliott P; McClure, Marcella A; Johnson, Kiki; Cleveland, Sean; Hunt, Kristopher A; Fields, Matthew W

    2015-08-03

    Although many Archaea have AMP-Acs (acetyl-coenzyme A synthetase) and ADP-Acs, the extant methanogenic genus Methanosarcina is the only identified Archaeal genus that can utilize acetate via acetate kinase (Ack) and phosphotransacetylase (Pta). Despite the importance of ack as the potential urkinase in the ASKHA phosphotransferase superfamily, an origin hypothesis does not exist for the acetate kinase in Bacteria, Archaea, or Eukarya. Here we demonstrate that Archaeal AMP-Acs and ADP-Acs contain paralogous ATPase motifs previously identified in Ack, which demonstrate a novel relation between these proteins in Archaea. The identification of ATPase motif conservation and resulting structural features in AMP- and ADP-acetyl-CoA synthetase proteins in this study expand the ASKHA superfamily to include acetyl-CoA synthetase. Additional phylogenetic analysis showed that Pta and MaeB sequences had a common ancestor, and that the Pta lineage within the halophilc archaea was an ancestral lineage. These results suggested that divergence of a duplicated maeB within an ancient halophilic, archaeal lineage formed a putative pta ancestor. These results provide a potential scenario for the establishment of the Ack/Pta pathway and provide novel insight into the evolution of acetate metabolism for all three domains of life.

  3. Vertical profiles of community abundance and diversity of anaerobic methanotrophic archaea (ANME) and bacteria in a simple waste landfill in north China.

    Science.gov (United States)

    Dong, Jun; Ding, Linjie; Wang, Xu; Chi, Zifang; Lei, Jiansen

    2015-03-01

    Anaerobic methane oxidation (AMO) is considered to be an important sink of CH4 in habitats as marine sediments. But, few studies focused on AMO in landfills which may be an important sink of CH4 derived from waste fermentation. To show evidence of AMO and to uncover function anaerobic methanotroph (ANME) community in landfill, different age waste samples were collected in Jinqianpu landfill located in north China. Through high-throughput sequencing, Methanomicrobiales and Methanosarcinales archaea associated with ANME and reverse methanogenic archaea of Methanosarcina and Methanobacterium were detected. Sulfate-reducing bacteria (SRB) (Desulfobulbus and Desulfococcus) which could couple with ANME-conducting AMO were also found. But, the community structure of ANME had no significant difference with depths. From the results of investigation, we can come to a conclusion that sulfate-dependent anaerobic methane oxidation (SR-DAMO) would be the dominant AMO process in the landfill, while iron-dependent anaerobic methane oxidation (M/IR-DAMO) process was weak though concentration of ferric iron was large in the landfill. Denitrification-dependent anaerobic methane oxidation (NR-DAMO) was negative because of lack of nitrate and relevant function microorganisms in the landfill. Results also indicate that CH4 mitigation would have higher potential by increasing electron acceptor contents and promoting the growth of relevant function microorganisms.

  4. Functionally redundant but dissimilar microbial communities within biogas reactors treating maize silage in co-fermentation with sugar beet silage.

    Science.gov (United States)

    Langer, Susanne G; Ahmed, Sharif; Einfalt, Daniel; Bengelsdorf, Frank R; Kazda, Marian

    2015-09-01

    Numerous observations indicate a high flexibility of microbial communities in different biogas reactors during anaerobic digestion. Here, we describe the functional redundancy and structural changes of involved microbial communities in four lab-scale continuously stirred tank reactors (CSTRs, 39°C, 12 L volume) supplied with different mixtures of maize silage (MS) and sugar beet silage (SBS) over 80 days. Continuously stirred tank reactors were fed with mixtures of MS and SBS in volatile solid ratios of 1:0 (Continuous Fermenter (CF) 1), 6:1 (CF2), 3:1 (CF3), 1:3 (CF4) with equal organic loading rates (OLR 1.25 kgVS m(-3)  d(-1) ) and showed similar biogas production rates in all reactors. The compositions of bacterial and archaeal communities were analysed by 454 amplicon sequencing approach based on 16S rRNA genes. Both bacterial and archaeal communities shifted with increasing amounts of SBS. Especially pronounced were changes in the archaeal composition towards Methanosarcina with increasing proportion of SBS, while Methanosaeta declined simultaneously. Compositional shifts within the microbial communities did not influence the respective biogas production rates indicating that these communities adapted to environmental conditions induced by different feedstock mixtures. The diverse microbial communities optimized their metabolism in a way that ensured efficient biogas production.

  5. Dynamic variation of the microbial community structure during the long-time mono-fermentation of maize and sugar beet silage.

    Science.gov (United States)

    Klang, Johanna; Theuerl, Susanne; Szewzyk, Ulrich; Huth, Markus; Tölle, Rainer; Klocke, Michael

    2015-09-01

    This study investigated the development of the microbial community during a long-term (337 days) anaerobic digestion of maize and sugar beet silage, two feedstocks that significantly differ in their chemical composition. For the characterization of the microbial dynamics, the community profiling method terminal restriction fragment length polymorphism (TRFLP) in combination with a cloning-sequencing approach was applied. Our results revealed a specific adaptation of the microbial community to the supplied feedstocks. Based on the high amount of complex compounds, the anaerobic conversion rate of maize silage was slightly lower compared with the sugar beet silage. It was demonstrated that members from the phylum Bacteroidetes are mainly involved in the degradation of low molecular weight substances such as sugar, ethanol and acetate, the main compounds of the sugar beet silage. It was further shown that species of the genus Methanosaeta are highly sensitive against sudden stress situations such as a strong decrease in the ammonium nitrogen (NH₄(+)-N) concentration or a drop of the pH value. In both cases, a functional compensation by members of the genera Methanoculleus and/or Methanosarcina was detected. However, the overall biomass conversion of both feedstocks proceeded efficiently as a steady state between acid production and consumption was recorded, which further resulted in an equal biogas yield.

  6. Impact of aluminum chloride on process performance and microbial community structure of granular sludge in an upflow anaerobic sludge blanket reactor for natural rubber processing wastewater treatment.

    Science.gov (United States)

    Thanh, Nguyen Thi; Watari, Takahiro; Thao, Tran Phuong; Hatamoto, Masashi; Tanikawa, Daisuke; Syutsubo, Kazuaki; Fukuda, Masao; Tan, Nguyen Minh; Anh, To Kim; Yamaguchi, Takashi; Huong, Nguyen Lan

    In this study, granular sludge formation was carried out using an aluminum chloride supplement in an upflow anaerobic sludge blanket (UASB) reactor treating natural rubber processing wastewater. Results show that during the first 75 days after the start-up of the UASB reactor with an organic loading rate (OLR) of 2.65 kg-COD·m(-3)·day(-1), it performed stably with a removal of 90% of the total chemical oxygen demand (COD) and sludge still remained in small dispersed flocs. However, after aluminum chloride was added at a concentration of 300 mg·L(-1) and the OLR range was increased up to 5.32 kg-COD·m(-3)·day(-1), the total COD removal efficiency rose to 96.5 ± 2.6%, with a methane recovery rate of 84.9 ± 13.4%, and the flocs began to form granules. Massively parallel 16S rRNA gene sequencing of the sludge retained in the UASB reactor showed that total sequence reads of Methanosaeta sp. and Methanosarcina sp., reported to be the key organisms for granulation, increased after 311 days of operation. This indicates that the microbial community structure of the retained sludge in the UASB reactor at the end of the experiment gave a good account of itself in not only COD removal, but also granule formation.

  7. The microbial community of a passive biochemical reactor treating arsenic, zinc and sulfate-rich seepage

    Directory of Open Access Journals (Sweden)

    Susan Anne Baldwin

    2015-03-01

    Full Text Available Sulfidogenic biochemical reactors for metal removal that use complex organic carbon have been shown to be effective in laboratory studies, but their performance in the field is highly variable. Successful operation depends on the types of microorganisms supported by the organic matrix, and factors affecting the community composition are unknown. A molecular survey of a field-based biochemical reactor that had been removing zinc and arsenic for over six years revealed that the microbial community was dominated by methanogens related to Methanocorpusculum sp. and Methanosarcina sp., which co-occurred with Bacteroidetes environmental groups, such as Vadin HA17, in places where the organic matter was more degraded. The metabolic potential for organic matter decomposition by Ruminococcaceae was prevalent in samples with more pyrolysable carbon. Rhodobium- and Hyphomicrobium-related genera within the Rhizobiales Order that have the metabolic potential for dark hydrogen fermentation and methylotrophy, and unclassified Comamonadaceae were the dominant Proteobacteria. The unclassified environmental group Sh765B-TzT-29 was an important Delta-Proteobacteria group in this BCR, that co-occurred with the dominant Rhizobiales OTUs. Organic matter degradation is one driver for shifting the microbial community composition and therefore possibly the performance of these bioreactors over time.

  8. Microbial Community Succession During Lactate Amendment of Chromium Contaminated Groundwater Reveals a Predominance of Pelosinus spp.

    Energy Technology Data Exchange (ETDEWEB)

    Mosher, Jennifer J [ORNL; Phelps, Tommy Joe [ORNL; Drake, Meghan M [ORNL; Campbell, James H [ORNL; Moberly, James G [ORNL; Schadt, Christopher Warren [ORNL; Podar, Mircea [ORNL; Brown, Steven D [ORNL; Hazen, Terry [Lawrence Berkeley National Laboratory (LBNL); Arkin, Adam [Lawrence Berkeley National Laboratory (LBNL); Palumbo, Anthony Vito [ORNL; Faybishenko, Boris A [Lawrence Berkeley National Laboratory (LBNL); Elias, Dwayne A [ORNL

    2012-01-01

    Microbial community structure and metabolism in contaminated ecosystems are potentially controlled not only by the different populations within the community, but a myriad of dynamic physicochemical parameters as well. The goal of the current work was to determine the impact of organic acid enrichment, in this case lactate, on the succession of the native microbial community from a contaminated groundwater aquifer. Triplicate anaerobic, continuous-flow glass reactors were inoculated with Hanford 100-H groundwater and incubated for 95 days to obtain a stable, enriched community. The microbial community experienced a shift in the population dynamics over time to eventually form a community with far less diversity than the original. The final community was dominated by Pelosinus spp. and to a lesser degree, Acetobacterium spp. with small amounts of other bacteria and archaea including methanogens. The resultant diversity was far decreased from 63 genera within 12 phyla to 11 bacterial genera (from three phyla) and 2 archaeal genera (from one phylum). Isolation efforts were successful in attaining new species of Pelosinus and known members of Methanosarcina barkerii along with several sulfate- and Fe(III)- reducing consortia members. The continuous-flow reactors allowed for testing physiochemical factors with microbial community dynamics on a smaller, replicable, scale while also facilitating the isolation of several previously uncultured community members. These lab-scale simulations will presumably allow for a deeper understanding of the community metabolism with specific carbon amendments that can inform future in situ efforts.

  9. Low Pressure Tolerance by Methanogens in an Aqueous Environment: Implications for Subsurface Life on Mars

    Science.gov (United States)

    Mickol, R. L.; Kral, T. A.

    2016-09-01

    The low pressure at the surface of Mars (average: 6 mbar) is one potentially biocidal factor that any extant life on the planet would need to endure. Near subsurface life, while shielded from ultraviolet radiation, would also be exposed to this low pressure environment, as the atmospheric gas-phase pressure increases very gradually with depth. Few studies have focused on low pressure as inhibitory to the growth or survival of organisms. However, recent work has uncovered a potential constraint to bacterial growth below 25 mbar. The study reported here tested the survivability of four methanogen species (Methanothermobacter wolfeii, Methanosarcina barkeri, Methanobacterium formicicum, Methanococcus maripaludis) under low pressure conditions approaching average martian surface pressure (6 mbar - 143 mbar) in an aqueous environment. Each of the four species survived exposure of varying length (3 days - 21 days) at pressures down to 6 mbar. This research is an important stepping-stone to determining if methanogens can actively metabolize/grow under these low pressures. Additionally, the recently discovered recurring slope lineae suggest that liquid water columns may connect the surface to deeper levels in the subsurface. If that is the case, any organism being transported in the water column would encounter the changing pressures during the transport.

  10. Archaeal communities associated with roots of the common reed (Phragmites australis) in Beijing Cuihu Wetland.

    Science.gov (United States)

    Liu, Yin; Li, Hong; Liu, Qun Fang; Li, Yan Hong

    2015-05-01

    The richness, phylogeny and composition of archaeal community associated with the roots of common reed (Phragmites australis) growing in the Beijing Cuihu Wetland, China was investigated using a 16S rDNA library. In total, 235 individual sequences were collected, and a phylogenetic analysis revealed that 69.4 and 11.5 % of clones were affiliated with the Euryarchaeota and the Crenarchaeota, respectively. In Euryarchaeota, the archaeal community was dominated by species in following genera: Methanobacterium in the order Methanobacteriales (60.7 %); Methanoregula and Methanospirillum in the order Methanomicrobiales (20.2 %), and Methanomethylovorans, Methanosarcina and Methanosaeta in the order Methanosarcinales (17.2 %). Of 27 sequences assigned to uncultured Crenarchaeota, 22 were grouped into Group 1.3, and five grouped into Group 1.1b. Hence, the archaeal communities associated with reed roots are largely involved in methane production, and, to a lesser extent, in ammonia oxidization. Quantification of the archaeal amoA gene indicated that ammonia oxidizing archaea were more numerous in the rhizosphere soil than in the root tissue or surrounding water. A total of 19.1 % of the sequences were unclassified, suggesting that many unidentified archaea are probably involved in the reed wetland ecosystem.

  11. Cloning and sequencing of the genes coding for the A and B subunits of vacuolar-type Na(+)-ATPase from Enterococcus hirae. Coexistence of vacuolar- and F0F1-type ATPases in one bacterial cell.

    Science.gov (United States)

    Takase, K; Yamato, I; Kakinuma, Y

    1993-06-05

    The eubacterium Enterococcus hirae ATCC 9790 possesses a H(+)-translocating ATPase, and the deduced amino acid sequences of the genes coding for this enzyme have indicated that it is a typical F0F1-type ATPase (Shibata, C., Ehara, T., Tomura, K., Igarashi, K., and Kobayashi, H. (1992) J. Bacteriol. 174, 6117-6124). We cloned the ntpA and ntpB genes coding for the A and B subunits, respectively, of Na(+)-translocating ATPase from the same bacterium, and the full amino acid sequences of the two subunits were deduced from the nucleotide sequence. The A (593 amino acid residues) and B (458 amino acid residues) subunits were highly homologous (48-60% identical) to the A (large or alpha) and the B (small or beta) subunits, respectively, of vacuolar-type H(+)-ATPases which have been found in eukaryotic endomembrane systems (Neurospora crassa, Saccharomyces cerevisiae, Arabidopsis thaliana, and carrot) and archaebacterial cell membranes (Sulfolobus acidocaldarius and Methanosarcina barkeri). The A and B subunits of Na(+)-ATPase showed about 23-28% identities with the beta and alpha subunits of E. hirae F1-ATPase and of Escherichia coli F1-ATPase, respectively. These results indicate that E. hirae Na(+)-ATPase belongs to the vacuolar-type ATPase. This is the first demonstration that both genes for V- and F-type ATPases are functionally expressed in one bacterial cell.

  12. Characterization of wheat straw-degrading anaerobic alkali-tolerant mixed cultures from soda lake sediments by molecular and cultivation techniques.

    Science.gov (United States)

    Porsch, Katharina; Wirth, Balázs; Tóth, Erika M; Schattenberg, Florian; Nikolausz, Marcell

    2015-09-01

    Alkaline pretreatment has the potential to enhance the anaerobic digestion of lignocellulosic biomass to biogas. However, the elevated pH of the substrate may require alkalitolerant microbial communities for an effective digestion. Three mixed anaerobic lignocellulolytic cultures were enriched from sediments from two soda lakes with wheat straw as substrate under alkaline (pH 9) mesophilic (37°C) and thermophilic (55°C) conditions. The gas production of the three cultures ceased after 4 to 5 weeks, and the produced gas was composed of carbon dioxide and methane. The main liquid intermediates were acetate and propionate. The physiological behavior of the cultures was stable even after several transfers. The enrichment process was also followed by molecular fingerprinting (terminal restriction fragment length polymorphism) of the bacterial 16S rRNA gene and of the mcrA/mrtA functional gene for methanogens. The main shift in the microbial community composition occurred between the sediment samples and the first enrichment, whereas the structure was stable in the following transfers. The bacterial communities mainly consisted of Sphingobacteriales, Clostridiales and Spirochaeta, but differed at genus level. Methanothermobacter and Methanosarcina genera and the order Methanomicrobiales were predominant methanogenes in the obtained cultures. Additionally, single cellulolytic microorganisms were isolated from enrichment cultures and identified as members of the alkaliphilic or alkalitolerant genera. The results show that anaerobic alkaline habitats harbor diverse microbial communities, which can degrade lignocellulose effectively and are therefore a potential resource for improving anaerobic digestion.

  13. Stimulation of methane generation from nonproductive coal by addition of nutrients or a microbial consortium

    Science.gov (United States)

    Jones, Elizabeth J.P.; Voytek, Mary A.; Corum, Margo D.; Orem, William H.

    2010-01-01

    Biogenic formation of methane from coal is of great interest as an underexploited source of clean energy. The goal of some coal bed producers is to extend coal bed methane productivity and to utilize hydrocarbon wastes such as coal slurry to generate new methane. However, the process and factors controlling the process, and thus ways to stimulate it, are poorly understood. Subbituminous coal from a nonproductive well in south Texas was stimulated to produce methane in microcosms when the native population was supplemented with nutrients (biostimulation) or when nutrients and a consortium of bacteria and methanogens enriched from wetland sediment were added (bioaugmentation). The native population enriched by nutrient addition included Pseudomonas spp., Veillonellaceae, and Methanosarcina barkeri. The bioaugmented microcosm generated methane more rapidly and to a higher concentration than the biostimulated microcosm. Dissolved organics, including long-chain fatty acids, single-ring aromatics, and long-chain alkanes accumulated in the first 39 days of the bioaugmented microcosm and were then degraded, accompanied by generation of methane. The bioaugmented microcosm was dominated by Geobacter sp., and most of the methane generation was associated with growth of Methanosaeta concilii. The ability of the bioaugmentation culture to produce methane from coal intermediates was confirmed in incubations of culture with representative organic compounds. This study indicates that methane production could be stimulated at the nonproductive field site and that low microbial biomass may be limiting in situ methane generation. In addition, the microcosm study suggests that the pathway for generating methane from coal involves complex microbial partnerships.

  14. Microbial composition and characterization of prevalent methanogens and acetogens isolated from syntrophic methanogenic granules

    Energy Technology Data Exchange (ETDEWEB)

    Wu Weimin (Michigan Biotechnology Inst., Lansing, MI (United States) Dept. of Civil and Environmental Engineering, Michigan State Univ., East Lansing, MI (United States) Dept. of Biochemistry, Michigan State Univ., East Lansing (United States)); Jain, M.K. (Michigan Biotechnology Inst., Lansing, MI (United States) Dept. of Animal Science, Michigan State Univ., East Lansing, MI (United States)); Conway de Macario, E. (Wadsworth Center for Laboratories and Research, New York State Dept. of Health, and School of Public Health, State Univ. of New York, NY (United States)); Thiele, J.H. (Michigan Biotechnology Inst., Lansing, MI (United States) Dept. of Biochemistry, Michigan State Univ., East Lansing, MI (United States)); Zeikus, J.G. (Michigan Biotechnology Inst., Lansing, MI (United States) Dept. of Biochemistry, Michigan State Univ., East Lansing, MI (United States) Dept. of Microbiology and Public Health, Michigan State Univ., East Lansing, MI (United States))

    1992-11-01

    The microbial species composition of methanogenic granules developed on an acetate-propionate-butyrate mixture was characterized. The granules contained high numbers of adhesive methanogens (10[sup 12]/g dry weight) and butyrate-, isobutyrate-, and propionate-degrading synthrophic acetogens (10[sup 11]/g dry weight), but low numbers of hydrolytic-fermentative bacteria (10[sup 9]/g dry weight). Prevalent methanogens in the granules included: Methanobacterium formicicum strain T1N and RF, Methanosarcina mazei strain T18, Methanospirillum hungatei strain BD, and a non-filamentous, bamboo-shaped rod species, Methanothrix/Methanosaeta-like strain M7. Prevalent syntrophic acetogens included: A butyrate-degrading Syntrophospora bryantii-like strain BH, a butyrate-isobutyrate degrading non-spore-forming rod, strain IB, a propionate-degrading spore-forming oval-shaped species, strain PT, and a propionate-degrading non-spore-forming sulfate-reducing rod species, strain PW, which was able to grow syntrophically with an H[sub 2]-utilizing methanogen. Sulfate-reducing bacteria did not play a significant role in the metabolism of H[sub 2], formate, acetate and butyrate but they were involved in propionate degradation. (orig.).

  15. Conductive Fe3O4 nanoparticles accelerate syntrophic methane production from butyrate oxidation in two different lake sediments

    Directory of Open Access Journals (Sweden)

    Jianchao Zhang

    2016-08-01

    Full Text Available Syntrophic methanogenesis is an essential link in the global carbon cycle and a key bioprocess for the disposal of organic waste and production of biogas. Recent studies suggest direct interspecies electron transfer (DIET is involved in electron exchange in methanogenesis occurring in paddy soils, anaerobic digesters and specific co-cultures with Geobacter. In this study, we evaluate the possible involvement of DIET in the syntrophic oxidation of butyrate in the enrichments from two lake sediments (an urban lake and a natural lake. The results showed that the production of CH4 was significantly accelerated in the presence of conductive nanoscale Fe3O4 or carbon nanotubes (CNTs in the sediment enrichments. Observations made with fluorescence in situ hybridization (FISH and scanning electron microscope (SEM indicated that microbial aggregates were formed in the enrichments. It appeared that the average cell-to-cell distance in aggregates in nanomaterial-amended enrichments was larger than that in aggregates in the non-amended control. These results suggested that DIET-mediated syntrophic methanogenesis could occur in the lake sediments in the presence of conductive materials. Microbial community analysis of the enrichments revealed that the genera of Syntrophomonas, Sulfurospirillum, Methanosarcina and Methanoregula were responsible for syntrophic oxidation of butyrate in lake sediment samples. The mechanism for the conductive-material-facilitated DIET in butyrate syntrophy deserves further investigation.

  16. Expanding the Diet for DIET: Electron Donors Supporting Direct Interspecies Electron Transfer (DIET in Defined Co-Cultures

    Directory of Open Access Journals (Sweden)

    Li-YIng eWang

    2016-03-01

    Full Text Available Direct interspecies electron transfer (DIET has been recognized as an alternative to interspecies H2 transfer as a mechanism for syntrophic growth, but previous studies on DIET with defined co-cultures have only documented DIET with ethanol as the electron donor in the absence of conductive materials. Co-cultures of Geobacter metallireducens and Geobacter sulfurreducens metabolized propanol, butanol, propionate, and butyrate with the reduction of fumarate to succinate. G. metallireducens utilized each of these substrates whereas only electrons available from DIET supported G. sulfurreducens respiration. A co-culture of G. metallireducens and a strain of G. sulfurreducens that could not metabolize acetate oxidized acetate with fumarate as the electron acceptor, demonstrating that acetate can also be syntrophically metabolized via DIET. A co-culture of G. metallireducens and Methanosaeta harundinacea previously shown to syntrophically convert ethanol to methane via DIET metabolized propanol or butanol as the sole electron donor, but not propionate or butyrate. The stoichiometric accumulation of propionate or butyrate in the propanol- or butanol-fed cultures demonstrated that M. harundinaceae could conserve energy to support growth solely from electrons derived from DIET. Co-cultures of G. metallireducens and Methanosarcina barkeri could also incompletely metabolize propanol and butanol and did not metabolize propionate or butyrate as sole electron donors. These results expand the range of substrates that are known to be syntrophically metabolized through DIET, but suggest that claims of propionate and butyrate metabolism via DIET in mixed microbial communities warrant further validation.

  17. Characteristic microbial community of a dry thermophilic methanogenic digester: its long-term stability and change with feeding.

    Science.gov (United States)

    Tang, Yue-Qin; Ji, Pan; Hayashi, Junpei; Koike, Yoji; Wu, Xiao-Lei; Kida, Kenji

    2011-09-01

    Thermophilic dry anaerobic digestion of sludge for cellulose methanization was acclimated at 53 °C for nearly 5 years using a waste paper-based medium. The stability of the microbial community structure and the microbial community responsible for the cellulose methanization were studied by 16S rRNA gene-based clone library analysis. The microbial community structure remained stable during the long-term acclimation period. Hydrogenotrophic methanogens dominated in methanogens and Methanothermobacter, Methanobacterium, Methanoculleus, and Methanosarcina were responsible for the methane production. Bacteria showed relatively high diversity and distributed mainly in the phyla Firmicutes, Bacteroidetes, and Synergistetes. Ninety percent of operational taxonomic units (OTUs) were affiliated with the phylum Firmicutes, indicating the crucial roles of this phylum in the digestion. Relatives of Clostridium stercorarium, Clostridium thermocellum, and Halocella cellulosilytica were dominant cellulose degraders. The acclimated stable sludge was used to treat garbage stillage discharged from a fuel ethanol production process, and the shift of microbial communities with the change of feed was analyzed. Both archaeal and bacterial communities had obviously changed: Methanoculleus spp. and Methanothermobacter spp. and the protein- and fatty acid-degrading bacteria became dominant. Accumulation of ammonia as well as volatile fatty acids led to the inhibition of microbial activity and finally resulted in the deterioration of methane fermentation of the garbage stillage.

  18. Significance of ether lipids derived from methanogens in sediments from modern and ancient basins

    Energy Technology Data Exchange (ETDEWEB)

    Shouten, S.; Sinninghe Damste, J.S.; Hoefs, M.J.L.; De Leeuw, J.W. (Netherlands Institute for Sea Research, Texel (Netherlands) Netherlands and Utrecht Univ. (Netherlands))

    1996-01-01

    High amounts of biphytanyl carbon skeletons with 0-3 cyclopentyl rings and low amounts of 2, 6, 10, 14, 18-pentamethyleicosane (PME) were detected after treatment with hydrogen iodide of fractions of several sediment extracts. These compounds are biosynthesized by archaebacteria only. Since the sediments were deposited in normal marine environments, these lipids are derived from methanogenic bacteria and more specifically Methanosarcina barkeri. In recent sediments from in and below the extensive oxygen-minimum zone in the Arabian Sea high amounts were detected in surface sediment samples where oxygen is still present. Our data suggest that these ether-bound compounds may be indicators for methane-production in the upper part of the water column and possibly an extensive oxygen-minimum zone. The concentrations varied strongly (2 orders of magnitude) in the Miocene Monterey Formation suggesting significant variations in water column methanogenic activity. The stable carbon isotopic compositions of the acyclic and cyclic biphytane carbon skeletons are several per mil heavier than those of compounds derived from photoautotrophic organisms whereas the stable carbon isotopic compositions of PME are several per mil lighter. This indicates the presence of different species of methanogens utilizing different carbon substrates.

  19. Significance of ether lipids derived from methanogens in sediments from modern and ancient basins

    Energy Technology Data Exchange (ETDEWEB)

    Shouten, S.; Sinninghe Damste, J.S.; Hoefs, M.J.L.; De Leeuw, J.W. [Netherlands Institute for Sea Research, Texel (Netherlands)]|[Netherlands and Utrecht Univ. (Netherlands)

    1996-12-31

    High amounts of biphytanyl carbon skeletons with 0-3 cyclopentyl rings and low amounts of 2, 6, 10, 14, 18-pentamethyleicosane (PME) were detected after treatment with hydrogen iodide of fractions of several sediment extracts. These compounds are biosynthesized by archaebacteria only. Since the sediments were deposited in normal marine environments, these lipids are derived from methanogenic bacteria and more specifically Methanosarcina barkeri. In recent sediments from in and below the extensive oxygen-minimum zone in the Arabian Sea high amounts were detected in surface sediment samples where oxygen is still present. Our data suggest that these ether-bound compounds may be indicators for methane-production in the upper part of the water column and possibly an extensive oxygen-minimum zone. The concentrations varied strongly (2 orders of magnitude) in the Miocene Monterey Formation suggesting significant variations in water column methanogenic activity. The stable carbon isotopic compositions of the acyclic and cyclic biphytane carbon skeletons are several per mil heavier than those of compounds derived from photoautotrophic organisms whereas the stable carbon isotopic compositions of PME are several per mil lighter. This indicates the presence of different species of methanogens utilizing different carbon substrates.

  20. Reduction of Fe(III) oxides by phylogenetically and physiologically diverse thermophilic methanogens.

    Science.gov (United States)

    Yamada, Chihaya; Kato, Souichiro; Kimura, Satoshi; Ishii, Masaharu; Igarashi, Yasuo

    2014-09-01

    Three thermophilic methanogens (Methanothermobacter thermautotrophicus, Methanosaeta thermophila, and Methanosarcina thermophila) were investigated for their ability to reduce poorly crystalline Fe(III) oxides (ferrihydrite) and the inhibitory effects of ferrihydrite on their methanogenesis. This study demonstrated that Fe(II) generation from ferrihydrite occurs in the cultures of the three thermophilic methanogens only when H2 was supplied as the source of reducing equivalents, even in the cultures of Mst. thermophila that do not grow on and produce CH4 from H2/CO2. While supplementation of ferrihydrite resulted in complete inhibition or suppression of methanogenesis by the thermophilic methanogens, ferrihydrite reduction by the methanogens at least partially alleviates the inhibitory effects. Microscopic and crystallographic analyses on the ferrihydrite-reducing Msr. thermophila cultures exhibited generation of magnetite on its cell surfaces through partial reduction of ferrihydrite. These findings suggest that at least certain thermophilic methanogens have the ability to extracellularly transfer electrons to insoluble Fe(III) compounds, affecting their methanogenic activities, which would in turn have significant impacts on materials and energy cycles in thermophilic anoxic environments. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  1. Pretreatment of coking wastewater using anaerobic sequencing batch reactor (ASBR)

    Institute of Scientific and Technical Information of China (English)

    LI Bing; SUN Ying-lan; LI Yu-ying

    2005-01-01

    A laboratory-scale anaerobic sequencing batch reactor (ASBR) was used to pretreat coking wastewater. Inoculated anaerobic granular biomass was acclimated for 225 d to the coking wastewater, and then the biochemical methane potential (BMP)of the coking wastewater in the acclimated granular biomass was measured. At the same time, some fundamental technological factors, such as the filling time and the reacting time ratio (tf/tr), the mixing intensity and the intermittent mixing mode, that affect anaerobic pretreatment of coking wastewater with ASBR, were evaluated through orthogonal tests. The COD removal efficiency reached 38%~50% in the stable operation period with the organic loading rate of 0.37~0.54 kg COD/(m3.d) at the optimum conditions of tf/tr, the mixing intensity and the intermittent mixing mode. In addition, the biodegradability of coking wastewater distinctly increased after the pretreatment using ASBR. At the end of the experiment, the microorganism forms on the granulated sludge in the ASBR were observed using SEM (scanning electron microscope) and fluoroscope. The results showed that the dominant microorganism on the granular sludge was Methanosaeta instead of Methanosarcina dominated on the inoculated sludge.

  2. Model of a DNA-protein complex of the architectural monomeric protein MC1 from Euryarchaea.

    Directory of Open Access Journals (Sweden)

    Françoise Paquet

    Full Text Available In Archaea the two major modes of DNA packaging are wrapping by histone proteins or bending by architectural non-histone proteins. To supplement our knowledge about the binding mode of the different DNA-bending proteins observed across the three domains of life, we present here the first model of a complex in which the monomeric Methanogen Chromosomal protein 1 (MC1 from Euryarchaea binds to the concave side of a strongly bent DNA. In laboratory growth conditions MC1 is the most abundant architectural protein present in Methanosarcina thermophila CHTI55. Like most proteins that strongly bend DNA, MC1 is known to bind in the minor groove. Interaction areas for MC1 and DNA were mapped by Nuclear Magnetic Resonance (NMR data. The polarity of protein binding was determined using paramagnetic probes attached to the DNA. The first structural model of the DNA-MC1 complex we propose here was obtained by two complementary docking approaches and is in good agreement with the experimental data previously provided by electron microscopy and biochemistry. Residues essential to DNA-binding and -bending were highlighted and confirmed by site-directed mutagenesis. It was found that the Arg25 side-chain was essential to neutralize the negative charge of two phosphates that come very close in response to a dramatic curvature of the DNA.

  3. Characterization of the COD removal, electricity generation, and bacterial communities in microbial fuel cells treating molasses wastewater.

    Science.gov (United States)

    Lee, Yun-Yeong; Kim, Tae G; Cho, Kyung-Suk

    2016-11-09

    The chemical oxygen demand (COD) removal, electricity generation, and microbial communities were compared in 3 types of microbial fuel cells (MFCs) treating molasses wastewater. Single-chamber MFCs without and with a proton exchange membrane (PEM), and double-chamber MFC were constructed. A total of 10,000 mg L(-1) COD of molasses wastewater was continuously fed. The COD removal, electricity generation, and microbial communities in the two types of single-chamber MFCs were similar, indicating that the PEM did not enhance the reactor performance. The COD removal in the single-chamber MFCs (89-90%) was higher than that in the double-chamber MFC (50%). However, electricity generation in the double-chamber MFC was higher than that in the single-chamber MFCs. The current density (80 mA m(-2)) and power density (17 mW m(-2)) in the double-chamber MFC were 1.4- and 2.2-times higher than those in the single-chamber MFCs, respectively. The bacterial community structures in single- and double-chamber MFCs were also distinguishable. The amount of Proteobacteria in the double-chamber MFC was 2-3 times higher than those in the single-chamber MFCs. For the archaeal community, Methanothrix (96.4%) was remarkably dominant in the single-chamber MFCs, but Methanobacterium (35.1%), Methanosarcina (28.3%), and Methanothrix (16.2%) were abundant in the double-chamber MFC.

  4. A Thermodynamically-consistent FBA-based Approach to Biogeochemical Reaction Modeling

    Science.gov (United States)

    Shapiro, B.; Jin, Q.

    2015-12-01

    Microbial rates are critical to understanding biogeochemical processes in natural environments. Recently, flux balance analysis (FBA) has been applied to predict microbial rates in aquifers and other settings. FBA is a genome-scale constraint-based modeling approach that computes metabolic rates and other phenotypes of microorganisms. This approach requires a prior knowledge of substrate uptake rates, which is not available for most natural microbes. Here we propose to constrain substrate uptake rates on the basis of microbial kinetics. Specifically, we calculate rates of respiration (and fermentation) using a revised Monod equation; this equation accounts for both the kinetics and thermodynamics of microbial catabolism. Substrate uptake rates are then computed from the rates of respiration, and applied to FBA to predict rates of microbial growth. We implemented this method by linking two software tools, PHREEQC and COBRA Toolbox. We applied this method to acetotrophic methanogenesis by Methanosarcina barkeri, and compared the simulation results to previous laboratory observations. The new method constrains acetate uptake by accounting for the kinetics and thermodynamics of methanogenesis, and predicted well the observations of previous experiments. In comparison, traditional methods of dynamic-FBA constrain acetate uptake on the basis of enzyme kinetics, and failed to reproduce the experimental results. These results show that microbial rate laws may provide a better constraint than enzyme kinetics for applying FBA to biogeochemical reaction modeling.

  5. Conductive Fe3O4 Nanoparticles Accelerate Syntrophic Methane Production from Butyrate Oxidation in Two Different Lake Sediments

    Science.gov (United States)

    Zhang, Jianchao; Lu, Yahai

    2016-01-01

    Syntrophic methanogenesis is an essential link in the global carbon cycle and a key bioprocess for the disposal of organic waste and production of biogas. Recent studies suggest direct interspecies electron transfer (DIET) is involved in electron exchange in methanogenesis occurring in paddy soils, anaerobic digesters, and specific co-cultures with Geobacter. In this study, we evaluate the possible involvement of DIET in the syntrophic oxidation of butyrate in the enrichments from two lake sediments (an urban lake and a natural lake). The results showed that the production of CH4 was significantly accelerated in the presence of conductive nanoscale Fe3O4 or carbon nanotubes in the sediment enrichments. Observations made with fluorescence in situ hybridization and scanning electron microscope indicated that microbial aggregates were formed in the enrichments. It appeared that the average cell-to-cell distance in aggregates in nanomaterial-amended enrichments was larger than that in aggregates in the non-amended control. These results suggested that DIET-mediated syntrophic methanogenesis could occur in the lake sediments in the presence of conductive materials. Microbial community analysis of the enrichments revealed that the genera of Syntrophomonas, Sulfurospirillum, Methanosarcina, and Methanoregula were responsible for syntrophic oxidation of butyrate in lake sediment samples. The mechanism for the conductive-material-facilitated DIET in butyrate syntrophy deserves further investigation. PMID:27597850

  6. Microbial Ecology of Anaerobic Digesters: The Key Players of Anaerobiosis

    Directory of Open Access Journals (Sweden)

    Fayyaz Ali Shah

    2014-01-01

    Full Text Available Anaerobic digestion is the method of wastes treatment aimed at a reduction of their hazardous effects on the biosphere. The mutualistic behavior of various anaerobic microorganisms results in the decomposition of complex organic substances into simple, chemically stabilized compounds, mainly methane and CO2. The conversions of complex organic compounds to CH4 and CO2 are possible due to the cooperation of four different groups of microorganisms, that is, fermentative, syntrophic, acetogenic, and methanogenic bacteria. Microbes adopt various pathways to evade from the unfavorable conditions in the anaerobic digester like competition between sulfate reducing bacteria (SRB and methane forming bacteria for the same substrate. Methanosarcina are able to use both acetoclastic and hydrogenotrophic pathways for methane production. This review highlights the cellulosic microorganisms, structure of cellulose, inoculum to substrate ratio, and source of inoculum and its effect on methanogenesis. The molecular techniques such as DGGE (denaturing gradient gel electrophoresis utilized for dynamic changes in microbial communities and FISH (fluorescent in situ hybridization that deal with taxonomy and interaction and distribution of tropic groups used are also discussed.

  7. Methane production and microbial community structure for alkaline pretreated waste activated sludge.

    Science.gov (United States)

    Sun, Rui; Xing, Defeng; Jia, Jianna; Zhou, Aijuan; Zhang, Lu; Ren, Nanqi

    2014-10-01

    Alkaline pretreatment was studied to analyze the influence on waste activated sludge (WAS) reduction, methane production and microbial community structure during anaerobic digestion. Methane production from alkaline pretreated sludge (A-WAS) (pH = 12) increased from 251.2 mL/Ld to 362.2 mL/Ld with the methane content of 68.7% compared to raw sludge (R-WAS). Sludge reduction had been improved, and volatile suspended solids (VSS) removal rate and protein reduction had increased by ∼ 10% and ∼ 35%, respectively. The bacterial and methanogenic communities were analyzed using 454 pyrosequencing and clone libraries of 16S rRNA gene. Remarkable shifts were observed in microbial community structures after alkaline pretreatment, especially for Archaea. The dominant methanogenic population changed from Methanosaeta for R-WAS to Methanosarcina for A-WAS. In addition to the enhancement of solubilization and hydrolysis of anaerobic digestion of WAS, alkaline pretreatment showed significant impacts on the enrichment and syntrophic interactions between microbial communities.

  8. Thermo-alkaline pretreatment of waste activated sludge at low-temperatures: effects on sludge disintegration, methane production, and methanogen community structure.

    Science.gov (United States)

    Kim, Jaai; Yu, Youngseob; Lee, Changsoo

    2013-09-01

    Low-temperature thermo-alkaline pretreatment of waste activated sludge (WAS) was studied, within the region of 0-0.2 M NaOH and 60-90°C, for the effects of NaOH concentration and temperature on sludge degradability in anaerobic digestion (AD). Significant disintegration of sludge solids (up to 75.6%) and an increase in methane production (up to 70.6%) were observed in the pretreatment trials. Two quadratic models were successfully generated by response surface analysis (R(2)>0.9, pmethane production (MP) respond to changes in the pretreatment conditions. The maximum responses of SD (77.8%) and MP (73.9% increase over the control) were shown at [0.16 M NaOH, 90°C] and [0.10 M NaOH, 73.7°C], respectively. NaOH addition showed a significant influence on the evolution of methanogen community structure during AD, whereas temperature did not. Aceticlastic Methanosaeta and Methanosarcina speceies were likely the major methanogens.

  9. Temperature impacts differentially on the methanogenic food web of cellulose-supplemented peatland soil.

    Science.gov (United States)

    Schmidt, Oliver; Horn, Marcus A; Kolb, Steffen; Drake, Harold L

    2015-03-01

    The impact of temperature on the largely unresolved intermediary ecosystem metabolism and associated unknown microbiota that link cellulose degradation and methane production in soils of a moderately acidic (pH 4.5) fen was investigated. Supplemental [(13) C]cellulose stimulated the accumulation of propionate, acetate and carbon dioxide as well as initial methane production in anoxic peat soil slurries at 15°C and 5°C. Accumulation of organic acids at 15°C was twice as fast as that at 5°C. 16S rRNA [(13) C]cellulose stable isotope probing identified novel unclassified Bacteria (79% identity to the next cultured relative Fibrobacter succinogenes), unclassified Bacteroidetes (89% identity to Prolixibacter bellariivorans), Porphyromonadaceae, Acidobacteriaceae and Ruminococcaceae as main anaerobic degraders of cellulose-derived carbon at both 15°C and 5°C. Holophagaceae and Spirochaetaceae were more abundant at 15°C. Clostridiaceae dominated the degradation of cellulose-derived carbon only at 5°C. Methanosarcina was the dominant methanogenic taxa at both 15°C and 5°C. Relative abundance of Methanocella increased at 15°C whereas that of Methanoregula and Methanosaeta increased at 5°C. Thaumarchaeota closely related to Nitrosotalea (presently not known to grow anaerobically) were abundant at 5°C but absent at 15°C indicating that Nitrosotalea sp. might be capable of anaerobic growth at low temperatures in peat.

  10. Thermophilic anaerobic co-digestion of garbage, screened swine and dairy cattle manure.

    Science.gov (United States)

    Liu, Kai; Tang, Yue-Qin; Matsui, Toru; Morimura, Shigeru; Wu, Xiao-Lei; Kida, Kenji

    2009-01-01

    Methane fermentation characteristics of garbage, swine manure (SM), dairy cattle manure (DCM) and mixtures of these wastes were studied. SM and DCM showed much lower volatile total solid (VTS) digestion efficiencies and methane yield than those of garbage. VTS digestion efficiency of SM was significantly increased when it was co-digested with garbage (Garbage: SM=1:1). Co-digestion of garbage, SM and DCM with respect to the relative quantity of each waste discharged in the Kikuchi (1: 16: 27) and Aso (1: 19: 12) areas indicated that co-digestion with garbage would improve the digestion characteristic of SM and DCM as far as the ratio of DCM in the wastes was maintained below a certain level. When the mixed waste (Garbage: SM: DCM=1:19:12) was treated using a thermophilic UAF reactor, methanogens responsible for the methane production were Methanoculleus and Methanosarcina species. Bacterial species in the phylum Firmicutes were dominant bacteria responsible for the digestion of these wastes. As the percentage of garbage in the mixed wastes used in this study was low (2-3%) and the digestion efficiency of DCM was obviously improved, the co-digestion of SM and DCM with limited garbage was a prospective method to treat the livestock waste effectively and was an attractive alternative technology for the construction of a sustainable environment and society in stock raising area.

  11. Assessment of microbial communities associated with fermentative-methanogenic biodegradation of aromatic hydrocarbons in groundwater contaminated with a biodiesel blend (B20).

    Science.gov (United States)

    Ramos, Débora Toledo; da Silva, Márcio Luís Busi; Nossa, Carlos Wolfgang; Alvarez, Pedro J J; Corseuil, Henry Xavier

    2014-09-01

    A controlled field experiment was conducted to assess the potential for fermentative-methanogenic biostimulation (by ammonium-acetate injection) to enhance biodegradation of benzene, toluene, ethylbenzene and xylenes (BTEX) as well as polycyclic aromatic hydrocarbons (PAHs) in groundwater contaminated with biodiesel B20 (20:80 v/v soybean biodiesel and diesel). Changes in microbial community structure were assessed by pyrosequencing 16S rRNA analyses. BTEX and PAH removal began 0.7 year following the release, concomitantly with the increase in the relative abundance of Desulfitobacterium and Geobacter spp. (from 5 to 52.7 % and 15.8 to 37.3 % of total Bacteria 16S rRNA, respectively), which are known to anaerobically degrade hydrocarbons. The accumulation of anaerobic metabolites acetate and hydrogen that could hinder the thermodynamic feasibility of BTEX and PAH biotransformations under fermentative/methanogenic conditions was apparently alleviated by the growing predominance of Methanosarcina. This suggests the importance of microbial population shifts that enrich microorganisms capable of interacting syntrophically to enhance the feasibility of fermentative-methanogenic bioremediation of biodiesel blend releases.

  12. The effect of managing nutrients in the performance of anaerobic digesters of municipal wastewater treatment plants.

    Science.gov (United States)

    Demirer, Sibel Uludag; Taskin, Bilgin; Demirer, Goksel N; Duran, Metin

    2013-09-01

    Is it possible to create conditions in the anaerobic digesters to control nutrients without changing the performance of a reactor? This study investigates an answer for this question. To this purpose, anaerobic reactors are operated at high concentrations of Mg(2+) ion to harvest the nutrient ions (NH4 (+) and PO4 (3-)) in the form of struvite, that is, magnesium ammonium phosphate. The effects of this modification on the anaerobic digestion of sewage sludge were investigated in terms of chemical oxygen demand (COD) removal and cumulative CH4 production as well as the changes in the biological diversity. The results showed that approximately 50 % of the nutrients (NH4 (+) and PO4 (3-)) were removed regardless of the method adopted for the addition of Mg(2+) ion, slug or daily dosing. The numbers of Methanosaeta and Methanosarcina in the samples withdrawn prior to and after the addition of Mg(2+) did not show significant difference according to the results obtained from qPCR analyses. The research results showed that the addition of Mg(2+) into the anaerobic digesters in municipal wastewater treatment facilities may help to remove the nutrients from the effluent while recovering in their solid forms.

  13. Microwave radiation and reactor design influence microbial communities during methane fermentation.

    Science.gov (United States)

    Cydzik-Kwiatkowska, Agnieszka; Zieliński, Marcin; Jaranowska, Paulina

    2012-09-01

    The effect of reactor design and method of heating on the efficiency of methane fermentation and composition of microbial communities, especially methanogenic Archaea, were determined. The research was carried out using submerge- and trickling-bed reactors fed with wastewater and the heat supply into the reactors included a convection heating method and microwave radiation. The polymerase chain reaction-denaturing gradient gel electrophoresis and relative real-time PCR were used in order to assess the biofilm communities. The best fermentation results and the highest abundance of methanogenic Archaea in biomass were observed in microwave heated trickling-bed reactors. The research proved that in reactors of identical design, the application of microwaves enabled a higher fermentation efficiency to be obtained and simultaneously increased the diversity of methanogenic Archaea communities that favors process stability. All the identified sequences of Archaea belonged to Methanosarcina sp., suggesting that species from this genera are susceptible to non-thermal effects of microwaves. There were no effects from microwaves on the bacterial communities in both types of reactors, however, the bacterial species composition varied in the reactors of different design.

  14. Quantitative and qualitative transitions of methanogen community structure during the batch anaerobic digestion of cheese-processing wastewater.

    Science.gov (United States)

    Lee, Changsoo; Kim, Jaai; Shin, Seung Gu; O'Flaherty, Vincent; Hwang, Seokhwan

    2010-08-01

    Qualitative and quantitative shifts in methanogen community structure, associated with process performance data, were investigated during the batch anaerobic digestion of a cheese-processing wastewater, whey permeate. Denaturing gradient gel electrophoresis (DGGE) and real-time PCR techniques were applied to obtain qualitative and quantitative microbial data sets, respectively, based on methanogen 16S rRNA genes. Throughout the operation, dynamic variations in both qualitative and quantitative community structures were observed, with repeated shifts in dominance between the aceticlastic Methanosarcinaceae (suggested mainly by the detection of a Methanosarcina-like population) and the hydrogenotrophic Methanomicrobiales (suggested mainly by the detection of a Methanofollis-like population). This trend corresponded well to the diauxic utilization of acetate and longer-chain fatty acids (C(3)-C(6)), mainly propionate. Joint-plot non-metric multidimensional scaling (NMS) analysis demonstrated that the qualitative and quantitative community shifts had significant correlations with the composition of residual organic acids and the methane production rate, respectively. This suggests the potential use of microbial community shift analysis as an indicative tool for diagnosing anaerobic digestion processes. The results suggest that more attention should be directed to quantitative, as well as qualitative, approaches for a better understanding of anaerobic digestion, particularly in terms of biogas production efficiency, under dynamic and transitional conditions.

  15. Electromicrobiology: Electron Transfer via Biowires in Nature and Practical Applications

    Directory of Open Access Journals (Sweden)

    Lovley Derek

    2016-01-01

    Full Text Available One of the most exciting developments in the field of electromicrobiology has been the discovery of electrically conductive pili (e-pili in Geobacter species that transport electrons with a metallic-like mechanism. The e-pili are essential for extracellular electron transport to Fe(III oxides and longrange electron transport through the conductive biofilms that form on the anodes of microbial fuel cells. The e-pili also facilitate direct interspecies electron transfer between Geobacter and Methanosaeta or Methanosarcina species. Metatranscriptomic studies have demonstrated that Geobacter/Methanosaeta DIET is an important process in anaerobic digesters converting brewery wastes to methane. Increasing e-pili expression through genetic modification of regulatory systems or adaptive evolution yields strains with enhanced rates of extracellular electron transfer. Measurement of the conductivity of individual e-pili has demonstrated that they have conductivities higher than those of a number of synthetic conducting organic polymers. Multiple lines of evidence have demonstrated that aromatic amino acids play an important role in the electron transport along e-pili, suggesting opportunities to tune e-pili conductivity via genetic manipulation of the amino acid composition of e-pili. It is expected that e-pili will be harnessed to improve microbe-electrode processes such as microbial electrosynthesis and for the development of novel biosensors. Also, e-pili show promise as a sustainable ‘green’ replacement for electronic materials that contain toxic components and/or are produced with harsh chemicals.

  16. Microbial communities change in an anaerobic digestion after application of microbial electrolysis cells.

    Science.gov (United States)

    Lee, Beom; Park, Jun-Gyu; Shin, Won-Beom; Tian, Dong-Jie; Jun, Hang-Bae

    2017-06-01

    Microbial electrolysis cells (MECs) are being studied to improve the efficiency of anaerobic digesters and biogas production. In the present study, we investigated the effects of electrochemical reactions in AD-MEC (anaerobic digester combined with MECs) on changes in the microbial communities of bulk sludge through 454-pyrosequencing analysis, as well as the effect of these changes on anaerobic digestion. Methanobacterium beijingense and Methanobacterium petrolearium were the dominant archaeal species in AD, while Methanosarcina thermophila and Methanobacterium formicicum were dominant in AD-MEC at steady-state. There were no substantial differences in dominant bacterial species. Clostridia class was more abundant than Bacteroidia class in both reactors. Compared to AD, AD-MEC showed a 40% increase in overall bacterial population, increasing the removal of organic matters and the conversion of volatile fatty acids (VFAs). Thus, the MEC reaction more effectively converts organic matters to VFAs and activates microbial communities favorable for methane production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Digestion performance and microbial community in full-scale methane fermentation of stillage from sweet potato-shochu production.

    Science.gov (United States)

    Kobayashi, Tsutomu; Tang, Yueqin; Urakami, Toyoshi; Morimura, Shigeru; Kida, Kenji

    2014-02-01

    Sweet potato shochu is a traditional Japanese spirit produced mainly in the South Kyushu area in Japan. The amount of stillage reaches approximately 8 x 10(5) tons per year. Wastewater mainly containing stillage from the production of sweet potato-shochu was treated thermophilically in a full-scale treatment plant using fixed-bed reactors (8 reactors x 283 m3). Following the addition of Ni2+ and Co2+, the reactors have been stably operated for six years at a high chemical oxygen demand (COD) loading rate of 14 kg/(m3 x day). Analysis of coenzyme content and microbial communities indicated that similar microbial communities were present in the liquid phase and on the fiber carriers installed in reactors. Bacteria in the phyla Firmicutes as well as Bacteroidetes were dominant bacteria, and Methanosarcina thermophila as well as Methanothermobacter crinale were dominant methanogens in the reactors. This study reveals that stillage from sweet potato-shochu production can be treated effectively in a full-scale fixed-bed reactor under thermophilic conditions with the help of Ni2+ and Co2+. The high diversity of bacterial community and the coexistence of both aceticlastic and hydrogenotrophic methanogens contributed to the excellent fermentation performance.

  18. Syntrophic microbial communities on straw as biofilm carrier increase the methane yield of a biowaste-digesting biogas reactor

    Directory of Open Access Journals (Sweden)

    Frank R. Bengelsdorf

    2015-08-01

    Full Text Available Biogas from biowaste can be an important source of renewable energy, but the fermentation process of low-structure waste is often unstable. The present study uses a full-scale biogas reactor to test the hypothesis that straw as an additional biofilm carrier will increase methane yield; and this effect is mirrored in a specific microbial community attached to the straw. Better reactor performance after addition of straw, at simultaneously higher organic loading rate and specific methane yield confirmed the hypothesis. The microbial communities on straw as a biofilm carrier and of the liquid reactor content were investigated using 16S rDNA amplicon sequencing by means of 454 pyrosequencing technology. The results revealed high diversity of the bacterial communities in the liquid reactor content as well as the biofilms on the straw. The most abundant archaea in all samples belonged to the genera Methanoculleus and Methanosarcina. Addition of straw resulted in a significantly different microbial community attached to the biofilm carrier. The bacterium Candidatus Cloacamonas acidaminovorans and methanogenic archaea of the genus Methanoculleus dominated the biofilm on straw. Syntrophic interactions between the hydrogenotrophic Methanoculleus sp. and members of the hydrogen-producing bacterial community within biofilms may explain the improved methane yield. Thus, straw addition can be used to improve and to stabilize the anaerobic process in substrates lacking biofilm-supporting structures.

  19. Impact of different antibiotics on methane production using waste-activated sludge: mechanisms and microbial community dynamics.

    Science.gov (United States)

    Mustapha, Nurul Asyifah; Sakai, Kenji; Shirai, Yoshihito; Maeda, Toshinari

    2016-11-01

    Anaerobic digestion is an effective method for reducing the by-product of waste-activated sludge (WAS) from wastewater treatment plants and for producing bioenergy from WAS. However, only a limited number of studies have attempted to improve anaerobic digestion by targeting the microbial interactions in WAS. In this study, we examined whether different antibiotics positively, negatively, or neutrally influence methane fermentation by evaluating changes in the microbial community and functions in WAS. Addition of azithromycin promoted the microbial communities related to the acidogenic and acetogenic stages, and a high concentration of soluble proteins and a high activity of methanogens were detected. Chloramphenicol inhibited methane production but did not affect the bacteria that contribute to the hydrolysis, acidogenesis, and acetogenesis digestion stages. The addition of kanamycin, which exhibits the same methane productivity as a control (antibiotic-free WAS), did not affect all of the microbial communities during anaerobic digestion. This study demonstrates the simultaneous functions and interactions of diverse bacteria and methanogenic Archaea in different stages of the anaerobic digestion of WAS. The ratio of Caldilinea, Methanosarcina, and Clostridium may correspond closely to the trend of methane production in each antibiotic. The changes in microbial activities and function by antibiotics facilitate a better understanding of bioenergy production.

  20. Acid resistance of methanogenic bacteria in a two-stage anaerobic process treating high concentration methanol Wastewater

    Institute of Scientific and Technical Information of China (English)

    ZHOU Xuefei; REN Nanqi

    2007-01-01

    In this study,the two-stage upflow anaerobic sludge blanket(UASB)system and batch experiments were employed to evaluate the performance of anaerobic digestion for the treatment of high concentration methanol wastewater.The acid resistance of granular sludge and methanogenic bacteria and their metabolizing activity were investigated.The results show that the pH of the first UASB changed from 4.9 to 5.8 and 5.5 to 6.2 for the second reactor.Apparently,these were not the advisable pH levels that common metha nogenic bacteria could accept.The methanogenic bacteria of the system,viz.Methanosarcina barkeri,had some acid resistance and could still degrade methanol at pH 5.0.If the methanogenic bacteria were trained further,their acid resistance would be improved somewhat.Granular sludge of the system could protect the methanogenic bacteria within its body against the impact of the acidic environment and make them degrade methanol at pH 4.5.The performance of granular sludge was attributed to its structure,bacteria species,and the distribution of bacterium inside the granule.

  1. Microbial community dynamics in continuous microbial fuel cells fed with synthetic wastewater and pig slurry.

    Science.gov (United States)

    Sotres, Ana; Tey, Laura; Bonmatí, August; Viñas, Marc

    2016-10-01

    Two-chambered microbial fuel cells (MFCs) operating with synthetic wastewater and pig slurry were assessed. Additionally, the use of 2-bromoethanesulfonate (BES-Inh) was studied. The synthetic wastewater-fed MFC (MFCSW) showed a maximum power density (PDmax) of 2138mWm(-3), and the addition of BES-Inh (10mM) did not show any improvement in its performance (PDmax=2078mWm(-3)). When pig slurry was used as feed (MFCPS), PDmax increased up to 5623mWm(-3). The microbial community composition was affected by the type of substrate used. While, Pseudomonadaceae and Clostridiaceae were the most representative families within the acetate-based medium, Flavobacteriaceae, Chitinophagaceae, Comamonadaceae and Nitrosomonadaceae were predominant when pig slurry was used as feed. Otherwise, only the Eubacterial microbial community composition was strongly modified when adding BES-Inh, thus leading to an enrichment of the Bacteroidetes phylum. Oppositely, the Archaeal community was less affected by the addition of BES-Inh, and Methanosarcina sp., arose as the predominant family in both situations. Despite all the differences in microbial communities, 6 operational taxonomic units (OTUs) belonging to Bacteroidetes (Porphyromonadaceae and Marinilabiaceae) and Firmicutes (Clostridiales) were found to be common to both MFCs, also for different contents of COD and N-NH4(+), and therefore could be considered as the bioanode core microbiome.

  2. Shifts in methanogenic community composition and methane fluxes along the degradation of discontinuous permafrost.

    Science.gov (United States)

    Liebner, Susanne; Ganzert, Lars; Kiss, Andrea; Yang, Sizhong; Wagner, Dirk; Svenning, Mette M

    2015-01-01

    The response of methanogens to thawing permafrost is an important factor for the global greenhouse gas budget. We tracked methanogenic community structure, activity, and abundance along the degradation of sub-Arctic palsa peatland permafrost. We observed the development of pronounced methane production, release, and abundance of functional (mcrA) methanogenic gene numbers following the transitions from permafrost (palsa) to thaw pond structures. This was associated with the establishment of a methanogenic community consisting both of hydrogenotrophic (Methanobacterium, Methanocellales), and potential acetoclastic (Methanosarcina) members and their activity. While peat bog development was not reflected in significant changes of mcrA copy numbers, potential methane production, and rates of methane release decreased. This was primarily linked to a decline of potential acetoclastic in favor of hydrogenotrophic methanogens. Although palsa peatland succession offers similarities with typical transitions from fen to bog ecosystems, the observed dynamics in methane fluxes and methanogenic communities are primarily attributed to changes within the dominant Bryophyta and Cyperaceae taxa rather than to changes in peat moss and sedge coverage, pH and nutrient regime. Overall, the palsa peatland methanogenic community was characterized by a few dominant operational taxonomic units (OTUs). These OTUs seem to be indicative for methanogenic species that thrive in terrestrial organic rich environments. In summary, our study shows that after an initial stage of high methane emissions following permafrost thaw, methane fluxes, and methanogenic communities establish that are typical for northern peat bogs.

  3. Red mud enhances methanogenesis with the simultaneous improvement of hydrolysis-acidification and electrical conductivity.

    Science.gov (United States)

    Ye, Jie; Hu, Andong; Ren, Guoping; Zhou, Ting; Zhang, Guangming; Zhou, Shungui

    2017-08-12

    The role of red mud in the improvement of methanogenesis during sludge anaerobic digestion was innovatively investigated in this study. The results demonstrated that the addition of 20g/L red mud resulted in a 35.5% increase in methane accumulation. Red mud effectively promoted the hydrolysis-acidification of organic compounds in the sludge, which resulted in the increase of protein, polysaccharide, and VFAs by 5.1-94.5%. The activities of key enzymes were improved by 41.4-257.3%. Electrochemical measurements presented direct evidence that the electrical conductivity was significantly improved with red mud. More conductive magnetite was formed during the secondary mineralization after Fe(III) reduction by Fe (III)-reducing genes such as Clostridiaceae and Ruminococcaceae. The higher conductivity enhanced the electron transfer between the syntrophic bacteria (Geobacteraceae) and methanogens (Methanosaeta and Methanosarcina), and then improved the methanogenesis. This research provides a novel perspective on the synergism between sludge and red mud for methane production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Archaeosomes varying in lipid composition differ in receptor-mediated endocytosis and differentially adjuvant immune responses to entrapped antigen

    Directory of Open Access Journals (Sweden)

    G. Dennis Sprott

    2003-01-01

    Full Text Available Archaeosomes prepared from total polar lipids extracted from six archaeal species with divergent lipid compositions had the capacity to deliver antigen for presentation via both MHC class I and class II pathways. Lipid extracts from Halobacterium halobium and from Halococcus morrhuae strains 14039 and 16008 contained archaetidylglycerol methylphosphate and sulfated glycolipids rich in mannose residues, and lacked archaetidylserine, whereas the opposite was found in Methanobrevibacter smithii, Methanosarcina mazei and Methanococcus jannaschii. Annexin V labeling revealed a surface orientation of phosphoserine head groups in M. smithii, M. mazei and M. jannaschii archaeosomes. Uptake of rhodamine-labeled M. smithii or M. jannaschii archaeosomes by murine peritoneal macrophages was inhibited by unlabeled liposomes containing phosphatidylserine, by the sulfhydryl inhibitor N-ethylmaleimide, and by ATP depletion using azide plus fluoride, but not by H. halobium archaeosomes. In contrast, N-ethylmaleimide failed to inhibit uptake of the four other rhodamine-labeled archaeosome types, and azide plus fluoride did not inhibit uptake of H. halobium or H. morrhuae archaeosomes. These results suggest endocytosis of archaeosomes rich in surface-exposed phosphoserine head groups via a phosphatidylserine receptor, and energy-independent surface adsorption of certain other archaeosome composition classes. Lipid composition affected not only the endocytic mechanism, but also served to differentially modulate the activation of dendritic cells. The induction of IL-12 secretion from dendritic cells exposed to H. morrhuae 14039 archaeosomes was striking compared with cells exposed to archaeosomes from 16008. Thus, archaeosome types uniquely modulate antigen delivery and dendritic cell activation.

  5. Utilization of high-strength wastewater for the production of biogas as a renewable energy source using hybrid upflow anaerobic sludge blanket (HUASB) reactor

    Energy Technology Data Exchange (ETDEWEB)

    Shivayogimath, C.B.; Ramanujam, T.K.

    1998-07-01

    Anaerobic digestion of distillery spentwash, a high-strength wastewater, was studied using a hybrid upflow anaerobic sludge blanket (HUASB) reactor for 240 days under ambient conditions. The HUASB reactor combined an open volume in the bottom two-thirds of the reactor for sludge blanket and polypropylene pall rings packing in the upper one-third of the reactor. The aim of the study was to achieve optimum biogas production and waste treatment. Using non-granular anaerobic sewage sludge as seed, the start-up of the HUASB reactor was successfully completed, with the production of active bacterial granules of 1--2 mm size, within 90 days. Examination of the bacterial granules under scanning electron microscope (SEM) revealed that Methanothrix like microorganisms were the dominant species besides Methanosarcina. An organic loading of 24 kg COD/m{sup 3}d at a low hydraulic retention time (HRT) of 6 hours was achieved with 82% reduction in COD. Biogas with high methane content (80%) was produced at these loadings. The specific biogas yield was 0.36 m{sup 3} CH{sub 4}/kg COD. Packing in the upper third of the reactor was very efficient as a gas-solid separator (GSS); and in addition it retained the biomass.

  6. Enhanced treatment of Fischer-Tropsch (F-T) wastewater using the up-flow anaerobic sludge blanket coupled with bioelectrochemical system: Effect of electric field.

    Science.gov (United States)

    Wang, Dexin; Han, Hongjun; Han, Yuxing; Li, Kun; Zhu, Hao

    2017-05-01

    The coupling of bioelectrochemical system (BES) with an up-flow anaerobic sludge blanket (UASB) was established for enhanced Fischer-Tropsch (F-T) wastewater treatment while the UASB (control group) was operated in parallel. The presence of electric field could offer system a more reductive micro-environment that lower the ORP values and maintain the appropriate pH range, resulting in the higher chemical oxygen demand (COD) removal efficiency and methane production for BES-UASB (86.8% and 2.31±0.1L/(L·d)) while those values in control group were 72.1% and 1.77±0.08L/(L·d). In addition, the coupled system could promote sludge granulation to perform a positive effect on maintaining stability of pollutants removal. The high-throughput 16S rRNA gene pyrosequencing in this study further confirmed that the promoting direct interspecies electron transfer (DIET) between Geobacter and Methanosarcina might be established in BES-UASB to improve the syntrophic degradation of propionate and butyrate, finally facilitated completely methane production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Biogas by two-stage microbial anaerobic and semi-continuous digestion of Chinese cabbage waste

    Institute of Scientific and Technical Information of China (English)

    Xiaoying Dong; Lijie Shao; Yan Wang; Wei Kou; Yanxin Cao; Dalei Zhang

    2015-01-01

    Anaerobic digestion of Chinese cabbage waste was investigated through a pilot-scale two-stage digester at a mesophilic temperature of 37 °C. In the acidification digester, the main product was acetic acid, with the maxi-mum concentration of 4289 mg·L-1 on the fourth day, accounting for 50.32%of total volatile fatty acids. The oxidation reduction potential (ORP) and NH4+-N level decreased gradual y with hydraulic retention time (HRT) of acidification. In the second digestion phase, the maximum methanogenic bacterial concentration reached 9.6 × 1010 ml-1 at the organic loading rate (OLR) of 3.5–4 kg VS·m-3, with corresponding HRT of 12–16 days. Accordingly, the optimal biogas production was 0.62 m3·(kg VS)-1, with methane content of 65%–68%. ORP and NH4+-N levels in the methanizer remained between-500 and-560 mV and 2000–4500 mg·L-1, respec-tively. Methanococcus and Methanosarcina served as the main methanogens in the anaerobic digester.

  8. Diversity of prokaryotes at a shallow submarine vent of Panarea Island (Italy by high-throughput sequencing

    Directory of Open Access Journals (Sweden)

    Teresa L. Maugeri

    2013-09-01

    Full Text Available To determine microbial community composition and possible key microbial processes in the shallow-sea hydrothermal vent system off Panarea Island (Italy, we examined bacterial and archaeal communities of sediment and fluid samples from a hot vent by 16S rDNA Illumina sequencing technique. Both high abundant (>1% of total sequences, low abundant (from 0.1 to <1% and rare (< 0.1% phylogenetic groups were responsible for the distinct prokaryotic communities characterizing the heated sediment and fluid. The bacterial and archaeal communities from sediment were dominated by sequences affiliated with Rhodovulum genus (Alphaproteobacteria, including phototrophic ferrous-iron-oxidizing purple bacteria, Thiohalospira and Thiomicrospira (Gammaproteobacteria, typically involved in the sulphur cycle, and Methanococcus (Euryarchaeota. Fluid communities were dominated by anoxygenic phototrophic members of Chlorobium, followed by Thiomicrospira (Gammaproteobacteria, Sulfurimonas, Arcobacter and Sulfurospirillum (Epsilonproteobacteria, and Methanosarcina (Euryarchaeota. Obtained sequences were affiliated with prokaryotes taking a key part in the carbon, iron and sulphur cycling at the shallow hydrothermal system off Panarea Island. Despite the huge sequencing efforts, a great number of Bacteria and Archaea still remains unaffiliated at genus level, indicating that Black Point vent represents a hotspot of prokaryotic diversity.

  9. Impact of the substrate loading regime and phosphoric acid supplementation on performance of biogas reactors and microbial community dynamics during anaerobic digestion of chicken wastes.

    Science.gov (United States)

    Belostotskiy, Dmitry E; Ziganshina, Elvira E; Siniagina, Maria; Boulygina, Eugenia A; Miluykov, Vasili A; Ziganshin, Ayrat M

    2015-10-01

    This study evaluates the effects of increasing organic loading rate (OLR) and decreasing hydraulic retention time (HRT) as well as phosphoric acid addition on mesophilic reactors' performance and biogas production from chicken wastes. Furthermore, microbial community composition in reactors was characterized by a 16S rRNA gene-based pyrosequencing analysis. Each step of increasing OLR impacted on the activity of microorganisms what caused a temporary decrease in biogas production. The addition of phosphoric acid resulted in the increased biogas production with values between 361 and 447 mL g(VS)(-1) from day 61 to day 74 compared to control reactor (309-350 mL g(VS)(-1)). With reactors' operation, Bacteroidetes phylotypes were noticeably replaced with Firmicutes representatives, and significant increase of Clostridium sp. was identified. Within Euryarchaeota, Methanosarcina sp. dominated in all analyzed samples, in which high ammonium levels were detected (3.4-4.9 NH4(+)-N g L(-1)). These results can help in better understanding the anaerobic digestion process of simultaneously ammonium/phosphate-rich substrates.

  10. Improved Monitoring of Semi-Continuous Anaerobic Digestion of Sugarcane Waste: Effects of Increasing Organic Loading Rate on Methanogenic Community Dynamics

    Directory of Open Access Journals (Sweden)

    Athaydes Francisco Leite

    2015-09-01

    Full Text Available The anaerobic digestion of filter cake and its co-digestion with bagasse, and the effect of gradual increase of the organic loading rate (OLR from start-up to overload were investigated. Understanding the influence of environmental and technical parameters on the development of particular methanogenic pathway in the biogas process was an important aim for the prediction and prevention of process failure. The rapid accumulation of volatile organic acids at high OLR of 3.0 to 4.0 gvs·L−1·day−1 indicated strong process inhibition. Methanogenic community dynamics of the reactors was monitored by stable isotope composition of biogas and molecular biological analysis. A potential shift toward the aceticlastic methanogenesis was observed along with the OLR increase under stable reactor operating conditions. Reactor overloading and process failure were indicated by the tendency to return to a predominance of hydrogenotrophic methanogenesis with rising abundances of the orders Methanobacteriales and Methanomicrobiales and drop of the genus Methanosarcina abundance.

  11. Improved Monitoring of Semi-Continuous Anaerobic Digestion of Sugarcane Waste: Effects of Increasing Organic Loading Rate on Methanogenic Community Dynamics

    Science.gov (United States)

    Leite, Athaydes Francisco; Janke, Leandro; Lv, Zuopeng; Harms, Hauke; Richnow, Hans-Hermann; Nikolausz, Marcell

    2015-01-01

    The anaerobic digestion of filter cake and its co-digestion with bagasse, and the effect of gradual increase of the organic loading rate (OLR) from start-up to overload were investigated. Understanding the influence of environmental and technical parameters on the development of particular methanogenic pathway in the biogas process was an important aim for the prediction and prevention of process failure. The rapid accumulation of volatile organic acids at high OLR of 3.0 to 4.0 gvs·L−1·day−1 indicated strong process inhibition. Methanogenic community dynamics of the reactors was monitored by stable isotope composition of biogas and molecular biological analysis. A potential shift toward the aceticlastic methanogenesis was observed along with the OLR increase under stable reactor operating conditions. Reactor overloading and process failure were indicated by the tendency to return to a predominance of hydrogenotrophic methanogenesis with rising abundances of the orders Methanobacteriales and Methanomicrobiales and drop of the genus Methanosarcina abundance. PMID:26404240

  12. Effect of increasing total solids contents on anaerobic digestion of food waste under mesophilic conditions: performance and microbial characteristics analysis.

    Directory of Open Access Journals (Sweden)

    Jing Yi

    Full Text Available The total solids content of feedstocks affects the performances of anaerobic digestion and the change of total solids content will lead the change of microbial morphology in systems. In order to increase the efficiency of anaerobic digestion, it is necessary to understand the role of the total solids content on the behavior of the microbial communities involved in anaerobic digestion of organic matter from wet to dry technology. The performances of mesophilic anaerobic digestion of food waste with different total solids contents from 5% to 20% were compared and the microbial communities in reactors were investigated using 454 pyrosequencing technology. Three stable anaerobic digestion processes were achieved for food waste biodegradation and methane generation. Better performances mainly including volatile solids reduction and methane yield were obtained in the reactors with higher total solids content. Pyrosequencing results revealed significant shifts in bacterial community with increasing total solids contents. The proportion of phylum Chloroflexi decreased obviously with increasing total solids contents while other functional bacteria showed increasing trend. Methanosarcina absolutely dominated in archaeal communities in three reactors and the relative abundance of this group showed increasing trend with increasing total solids contents. These results revealed the effects of the total solids content on the performance parameters and the behavior of the microbial communities involved in the anaerobic digestion of food waste from wet to dry technologies.

  13. Effect of Increasing Total Solids Contents on Anaerobic Digestion of Food Waste under Mesophilic Conditions: Performance and Microbial Characteristics Analysis

    Science.gov (United States)

    Jin, Jingwei; Dai, Xiaohu

    2014-01-01

    The total solids content of feedstocks affects the performances of anaerobic digestion and the change of total solids content will lead the change of microbial morphology in systems. In order to increase the efficiency of anaerobic digestion, it is necessary to understand the role of the total solids content on the behavior of the microbial communities involved in anaerobic digestion of organic matter from wet to dry technology. The performances of mesophilic anaerobic digestion of food waste with different total solids contents from 5% to 20% were compared and the microbial communities in reactors were investigated using 454 pyrosequencing technology. Three stable anaerobic digestion processes were achieved for food waste biodegradation and methane generation. Better performances mainly including volatile solids reduction and methane yield were obtained in the reactors with higher total solids content. Pyrosequencing results revealed significant shifts in bacterial community with increasing total solids contents. The proportion of phylum Chloroflexi decreased obviously with increasing total solids contents while other functional bacteria showed increasing trend. Methanosarcina absolutely dominated in archaeal communities in three reactors and the relative abundance of this group showed increasing trend with increasing total solids contents. These results revealed the effects of the total solids content on the performance parameters and the behavior of the microbial communities involved in the anaerobic digestion of food waste from wet to dry technologies. PMID:25051352

  14. Microbial Ecology of Anaerobic Digesters: The Key Players of Anaerobiosis

    Science.gov (United States)

    Ali Shah, Fayyaz; Mahmood, Qaisar; Maroof Shah, Mohammad; Pervez, Arshid; Ahmad Asad, Saeed

    2014-01-01

    Anaerobic digestion is the method of wastes treatment aimed at a reduction of their hazardous effects on the biosphere. The mutualistic behavior of various anaerobic microorganisms results in the decomposition of complex organic substances into simple, chemically stabilized compounds, mainly methane and CO2. The conversions of complex organic compounds to CH4 and CO2 are possible due to the cooperation of four different groups of microorganisms, that is, fermentative, syntrophic, acetogenic, and methanogenic bacteria. Microbes adopt various pathways to evade from the unfavorable conditions in the anaerobic digester like competition between sulfate reducing bacteria (SRB) and methane forming bacteria for the same substrate. Methanosarcina are able to use both acetoclastic and hydrogenotrophic pathways for methane production. This review highlights the cellulosic microorganisms, structure of cellulose, inoculum to substrate ratio, and source of inoculum and its effect on methanogenesis. The molecular techniques such as DGGE (denaturing gradient gel electrophoresis) utilized for dynamic changes in microbial communities and FISH (fluorescent in situ hybridization) that deal with taxonomy and interaction and distribution of tropic groups used are also discussed. PMID:24701142

  15. Dry anaerobic digestion of food waste and cardboard at different substrate loads, solid contents and co-digestion proportions.

    Science.gov (United States)

    Capson-Tojo, Gabriel; Trably, Eric; Rouez, Maxime; Crest, Marion; Steyer, Jean-Philippe; Delgenès, Jean-Philippe; Escudié, Renaud

    2017-06-01

    The increasing food waste production calls for developing efficient technologies for its treatment. Anaerobic processes provide an effective waste valorization. The influence of the initial substrate load on the performance of batch dry anaerobic co-digestion reactors treating food waste and cardboard was investigated. The load was varied by modifying the substrate to inoculum ratio (S/X), the total solids content and the co-digestion proportions. The results showed that the S/X was a crucial parameter. Within the tested values (0.25, 1 and 4gVS·gVS(-1)), only the reactors working at 0.25 produced methane. Methanosarcina was the main archaea, indicating its importance for efficient methanogenesis. Acidogenic fermentation was predominant at higher S/X, producing hydrogen and other metabolites. Higher substrate conversions (≤48%) and hydrogen yields (≤62mL·gVS(-1)) were achieved at low loads. This study suggests that different value-added compounds can be produced in dry conditions, with the initial substrate load as easy-to-control operational parameter. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Seryl-tRNA Synthetases from Methanogenic Archaea: Suppression of Bacterial Amber Mutation and Heterologous Toxicity

    Directory of Open Access Journals (Sweden)

    Drasko Boko

    2010-01-01

    Full Text Available Methanogenic archaea possess unusual seryl-tRNA synthetases (SerRS, evolutionarily distinct from the SerRSs found in other archaea, eucaryotes and bacteria. Our recent X-ray structural analysis of Methanosarcina barkeri SerRS revealed an idiosyncratic N-terminal domain and catalytic zinc ion in the active site. To shed further light on substrate discrimination by methanogenic-type SerRS, we set up to explore in vivo the interaction of methanogenic-type SerRSs with their cognate tRNAs in Escherichia coli or Saccharomyces cerevisiae. The expression of various methanogenic-type SerRSs was toxic for E. coli, resulting in the synthesis of erroneous proteins, as revealed by β-galactosidase stability assay. Although SerRSs from methanogenic archaea recognize tRNAsSer from all three domains of life in vitro, the toxicity presumably precluded the complementation of endogenous SerRS function in both, E. coli and S. cerevisiae. However, despite the observed toxicity, coexpression of methanogenic-type SerRS with its cognate tRNA suppressed bacterial amber mutation.

  17. Microbial community shifts and biogas conversion computation during steady, inhibited and recovered stages of thermophilic methane fermentation on chicken manure with a wide variation of ammonia.

    Science.gov (United States)

    Niu, Qigui; Qiao, Wei; Qiang, Hong; Li, Yu-You

    2013-10-01

    The thermophilic methane fermentation of chicken manure (10% TS) was investigated within a wide range of ammonia. Microbiological analysis showed significant shifts in Archaeal and Bacterial proportions with VFA accmulation and CH4 formation before and after inhibition. VFA accumulated sharply with lower methane production, 0.29 L/g VS, than during the steady stage, 0.32 L/g VS. Biogas production almost ceased with the synergy inhibition of TAN (8000 mg/L) and VFA (25,000 mg/L). Hydrogenotrophic Methanothermobacter thermautotrophicus str. was the dominate archaea with 95% in the inhibition stage and 100% after 40 days recovery compared to 9.3% in the steady stage. Aceticlastic Methanosarcina was not encountered with coincided phenomenal of high VFA in the inhibition stage as well as recovery stage. Evaluation of the microbial diversity and functional bacteria indicated the dominate phylum of Firmicutes were 94.74% and 84.4% with and without inhibition. The microbial community shifted significantly with elevated ammonia concentration affecting the performance. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Hydrogenotrophic methanogens dominate in biogas reactors fed with defined substrates.

    Science.gov (United States)

    Kampmann, K; Ratering, S; Baumann, R; Schmidt, M; Zerr, W; Schnell, S

    2012-09-01

    Methanogenic communities in 200L biogas reactors containing liquid manure were investigated for 33 d. The reactors were consecutively fed with casein, starch and cream. Real-time PCR with primers targeting the gene for methyl coenzyme-M reductase (mcrA) resulted in copy numbers of up to 2.1×10(9) g dry mass(-1). Single strand conformation polymorphism (SSCP) analysis revealed a stable community consisting of few hydrogenotrophic methanogens. One of the two most abundant species was closely related to Methanospirillum hungatei, whereas the other one was only distantly related to other methanogens, with Methanopyrus kandleri being the closest cultivated relative. Most probable number (MPN) cultivations were accomplished with a sample from a 600 m(3) reactor from which all manures used in the experiments originated, and equal cell counts of ca. 10(9) g dry mass(-1) were found for cultivations with acetate, H(2) and methanol. SSCP analysis of these samples and sequencing of the DNA bands identified different hydrogenotrophic methanogens in all samples, and acetoclastic methanogens closely related to Methanosarcina mazei in the samples cultivated with acetate and methanol. As the acetoclastic species were not found in any other SSCP sample, it was supposed that the ammonia values in the manure of the laboratory biogas reactor, which ranged from 2.48 to 3.61 g NH(4)-NL(-1), inhibited the growth of the acetoclastic methanogens.

  19. Performance and microbial community analysis of two-stage process with extreme thermophilic hydrogen and thermophilic methane production from hydrolysate in UASB reactors.

    Science.gov (United States)

    Kongjan, Prawit; O-Thong, Sompong; Angelidaki, Irini

    2011-03-01

    The two-stage process for extreme thermophilic hydrogen and thermophilic methane production from wheat straw hydrolysate was investigated in up-flow anaerobic sludge bed (UASB) reactors. Specific hydrogen and methane yields of 89 ml-H(2)/g-VS (190 ml-H(2)/g-sugars) and 307 ml-CH(4)/g-VS, respectively were achieved simultaneously with the overall VS removal efficiency of 81% by operating with total hydraulic retention time (HRT) of 4 days . The energy conversion efficiency was dramatically increased from only 7.5% in the hydrogen stage to 87.5% of the potential energy from hydrolysate, corresponding to total energy of 13.4 kJ/g-VS. Dominant hydrogen-producing bacteria in the H(2)-UASB reactor were Thermoanaerobacter wiegelii, Caldanaerobacter subteraneus, and Caloramator fervidus. Meanwhile, the CH(4)-UASB reactor was dominated with methanogens of Methanosarcina mazei and Methanothermobacter defluvii. The results from this study suggest the two stage anaerobic process can be effectively used for energy recovery and for stabilization of hydrolysate at anaerobic conditions.

  20. Enhanced waste activated sludge digestion using a submerged anaerobic dynamic membrane bioreactor: performance, sludge characteristics and microbial community

    Science.gov (United States)

    Yu, Hongguang; Wang, Zhiwei; Wu, Zhichao; Zhu, Chaowei

    2016-02-01

    Anaerobic digestion (AD) plays an important role in waste activated sludge (WAS) treatment; however, conventional AD (CAD) process needs substantial improvements, especially for the treatment of WAS with low solids content and poor anaerobic biodegradability. Herein, we propose a submerged anaerobic dynamic membrane bioreactor (AnDMBR) for simultaneous WAS thickening and digestion without any pretreatment. During the long-term operation, the AnDMBR exhibited an enhanced sludge reduction and improved methane production over CAD process. Moreover, the biogas generated in the AnDMBR contained higher methane content than CAD process. Stable carbon isotopic signatures elucidated the occurrence of combined methanogenic pathways in the AnDMBR process, in which hydrogenotrophic methanogenic pathway made a larger contribution to the total methane production. It was also found that organic matter degradation was enhanced in the AnDMBR, thus providing more favorable substrates for microorganisms. Pyrosequencing revealed that Proteobacteria and Bacteroidetes were abundant in bacterial communities and Methanosarcina and Methanosaeta in archaeal communities, which played an important role in the AnDMBR system. This study shed light on the enhanced digestion of WAS using AnDMBR technology.

  1. Potential and existing mechanisms of enteric methane production in ruminants

    Directory of Open Access Journals (Sweden)

    Junyi Qiao

    2014-10-01

    Full Text Available Enteric methane (CH4 emissions in ruminants have attracted considerable attention due to their impact on greenhouse gases and the contribution of agricultural practices to global warming. Over the last two decades, a number of approaches have been adopted to mitigate CH4 emissions. However, the mechanisms of methanogenesis have still not been fully defined. According to the genome sequences of M. ruminantium in the rumen and of M. AbM4 in the abomasum, the pathways of carbon dioxide (CO2 reduction and formate oxidation to CH4 have now been authenticated in ruminants. Furthermore, in the light of species or genera description of methanogens, the precursors of methanogenesis discovered in the rumen and research advances in related subjects, pathways of acetate dissimilation via Methanosarcina and Methanosaeta as well as metabolism of methanol to CH4 might be present in the rumen, although neither process has yet been experimentally demonstrated in the rumen. Herein the research advances in methanogenesic mechanisms including existing and potential mechanisms are reviewed in detail. In addition, further research efforts to understand the methanogenesis mechanism should focus on isolation and identification of more specific methanogens, and their genome sequences. Such increased knowledge will provide benefits in terms of improved dietary energy utilization and a reduced contribution of enteric CH4 emissions to total global greenhouse gas emissions from the ruminant production system.

  2. Biogas production and microbial community shift through neutral pH control during the anaerobic digestion of pig manure.

    Science.gov (United States)

    Zhou, Jun; Zhang, Rui; Liu, Fenwu; Yong, Xiaoyu; Wu, Xiayuan; Zheng, Tao; Jiang, Min; Jia, Honghua

    2016-10-01

    Laboratory-scale reactors, in which the pH could be auto-adjusted, were employed to investigate the mesophilic methane fermentation with pig manure (7.8% total solids) at pH 6.0, 7.0, and 8.0. Results showed that the performance of anaerobic digestion was strongly dependent on pH value. Biogas production and methane content at neutral pH 7.0 were significantly higher (16,607mL, 51.81%) than those at pH 6.0 (6916mL, 42.9%) and 8.0 (9739mL, 35.6%). Denaturing gradient gel electrophoresis fingerprinting and Shannon's index indicated that the samples contained highly diverse microbial communities. The major genus at pH 7.0 was Methanocorpusculum, compared with that was Methanosarcina at both pH 6.0 and 8.0. Our research revealed that cultures maintained at pH 7.0 could support increased biogas production, which has significant implications for the scale-up biogas engineering.

  3. Effect of magnetic field enhancement of the photocurrent in ferromagnetic metal-dielectric heterostructures SiO{sub 2}(Co)/GaAs

    Energy Technology Data Exchange (ETDEWEB)

    Pavlov, V. V.; Lutsev, L. V.; Usachev, P. A.; Astretsov, A. A.; Pisarev, R. V. [Ioffe Physical-Technical Institute, Russian Academy of Sciences, 194021 St. Petersburg (Russian Federation); Stognij, A. I.; Novitskii, N. N. [Scientific and Practical Materials Research Centre, National Academy of Sciences of Belarus, 220072 Minsk (Belarus)

    2015-04-13

    Heterostructures of silicon dioxide films containing cobalt nanoparticles SiO{sub 2}(Co) grown on GaAs substrate exhibit at room temperature high values of magnetic field enhancement of photocurrent in the vicinity and above the GaAs bandgap of ∼1.4 eV. For photon energies E above the GaAs bandgap, the photocurrent significantly increases, while the avalanche process is suppressed by the magnetic field, and the current flowing through the heterostructure decreases. The photocurrent is enhanced in the SiO{sub 2}(Co 60 at. %)/GaAs heterostructure at the magnetic field H = 1.65 kOe by a factor of about ten for the photon energy E = 1.5 eV. This phenomenon is explained by a model describing electronic transitions in magnetic fields with the spin-dependent recombination process at deep impurity centers in the GaAs interface region.

  4. Three-dimensional architecture of lithium-anodes made from graphite fibers coated with thin-films of silicon oxycarbide: Design, performance and manufacturability

    Science.gov (United States)

    Saleh, Ibrahim; Raj, Rishi

    2016-04-01

    Silicon oxycarbide (SiCO) is an amorphous molecular network of Sisbnd Csbnd O tetrahedra anchored to graphene-like carbon. The graphene forms a three dimensional cellular network with a domain size of ∼5 nm. Therefore nanometer thick films of SiCO grown on graphite may be expected to have unusual behavior. We grow these films on a bed of commercially available graphite fibers that serve the dual function of a current collector. The electrochemical behavior of the composite is measured as a function of the thickness of the SiCO films. Thick films approach the typical behavior of bulk SiCO (which has three times the capacity of graphite, but suffers from poor first cycle efficiency). However, films, approximately 100 nm thick, show high first cycle efficiency as well as high capacity. The composite performs better than the prediction from the rule-of-mixtures, which further substantiates the unusual behavior of the thin-film architecture. The Raman spectra of these thin films also differ from bulk SiCO. The development of thin graphite fibers, with a high surface to volume ratio that have the same capacity as the current graphite-powder technology, coupled with manufacturing of these thin-films by a liquid-polymer precursor based process, can propel these results toward commercialization.

  5. Autocatalytic growth of Co on pure Co surfaces using Co{sub 2}(CO){sub 8} precursor

    Energy Technology Data Exchange (ETDEWEB)

    Cordoba, R.; Sese, J.; Ibarra, M.R. [Laboratorio de Microscopias Avanzadas (LMA), Instituto de Nanociencia de Aragon (INA), Universidad de Zaragoza, E-50018 Zaragoza (Spain); Departamento de Fisica de la Materia Condensada, Universidad de Zaragoza, E-50009 Zaragoza (Spain); De Teresa, J.M., E-mail: deteresa@unizar.es [Laboratorio de Microscopias Avanzadas (LMA), Instituto de Nanociencia de Aragon (INA), Universidad de Zaragoza, E-50018 Zaragoza (Spain); Departamento de Fisica de la Materia Condensada, Universidad de Zaragoza, E-50009 Zaragoza (Spain); Instituto de Ciencia de Materiales de Aragon (ICMA), Facultad de Ciencias, Universidad de Zaragoza-CSIC, E-50009 Zaragoza (Spain)

    2012-12-15

    Highlights: Black-Right-Pointing-Pointer We investigate the autocatalytic growth of Co using Co{sub 2}(CO){sub 8} precursor. Black-Right-Pointing-Pointer On Si wafers and Co grown by FEBID, no role is played by autocatalytic growth. Black-Right-Pointing-Pointer On Co films grown by sputtering, Co grows autocatalytically. Black-Right-Pointing-Pointer Implications of the results on Co by FEBID are discussed. - Abstract: The autocatalytic growth of Co on different surfaces using the Co{sub 2}(CO){sub 8} precursor is investigated. It is observed that Co{sub 2}(CO){sub 8} molecules dissociate spontaneously on pure Co surfaces grown by sputtering, forming a pure Co film. The microstructure of this film consists of Co nanocrystals with size below 100 nm. However, when the same type of experiment is done on a Co surface grown by focused-electron-beam induced deposition there is no autocatalytic growth of Co. On other surfaces such as Si substrates and Al films grown by sputtering, the spontaneous dissociation of the Co{sub 2}(CO){sub 8} molecules does not occur. The origin and implications of these results are discussed.

  6. Fourfold magnetic anisotropy, coercivity and magnetization reversal of Co/V bilayers grown on MgO(0 0 1)

    Energy Technology Data Exchange (ETDEWEB)

    Calleja, J F [Departamento de Fisica, Facultad de Ciencias, Universidad de Oviedo, Calvo Sotelo s/n, 33007 Oviedo (Spain); Muro, M GarcIa del [Departament de Fisica Fonamental and Institut de Nanociencia i Nanotecnologia IN2UB de la Universitat de Barcelona, MartIi Franques, 1, E-08028 Barcelona (Spain); Presa, B [Departamento de Fisica, Facultad de Ciencias, Universidad de Oviedo, Calvo Sotelo s/n, 33007 Oviedo (Spain); Matarranz, R [Departamento de Fisica, Facultad de Ciencias, Universidad de Oviedo, Calvo Sotelo s/n, 33007 Oviedo (Spain); Corrales, J A [Departmento de Informatica, Universidad de Oviedo, Edificio Departamental 1, Campus de Viesques s/n, 33204 Gijon (Spain); Labarta, A [Departament de Fisica Fonamental and Institut de Nanociencia i Nanotecnologia IN2UB de la Universitat de Barcelona, MartIi Franques, 1, E-08028 Barcelona (Spain); Contreras, M C [Departamento de Fisica, Facultad de Ciencias, Universidad de Oviedo, Calvo Sotelo s/n, 33007 Oviedo (Spain)

    2007-11-21

    Magnetic anisotropy and magnetization reversal of Al/Co/V/MgO(0 0 1) thin films have been investigated. The films were fabricated by magnetron sputtering. The roles of both Co and V layers thicknesses have been studied. Magnetic characterization has been carried out by transverse susceptibility (TS) measurements and hysteresis loops. Cobalt is grown in the hcp structure on V with the c axis parallel to the film plane. Two types of hcp Co crystal are grown with the c axes perpendicular to each other. This structure gives rise to a fourfold magnetic anisotropy. When the V layer thickness is below 40 A a superimposed uniaxial anisotropy develops, the effect of which is a depression in the TS, in agreement with theoretical calculations. This uniaxial anisotropy is induced by the substrate and due to a discontinuous growth of the V layer. For hcp Co grown on V, the magnetic anisotropy rapidly increases with Co layer thickness. In this case, unexpected shifted hysteresis loops along the hard axes were observed when the films were not saturated. This has been explained by taking into account the magnetization reversal along the hard axis: it proceeds via magnetization rotation of some portions of the film at high fields, and by domain wall motion of the rest of the film at lower field values.

  7. 海岸带沉积物环境中甲烷代谢菌的富集培养及混合菌群分析%Enrichment cultivation of methane metabolic bacteria in the environment of sediments from the coastal zones and analysis of mixed bacteria communities

    Institute of Scientific and Technical Information of China (English)

    于丽波; 姜丽晶; 汤熙翔

    2011-01-01

    结合Hungate无氧分离技术,对海岸带沉积物中甲烷代谢菌进行了富集培养,分别为珠江口的甲烷产生菌和九龙江口的甲烷氧化菌及其他甲基氧化菌.其研究结果表明:在珠江口淇澳岛海岸带沉积物中,甲烷八叠球菌属(Methanosarcina)为优势菌株,分布于沉积物的上、中、下3个层位,并发现了部分序列与不可培养的泉古菌门( Crenarchaeota)的杂色泉古菌(miscellaneous crenarchaeotic group,MCG)的相似度为90% ~99%.在九龙江口的海岸带沉积物环境中,噬甲基菌属(Methylophaga)为优势菌群,在富集产物菌群多样性中占60% ~99%;还有一些相似度较低(为95% ~97%)的菌群,为潜在的新种.%Methane ( CH4 ) is one of the main " greenhouse gases" responsible for global wanning. Most methane in the atmosphere is the result of the interaction between Methanogens and Methanotrophs. Therefore, it is of great significance for both energy and environment to research Methanogens and Methanotrophs. In this study, enrichment and biodiversity analyses were conducted on bacteria related to methane metabolism from the coastal zone sediments. The Methanogens from Qi' ao Island in the Pearl River Estuary, China and the Methanotrophs and other methyl-oxidizing bacteria from the Jiulongjiang River Estuary, China were cultivated. The main experimental methods included the Hungate anaerobic technique for enrichment and isolation as well as a molecular biological technique for biodiversity analysis was made on the enrichment products. The current study indicated that Methanosarcina was the predominant genus identified throughout the three layers of sediments found in the seashore area of Qi' ao Island in Pearl River Estuary. These bacteria could utilize propionate and lactate as substrates in addition to utilizing methanol, H2/CO2, mono-methylamine ( MM A) , double-methyla-mine( DMA) , trimethylamine( TMA) , as previously reported. The uncultured

  8. The isolation and characterization of new C. thermocellum strains and the evaluation of multiple anaerobic digestion systems

    Science.gov (United States)

    Lv, Wen

    with system performances. Methanosarcina and Methanobacterium were the most important methanogenic genera in the digesters where intense hydrolysis/acidogenesis and methanogenesis occurred, while Methanosaeta established itself in the mesophilic digesters with sufficient retention time and low concentrations of volatile fatty acids (VFA). The populations of all the quantified methanogen genera (Methanobacterium, Methanosarcina, Methanosaeta , and Methanoculleus) were inversely correlated or associated with high concentrations of VFA. The results of DGGE and qPCR were confirmed and improved by the pyrosequencing data (Chapter 8). Different operation conditions led to the development of different microbial communities that resulted in the functional differences among AD systems. The bacterial community tended to be more diverse in the digesters with more lenient conditions. Firmicutes was a major phylum in each AD system and might be associated with system performance. Chloroflexi was a major phylum in each thermophilic digester with balanced hydrolysis/acidogenesis and methanogenesis, so it might be indicative of efficient operations of thermophilic digesters. Thermotogae only appeared as a major phylum in the AT-TPAD system and might be important to its performance. The results of my studies had impacts on the development of renewable bioenergy. On one hand, the two new thermophilic cellulolytic isolates may be further evaluated for development of CBP strains. On the other hand, the series of comparative and integrated studies of different AD systems provided new knowledge that may guide future research and development of AD systems, particularly TPAD systems. Furthermore, the correlation between system performances and microbial communities may help improve design and operation of AD in general.

  9. 升流式厌氧污泥床和连续流搅拌槽式反应器的废水处理效能及产甲烷菌群组成的对比分析%Comparative Analysis of the Efficiency and the Methanogens Composition in Upflow Anaerobic Sludge Blanket and Continuous Stirred-Tank Reactor

    Institute of Scientific and Technical Information of China (English)

    张立国; 李建政; 班巧英; 许一平

    2012-01-01

    分别运行升流式厌氧污泥床(UASB)反应器和连续流搅拌槽式反应器(CSTR)并使其达到稳定运行状态,在有机负荷率(OLR)均为6.0kg·m-3·d-1的条件下,对比分析了二者在稳定期的运行特性和产甲烷菌群的组成.结果表明,UASB的化学需氧量(COD)去除率为95%,显著高于CSTR的COD去除率(84%).然而,CSTR系统中的活性污泥的比产甲烷速率(315L·kg-1·d-1)和比COD去除率(0.85kg·kg-1·d-1)则显著高于UASB的260L·kg-1·d-1和0.67kg·kg-1·d-1.采用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)指纹分析技术对系统稳定期的活性污泥进行分析的结果表明,UASB系统的优势产甲烷菌为Methanosaeta concilii 和 Methanospirillum hungatei,而CSTR系统中的优势产甲烷菌为Methanosarcina mazeii和Methanobacterium formicicum.污泥微生物群落组成及其代谢特征的不同是造成厌氧处理系统效能差异的内在原因.UASB和CSTR在COD去除效能和污泥比活性方面各有所长,在实际应用中,须根据废水水质和预期处理程度合理选用.%The efficiency and the methanogens composition in an Upflow Anaerobic Sludge Blanket (UASB) reactor and a Continuous Stirred-Tank Reactor (CSTR) are investigated after achieving steady states at the same Organic Loading Rate (OLR) of 6.0kg· m-3 · d-1. The results show that the average removal rate of COD reaches 95% in the UASB, significantly higher than 84% of the CSTR. However, the specific methane production rate and the specific COD removal rate of the activated sludge are SlSL·kg-1·d-1 and 0.85kg·kg-1·d-1, respectively, in the CSTR, notably higher than those of the UASB of 260L·kg-1·d-1 and 0.67kg· kg-1·d-1, respectively. The analysis of the methanogens composition of the activated sludge by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) indicates that Methanosaeta concilii and Methanospirillum hungatei are the dominant methanogens in the UASB, while

  10. Development of biofilm in anaerobic reactors treating wastewater from coffee grain processing Desenvolvimento de biofilme em reatores anaeróbios tratando água residuária do processamento dos frutos do cafeeiro

    Directory of Open Access Journals (Sweden)

    Fátima R. L. Fia

    2010-02-01

    Full Text Available In recent decades the use of anaerobic fixed bed reactors has been established in Brazil for the treatment of different effluents. As the capability of retaining microorganisms by support media (fixed bed is a factor influencing the performance of these reactors, the present study aims at evaluating the influence of three fixed bed on the effectiveness of treating an effluent with high pollution potential: wastewater from coffee grain processing (WCP, with organic matter concentrations varying from 812 to 5320 mg L-1 in the form of chemical oxygen demand (COD. Support media used for the immobilization of biomass were: blast furnace slag, polyurethane foam and #2 crushed stone with porosities of 53, 95 and 48%, respectively. The mean efficiency of COD removal in the reactor filled with polyurethane foam was 80%, attributed to its higher porosity index, which also provided greater retention and fixation of biomass which, when quantified as total volatile solids, was found to be 1301 mg g-1 of foam. The biofilm was made up of various microorganisms, including rod, curved rods, cocci, filaments and morphologies similar to Methanosaeta sp. and Methanosarcina sp.Nas últimas décadas tem-se registrado, no Brasil, o uso de reatores anaeróbios de leito fixo para o tratamento de diversos tipos de efluentes. Uma vez que a capacidade de retenção de micro-organismos pelo meio suporte (leito fixo é fator de influência no desempenho desses reatores, buscou-se, com a realização do presente estudo, avaliar a influência do leito fixo na eficiência de três unidades tratando um efluente com elevado potencial poluidor: água residuária do processamento dos frutos do cafeeiro (ARC, com concentração de matéria orgânica variando entre 812 e 5.320 mg L-1 na forma de DQO. Os tipos de suporte utilizados na imobilização da biomassa foram: escória de alto-forno, espuma de poliuretano e brita nº 2, com índice de vazios de 53, 95 e 48%, respectivamente. A

  11. Power Stroke Angular Velocity Profiles of Archaeal A-ATP Synthase Versus Thermophilic and Mesophilic F-ATP Synthase Molecular Motors.

    Science.gov (United States)

    Sielaff, Hendrik; Martin, James; Singh, Dhirendra; Biuković, Goran; Grüber, Gerhard; Frasch, Wayne D

    2016-12-02

    The angular velocities of ATPase-dependent power strokes as a function of the rotational position for the A-type molecular motor A3B3DF, from the Methanosarcina mazei Gö1 A-ATP synthase, and the thermophilic motor α3β3γ, from Geobacillus stearothermophilus (formerly known as Bacillus PS3) F-ATP synthase, are resolved at 5 μs resolution for the first time. Unexpectedly, the angular velocity profile of the A-type was closely similar in the angular positions of accelerations and decelerations to the profiles of the evolutionarily distant F-type motors of thermophilic and mesophilic origins, and they differ only in the magnitude of their velocities. M. mazei A3B3DF power strokes occurred in 120° steps at saturating ATP concentrations like the F-type motors. However, because ATP-binding dwells did not interrupt the 120° steps at limiting ATP, ATP binding to A3B3DF must occur during the catalytic dwell. Elevated concentrations of ADP did not increase dwells occurring 40° after the catalytic dwell. In F-type motors, elevated ADP induces dwells 40° after the catalytic dwell and slows the overall velocity. The similarities in these power stroke profiles are consistent with a common rotational mechanism for A-type and F-type rotary motors, in which the angular velocity is limited by the rotary position at which ATP binding occurs and by the drag imposed on the axle as it rotates within the ring of stator subunits.

  12. Wetland restoration and methanogenesis: the activity of microbial populations and competition for substrates at different temperatures

    Directory of Open Access Journals (Sweden)

    V. Jerman

    2009-02-01

    Full Text Available Ljubljana marsh in Slovenia is a 16 000 ha area of partly drained fen, intended to be flooded to restore its ecological functions. The resultant water-logging may create anoxic conditions, eventually stimulating production and emission of methane, the most important greenhouse gas next to carbon dioxide. We examined the upper layer (~30 cm of Ljubljana marsh soil for microbial processes that would predominate in water-saturated conditions, focusing on the potential for iron reduction, carbon mineralization (CO2 and CH4 production, and methane emission. Methane emission from water-saturated microcosms was near minimum detectable levels even after extended periods of flooding (>5 months. Methane production in anoxic soil slurries started only after a lag period and was inversely related to iron reduction, which suggested that iron reduction out-competed methanogenesis for electron donors, such as H2 and acetate. Methane production was observed only in samples incubated at 14–38°C. At the beginning of methanogenesis, acetoclastic methanogenesis dominated. In accordance with the preferred substrate, most (91% mcrA (encoding the methyl coenzyme-M reductase, a key gene in methanogenesis clone sequences could be affiliated to the acetoclastic genus Methanosarcina. No methanogens were detected in the original soil. However, a diverse community of iron-reducing Geobacteraceae was found. Our results suggest that methane emission can remain transient and low if water-table fluctuations allow re-oxidation of ferrous iron, sustaining iron reduction as the most important process in terminal carbon mineralization.

  13. Wetland restoration and methanogenesis: the activity of microbial populations and competition for substrates at different temperatures

    Directory of Open Access Journals (Sweden)

    V. Jerman

    2009-06-01

    Full Text Available Ljubljana marsh in Slovenia is a 16 000 ha area of partly drained fen, intended to be flooded to restore its ecological functions. The resultant water-logging may create anoxic conditions, eventually stimulating production and emission of methane, the most important greenhouse gas next to carbon dioxide. We examined the upper layer (~30 cm of Ljubljana marsh soil for microbial processes that would predominate in water-saturated conditions, focusing on the potential for iron reduction, carbon mineralization (CO2 and CH4 production, and methane emission. Methane emission from water-saturated microcosms was near minimum detectable levels even after extended periods of flooding (>5 months. Methane production in anoxic soil slurries started only after a lag period of 84 d at 15°C and a minimum of 7 d at 37°C, the optimum temperature for methanogenesis. This lag was inversely related to iron reduction, which suggested that iron reduction out-competed methanogenesis for electron donors, such as H2 and acetate. Methane production was observed only in samples incubated at 14–38°C. At the beginning of methanogenesis, acetoclastic methanogenesis dominated. In accordance with the preferred substrate, most (91% mcrA (encoding the methyl coenzyme-M reductase, a key gene in methanogenesis clone sequences could be affiliated to the acetoclastic genus Methanosarcina. No methanogens were detected in the original soil. However, a diverse community of iron-reducing Geobacteraceae was found. Our results suggest that methane emission can remain transient and low if water-table fluctuations allow re-oxidation of ferrous iron, sustaining iron reduction as the most important process in terminal carbon mineralization.

  14. Structural basis of the oxidative activation of the carboxysomal [gamma]-carbonic anhydrase, CcmM

    Energy Technology Data Exchange (ETDEWEB)

    Peña, Kerry L.; Castel, Stephane E.; de Araujo, Charlotte; Espie, George S.; Kimber, Matthew S. (Guelph); (Toronto)

    2010-04-26

    Cyanobacterial RuBisCO is sequestered in large, icosahedral, protein-bounded microcompartments called carboxysomes. Bicarbonate is pumped into the cytosol, diffuses into the carboxysome through small pores in its shell, and is then converted to CO{sub 2} by carbonic anhydrase (CA) prior to fixation. Paradoxically, many {beta}-cyanobacteria, including Thermosynechococcus elongatus BP-1, lack the conventional carboxysomal {beta}-CA, ccaA. The N-terminal domain of the carboxysomal protein CcmM is homologous to {gamma}-CA from Methanosarcina thermophila (Cam) but recombinant CcmM derived from ccaA-containing cyanobacteria show no CA activity. We demonstrate here that either full length CcmM from T. elongatus, or a construct truncated after 209 residues (CcmM209), is active as a CA - the first catalytically active bacterial {gamma}-CA reported. The 2.0 {angstrom} structure of CcmM209 reveals a trimeric, left-handed {beta}-helix structure that closely resembles Cam, except that residues 198-207 form a third {alpha}-helix stabilized by an essential Cys194-Cys200 disulfide bond. Deleting residues 194-209 (CcmM193) results in an inactive protein whose 1.1 {angstrom} structure shows disordering of the N- and C-termini, and reorganization of the trimeric interface and active site. Under reducing conditions, CcmM209 is similarly partially disordered and inactive as a CA. CcmM protein in fresh E. coli cell extracts is inactive, implying that the cellular reducing machinery can reduce and inactivate CcmM, while diamide, a thiol oxidizing agent, activates the enzyme. Thus, like membrane-bound eukaryotic cellular compartments, the {beta}-carboxysome appears to be able to maintain an oxidizing interior by precluding the entry of thioredoxin and other endogenous reducing agents.

  15. Groundwater ecosystem resilience to organic contaminations: microbial and geochemical dynamics throughout the 5-year life cycle of a surrogate ethanol blend fuel plume.

    Science.gov (United States)

    Ma, Jie; Nossa, Carlos W; Alvarez, Pedro J J

    2015-09-01

    The capacity of groundwater ecosystem to recover from contamination by organic chemicals is a vital concern for environmental scientists. A pilot-scale aquifer system was used to investigate the long-term dynamics of contaminants, groundwater geochemistry, and microbial community structure (by 16S rRNA gene pyrosequencing and quantitative real-time PCR) throughout the 5-year life cycle of a surrogate ethanol blend fuel plume (10% ethanol + 50 mg/L benzene + 50 mg/L toluene). Two-year continuous ethanol-blended release significantly changed the groundwater geochemistry (resulted in anaerobic, low pH, and organotrophic conditions) and increased bacterial and archaeal populations by 82- and 314-fold respectively. Various anaerobic heterotrophs (fermenters, acetogens, methanogens, and hydrocarbon degraders) were enriched. Two years after the release was shut off, all contaminants and their degradation byproducts disappeared and groundwater geochemistry completely restored to the pre-release states (aerobic, neutral pH, and oligotrophic). Bacterial and archaeal populations declined by 18- and 45-fold respectively (relative to the time of shut off). Microbial community structure reverted towards the pre-release states and alpha diversity indices rebounded, suggesting the resilience of microbial community to ethanol blend releases. We also found shifts from O2-sensitive methanogens (e.g., Methanobacterium) to methanogens that are not so sensitive to O2 (e.g., Methanosarcina and Methanocella), which is likely to contribute to the persistence of methanogens and methane generation following the source removal. Overall, the rapid disappearance of contaminants and their metabolites, rebound of geochemical footprints, and resilience of microbial community unequivocally document the natural capacity of groundwater ecosystem to attenuate and recover from a large volume of catastrophic spill of ethanol-based biofuel.

  16. Metabolic flexibility as a major predictor of spatial distribution in microbial communities.

    Directory of Open Access Journals (Sweden)

    Franck Carbonero

    Full Text Available A better understand the ecology of microbes and their role in the global ecosystem could be achieved if traditional ecological theories can be applied to microbes. In ecology organisms are defined as specialists or generalists according to the breadth of their niche. Spatial distribution is often used as a proxy measure of niche breadth; generalists have broad niches and a wide spatial distribution and specialists a narrow niche and spatial distribution. Previous studies suggest that microbial distribution patterns are contrary to this idea; a microbial generalist genus (Desulfobulbus has a limited spatial distribution while a specialist genus (Methanosaeta has a cosmopolitan distribution. Therefore, we hypothesise that this counter-intuitive distribution within generalist and specialist microbial genera is a common microbial characteristic. Using molecular fingerprinting the distribution of four microbial genera, two generalists, Desulfobulbus and the methanogenic archaea Methanosarcina, and two specialists, Methanosaeta and the sulfate-reducing bacteria Desulfobacter were analysed in sediment samples from along a UK estuary. Detected genotypes of both generalist genera showed a distinct spatial distribution, significantly correlated with geographic distance between sites. Genotypes of both specialist genera showed no significant differential spatial distribution. These data support the hypothesis that the spatial distribution of specialist and generalist microbes does not match that seen with specialist and generalist large organisms. It may be that generalist microbes, while having a wider potential niche, are constrained, possibly by intrageneric competition, to exploit only a small part of that potential niche while specialists, with far fewer constraints to their niche, are more capable of filling their potential niche more effectively, perhaps by avoiding intrageneric competition. We suggest that these counter-intuitive distribution

  17. Evaluation of Biogas Production Performance and Dynamics of the Microbial Community in Different Straws.

    Science.gov (United States)

    Li, Xue; Liu, Yan-Hua; Zhang, Xin; Ge, Chang-Ming; Piao, Ren-Zhe; Wang, Wei-Dong; Cui, Zong-Jun; Zhao, Hong-Yan

    2017-03-28

    The development and utilization of crop straw biogas resources can effectively alleviate the shortage of energy, environmental pollution, and other issues. This study performed a continuous batch test at 35°C to assess the methane production potential and volatile organic acid contents using the modified Gompertz equation. Illumina MiSeq platform sequencing, which is a sequencing method based on sequencing-by-synthesis, was used to compare the archaeal community diversity, and denaturing gradient gel electrophoresis (DGGE) was used to analyze the bacterial community diversity in rice straw, dry maize straw, silage maize straw, and tobacco straw. The results showed that cumulative gas production values for silage maize straw, rice straw, dry maize straw, and tobacco straw were 4,870, 4,032.5, 3,907.5, and 3,628.3 ml/g ·VS , respectively, after 24 days. Maximum daily gas production values of silage maize straw and rice straw were 1,025 and 904.17 ml/g ·VS, respectively, followed by tobacco straw and dry maize straw. The methane content of all four kinds of straws was > 60%, particularly that of silage maize straw, which peaked at 67.3%. Biogas production from the four kinds of straw was in the order silage maize straw > rice straw > dry maize straw > tobacco straw, and the values were 1,166.7, 1,048.4, 890, and 637.4 ml/g ·VS, respectively. The microbial community analysis showed that metabolism was mainly carried out by acetate-utilizing methanogens, and that Methanosarcina was the dominant archaeal genus in the four kinds of straw, and the DGGE bands belonged to the phyla Firmicutes, Bacteroidetes, and Chloroflexi. Silage maize is useful for biogas production because it contains four kinds of straw.

  18. Comparative genomic analyses of nickel, cobalt and vitamin B12 utilization

    Directory of Open Access Journals (Sweden)

    Gelfand Mikhail S

    2009-02-01

    Full Text Available Abstract Background Nickel (Ni and cobalt (Co are trace elements required for a variety of biological processes. Ni is directly coordinated by proteins, whereas Co is mainly used as a component of vitamin B12. Although a number of Ni and Co-dependent enzymes have been characterized, systematic evolutionary analyses of utilization of these metals are limited. Results We carried out comparative genomic analyses to examine occurrence and evolutionary dynamics of the use of Ni and Co at the level of (i transport systems, and (ii metalloproteomes. Our data show that both metals are widely used in bacteria and archaea. Cbi/NikMNQO is the most common prokaryotic Ni/Co transporter, while Ni-dependent urease and Ni-Fe hydrogenase, and B12-dependent methionine synthase (MetH, ribonucleotide reductase and methylmalonyl-CoA mutase are the most widespread metalloproteins for Ni and Co, respectively. Occurrence of other metalloenzymes showed a mosaic distribution and a new B12-dependent protein family was predicted. Deltaproteobacteria and Methanosarcina generally have larger Ni- and Co-dependent proteomes. On the other hand, utilization of these two metals is limited in eukaryotes, and very few of these organisms utilize both of them. The Ni-utilizing eukaryotes are mostly fungi (except saccharomycotina and plants, whereas most B12-utilizing organisms are animals. The NiCoT transporter family is the most widespread eukaryotic Ni transporter, and eukaryotic urease and MetH are the most common Ni- and B12-dependent enzymes, respectively. Finally, investigation of environmental and other conditions and identity of organisms that show dependence on Ni or Co revealed that host-associated organisms (particularly obligate intracellular parasites and endosymbionts have a tendency for loss of Ni/Co utilization. Conclusion Our data provide information on the evolutionary dynamics of Ni and Co utilization and highlight widespread use of these metals in the three

  19. The Genome Sequence of the psychrophilic archaeon, Methanococcoides burtonii: the Role of Genome Evolution in Cold-adaptation

    Energy Technology Data Exchange (ETDEWEB)

    Allen, Michelle A.; Lauro, Federico M.; Williams, Timothy J.; Burg, Dominic; Siddiqui, Khawar S.; De Francisci, David; Chong, Kevin W.Y.; Pilak, Oliver; Chew, Hwee H.; De Maere, Matthew Z.; Ting, Lily; Katrib, Marilyn; Ng, Charmaine; Sowers, Kevin R.; Galperin, Michael Y.; Anderson, Iain J.; Ivanova, Natalia; Dalin, Eileen; Martinez, Michelle; Lapidus, Alla; Hauser, Loren; Land, Miriam; Thomas, Torsten; Cavicchioli, Ricardo

    2009-04-01

    Psychrophilic archaea are abundant and perform critical roles throughout the Earth's expansive cold biosphere. Here we report the first complete genome sequence for a psychrophilic methanogenic archaeon, Methanococcoides burtonii. The genome sequence was manually annotated including the use of a five tiered Evidence Rating system that ranked annotations from Evidence Rating (ER) 1 (gene product experimentally characterized from the parent organism) to ER5 (hypothetical gene product) to provide a rapid means of assessing the certainty of gene function predictions. The genome is characterized by a higher level of aberrant sequence composition (51%) than any other archaeon. In comparison to hyper/thermophilic archaea which are subject to selection of synonymous codon usage, M. burtonii has evolved cold adaptation through a genomic capacity to accommodate highly skewed amino acid content, while retaining codon usage in common with its mesophilic Methanosarcina cousins. Polysaccharide biosynthesis genes comprise at least 3.3% of protein coding genes in the genome, and Cell wall/membrane/envelope biogenesis COG genes are over-represented. Likewise, signal transduction (COG category T) genes are over-represented and M. burtonii has a high 'IQ' (a measure of adaptive potential) compared to many methanogens. Numerous genes in these two over-represented COG categories appear to have been acquired from {var_epsilon}- and {delta}-proteobacteria, as do specific genes involved in central metabolism such as a novel B form of aconitase. Transposases also distinguish M. burtonii from other archaea, and their genomic characteristics indicate they play an important role in evolving the M. burtonii genome. Our study reveals a capacity for this model psychrophile to evolve through genome plasticity (including nucleotide skew, horizontal gene transfer and transposase activity) that enables adaptation to the cold, and to the biological and physical changes that have

  20. Changing Feeding Regimes To Demonstrate Flexible Biogas Production: Effects on Process Performance, Microbial Community Structure, and Methanogenesis Pathways.

    Science.gov (United States)

    Mulat, Daniel Girma; Jacobi, H Fabian; Feilberg, Anders; Adamsen, Anders Peter S; Richnow, Hans-Hermann; Nikolausz, Marcell

    2015-10-23

    Flexible biogas production that adapts biogas output to energy demand can be regulated by changing feeding regimes. In this study, the effect of changes in feeding intervals on process performance, microbial community structure, and the methanogenesis pathway was investigated. Three different feeding regimes (once daily, every second day, and every 2 h) at the same organic loading rate were studied in continuously stirred tank reactors treating distiller's dried grains with solubles. A larger amount of biogas was produced after feeding in the reactors fed less frequently (once per day and every second day), whereas the amount remained constant in the reactor fed more frequently (every 2 h), indicating the suitability of the former for the flexible production of biogas. Compared to the conventional more frequent feeding regimes, a methane yield that was up to 14% higher and an improved stability of the process against organic overloading were achieved by employing less frequent feeding regimes. The community structures of bacteria and methanogenic archaea were monitored by terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA and mcrA genes, respectively. The results showed that the composition of the bacterial community varied under the different feeding regimes, and the observed T-RFLP patterns were best explained by the differences in the total ammonia nitrogen concentrations, H2 levels, and pH values. However, the methanogenic community remained stable under all feeding regimes, with the dominance of the Methanosarcina genus followed by that of the Methanobacterium genus. Stable isotope analysis showed that the average amount of methane produced during each feeding event by acetoclastic and hydrogenotrophic methanogenesis was not influenced by the three different feeding regimes.

  1. Spatial variations of community structures and methane cycling across a transect of Lei-Gong-Hou mud volcanoes in eastern Taiwan.

    Science.gov (United States)

    Wang, Pei-Ling; Chiu, Yi-Ping; Cheng, Ting-Wen; Chang, Yung-Hsin; Tu, Wei-Xain; Lin, Li-Hung

    2014-01-01

    This study analyzed cored sediments retrieved from sites distributed across a transect of the Lei-Gong-Hou mud volcanoes in eastern Taiwan to uncover the spatial distributions of biogeochemical processes and community assemblages involved in methane cycling. The profiles of methane concentration and carbon isotopic composition revealed various orders of the predominance of specific methane-related metabolisms along depth. At a site proximal to the bubbling pool, the methanogenic zone was sandwiched by the anaerobic methanotrophic zones. For two sites distributed toward the topographic depression, the methanogenic zone overlaid the anaerobic methanotrophic zone. The predominance of anaerobic methanotrophy at specific depth intervals is supported by the enhanced copy numbers of the ANME-2a 16S rRNA gene and coincides with high dissolved Fe/Mn concentrations and copy numbers of the Desulfuromonas/Pelobacter 16S rRNA gene. Assemblages of 16S rRNA and mcrA genes revealed that methanogenesis was mediated by Methanococcoides and Methanosarcina. pmoA genes and a few 16S rRNA genes related to aerobic methanotrophs were detected in limited numbers of subsurface samples. While dissolved Fe/Mn signifies the presence of anaerobic metabolisms near the surface, the correlations between geochemical characteristics and gene abundances, and the absence of aerobic methanotrophs in top sediments suggest that anaerobic methanotrophy is potentially dependent on iron/manganese reduction and dominates over aerobic methanotrophy for the removal of methane produced in situ or from a deep source. Near-surface methanogenesis contributes to the methane emissions from mud platform. The alternating arrangements of methanogenic and methanotrophic zones at different sites suggest that the interactions between mud deposition, evaporation, oxidation and fluid transport modulate the assemblages of microbial communities and methane cycling in different compartments of terrestrial mud volcanoes.

  2. The Effect of Higher Sludge Recycling Rate on Anaerobic Treatment of Palm Oil Mill Effluent in a Semi-Commercial Closed Digester for Renewable Energy

    Directory of Open Access Journals (Sweden)

    Alawi Sulaiman

    2009-01-01

    Full Text Available Problem statement: A 500 m3 semi-commercial closed anaerobic digester was constructed for Palm Oil Mill Effluent (POME treatment and methane gas capture for renewable energy. During the start-up operation period, the Volatile Fatty Acids (VFA accumulation could not be controlled and caused instability on the system. Approach: A settling tank was installed and sludge was recycled as to provide a balanced microorganisms population for the treatment of POME and methane gas production. The effect of sludge recycling rate was studied by applying Organic Loading Rates (OLR (between 1.0 and 10.0 kgCOD m-3 day-1 at different sludge recycling rates (6, 12 and 18 m3 day-1. Results: At sludge recycling rate of 18 m3 day-1, the maximum OLR was 10.0 kgCOD m-3 day-1 with biogas and methane productivity of 1.5 and 0.9 m3 m-3 day-1, respectively. By increasing the sludge recycling rate the VFA concentration was controlled below its inhibitory limit (1000 mg L-1 and the COD removal efficiency recorded was above 95% which indicated good treatment performance for the digester. Two methanogens species (Methanosarcina sp. and Methanosaeta concilii had been identified from sludge samples obtained from the digester and recycled stream. Conclusion: By increasing the sludge recycling rate upon higher application of OLR, the treatment process was kept stable with high COD removal efficiency. The biogas and methane productivity were initially improved but reduced once OLR and recycling rate were increased to 10.0 kg COD m3 day-1 and 18 m3 day-1 respectively.

  3. Biotic and abiotic processes contribute to successful anaerobic degradation of cyanide by UASB reactor biomass treating brewery waste water.

    Science.gov (United States)

    Novak, Domen; Franke-Whittle, Ingrid H; Pirc, Elizabeta Tratar; Jerman, Vesna; Insam, Heribert; Logar, Romana Marinšek; Stres, Blaž

    2013-07-01

    In contrast to the general aerobic detoxification of industrial effluents containing cyanide, anaerobic cyanide degradation is not well understood, including the microbial communities involved. To address this knowledge gap, this study measured anaerobic cyanide degradation and the rearrangements in bacterial and archaeal microbial communities in an upflow anaerobic sludge blanket (UASB) reactor biomass treating brewery waste water using bio-methane potential assays, molecular profiling, sequencing and microarray approaches. Successful biogas formation and cyanide removal without inhibition were observed at cyanide concentrations up to 5 mg l(-1). At 8.5 mg l(-1) cyanide, there was a 22 day lag phase in microbial activity, but subsequent methane production rates were equivalent to when 5 mg l(-1) was used. The higher cumulative methane production in cyanide-amended samples indicated that part of the biogas was derived from cyanide degradation. Anaerobic degradation of cyanide using autoclaved UASB biomass proceeded at a rate more than two times lower than when UASB biomass was not autoclaved, indicating that anaerobic cyanide degradation was in fact a combination of simultaneous abiotic and biotic processes. Phylogenetic analyses of bacterial and archaeal 16S rRNA genes for the first time identified and linked the bacterial phylum Firmicutes and the archaeal genus Methanosarcina sp. as important microbial groups involved in cyanide degradation. Methanogenic activity of unadapted granulated biomass was detected at higher cyanide concentrations than reported previously for the unadapted suspended biomass, making the aggregated structure and predominantly hydrogenotrophic nature of methanogenic community important features in cyanide degradation. The combination of brewery waste water and cyanide substrate was thus shown to be of high interest for industrial level anaerobic cyanide degradation. Copyright © 2013 Elsevier Ltd. All rights reserved.

  4. Effects of Spartina alterniflora invasion on the communities of methanogens and sulfate-reducing bacteria in estuarine marsh sediments

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    Jemaneh eZeleke

    2013-08-01

    Full Text Available The effect of plant invasion on the microorganisms of soil sediments is very important for estuary ecology. The community structures of methanogens and sulfate-reducing bacteria (SRB as a function of Spartina alterniflora invasion in Phragmites australis-vegetated sediments of the Dongtan wetland in the Yangtze River estuary, China, were investigated using 454 pyrosequencing and quantitative real-time PCR (qPCR of the methyl coenzyme M reductase A (mcrA and dissimilatory sulfite-reductase (dsrB genes. Sediment samples were collected from two replicate locations, and each location included three sampling stands each covered by monocultures of P. australis, S. alterniflora and both plants (transition stands, respectively. qPCR analysis revealed higher copy numbers of mcrA genes in sediments from S. alterniflora stands than P. australis stands (5- and 7.5-fold more in the spring and summer, respectively, which is consistent with the higher methane flux rates measured in the S. alterniflora stands (up to 8.01 ± 5.61 mg m-2 h-1. Similar trends were observed for SRB, and they were up to two orders of magnitude higher than the methanogens. Diversity indices indicated a lower diversity of methanogens in the S. alterniflora stands than the P. australis stands. In contrast, insignificant variations were observed in the diversity of SRB with the invasion. Although Methanomicrobiales and Methanococcales, the hydrogenotrophic methanogens, dominated in the salt marsh, Methanomicrobiales displayed a slight increase with the invasion and growth of S. alterniflora, whereas the later responded differently. Methanosarcina, the metabolically diverse methanogens, did not vary with the invasion of, but Methanosaeta, the exclusive acetate utilizers, appeared to increase with S. alterniflora invasion. In SRB, sequences closely related to the families Desulfobacteraceae and Desulfobulbaceae dominated in the salt marsh, although they displayed minimal changes with the S

  5. Comparative fecal metagenomics unveils unique functional capacity of the swine gut

    Directory of Open Access Journals (Sweden)

    Martinson John

    2011-05-01

    Full Text Available Abstract Background Uncovering the taxonomic composition and functional capacity within the swine gut microbial consortia is of great importance to animal physiology and health as well as to food and water safety due to the presence of human pathogens in pig feces. Nonetheless, limited information on the functional diversity of the swine gut microbiome is available. Results Analysis of 637, 722 pyrosequencing reads (130 megabases generated from Yorkshire pig fecal DNA extracts was performed to help better understand the microbial diversity and largely unknown functional capacity of the swine gut microbiome. Swine fecal metagenomic sequences were annotated using both MG-RAST and JGI IMG/M-ER pipelines. Taxonomic analysis of metagenomic reads indicated that swine fecal microbiomes were dominated by Firmicutes and Bacteroidetes phyla. At a finer phylogenetic resolution, Prevotella spp. dominated the swine fecal metagenome, while some genes associated with Treponema and Anareovibrio species were found to be exclusively within the pig fecal metagenomic sequences analyzed. Functional analysis revealed that carbohydrate metabolism was the most abundant SEED subsystem, representing 13% of the swine metagenome. Genes associated with stress, virulence, cell wall and cell capsule were also abundant. Virulence factors associated with antibiotic resistance genes with highest sequence homology to genes in Bacteroidetes, Clostridia, and Methanosarcina were numerous within the gene families unique to the swine fecal metagenomes. Other abundant proteins unique to the distal swine gut shared high sequence homology to putative carbohydrate membrane transporters. Conclusions The results from this metagenomic survey demonstrated the presence of genes associated with resistance to antibiotics and carbohydrate metabolism suggesting that the swine gut microbiome may be shaped by husbandry practices.

  6. Enzymological studies of one-carbon reactions in the pathway of acetate utilization by methanogenic bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Ferry, J.G.

    1991-12-31

    Several enzymes in the pathway of acetate conversion to methane and carbon dioxide have been purified from Methanosarcina thermophila. The mechanisms of these enzymes are under investigation utilizing biochemical, biophysical and molecular genetic approaches. Acetate kinase and phosphotransacetylase catalyzes the activation of acetate to acetyl-CoA. The primary structure of these enzymes will be determined through cloning and sequencing of the genes. Two protein components of the CO dehydrogenase complex are under investigations. The metal centers of each component have been characterized using EPR. Cloning and sequencing of the genes for the two subunits of each component is in progress. Results indicate that the Ni/Fe-S component cleaves the C-C and C-S bonds of acetyl-CoA followed by oxidation of the carbonyl group to carbon dioxide and transfer of the methyl group to the Co/Fe-S component. The enzymes and cofactors involved in transfer of the methyl group from the Co/Fe-S component to coenzyme M will be purified and characterized. Ferredoxin is an electron acceptor for the Ni/Fe-S component and also serves to reductively reactivate methylreductase which catalyzes the demethylation of methyl coenzyme M to methane. This ferredoxin is being characterized utilizing EPR and RR spectroscopic methods to determine the properties of the Fe-S centers. Genes encoding this and other ferredoxins have been cloned and sequenced to determine the primary structures. Carbonic anhydrase is being purified and characterized to determine the function of this enzyme in the pathway.

  7. Mechanistic enzymology of CO dehydrogenase from Clostridium thermoaceticum. Progress report, August 15, 1993--March 24, 1995

    Energy Technology Data Exchange (ETDEWEB)

    Ragsdale, S.W.

    1992-11-01

    The final steps in acetyl-CoA biosynthesis by anaerobic bacteria are performed by carbon monoxide dehydrogenase (CODH), a nickel/iron-sulfur protein. An important achievement was to establish conditions under which acetyl-CoA synthesis by purified enzymes equals the in vivo rate of acetate synthesis. Under these optimized conditions we established that the rate limiting step in the synthesis of acetyl-CoA from methyl-H{sub 4}folate, CO and CoA is likely to be the methylation of CODH by the methylated corrinoid/iron-sulfur protein. We then focused on stopped flow studies of this rate limiting transmethylation reaction and established its mechanism. We have studied the carbonylation of CODH by infrared and resonance Raman spectroscopy and determined that the [Ni-Fe{sup 3-4}S{sub 4}]-CO species which has been characterized by magnetic resonance methods can be described as [Ni-X-Fe{sub 3-4}S{sub 4}]-C{equivalent_to}O. We showed that this species is the catalytically competent precursor of the carbonyl group of acetyl-CoA. We have made progress in the synthesis of seleno-coenzyme A, which we will use to probe the binding of CoA to CODH. We also have compared the CODH from Methanosarcina thermophila with the C. thermoaceticum enzyme by EPR and electrochemical methods and found that the metal sites of these enzymes are remarkably similar given the evolutionary separation between archaea and bacteria domains.

  8. Enzymological studies of one-carbon reactions in the pathway of acetate utilization by methanogenic bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Ferry, J.G.

    1991-01-01

    Several enzymes in the pathway of acetate conversion to methane and carbon dioxide have been purified from Methanosarcina thermophila. The mechanisms of these enzymes are under investigation utilizing biochemical, biophysical and molecular genetic approaches. Acetate kinase and phosphotransacetylase catalyzes the activation of acetate to acetyl-CoA. The primary structure of these enzymes will be determined through cloning and sequencing of the genes. Two protein components of the CO dehydrogenase complex are under investigations. The metal centers of each component have been characterized using EPR. Cloning and sequencing of the genes for the two subunits of each component is in progress. Results indicate that the Ni/Fe-S component cleaves the C-C and C-S bonds of acetyl-CoA followed by oxidation of the carbonyl group to carbon dioxide and transfer of the methyl group to the Co/Fe-S component. The enzymes and cofactors involved in transfer of the methyl group from the Co/Fe-S component to coenzyme M will be purified and characterized. Ferredoxin is an electron acceptor for the Ni/Fe-S component and also serves to reductively reactivate methylreductase which catalyzes the demethylation of methyl coenzyme M to methane. This ferredoxin is being characterized utilizing EPR and RR spectroscopic methods to determine the properties of the Fe-S centers. Genes encoding this and other ferredoxins have been cloned and sequenced to determine the primary structures. Carbonic anhydrase is being purified and characterized to determine the function of this enzyme in the pathway.

  9. Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity

    Science.gov (United States)

    Yan, Jun; Im, Jeongdae; Yang, Yi; Löffler, Frank E.

    2013-01-01

    Dehalococcoides mccartyi strains are corrinoid-auxotrophic Bacteria and axenic cultures that require vitamin B12 (CN-Cbl) to conserve energy via organohalide respiration. Cultures of D. mccartyi strains BAV1, GT and FL2 grown with limiting amounts of 1 µg l−1 CN-Cbl quickly depleted CN-Cbl, and reductive dechlorination of polychlorinated ethenes was incomplete leading to vinyl chloride (VC) accumulation. In contrast, the same cultures amended with 25 µg l−1 CN-Cbl exhibited up to 2.3-fold higher dechlorination rates, 2.8–9.1-fold increased growth yields, and completely consumed growth-supporting chlorinated ethenes. To explore whether known cobamide-producing microbes supply Dehalococcoides with the required corrinoid cofactor, co-culture experiments were performed with the methanogen Methanosarcina barkeri strain Fusaro and two acetogens, Sporomusa ovata and Sporomusa sp. strain KB-1, as Dehalococcoides partner populations. During growth with H2/CO2, M. barkeri axenic cultures produced 4.2 ± 0.1 µg l−1 extracellular cobamide (factor III), whereas the Sporomusa cultures produced phenolyl- and p-cresolyl-cobamides. Neither factor III nor the phenolic cobamides supported Dehalococcoides reductive dechlorination activity suggesting that M. barkeri and the Sporomusa sp. cannot fulfil Dehalococcoides' nutritional requirements. Dehalococcoides dechlorination activity and growth occurred in M. barkeri and Sporomusa sp. co-cultures amended with 10 µM 5′,6′-dimethylbenzimidazole (DMB), indicating that a cobalamin is a preferred corrinoid cofactor of strains BAV1, GT and FL2 when grown with chlorinated ethenes as electron acceptors. Even though the methanogen and acetogen populations tested did not produce cobalamin, the addition of DMB enabled guided biosynthesis and generated a cobalamin that supported Dehalococcoides' activity and growth. Guided cobalamin biosynthesis may offer opportunities to sustain and enhance Dehalococcoides activity in contaminated

  10. EFFECT OF SULPHATE ON LOW-TEMPERATURE ANAEROBIC DIGESTION

    Directory of Open Access Journals (Sweden)

    Padhraig eMadden

    2014-07-01

    Full Text Available The effect of sulphate addition on the stability of, and microbial community behaviour in, low-temperature anaerobic expanded granular sludge bed-based bioreactors was investigated at 15°C. Efficient bioreactor performance was observed, with chemical oxygen demand removal efficiencies of >90%, and a mean SO42- removal rate of 98.3%. In situ methanogensis appeared unaffected at a COD:SO42- influent ratio of 8:1, and subsequently of 3:1, and was impacted marginally only when the COD: SO42- ratio was 1:2. . Specific methanogenic activity assays indicated a complex set of interactions between sulphate-reducing bacteria (SRB, methanogens and homoacetogenic bacteria. SO42- addition resulted in predominantly acetoclastic, rather than hydrogenotrophic, methanogenesis until >600 days of SO42--influenced bioreactor operation. Temporal microbial community development was monitored by denaturation gradient gel electrophoresis (DGGE of 16S rRNA genes. Fluorescence in situ hybridisations (FISH, qPCR and microsensor analysis were combined to investigate the distribution of microbial groups, and particularly SRB and methanogens, along the structure of granular biofilms. qPCR data indicated that sulphidogenic genes were present in methanogenic and sulfidogenic biofilms, indicating the potential for sulphate reduction even in bioreactors not exposed to SO42-. Although the architecture of methanogenic and sulphidogenic granules was similar, indicating the presence of SRB even in methanogenic systems, FISH with rRNA targets found that the SRB were more abundant in the sulphidogenic biofilms. Methanosaeta species were the predominant, keystone members of the archaeal community, with the complete absence of the Methanosarcina species in the experimental bioreactor by trial conclusion. Microsensor data suggested the ordered distribution of sulphate reduction and sulphide accumulation, even in methanogenic granules.

  11. Effect of sulfate on low-temperature anaerobic digestion.

    Science.gov (United States)

    Madden, Pádhraig; Al-Raei, Abdul M; Enright, Anne M; Chinalia, Fabio A; de Beer, Dirk; O'Flaherty, Vincent; Collins, Gavin

    2014-01-01

    The effect of sulfate addition on the stability of, and microbial community behavior in, low-temperature anaerobic expanded granular sludge bed-based bioreactors was investigated at 15°C. Efficient bioreactor performance was observed, with chemical oxygen demand (COD) removal efficiencies of >90%, and a mean SO(2-) 4 removal rate of 98.3%. In situ methanogensis appeared unaffected at a COD: SO(2-) 4 influent ratio of 8:1, and subsequently of 3:1, and was impacted marginally only when the COD: SO(2-) 4 ratio was 1:2. Specific methanogenic activity assays indicated a complex set of interactions between sulfate-reducing bacteria (SRB), methanogens and homoacetogenic bacteria. SO(2-) 4 addition resulted in predominantly acetoclastic, rather than hydrogenotrophic, methanogenesis until >600 days of SO(2-) 4-influenced bioreactor operation. Temporal microbial community development was monitored by denaturation gradient gel electrophoresis (DGGE) of 16S rRNA genes. Fluorescence in situ hybridizations (FISH), qPCR and microsensor analysis were combined to investigate the distribution of microbial groups, and particularly SRB and methanogens, along the structure of granular biofilms. qPCR data indicated that sulfidogenic genes were present in methanogenic and sulfidogenic biofilms, indicating the potential for sulfate reduction even in bioreactors not exposed to SO(2-) 4. Although the architecture of methanogenic and sulfidogenic granules was similar, indicating the presence of SRB even in methanogenic systems, FISH with rRNA targets found that the SRB were more abundant in the sulfidogenic biofilms. Methanosaeta species were the predominant, keystone members of the archaeal community, with the complete absence of the Methanosarcina species in the experimental bioreactor by trial conclusion. Microsensor data suggested the ordered distribution of sulfate reduction and sulfide accumulation, even in methanogenic granules.

  12. Microbial community structures in an integrated two-phase anaerobic bioreactor fed by fruit vegetable wastes and wheat straw

    Institute of Scientific and Technical Information of China (English)

    Chong Wang; Jiane Zuo; Xiaojie Chen; Wei Xing; Linan Xing; Peng Li; Xiangyang Lu

    2014-01-01

    The microbial community structures in an integrated two-phase anaerobic reactor (ITPAR) were investigated by 16S rDNA clone library technology.The 75 L reactor was designed with a 25 L rotating acidogenic unit at the top and a 50 L conventional upflow methanogenic unit at the bottom,with a recirculation connected to the two units.The reactor had been operated for 21 stages to co-digest fruit/vegetable wastes and wheat straw,which showed a very good biogas production and decomposition of cellulosic materials.The results showed that many kinds of cellulose and glycan decomposition bacteria related with Bacteroidales,Clostridiales and Syntrophobacterales were dominated in the reactor,with more bacteria community diversities in the acidogenic unit.The methanogens were mostly related with Methanosaeta,Methanosarcina,Methanoculleus,Methanospirillum and Methanobacterium; the predominating genus Methanosaeta,accounting for 40.5%,54.2%,73.6% and 78.7% in four samples from top to bottom,indicated a major methanogenesis pathway by acetoclastic methanogenesis in the methanogenic unit.The beta diversity indexes illustrated a more similar distribution of bacterial communities than that of methanogens between acidogenic unit and methanogenic unit.The differentiation of methanogenic community composition in two phases,as well as pH values and volatile fatty acid (VFA) concentrations confirmed the phase separation of the ITPAR.Overall,the results of this study demonstrated that the special designing of ITPAR maintained a sufficient number of methanogens,more diverse communities and stronger syntrophic assodations among microorganisms,which made two phase anaerobic digestion of cellulosic materials more efficient.

  13. Spatial variations of community structures and methane cycling across a transect of Lei-Gong-Hou mud volcanoes in eastern Taiwan

    Directory of Open Access Journals (Sweden)

    Pei-Ling eWang

    2014-03-01

    Full Text Available This study analyzed cored sediments retrieved from sites distributed across a transect of the Lei-Gong-Hou mud volcanoes in eastern Taiwan to uncover the spatial distributions of biogeochemical processes and community assemblages involved in methane cycling. The profiles of methane concentration and carbon isotopic composition revealed various orders of the predominance of specific methane-related metabolisms along depth. At a site proximal to the bubbling pool, the methanogenic zone was sandwiched by the anaerobic methanotrophic zones. For two sites distributed toward the topographic depression, the methanogenic zone overlaid the anaerobic methanotrophic zone. The predominance of anaerobic methanotrophy at specific depth intervals is supported by the enhanced copy numbers of the ANME-2a 16S rRNA gene and coincides with high dissolved Fe/Mn concentrations and copy numbers of the Desulfuromonas/Pelobacter 16S rRNA gene. Assemblages of 16S rRNA and mcrA genes revealed that methanogenesis was mediated by Methanococcoides and Methanosarcina. pmoA genes and a few 16S rRNA genes related to aerobic methanotrophs were detected in limited numbers of subsurface samples. While dissolved Fe/Mn signifies the presence of anaerobic metabolisms near the surface, the correlations between geochemical characteristics and gene abundances, and the absence of aerobic methanotrophs in top sediments suggest that anaerobic methanotrophy is potentially dependent on iron/manganese reduction and dominates over aerobic methanotrophy for the removal of methane produced in situ or from a deep source. Near-surface methanogenesis contributes to the methane emissions from mud platform. The alternating arrangements of methanogenic and methanotrophic zones at different sites suggest that the interactions between mud deposition, evaporation, oxidation and fluid transport modulate the assemblages of microbial communities and methane cycling in different compartments of terrestrial

  14. Comparing mesophilic and thermophilic anaerobic digestion of chicken manure: Microbial community dynamics and process resilience

    Energy Technology Data Exchange (ETDEWEB)

    Niu, Qigui; Takemura, Yasuyuki; Kubota, Kengo [Department of Civil and Environmental Engineering, Graduate School of Engineering Tohoku University, 6-6-06 Aza-Aoba, Aramaki, Aoba-ku, Sendai, Miyagi 980-8579 (Japan); Li, Yu-You, E-mail: yyli@epl1.civil.tohoku.ac.jp [Department of Civil and Environmental Engineering, Graduate School of Engineering Tohoku University, 6-6-06 Aza-Aoba, Aramaki, Aoba-ku, Sendai, Miyagi 980-8579 (Japan); Key Lab of Northwest Water Resource, Environment and Ecology, MOE, Xi’an University of Architecture and Technology, Xi’an (China)

    2015-09-15

    Highlights: • Microbial community dynamics and process functional resilience were investigated. • The threshold of TAN in mesophilic reactor was higher than the thermophilic reactor. • The recoverable archaeal community dynamic sustained the process resilience. • Methanosarcina was more sensitive than Methanoculleus on ammonia inhibition. • TAN and FA effects the dynamic of hydrolytic and acidogenic bacteria obviously. - Abstract: While methane fermentation is considered as the most successful bioenergy treatment for chicken manure, the relationship between operational performance and the dynamic transition of archaeal and bacterial communities remains poorly understood. Two continuous stirred-tank reactors were investigated under thermophilic and mesophilic conditions feeding with 10%TS. The tolerance of thermophilic reactor on total ammonia nitrogen (TAN) was found to be 8000 mg/L with free ammonia (FA) 2000 mg/L compared to 16,000 mg/L (FA1500 mg/L) of mesophilic reactor. Biomethane production was 0.29 L/gV S{sub in} in the steady stage and decreased following TAN increase. After serious inhibition, the mesophilic reactor was recovered successfully by dilution and washing stratagem compared to the unrecoverable of thermophilic reactor. The relationship between the microbial community structure, the bioreactor performance and inhibitors such as TAN, FA, and volatile fatty acid was evaluated by canonical correspondence analysis. The performance of methanogenic activity and substrate removal efficiency were changed significantly correlating with the community evenness and phylogenetic structure. The resilient archaeal community was found even after serious inhibition in both reactors. Obvious dynamics of bacterial communities were observed in acidogenic and hydrolytic functional bacteria following TAN variation in the different stages.

  15. Archaeal community diversity and abundance changes along a natural salinity gradient in estuarine sediments.

    Science.gov (United States)

    Webster, Gordon; O'Sullivan, Louise A; Meng, Yiyu; Williams, Angharad S; Sass, Andrea M; Watkins, Andrew J; Parkes, R John; Weightman, Andrew J

    2015-02-01

    Archaea are widespread in marine sediments, but their occurrence and relationship with natural salinity gradients in estuarine sediments is not well understood. This study investigated the abundance and diversity of Archaea in sediments at three sites [Brightlingsea (BR), Alresford (AR) and Hythe (HY)] along the Colne Estuary, using quantitative real-time PCR (qPCR) of 16S rRNA genes, DNA hybridization, Archaea 16S rRNA and mcrA gene phylogenetic analyses. Total archaeal 16S rRNA abundance in sediments were higher in the low-salinity brackish sediments from HY (2-8 × 10(7) 16S rRNA gene copies cm(-3)) than the high-salinity marine sites from BR and AR (2 × 10(4)-2 × 10(7) and 4 × 10(6)-2 × 10(7) 16S rRNA gene copies cm(-3), respectively), although as a proportion of the total prokaryotes Archaea were higher at BR than at AR or HY. Phylogenetic analysis showed that members of the 'Bathyarchaeota' (MCG), Thaumarchaeota and methanogenic Euryarchaeota were the dominant groups of Archaea. The composition of Thaumarchaeota varied with salinity, as only 'marine' group I.1a was present in marine sediments (BR). Methanogen 16S rRNA genes from low-salinity sediments at HY were dominated by acetotrophic Methanosaeta and putatively hydrogentrophic Methanomicrobiales, whereas the marine site (BR) was dominated by mcrA genes belonging to methylotrophic Methanococcoides, versatile Methanosarcina and methanotrophic ANME-2a. Overall, the results indicate that salinity and associated factors play a role in controlling diversity and distribution of Archaea in estuarine sediments.

  16. Detoxification of tar water by anaerobic treatment in an UASB reactor - A study of the degradation of phenolic compounds in a combined denitrifying and anaerobic UASB reactor

    Energy Technology Data Exchange (ETDEWEB)

    Skibsted Mogensen, A.; Schmidt, J.E.; Ahring, B.K. [Technical Univ., Dept. of Environmental Science and Engineering, Lyngby (Denmark)

    1998-08-01

    The digestion of pyrolysis condensate (PC) in two combined anaerobic and denitrifying upflow anaerobic sludge blanket (UASB) reactors was studied. A COD removal of 80% was achieved with an influent concentration of 1.43% PC{sub pH}. When the reactor was fed with 100% PC during a period of 10 days good reactor operation was observed. Despite less than one retention time of operation, the results indicated clearly, that PC could be used as substrate in the biogas process, even in very high concentrations. A combined anaerobic and denitrifying UASB reactor was successfully digesting 5.5% of wet oxidised PC, but further loading increments deteriorated the anaerobic digestion process. The detoxification of PC was studied by determining the degradation of phenols during reactor operation and the toxicity of PC was decreased more than 77 times witnessed through decreased inhibition of the nitrification process. Phenol, methyl and dimethyl phenols along with methoxyphenols were shown to be degraded within the reactor systems. Degradation rates for phenol and substituted phenols were determined by the reactor experiment indicating that the biomass was selective towards the substrates. Maximum growth rates and half saturation constants for phenol, 4-Methylphenol and 2-Methoxy-4-methylphenol were determined in batch experiments. The degradation rates of phenols determined in batches were significantly higher compared to degradation rates observed in the reactor systems digesting pyrolysis condensate. Determination of the population of methanogens revealed, that Methanosarcina was found only in one reactor, while Methanobacterium and Methanosaeta were found in reactors and inoculum. A UASB reactor was designed for the treatment of pyrolysis condensate at the gasification plant at Harbooere, Denmark. (au) 35 refs.

  17. Sensitivity and adaptability of methanogens to perchlorates: Implications for life on Mars

    Science.gov (United States)

    Kral, Timothy A.; Goodhart, Timothy H.; Harpool, Joshua D.; Hearnsberger, Christopher E.; McCracken, Graham L.; McSpadden, Stanley W.

    2016-01-01

    In 2008, the Mars Phoenix Lander discovered perchlorate at its landing site, and in 2012, the Curiosity rover confirmed the presence of perchlorate on Mars. The research reported here was designed to determine if certain methanogens could grow in the presence of three different perchlorate salt solutions. The methanogens tested were Methanothermobacter wolfeii, Methanosarcina barkeri, Methanobacterium formicicum and Methanococcus maripaludis. Media were prepared containing 0%, 0.5%, 1.0%, 2%, 5% and 10% wt/vol magnesium perchlorate, sodium perchlorate, or calcium perchlorate. Organisms were inoculated into their respective media followed by incubation at each organism's growth temperature. Methane production, commonly used to measure methanogen growth, was measured by gas chromatography of headspace gas samples. Methane concentrations varied with species and perchlorate salt tested. However, all four methanogens produced substantial levels of methane in the presence of up to 1.0% perchlorate, but not higher. The standard procedure for growing methanogens typically includes sodium sulfide, a reducing agent, to reduce residual molecular oxygen. However, the sodium sulfide may have been reducing the perchlorate, thus allowing for growth of the methanogens. To investigate this possibility, experiments were conducted where stainless steel nails were used instead of sodium sulfide as the reducing agent. Prior to the addition of perchlorate and inoculation, the nails were removed from the liquid medium. Just as in the prior experiments, the methanogens produced methane at comparable levels to those seen with sodium sulfide as the reductant, indicating that sodium sulfide did not reduce the perchlorate to any significant extent. Additionally, cells metabolizing in 1% perchlorate were transferred to 2%, cells metabolizing in 2% were transferred to 5%, and finally cells metabolizing in 5% were transferred to 10%. All four species produced methane at 2% and 5%, but not 10

  18. Field Evidence for Magnetite Formation by a Methanogenic Microbial Community

    Science.gov (United States)

    Rossbach, S.; Beaver, C. L.; Williams, A.; Atekwana, E. A.; Slater, L. D.; Ntarlagiannis, D.; Lund, A.

    2015-12-01

    The aged, subsurface petroleum spill in Bemidji, Minnesota, has been surveyed with magnetic susceptibility (MS) measurements. High MS values were found in the free-product phase around the fluctuating water table. Although we had hypothesized that high MS values are related to the occurrence of the mineral magnetite resulting from the activity of iron-reducing bacteria, our microbial analysis pointed to the presence of a methanogenic microbial community at the locations and depths of the highest MS values. Here, we report on a more detailed microbial analysis based on high-throughput sequencing of the 16S rRNA gene of sediment samples from four consecutive years. In addition, we provide geochemical data (FeII/FeIII concentrations) to refine our conceptual model of methanogenic hydrocarbon degradation at aged petroleum spills and demonstrate that the microbial induced changes of sediment properties can be monitored with MS. The methanogenic microbial community at the Bemidji site consisted mainly of the syntrophic, hydrocarbon-degrading Smithella and the hydrogenotrophic, methane-generating Methanoregula. There is growing evidence in the literature that not only Bacteria, but also some methanogenic Archaea are able to reduce iron. In fact, a recent study reported that the methanogen Methanosarcina thermophila produced magnetite during the reduction of ferrihydrite in a laboratory experiment when hydrogen was present. Therefore, our finding of high MS values and the presence of magnetite in the methanogenic zone of an aged, subsurface petroleum spill could very well be the first field evidence for magnetite formation during methanogenic hydrocarbon degradation.

  19. Link Between Capacity for Current Production and Syntrophic Growth in Geobacter species

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    Amelia-Elena eRotaru

    2015-07-01

    Full Text Available Electrodes are unnatural electron acceptors, and it is yet unknown how some Geobacter species evolved to use electrodes as terminal electron acceptors. Analysis of different Geobacter species revealed that they varied in their capacity for current production. G. metallireducens and G. hydrogenophilus generated high current densities (ca. 0.05 mA/cm2, comparable to G. sulfurreducens. G. bremensis, G. chapellei, G. humireducens, and G. uranireducens, produced much lower currents (ca. 0.05 mA/cm2 and G. bemidjiensis was previously found to not produce current. There was no correspondence between the effectiveness of current generation and Fe(III oxide reduction rates. Some high-current-density strains (G. metallireducens and G. hydrogenophilus reduced Fe(III-oxides as fast as some low-current-density strains (G. bremensis, G. humireducens, and G. uranireducens whereas other low-current-density strains (G. bemidjiensis and G. chapellei reduced Fe(III oxide as slowly as G. sulfurreducens, a high-current-density strain. However, there was a correspondence between the ability to produce higher currents and the ability to grow syntrophically. G. hydrogenophilius was found to grow in co-culture with Methanosarcina barkeri, which is capable of direct interspecies electron transfer (DIET, but not with Methanospirillium hungatei capable only of H2 or formate transfer. Conductive granular activated carbon (GAC stimulated metabolism of the G. hydrogenophilus - M. barkeri co-culture, consistent with electron exchange via DIET. These findings, coupled with the previous finding that G. metallireducens and G. sulfurreducens are also capable of DIET, suggest that evolution to optimize DIET has fortuitiously conferred the capability for high-density current production to some Geobacter species.

  20. Novel euryarchaeotal lineages detected on rice roots and in the anoxic bulk soil of flooded rice microcosms

    Energy Technology Data Exchange (ETDEWEB)

    Grosskopf, R.; Stubner, S.; Liesack, W. [Max-Planck-Inst. fuer Terrestrische Mikrobiologie, Marburg (Germany)

    1998-12-01

    Because excised, washed roots of rice (Oryza sativa) immediately produce CH{sub 4} when they are incubated under anoxic conditions, the authors employed a culture-independent molecular approach to identify the methanogenic microbial community present on roots of rice plants. Archaeal small-subunit rRNA-encoding genes were amplified directly from total root DNA by PCR and then cloned. Thirty-two archaeal rice root (ARR) gene clones were randomly selected, and the amplified primary structures of ca. 750 nucleotide sequence positions were compared. Only 10 of the environmental sequences were affiliated with known methanogens; 5 were affiliated with Methanosarcina spp., and 5 were affiliated with Methanobacterium spp. The remaining 22 ARR gene clones formed four distinct lineages (rice clusters I through IV) which were not closely related to any known cultured member of the Archaea. Rice clusters I and II formed distinct clades within the phylogenetic radiation of the orders Methanosarcinales and Methanomicrobiales. Rice cluster I was novel, and rice cluster II was closely affiliated with environmental sequences obtained from bog peak in northern England. Rice cluster III occurred on the same branch as Thermoplasma acidophilum and marine group II but was only distantly related to these taxa. Rice cluster IV was a deep-branching crenarchaeotal assemblage that was closely related to clone pGrfC26, an environmental sequence recovered from a temperate marsh environment. The use of a domain-specific oligonucleotide probe in a fluorescent in situ hybridization analysis revealed that viable members of the Archaea were present on the surface of rice roots. In addition, the authors describe a novel euryarchaeotal main line of descent, designated rice cluster V, which was detected in anoxic rice paddy soil. These results indicate that there is an astonishing richness of archaeal diversity present on rice roots and in the surrounding paddy soil.

  1. Comparative genomic analyses of nickel, cobalt and vitamin B12 utilization

    Science.gov (United States)

    Zhang, Yan; Rodionov, Dmitry A; Gelfand, Mikhail S; Gladyshev, Vadim N

    2009-01-01

    Background Nickel (Ni) and cobalt (Co) are trace elements required for a variety of biological processes. Ni is directly coordinated by proteins, whereas Co is mainly used as a component of vitamin B12. Although a number of Ni and Co-dependent enzymes have been characterized, systematic evolutionary analyses of utilization of these metals are limited. Results We carried out comparative genomic analyses to examine occurrence and evolutionary dynamics of the use of Ni and Co at the level of (i) transport systems, and (ii) metalloproteomes. Our data show that both metals are widely used in bacteria and archaea. Cbi/NikMNQO is the most common prokaryotic Ni/Co transporter, while Ni-dependent urease and Ni-Fe hydrogenase, and B12-dependent methionine synthase (MetH), ribonucleotide reductase and methylmalonyl-CoA mutase are the most widespread metalloproteins for Ni and Co, respectively. Occurrence of other metalloenzymes showed a mosaic distribution and a new B12-dependent protein family was predicted. Deltaproteobacteria and Methanosarcina generally have larger Ni- and Co-dependent proteomes. On the other hand, utilization of these two metals is limited in eukaryotes, and very few of these organisms utilize both of them. The Ni-utilizing eukaryotes are mostly fungi (except saccharomycotina) and plants, whereas most B12-utilizing organisms are animals. The NiCoT transporter family is the most widespread eukaryotic Ni transporter, and eukaryotic urease and MetH are the most common Ni- and B12-dependent enzymes, respectively. Finally, investigation of environmental and other conditions and identity of organisms that show dependence on Ni or Co revealed that host-associated organisms (particularly obligate intracellular parasites and endosymbionts) have a tendency for loss of Ni/Co utilization. Conclusion Our data provide information on the evolutionary dynamics of Ni and Co utilization and highlight widespread use of these metals in the three domains of life, yet only a

  2. Microbial Community Response to Simulated Petroleum Seepage in Caspian Sea Sediments

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    Katrin Knittel

    2017-04-01

    Full Text Available Anaerobic microbial hydrocarbon degradation is a major biogeochemical process at marine seeps. Here we studied the response of the microbial community to petroleum seepage simulated for 190 days in a sediment core from the Caspian Sea using a sediment-oil-flow-through (SOFT system. Untreated (without simulated petroleum seepage and SOFT sediment microbial communities shared 43% bacterial genus-level 16S rRNA-based operational taxonomic units (OTU0.945 but shared only 23% archaeal OTU0.945. The community differed significantly between sediment layers. The detection of fourfold higher deltaproteobacterial cell numbers in SOFT than in untreated sediment at depths characterized by highest sulfate reduction rates and strongest decrease of gaseous and mid-chain alkane concentrations indicated a specific response of hydrocarbon-degrading Deltaproteobacteria. Based on an increase in specific CARD-FISH cell numbers, we suggest the following groups of sulfate-reducing bacteria to be likely responsible for the observed decrease in aliphatic and aromatic hydrocarbon concentration in SOFT sediments: clade SCA1 for propane and butane degradation, clade LCA2 for mid- to long-chain alkane degradation, clade Cyhx for cycloalkanes, pentane and hexane degradation, and relatives of Desulfobacula for toluene degradation. Highest numbers of archaea of the genus Methanosarcina were found in the methanogenic zone of the SOFT core where we detected preferential degradation of long-chain hydrocarbons. Sequencing of masD, a marker gene for alkane degradation encoding (1-methylalkylsuccinate synthase, revealed a low diversity in SOFT sediment with two abundant species-level MasD OTU0.96.

  3. The Effects of Perchlorates on the Permafrost Methanogens: Implication for Autotrophic Life on Mars

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    Viktoria Shcherbakova

    2015-09-01

    Full Text Available The terrestrial permafrost represents a range of possible cryogenic extraterrestrial ecosystems on Earth-like planets without obvious surface ice, such as Mars. The autotrophic and chemolithotrophic psychrotolerant methanogens are more likely than aerobes to function as a model for life forms that may exist in frozen subsurface environments on Mars, which has no free oxygen, inaccessible organic matter, and extremely low amounts of unfrozen water. Our research on the genesis of methane, its content and distribution in permafrost horizons of different ages and origin demonstrated the presence of methane in permanently frozen fine-grained sediments. Earlier, we isolated and described four strains of methanogenic archaea of Methanobacterium and Methanosarcina genera from samples of Pliocene and Holocene permafrost from Eastern Siberia. In this paper we study the effect of sodium and magnesium perchlorates on growth of permafrost and nonpermafrost methanogens, and present evidence that permafrost hydogenotrophic methanogens are more resistant to the chaotropic agent found in Martian soil. In this paper we study the effect of sodium and magnesium perchlorates on the growth of permafrost and nonpermafrost methanogens, and present evidence that permafrost hydogenotrophic methanogens are more resistant to the chaotropic agent found in Martian soil. Furthermore, as shown in the studies strain M2T M. arcticum, probably can use perchlorate anion as an electron acceptor in anaerobic methane oxidation. Earth’s subzero subsurface environments are the best approximation of environments on Mars, which is most likely to harbor methanogens; thus, a biochemical understanding of these pathways is expected to provide a basis for designing experiments to detect autotrophic methane-producing life forms on Mars.

  4. Comprehensive microbial analysis of combined mesophilic anaerobic-thermophilic aerobic process treating high-strength food wastewater.

    Science.gov (United States)

    Jang, Hyun Min; Ha, Jeong Hyub; Park, Jong Moon; Kim, Mi-Sun; Sommer, Sven G

    2015-04-15

    A combined mesophilic anaerobic-thermophilic aerobic process was used to treat high-strength food wastewater in this study. During the experimental period, most of solid residue from the mesophilic anaerobic reactor (R1) was separated by centrifugation and introduced into the thermophilic aerobic reactor (R2) for further digestion. Then, thermophilic aerobically-digested sludge was reintroduced into R1 to enhance reactor performance. The combined process was operated with two different Runs: Run I with hydraulic retention time (HRT) = 40 d (corresponding OLR = 3.5 kg COD/m(3) d) and Run II with HRT = 20 d (corresponding OLR = 7 kg COD/m(3)). For a comparison, a single-stage mesophilic anaerobic reactor (R3) was operated concurrently with same OLRs and HRTs as the combined process. During the overall digestion, all reactors showed high stability without pH control. The combined process demonstrated significantly higher organic matter removal efficiencies (over 90%) of TS, VS and COD and methane production than did R3. Quantitative real-time PCR (qPCR) results indicated that higher populations of both bacteria and archaea were maintained in R1 than in R3. Pyrosequencing analysis revealed relatively high abundance of phylum Actinobacteria in both R1 and R2, and a predominance of phyla Synergistetes and Firmicutes in R3 during Run II. Furthermore, R1 and R2 shared genera (Prevotella, Aminobacterium, Geobacillus and Unclassified Actinobacteria), which suggests synergy between mesophilic anaerobic digestion and thermophilic aerobic digestion. For archaea, in R1 methanogenic archaea shifted from genus Methanosaeta to Methanosarcina, whereas genera Methanosaeta, Methanobacterium and Methanoculleus were predominant in R3. The results demonstrated dynamics of key microbial populations that were highly consistent with an enhanced reactor performance of the combined process.

  5. Structural basis for the site-specific incorporation of lysine derivatives into proteins.

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    Veronika Flügel

    Full Text Available Posttranslational modifications (PTMs of proteins determine their structure-function relationships, interaction partners, as well as their fate in the cell and are crucial for many cellular key processes. For instance chromatin structure and hence gene expression is epigenetically regulated by acetylation or methylation of lysine residues in histones, a phenomenon known as the 'histone code'. Recently it was shown that these lysine residues can furthermore be malonylated, succinylated, butyrylated, propionylated and crotonylated, resulting in significant alteration of gene expression patterns. However the functional implications of these PTMs, which only differ marginally in their chemical structure, is not yet understood. Therefore generation of proteins containing these modified amino acids site specifically is an important tool. In the last decade methods for the translational incorporation of non-natural amino acids using orthogonal aminoacyl-tRNA synthetase (aaRS:tRNAaaCUA pairs were developed. A number of studies show that aaRS can be evolved to use non-natural amino acids and expand the genetic code. Nevertheless the wild type pyrrolysyl-tRNA synthetase (PylRS from Methanosarcina mazei readily accepts a number of lysine derivatives as substrates. This enzyme can further be engineered by mutagenesis to utilize a range of non-natural amino acids. Here we present structural data on the wild type enzyme in complex with adenylated ε-N-alkynyl-, ε-N-butyryl-, ε-N-crotonyl- and ε-N-propionyl-lysine providing insights into the plasticity of the PylRS active site. This shows that given certain key features in the non-natural amino acid to be incorporated, directed evolution of this enzyme is not necessary for substrate tolerance.

  6. Metabolic engineering for the high-yield production of isoprenoid-based C₅ alcohols in E. coli.

    Science.gov (United States)

    George, Kevin W; Thompson, Mitchell G; Kang, Aram; Baidoo, Edward; Wang, George; Chan, Leanne Jade G; Adams, Paul D; Petzold, Christopher J; Keasling, Jay D; Lee, Taek Soon

    2015-06-08

    Branched five carbon (C5) alcohols are attractive targets for microbial production due to their desirable fuel properties and importance as platform chemicals. In this study, we engineered a heterologous isoprenoid pathway in E. coli for the high-yield production of 3-methyl-3-buten-1-ol, 3-methyl-2-buten-1-ol, and 3-methyl-1-butanol, three C5 alcohols that serve as potential biofuels. We first constructed a pathway for 3-methyl-3-buten-1-ol, where metabolite profiling identified NudB, a promiscuous phosphatase, as a likely pathway bottleneck. We achieved a 60% increase in the yield of 3-methyl-3-buten-1-ol by engineering the Shine-Dalgarno sequence of nudB, which increased protein levels by 9-fold and reduced isopentenyl diphosphate (IPP) accumulation by 4-fold. To further optimize the pathway, we adjusted mevalonate kinase (MK) expression and investigated MK enzymes from alternative microbes such as Methanosarcina mazei. Next, we expressed a fusion protein of IPP isomerase and the phosphatase (Idi1~NudB) along with a reductase (NemA) to diversify production to 3-methyl-2-buten-1-ol and 3-methyl-1-butanol. Finally, we used an oleyl alcohol overlay to improve alcohol recovery, achieving final titers of 2.23 g/L of 3-methyl-3-buten-1-ol (~70% of pathway-dependent theoretical yield), 150 mg/L of 3-methyl-2-buten-1-ol, and 300 mg/L of 3-methyl-1-butanol.

  7. Towards Reassignment of the Methionine Codon AUG to Two Different Noncanonical Amino Acids in Bacterial Translation

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    Alessandro De Simone

    2016-06-01

    Full Text Available Genetic encoding of noncanonical amino acids (ncAAs through sense codon reassignment is an efficient tool for expanding the chemical functionality of proteins. Incorporation of multiple ncAAs, however, is particularly challenging. This work describes the first attempts to reassign the sense methionine (Met codon AUG to two different ncAAs in bacterial protein translation. Escherichia coli methionyl-tRNA synthetase (MetRS charges two tRNAs with Met: tRNAfMet initiates protein synthesis (starting AUG codon, whereas elongator tRNAMet participates in protein elongation (internal AUG codon(s. Preliminary in vitro experiments show that these tRNAs can be charged with the Met analogues azidohomoalanine (Aha and ethionine (Eth by exploiting the different substrate specificities of EcMetRS and the heterologous MetRS / tRNAMet pair from the archaeon Sulfolobus acidocaldarius, respectively. Here, we explored whether this configuration would allow a differential decoding during in vivo protein initiation and elongation. First, we eliminated the elongator tRNAMet from a methionine auxotrophic E. coli strain, which was then equipped with a rescue plasmid harboring the heterologous pair. Although the imported pair was not fully orthogonal, it was possible to incorporate preferentially Eth at internal AUG codons in a model protein, suggesting that in vivo AUG codon reassignment is possible. To achieve full orthogonality during elongation, we imported the known orthogonal pair of Methanosarcina mazei pyrrolysyl-tRNA synthetase (PylRS / tRNAPyl and devised a genetic selection system based on the suppression of an amber stop codon in an important glycolytic gene, pfkA, which restores enzyme functionality and normal cellular growth. Using an evolved PylRS able to accept Met analogues, it should be possible to reassign the AUG codon to two different ncAAs by using directed evolution. This work is licensed under a Creative Commons Attribution 4.0 International

  8. Characterization of Sac10a, a hyperthermophile DNA-binding protein from Sulfolobus acidocaldarius.

    Science.gov (United States)

    Edmondson, Stephen P; Kahsai, Mebrahtu A; Gupta, Ramesh; Shriver, John W

    2004-10-19

    Sac10a is a member of a group of basic DNA-binding proteins thought to be important in chromatin structure and regulation in the archaeon Sulfolobus. We describe here the isolation, gene identification, and biophysical characterization of native Sac10a. The protein exists as a 23.8 kDa homodimer at pH 7 and unfolds with a T degrees of 122 degrees C. Dissociation of the dimer into folded globular subunits is promoted by decreased pH and salt concentration. Thermal unfolding of the monomeric subunits occurred with two transitions, indicating two independent domains. The dimer demonstrated a high affinity for duplex poly(dAdT) with a K(D) of 5 x 10(-)(10) M and a site size of 17 bp (in 0.15 M KCl, pH 7), with only weak binding (K(D) > 5 x 10(-)(6) M) to poly(dA)-poly(dT), poly(dGdC), poly(dG)-poly(dC), and Escherichia coli DNA under similar conditions. Binding to poly(dAdT) resulted in distortions in the DNA duplex that were consistent with overwinding as indicated by inversion of the CD spectrum of the DNA. The monomeric subunits are predicted to adopt a winged helix DNA-binding motif which dimerizes through formation of a two-stranded coiled coil involving an extended C-terminal helix with more than four heptad repeats (about 45 A in length). This is the first example of the conserved archaeal transcription regulator domain COG3432 to be characterized. Sequences for homologous proteins containing both COG3432 and predicted coiled coil domains occur in the genomes of both crenarchaeota (Sulfolobus, Pyrobaculum, Aeropyrum) and euryarchaeota (Methanosarcina, Methanococcus, Archaeoglobus, Thermoplasma), with multiple genes in some species. Sac10a shows no sequence similarity to the other Sulfolobus chromatin proteins Sac7d, Sac8, Sso10b2, and Alba.

  9. Effects of feeding time and organic loading in an anaerobic sequencing batch biofilm reactor (ASBBR) treating diluted whey.

    Science.gov (United States)

    Damasceno, Leonardo H S; Rodrigues, José A D; Ratusznei, Suzana M; Zaiat, Marcelo; Foresti, Eugênio

    2007-12-01

    An investigation was carried out on the performance of an anaerobic sequencing batch biofilm reactor (ASBBR) treating diluted cheese whey when submitted to different feed strategies and volumetric organic loads (VOL). Polyurethane foam cubes were used as support for biomass immobilization and stirring was provided by helix impellers. The reactor with a working volume of 3 L treated 2 L of wastewater in 8-h cycles at 500 rpm and 30 degrees C. The organic loads applied were 2, 4, 8 and 12 g COD L(-1) d(-1), obtained by increasing the feed concentration. Alkalinity was supplemented at a ratio of 50% NaHCO(3)/COD. For each organic load applied three feed strategies were tested: (a) batch operation with 8-h cycle; (b) 2-h fed-batch operation followed by 6-h batch; and (c) 4-h fed-batch followed by 4-h batch. The 2-h fed-batch operation followed by 6-h batch presented the best results for the organic loads of 2 and 4 g COD L(-1) d(-1), whereas the 4-h fed-batch operation followed by 4-h batch presented results slightly inferior for the same organic loads and the best results at organic loads of 8 and 12 g COD L(-1) d(-1). The concentration of total volatile acids varied with fill time. For the higher fill times maximum concentrations were obtained at the end of the cycle. Moreover, no significant difference was detected in the maximum concentration of total volatile acids for any of the investigated conditions. However, the maximum values of propionic acid tended to decrease with increasing fill time considering the same organic load. Microbiological analyses revealed the presence of Methanosaeta-like structures and methanogenic hydrogenotrophic-like fluorescent bacilli. No Methanosarcina-like structures were observed in the samples.

  10. Quantitative microbiological analysis of bacterial community shifts in a high-rate anaerobic bioreactor treating sulfite evaporator condensate.

    Science.gov (United States)

    Ney, U; Macario, A J; Conway de Macario, E; Aivasidis, A; Schoberth, S M; Sahm, H

    1990-08-01

    The bacterial population of a high-rate, anaerobic, fixed-bed loop reactor treating sulfite evaporator condensate from the pulp industry was studied over a 14-month period. This period was divided into seven cycles that included a startup at the beginning of each cycle. Some 82% of the total biomass was immobilized on and between the porous glass rings filling the reactor. The range of the total number of microorganisms in these biofilms was 2 x 10 to 7 x 10 cells per ml. Enumeration and characterization by microbiological methods and by phase-contrast, epifluorescence, and electron microscopy showed that the samples consisted mainly of the following methanogens: a Methanobacterium sp., a Methanosarcina sp., a Methanobrevibacter sp., and a Methanothrix sp., as well as furfural-degrading sulfate-reducing bacteria resembling Desulfovibrio furfuralis. Viable counts of hydrogenotrophic methanogens were relatively stable (mostly within the range of 3.2 x 10 to 7.5 x 10 cells per ml), but Methanobrevibacter cells increased from fixed bed into a second reactor vessel. Acetotrophic methanogens reached their highest numbers of 1.3 x 10 to 2.6 x 10 cells per ml in the last fermentation cycles. They showed a morphological shift from sarcinalike packets in early samples to single coccoid forms in later phases of the fermentation. Furfural-degrading sulfate reducers reached counts of 1 x 10 to 5.8 x 10 cells per ml. The distribution of the chief metabolic groups between free fluid and biofilms was analyzed in the fifth fermentation cycle: 4.5 times more furfural degraders were found in the free fluid than in the biofilms. In contrast, 5.8 times more acetotrophic and 16.6 times more hydrogenotrophic methanogens were found in the biofilms than in the free liquid. The data concerning time shifts of morphotypes among the trophic groups of methanogens corroborated the trends observed by using immunological assays on the same samples.

  11. Changing Feeding Regimes To Demonstrate Flexible Biogas Production: Effects on Process Performance, Microbial Community Structure, and Methanogenesis Pathways

    Science.gov (United States)

    Mulat, Daniel Girma; Jacobi, H. Fabian; Feilberg, Anders; Adamsen, Anders Peter S.; Richnow, Hans-Hermann

    2015-01-01

    Flexible biogas production that adapts biogas output to energy demand can be regulated by changing feeding regimes. In this study, the effect of changes in feeding intervals on process performance, microbial community structure, and the methanogenesis pathway was investigated. Three different feeding regimes (once daily, every second day, and every 2 h) at the same organic loading rate were studied in continuously stirred tank reactors treating distiller's dried grains with solubles. A larger amount of biogas was produced after feeding in the reactors fed less frequently (once per day and every second day), whereas the amount remained constant in the reactor fed more frequently (every 2 h), indicating the suitability of the former for the flexible production of biogas. Compared to the conventional more frequent feeding regimes, a methane yield that was up to 14% higher and an improved stability of the process against organic overloading were achieved by employing less frequent feeding regimes. The community structures of bacteria and methanogenic archaea were monitored by terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA and mcrA genes, respectively. The results showed that the composition of the bacterial community varied under the different feeding regimes, and the observed T-RFLP patterns were best explained by the differences in the total ammonia nitrogen concentrations, H2 levels, and pH values. However, the methanogenic community remained stable under all feeding regimes, with the dominance of the Methanosarcina genus followed by that of the Methanobacterium genus. Stable isotope analysis showed that the average amount of methane produced during each feeding event by acetoclastic and hydrogenotrophic methanogenesis was not influenced by the three different feeding regimes. PMID:26497462

  12. Activation of methanogenesis in arid biological soil crusts despite the presence of oxygen.

    Directory of Open Access Journals (Sweden)

    Roey Angel

    Full Text Available Methanogenesis is traditionally thought to occur only in highly reduced, anoxic environments. Wetland and rice field soils are well known sources for atmospheric methane, while aerated soils are considered sinks. Although methanogens have been detected in low numbers in some aerated, and even in desert soils, it remains unclear whether they are active under natural oxic conditions, such as in biological soil crusts (BSCs of arid regions. To answer this question we carried out a factorial experiment using microcosms under simulated natural conditions. The BSC on top of an arid soil was incubated under moist conditions in all possible combinations of flooding and drainage, light and dark, air and nitrogen headspace. In the light, oxygen was produced by photosynthesis. Methane production was detected in all microcosms, but rates were much lower when oxygen was present. In addition, the δ(13C of the methane differed between the oxic/oxygenic and anoxic microcosms. While under anoxic conditions methane was mainly produced from acetate, it was almost entirely produced from H(2/CO(2 under oxic/oxygenic conditions. Only two genera of methanogens were identified in the BSC-Methanosarcina and Methanocella; their abundance and activity in transcribing the mcrA gene (coding for methyl-CoM reductase was higher under anoxic than oxic/oxygenic conditions, respectively. Both methanogens also actively transcribed the oxygen detoxifying gene catalase. Since methanotrophs were not detectable in the BSC, all the methane produced was released into the atmosphere. Our findings point to a formerly unknown participation of desert soils in the global methane cycle.

  13. Performances of microbial fuel cells fed with rejected wastewater from BioCH4 and BioH2 processes treating molasses wastewater.

    Science.gov (United States)

    Lee, Yun-Yeong; Kim, Tae G; Cho, Kyung-Suk

    2016-01-01

    An integrated process involving conventional anaerobic digestion and microbial fuel cells (MFCs) has attracted attention recently to produce sustainable energy and to treat wastewater efficiently. To evaluate the possibility of CH4-producing process (BioCH4)-MFC or H2-producing process (BioH2)-MFC integrating systems, the MFC performances were investigated using rejected wastewater from a BioCH4 reactor (RWCH4) or BioH2 reactor (RWH2) treating molasses wastewater. When RWCH4 or RWH2 was fed into a single-chamber MFC reactor (designated as AC-MFCCH4 and AC-MFCH2, respectively) at different hydraulic retention times (HRT) of 1-7 d, both MFC systems showed maximum electricity production efficiencies at a HRT of 3 d. In the AC-MFCCH4 reactor, the average current density and average power density were 60.5 mA·m(-2) and 8.8 mW·m(-2), respectively. The AC-MFCH2 reactor generated an average current density of 71.4 mA·m(-2) and an average power density of 12.0 mW·m(-2). The COD removal rates were 45.7% in the AC-MFCCH4 reactor and 90.3% in the AC-MFCH2 reactor. There were no significant differences of the eubacterial community structures between the MFC systems, where Proteobacteria was remarkably dominant in both MFC systems. However, the archaeal community structures were significantly different where Methanothrix (89.3%) was remarkably dominant in the AC-MFCCH4 system, while Methanothrix (52.5%) and Methanosarcina (33.5%) were abundant in the AC-MFCH2 system. These findings demonstrate that the utilization of MFCs after the BioCH4 or BioH2 process is advantageous for energy recovery as well as COD removal from molasses wastewater.

  14. Laboratory-scale bioaugmentation relieves acetate accumulation and stimulates methane production in stalled anaerobic digesters.

    Science.gov (United States)

    Town, Jennifer R; Dumonceaux, Tim J

    2016-01-01

    An imbalance between acidogenic and methanogenic organisms during anaerobic digestion can result in increased accumulation of volatile fatty acids, decreased reactor pH, and inhibition of methane-producing Archaea. Most commonly the result of organic input overload or poor inoculum selection, these microbiological and biochemical changes severely hamper reactor performance, and there are a few tools available to facilitate reactor recovery. A small, stable consortium capable of catabolizing acetate and producing methane was propagated in vitro and evaluated as a potential bioaugmentation tool for stimulating methanogenesis in acidified reactors. Replicate laboratory-scale batch digesters were seeded with a combination of bioethanol stillage waste and a dairy manure inoculum previously observed to result in high volatile fatty acid accumulation and reactor failure. Experimental reactors were then amended with the acetoclastic consortium, and control reactors were amended with sterile culture media. Within 7 days, bioaugmented reactors had significantly reduced acetate accumulation and the proportion of methane in the biogas increased from 0.2 ± 0 to 74.4 ± 9.9 % while control reactors showed no significant reduction in acetate accumulation or increase in methane production. Organisms from the consortium were enumerated using specific quantitative PCR assays to evaluate their growth in the experimental reactors. While the abundance of hydrogenotrophic microorganisms remained stable during the recovery period, an acetoclastic methanogen phylogenetically similar to Methanosarcina sp. increased more than 100-fold and is hypothesized to be the primary contributor to reactor recovery. Genomic sequencing of this organism revealed genes related to the production of methane from acetate, hydrogen, and methanol.

  15. Bromeliad catchments as habitats for methanogenesis in tropical rainforest canopies

    Directory of Open Access Journals (Sweden)

    Shana K. Goffredi

    2011-12-01

    Full Text Available Tropical epiphytic plants within the family Bromeliaceae are unusual in that they possess foliage capable of retaining water and impounded material. This creates an acidic (pH 3.5-6.5 and anaerobic (< 1 ppm O2 environment suspended in the canopy. Results from a Costa Rican rainforest show that most bromeliads (n = 75/86 greater than ~20 cm in plant height or ~4-5 cm tank depth, showed presence of methanogens within the lower anoxic horizon of the tank. Archaea were dominated by methanogens (77-90% of recovered ribotypes and community structure, although variable, was generally comprised of a single type, closely related to either hydrogenotrophic Methanoregula or Methanocella, a specific clade of aceticlastic Methanosaeta, or Methanosarcina. Juvenile bromeliads, or those species, such as Guzmania, with shallow tanks, generally did not possess methanogens, as assayed by PCR specific for methanogen 16S rRNA genes, nor did artificial catchments (~ 100 ml volume, in place 6-12 months prior to sample collection. Methanogens were not detected in soil (n = 20, except in one case, in which the dominant ribotype was different from nearby bromeliads. Recovery of methyl coenzyme M reductase genes supported the occurrence of hydrogenotrophic and aceticlastic methanogens within bromeliad tanks, as well as the trend, via QPCR analysis of mcrA, of increased methanogenic capacity with increased plant height. Methane production rates of up to 300 nmol CH4 ml tank water -1 day-1 were measured in microcosm experiments. These results suggest that bromeliad-associated archaeal communities may play an important role in the cycling of carbon in neotropical forests.

  16. Wood ash amendment to biogas reactors as an alternative to landfilling? A preliminary study on changes in process chemistry and biology.

    Science.gov (United States)

    Podmirseg, Sabine M; Seewald, Martin S A; Knapp, Brigitte A; Bouzid, Ourdia; Biderre-Petit, Corinne; Peyret, Pierre; Insam, Heribert

    2013-08-01

    Wood ash addition to biogas plants represents an alternative to commonly used landfilling by improving the reactor performance, raising the pH and alleviating potential limits of trace elements. This study is the first on the effects of wood ash on reactor conditions and microbial communities in cattle slurry-based biogas reactors. General process parameters [temperature, pH, electrical conductivity, ammonia, volatile fatty acids, carbon/nitrogen (C/N), total solids (TS), volatile solids, and gas quantity and quality] were monitored along with molecular analyses of methanogens by polymerase chain reaction- denaturing gradient gel electrophoresis and modern microarrays (archaea and bacteria). A prompt pH rise was observed, as was an increase in C/N ratio and volatile fatty acids. Biogas production was inhibited, but recovered to even higher production rates and methane concentration after single amendment. High sulphur levels in the wood ash generated hydrogen sulphide and potentially hampered methanogenesis. Methanosarcina was the most dominant methanogen in all reactors; however, diversity was higher in ash-amended reactors. Bacterial groups like Firmicutes, Proteobacteria and Acidobacteria were favoured, which could improve the hydrolytic efficiency of the reactors. We recommend constant monitoring of the chemical composition of the used wood ash and suggest that ash amendment is adequate if added to the substrate at a rate low enough to allow adaptation of the microbiota (e.g. 0.25 g g(-1) TS). It could further help to enrich digestate with important nutrients, for example phosphorus, calcium and magnesium, but further experiments are required for the evaluation of wood ash concentrations that are tolerable for anaerobic digestion.

  17. Effect of organic loading on the microbiota in a temperature-phased anaerobic digestion (TPAD) system co-digesting dairy manure and waste whey.

    Science.gov (United States)

    Li, Yueh-Fen; Abraham, Christopher; Nelson, Michael C; Chen, Po-Hsu; Graf, Joerg; Yu, Zhongtang

    2015-10-01

    Temperature-phased anaerobic digestion (TPAD) has gained increasing attention because it provides the flexibility to operate digesters under conditions that enhance overall digester performance. However, research on impact of organic overloading rate (OLR) to microbiota of TPAD systems was limited. In this study, we investigated the composition and successions of the microbiota in both the thermophilic and the mesophilic digesters of a laboratory-scale TPAD system co-digesting dairy manure and waste whey before and during organic overloading. The thermophilic and the mesophilic digesters were operated at 50 and 35 °C, respectively, with a hydraulic retention time (HRT) of 10 days for each digester. High OLR (dairy manure with 5 % total solid and waste whey of ≥60.4 g chemical oxygen demand (COD)/l/day) resulted in decrease in pH and in biogas production and accumulation of volatile fatty acids (VFAs) in the thermophilic digester, while the mesophilic digester remained unchanged except a transient increase in biogas production. Both denaturant gradient gel electrophoresis (DGGE) and Illumina sequencing of 16S ribosomal RNA (rRNA) gene amplicons showed dramatic change in microbiota composition and profound successions of both bacterial and methanogenic communities. During the overloading, Thermotogae was replaced by Proteobacteria, while Methanobrevibacter and archaeon classified as WCHD3-02 grew in predominance at the expense of Methanoculleus in the thermophilic digester, whereas Methanosarcina dominated the methanogenic community, while Methanobacterium and Methanobrevibacter became less predominant in the mesophilic digester. Canonical correspondence analysis (CCA) revealed that digester temperature and pH were the most influential environmental factors that explained much of the variations of the microbiota in this TPAD system when it was overloaded.

  18. Biogas production and methanogenic archaeal community in mesophilic and thermophilic anaerobic co-digestion processes.

    Science.gov (United States)

    Yu, D; Kurola, J M; Lähde, K; Kymäläinen, M; Sinkkonen, A; Romantschuk, M

    2014-10-01

    Over 258 Mt of solid waste are generated annually in Europe, a large fraction of which is biowaste. Sewage sludge is another major waste fraction. In this study, biowaste and sewage sludge were co-digested in an anaerobic digestion reactor (30% and 70% of total wet weight, respectively). The purpose was to investigate the biogas production and methanogenic archaeal community composition in the anaerobic digestion reactor under meso- (35-37 °C) and thermophilic (55-57 °C) processes and an increasing organic loading rate (OLR, 1-10 kg VS m(-3) d(-1)), and also to find a feasible compromise between waste treatment capacity and biogas production without causing process instability. In summary, more biogas was produced with all OLRs by the thermophilic process. Both processes showed a limited diversity of the methanogenic archaeal community which was dominated by Methanobacteriales and Methanosarcinales (e.g. Methanosarcina) in both meso- and thermophilic processes. Methanothermobacter was detected as an additional dominant genus in the thermophilic process. In addition to operating temperatures, the OLRs, the acetate concentration, and the presence of key substrates like propionate also affected the methanogenic archaeal community composition. A bacterial cell count 6.25 times higher than archaeal cell count was observed throughout the thermophilic process, while the cell count ratio varied between 0.2 and 8.5 in the mesophilic process. This suggests that the thermophilic process is more stable, but also that the relative abundance between bacteria and archaea can vary without seriously affecting biogas production.

  19. Structural Properties of Alternate Monatomic Layered [Fe/Co]n Epitaxial Films on MgO Substrate

    Science.gov (United States)

    Chu, In Chang; Saki, Yoshinobu; Kawasaki, Shohei; Doi, Masaaki; Sahashi, Masashi

    2008-06-01

    Body-centered-cubic (bcc) Fe50Co50 material is reported to show a high bulk spin scattering coefficient on current perpendicular to plane-giant magneto-resistance (CPP-GMR) system. But the origin of that phenomenon does not make sure yet. We prepared artificially alternate monatomic layered (AML) [Fe/Co] 41 MLs epitaxial films (Ts: 75, 250 °C) by monatomic deposition method and investigated the topology of AML [Fe/Co]n epitaxial films on MgO substrate with different orientation (001), (011) by the scanning tunnel microscopy (STM) and reflection high energy electron diffraction (RHEED), which we could confirm Frank-van der Merwe (FM) growth mode for AML [Fe/Co]n on MgO(001) and Volmer-Weber (VW) growth mode for that on Mg(011). The roughness of surface, Ra (0.20 nm) of AML [Fe/Co] 41 MLs epitaxial film grown at 75 °C on MgO(001) is smaller than that (0.46 nm) of AML [Fe/Co] grown at 250 °C on MgO(001), which has the large terraces of over 50 nm (Ra: 0.17 nm), even though there are some valleys between large terraces. Moreover we confirmed the structural properties of trilayered epitaxial films with AML [Fe/Co]n (Ra: 0.18 nm) and Fe50Co50 alloy epitaxial film on Au electrode by RHEED before confirming the characteristics of CPP-GMR devices.

  20. Inhibitory Effect of Coumarin on Syntrophic Fatty Acid-Oxidizing and Methanogenic Cultures and Biogas Reactor Microbiomes.

    Science.gov (United States)

    Popp, Denny; Plugge, Caroline M; Kleinsteuber, Sabine; Harms, Hauke; Sträuber, Heike

    2017-07-01

    Coumarins are widely found in plants as natural constituents having antimicrobial activity. When considering plants that are rich in coumarins for biogas production, adverse effects on microorganisms driving the anaerobic digestion process are expected. Furthermore, coumarin derivatives, like warfarin, which are used as anticoagulating medicines, are found in wastewater, affecting its treatment. Coumarin, the structure common to all coumarins, inhibits the anaerobic digestion process. However, the details of this inhibition are still elusive. Here, we studied the impact of coumarin on acetogenesis and methanogenesis. First, coumarin was applied at four concentrations between 0.25 and 1 g · liter(-1) to pure cultures of the methanogens Methanosarcina barkeri and Methanospirillum hungatei, which resulted in up to 25% less methane production. Acetate production of syntrophic propionate- and butyrate-degrading cultures of Syntrophobacter fumaroxidans and Syntrophomonas wolfei was inhibited by 72% at a coumarin concentration of 1 g · liter(-1) Coumarin also inhibited acetogenesis and acetoclastic methanogenesis in a complex biogas reactor microbiome. When a coumarin-adapted microbiome was used, acetogenesis and methanogenesis were not inhibited. According to amplicon sequencing of bacterial 16S rRNA genes and mcrA genes, the communities of the two microbiomes were similar, although Methanoculleus was more abundant and Methanobacterium less abundant in the coumarin-adapted than in the nonadapted microbiome. Our results suggest that well-dosed feeding with coumarin-rich feedstocks to full-scale biogas reactors while keeping the coumarin concentrations below 0.5 g · liter(-1) will allow adaptation to coumarins by structural and functional community reorganization and coumarin degradation.IMPORTANCE Coumarins from natural and anthropogenic sources have an inhibitory impact on the anaerobic digestion process. Here, we studied in detail the adverse effects of the model

  1. Final Report Systems Level Analysis of the Function and Adaptive Responses of Methanogenic Consortia

    Energy Technology Data Exchange (ETDEWEB)

    Lovley, Derek R. [Univ. of Massachusetts, Amherst, MA (United States)

    2015-03-09

    The purpose of this research was to determine whether the syntrophic microbial associations that are central to the functioning of methane-producing terrestrial wetlands can be predictively modeled with coupled multi-species genome-scale metabolic models. Such models are important because methane is an important greenhouse gas and there is a need to predictively model how the methane-producing microbial communities will respond to environmental perturbations, such as global climate change. The research discovered that the most prodigious methane-producing microorganisms on earth participate in a previously unrecognized form of energy exchange. The methane-producers Methanosaeta and Methanosarcina forge biological electrical connections with other microbes in order to obtain electrons to reduce carbon dioxide to methane. This direct interspecies electron transfer (DIET) was demonstrated in complex microbial communities as well as in defined co-cultures. For example, metatranscriptomic analysis of gene expression in both natural communities and defined co-cultures demonstrated that Methanosaeta species highly expressed genes for the enzymes for the reduction of carbon dioxide to methane. Furthermore, Methanosaeta’s electron-donating partners highly expressed genes for the biological electrical connections known as microbial nanowires. A series of studies involving transcriptomics, genome resequencing, and analysis of the metabolism of a series of strains with targeted gene deletions, further elucidated the mechanisms and energetics of DIET in methane-producing co-cultures, as well as in a co-culture of Geobacter metallireducens and Geobacter sulfurreducens, which provided a system for studying DIET with two genetically tractable partners. Genome-scale modeling of DIET in the G. metallireducens/G. sulfurreducens co-culture suggested that DIET provides more energy to the electron-donating partner that electron exchange via interspecies hydrogen transfer, but that the

  2. Gene encoding γ-carbonic anhydrase is cotranscribed with argC and induced in response to stationary phase and high CO2 in Azospirillum brasilense Sp7

    Directory of Open Access Journals (Sweden)

    Mishra Mukti N

    2010-07-01

    Full Text Available Abstract Background Carbonic anhydrase (CA is a ubiquitous enzyme catalyzing the reversible hydration of CO2 to bicarbonate, a reaction underlying diverse biochemical and physiological processes. Gamma class carbonic anhydrases (γ-CAs are widespread in prokaryotes but their physiological roles remain elusive. At present, only γ-CA of Methanosarcina thermophila (Cam has been shown to have CA activity. Genome analysis of a rhizobacterium Azospirillum brasilense, revealed occurrence of ORFs encoding one β-CA and two γ-CAs. Results One of the putative γ-CA encoding genes of A. brasilense was cloned and overexpressed in E. coli. Electrometric assays for CA activity of the whole cell extracts overexpressing recombinant GCA1 did not show CO2 hydration activity. Reverse transcription-PCR analysis indicated that gca1 in A. brasilense is co-transcribed with its upstream gene annotated as argC, which encodes a putative N-acetyl-γ-glutamate-phosphate reductase. 5'-RACE also demonstrated that there was no transcription start site between argC and gca1, and the transcription start site located upstream of argC transcribed both the genes (argC-gca1. Using transcriptional fusions of argC-gca1 upstream region with promoterless lacZ, we further demonstrated that gca1 upstream region did not have any promoter and its transcription occurred from a promoter located in the argC upstream region. The transcription of argC-gca1 operon was upregulated in stationary phase and at elevated CO2 atmosphere. Conclusions This study shows lack of CO2 hydration activity in a recombinant protein expressed from a gene predicted to encode a γ-carbonic anhydrase in A. brasilense although it cross reacts with anti-Cam antibody raised against a well characterized γ-CA. The organization and regulation of this gene along with the putative argC gene suggests its involvement in arginine biosynthetic pathway instead of the predicted CO2 hydration.

  3. The membraneless bioelectrochemical reactor stimulates hydrogen fermentation by inhibiting methanogenic archaea.

    Science.gov (United States)

    Sasaki, Kengo; Morita, Masahiko; Sasaki, Daisuke; Ohmura, Naoya; Igarashi, Yasuo

    2013-08-01

    The membraneless bioelectrochemical reactor (Ml-BER) is useful for dark hydrogen fermentation. The effect of the electrochemical reaction on microorganisms in the Ml-BER was investigated using glucose as the substrate and compared with organisms in a membraneless non-bioelectrochemical reactor (Ml-NBER) and bioelectrochemical reactor (BER) with a proton exchange membrane. The potentials on the working electrode of the Ml-BER and BER with membrane were regulated to -0.9 V (versus Ag/AgCl) to avoid water electrolysis with a carbon electrode. The Ml-BER showed suppressed methane production (19.8 ± 9.1 mg-C·L(-1)·day(-1)) and increased hydrogen production (12.6 ± 3.1 mg-H·L(-1)·day(-1)) at pHout 6.2 ± 0.1, and the major intermediate was butyrate (24.9 ± 2.4 mM), suggesting efficient hydrogen fermentation. In contrast, the Ml-NBER showed high methane production (239.3 ± 17.9 mg-C·L(-1)·day(-1)) and low hydrogen production (0.2 ± 0.0 mg-H·L(-1)·day(-1)) at pHout 6.3 ± 0.1. In the cathodic chamber of the BER with membrane, methane production was high (276.3 ± 20.4 mg-C·L(-1)·day(-1)) (pHout, 7.2 ± 0.1). In the anodic chamber of the BER with membrane (anode-BER), gas production was low because of high lactate production (43.6 ± 1.7 mM) at pHout 5.0 ± 0.1. Methanogenic archaea were not detected in the Ml-BER and anode-BER. However, Methanosarcina sp. and Methanobacterium sp. were found in Ml-NBER. Prokaryotic copy numbers in the Ml-BER and Ml-NBER were similar, as were the bacterial community structures. Thus, the electrochemical reaction in the Ml-BER affected hydrogenotrophic and acetoclastic methanogens, but not the bacterial community.

  4. Sequence analysis corresponding to the PPE and PE proteins in Mycobacterium tuberculosis and other genomes

    Indian Academy of Sciences (India)

    Swathi Adindla; Lalitha Guruprasad

    2003-03-01

    Amino acid sequence analysis corresponding to the PPE proteins in H37Rv and CDC1551 strains of the Mycobacterium tuberculosis genomes resulted in the identification of a previously uncharacterized 225 amino acidresidue common region in 22 proteins. The pairwise sequence identities were as low as 18%. Conservation of amino acid residues was observed at fifteen positions that were distributed over the whole length of the region. The secondary structure corresponding to this region is predicted to be a mixture of -helices and -strands. Although the function is not known, proteins with this region specific to mycobacterial species may be associated with a common function. We further observed another group of 20 PPE proteins corresponding to the conserved C-terminal region comprising 44 amino acid residues with GFxGT and PxxPxxW sequence motifs. This region is preceded by a hydrophobic region, comprising 40–100 amino acid residues, that is flanked by charged amino acid residues. Identification of conserved regions described above may be useful to detect related proteins from other genomes and assist the design of suitable experiments to test their corresponding functions. Amino acid sequence analysis corresponding to the PE proteins resulted in the identification of tandem repeats comprising 41–43 amino acid residues in the C-terminal variable regions in two PE proteins (Rv0978 and Rv0980). These correspond to the AB repeats that were first identified in some proteins of the Methanosarcina mazei genome, and were demonstrated as surface antigens. We observed the AB repeats also in several other proteins of hitherto uncharacterized function in Archaea and Bacteria genomes. Some of these proteins are also associated with another repeat called the C-repeat or the PKD-domain comprising 85 amino acid residues. The secondary structure corresponding to the AB repeat is predicted mainly as 4 -strands. We suggest that proteins with AB repeats in Mycobacterium tuberculosis and

  5. The impact of permafrost-associated microorganisms on hydrate formation kinetics

    Science.gov (United States)

    Luzi-Helbing, Manja; Liebner, Susanne; Spangenberg, Erik; Wagner, Dirk; Schicks, Judith M.

    2016-04-01

    The relationship between gas hydrates, microorganisms and the surrounding sediment is extremely complex: On the one hand, microorganisms producing methane provide the prerequisite for gas hydrate formation. As it is known most of the gas incorporated into natural gas hydrates originates from biogenic sources. On the other hand, as a result of microbial activity gas hydrates are surrounded by a great variety of organic compounds which are not incorporated into the hydrate structure but may influence the formation or degradation process. For gas hydrate samples from marine environments such as the Gulf of Mexico a direct association between microbes and gas hydrates was shown by Lanoil et al. 2001. It is further assumed that microorganisms living within the gas hydrate stability zone produce biosurfactants which were found to enhance the hydrate formation process significantly and act as nucleation centres (Roger et al. 2007). Another source of organic compounds is sediment organic matter (SOM) originating from plant material or animal remains which may also enhance hydrate growth. So far, the studies regarding this relationship were focused on a marine environment. The scope of this work is to extend the investigations to microbes originating from permafrost areas. To understand the influence of microbial activity in a permafrost environment on the methane hydrate formation process and the stability conditions of the resulting hydrate phase we will perform laboratory studies. Thereby, we mimic gas hydrate formation in the presence and absence of methanogenic archaea (e.g. Methanosarcina soligelidi) and other psychrophilic bacteria isolated from permafrost environments of the Arctic and Antarctic to investigate their impact on hydrate induction time and formation rates. Our results may contribute to understand and predict the occurrences and behaviour of potential gas hydrates within or adjacent to the permafrost. Lanoil BD, Sassen R, La Duc MT, Sweet ST, Nealson KH

  6. Biological CO2 conversion to acetate in subsurface coal-sand formation using a high-pressure reactor system

    Science.gov (United States)

    Ohtomo, Y.; Ijiri, A.; Ikegawa, Y.; Tsutsumi, M.; Imachi, H.; Uramoto, G.; Hoshino, T.; Morono, Y.; Tanikawa, W.; Hirose, T.; Inagaki, F.

    2013-12-01

    belonged to a methylotrophic methanogen within the genus Methanosarcina. For the acetate-fed culture, no cell proliferation and methane-production were observed after two-years incubation. During the injection of CO2 and fluid, increase of dissolved CH4 concentration was observed, of which δ13CCH4 were constantly similar to those of the absorbed coal-bed methane (δ13CCBM, ~70‰), suggesting the enhanced gas recovery with fluid flow. The output volume of CO2 (ΣCO2out, 22.1 to 125.6 mM) was smaller than initial concentration (ΣCO2in, 138.38 mM), which can be explained by either adsorption on coal, formation of carbonate minerals, or microbial consumption. Increase of acetate concentration in the fluids was also observed, whereas δ13Cacetate depleted during experiment. Considering with the decrease of additive H2, it is most likely that homo-acetogenesis would occur during experiments, which is consistent with detection of Sporomusa-related 16S rRNA genes, homo-acetogenic bacterium, in cloning analysis of sandstone after experiment. Decrease of formate concentrations and increase of δ13Cformate indicate bacterial consumption of formate and isotopic fractionation. Our results suggest that CO2 injection to natural coal-sand formation stimulates homo-acetogenesis rather than methanogenesis, accompanied by biogenic CO2 conversion to acetate.

  7. Experimental methods for screening parameters influencing the growth to product yield (Y(x/CH4 of a biological methane production (BMP process performed with Methanothermobacter marburgensis

    Directory of Open Access Journals (Sweden)

    Sébastien Bernacchi

    2014-12-01

    Uncoupling of methanogenesis from growth of Methanosarcina barkeri by phosphate limitation. Appl Env

  8. Microbial succession within an anaerobic sequencing batch biofilm reactor (ASBBR treating cane vinasse at 55ºC

    Directory of Open Access Journals (Sweden)

    Maria Magdalena Ferreira Ribas

    2009-08-01

    grande diversidade de morfologias ao longo do experimento. A predominância de células como Methanosaeta foram observadas até a carga orgânica de 2,5 gDQO/L.d. Por outro lado, microrganismos como Methanosarcina foram a população arquéia predominante nas matrizes de espuma em taxas de carregamento orgânico acima de 3,3 gDQO/L.d. Este fato pode estar relacionado com maior taxa de utilização específica de acetato por esses microrganismos.

  9. Effect of temperature on performance and microbial community of thermophilic methane fermentation of kitchen waste%温度对厨余垃圾高温厌氧消化及微生物群落的影响

    Institute of Scientific and Technical Information of China (English)

    杨璐; 张影; 汤岳琴; 孙照勇; 黄玉莲; 木田建次

    2014-01-01

    The study aimed to investigate the effect of temperature on performance and microbial community of thermophilic methane fermentation of kitchen waste.Kitchen waste was treated using a completely stirred tank reactor (CSTR) at 53 ℃ and 60 ℃.Microbial community structure during treatment was analyzed using T-RFLP and 16S rRNA clone library methods.At the temperature of 53 ℃ and a VTS loading rate of 4 g/L/d,the treatment was stable without accumulation of VFA,with gas production approximately 900 mL/g-VTS.When the temperature was raised to 60 ℃,TOC and VFA accumulated sharply and the gas production decreased,with changes in both bacterial community and archaeal community.At 53 ℃,the acidproducing fermenting bacteria were dominant.When the temperature was increased to 60 ℃,however,the population of fatty acid-oxidizing,especially acetate-oxidizing bacteria increased significantly.The population of aceticlastic methanogen Methanosarcina decreased while hydrogenotrophic methangen Methanothermobacter increased at 60 ℃.The main pathway of methane production from acetate at 60 ℃ was through the acetate oxidization followed by hydrogenotrophic methanogenesis,which needs syntrophism of acetate-oxidizing bacteria and hydrogenotrophic methangens.The results showed that the operational temperature of 53 ℃ is suitable for thermophilic methane fermentation of kitchen waste.Increase of operational temperature to 60 ℃ would result in decrease of methane production through aceticlastic methanogenesis pathway,leading to accumulation of VFAs and deterioration of treatment.%使用全混流反应器(CSTR),研究53℃和60℃条件下厨余垃圾的处理效果,同时利用T-RFLP(末端限制性片段长度多态性)以及16S rRNA基因克隆文库的方法,对微生物群落结构进行跟踪解析.结果表明,在温度为53 ℃、挥发性总固体(VTS)负荷为4 g L-1 d-1时,处理性能稳定,有机酸积累少,产气率约为900 mL/g VTS;当温度升高至60℃

  10. 原核微生物菌群的空间分异增强秸秆-猪粪混合发酵效率%Spatial differentiation of prokaryotes enhancing performance of co-fermentation with straw and swine manure

    Institute of Scientific and Technical Information of China (English)

    李家宝; 芮俊鹏; 张时恒; 孙晓日; 闫志英; 刘晓风; 郑涛; 李香真

    2014-01-01

    that spatial distribution pattern of prokaryotes in the co-fermentation system might increase fermentation efficiency. Straw was digested by its associated microbiota, such as generaTreponema, ClostridiumⅢ, Alkaliflexusand Fibrobacter, providing substrates for the production of volatile fatty acids (VFAs). Propionic acid was most abundant in the sludge. The generaSyntrophomonas, Pelotomaculum,Methanoculleus, Methanosarcina andMethanosaeta metabolize VFAs into methane syntrophicallyvia both hydrogenotrophic and acetocalstic pathways. The generaAminobacterium and Cloacibacillus involved in amino acid degradation were much abundant in the sludge, indicating that protein was an important substrate for methanogenesis. These findings suggested that spatial distribution of microbiota, steady straw-associated degraders and dynamic microbial communities in the sludge were vital for the performance of biogas co-fermentation system.

  11. Functional analysis of archaeal MBF1 by complementation studies in yeast

    Directory of Open Access Journals (Sweden)

    Siebers Bettina

    2011-03-01

    Full Text Available Abstract Background Multiprotein-bridging factor 1 (MBF1 is a transcriptional co-activator that bridges a sequence-specific activator (basic-leucine zipper (bZIP like proteins (e.g. Gcn4 in yeast or steroid/nuclear-hormone receptor family (e.g. FTZ-F1 in insect and the TATA-box binding protein (TBP in Eukaryotes. MBF1 is absent in Bacteria, but is well- conserved in Eukaryotes and Archaea and harbors a C-terminal Cro-like Helix Turn Helix (HTH domain, which is the only highly conserved, classical HTH domain that is vertically inherited in all Eukaryotes and Archaea. The main structural difference between archaeal MBF1 (aMBF1 and eukaryotic MBF1 is the presence of a Zn ribbon motif in aMBF1. In addition MBF1 interacting activators are absent in the archaeal domain. To study the function and therefore the evolutionary conservation of MBF1 and its single domains complementation studies in yeast (mbf1Δ as well as domain swap experiments between aMBF1 and yMbf1 were performed. Results In contrast to previous reports for eukaryotic MBF1 (i.e. Arabidopsis thaliana, insect and human the two archaeal MBF1 orthologs, TMBF1 from the hyperthermophile Thermoproteus tenax and MMBF1 from the mesophile Methanosarcina mazei were not functional for complementation of an Saccharomyces cerevisiae mutant lacking Mbf1 (mbf1Δ. Of twelve chimeric proteins representing different combinations of the N-terminal, core domain, and the C-terminal extension from yeast and aMBF1, only the chimeric MBF1 comprising the yeast N-terminal and core domain fused to the archaeal C-terminal part was able to restore full wild-type activity of MBF1. However, as reported previously for Bombyx mori, the C-terminal part of yeast Mbf1 was shown to be not essential for function. In addition phylogenetic analyses revealed a common distribution of MBF1 in all Archaea with available genome sequence, except of two of the three Thaumarchaeota; Cenarchaeum symbiosum A and Nitrosopumilus maritimus

  12. The hydrogen concentration as parameter to identify natural attenuation processes of volatile chlorinated hydrocarbons in ground water; Die Wasserstoffkonzentration als Parameter zur Identifizierung des natuerlichen Abbaus von leichtfluechtigen Chlorkohlenwasserstoffen (LCKW) im Grundwasser

    Energy Technology Data Exchange (ETDEWEB)

    Alter, M.D.

    2006-06-15

    In this study, the hydrogen concentration as parameter to identify natural attenuation processes of volatile chlorinated hydrocarbons was investigated. The currently accepted and recommended bubble strip method for hydrogen sampling was optimized, and a storage method for hydrogen samples was developed. Furthermore batch experiments with a dechlorinating mixed culture and pure cultures were carried out to study H{sub 2}-concentrations of competing redox processes. The extraction of hydrogen from ground water was optimized by a reduced inlet diameter of the usually applied gas sampling bulbs, allowing a maximal turbulent ow and gas transfer. With a gas volume of 10 ml and flow rates of 50 to 140 ml/min, the course of extraction almost followed the theoretical course of equilibration. At flow rates > 100 ml/min a equilibrium of 98% was achieved within 20 min. Until recently it was generally accepted that hydrogen samples can be stored only for 2 hours and therefore have to be analyzed immediately in the eld. Here, it was shown that eld samples can be stored for 1-3 days until analysis. For the dechlorination of tetrachloroethene (PCE), a hydrogen threshold concentration of 1-2 nM was found with the dechlorinating mixed culture as well as with a pure culture of Sulfurospirillum multivorans in combination with another pure culture Methanosarcina mazei. No dechlorination was detectable below this concentration. With the dechlorinating mixed culture, this finding is valid for all successive dechlorination steps until ethene. The hydrogen threshold concentration for denitrification were below the detection limit of 0,2 nM with the dechlorinating mixed culture. A threshold concentration of 3,1-3,5 nM was found for sulphate reduction and a threshold of 7-9 nM H{sub 2} for hydrogenotrophic methanogenesis. This implies that the natural dechlorination at contaminated sites is preferred to competing processes like sulphate reduction and methanogenesis. The threshold

  13. 脉冲循环式渠槽厌氧反应器处理太湖腐熟蓝藻性能%Capability of pulse cycle corridor anaerobic reactor treating composted algae water

    Institute of Scientific and Technical Information of China (English)

    余亚琴; 吕锡武; 吴义锋; 许丽娟

    2013-01-01

    by a scanning electron microscope (SEM), the granular sludge was found in the stabilization stage with coupled growing micro-organisms, including methanosarcina, filamentous bacteria, and rod-shaped bacteria. Proteinase, coenzyme F420 and TTC-dehydrogenase activity of granular sludge from pulse cycle corridor anaerobic reactor had been in high level during the duration of the experiment. Results indicated that the pulse cycle corridor anaerobic reactor could effectively deal with the algae-laden water from the Taihu Lake. It provided important value for the biogas fermentation of algae.

  14. Sustainable resource recovery and energy conversion processes using microbial electrochemical technologies

    Science.gov (United States)

    Yates, Matthew D.

    , giving the greatest amount of control over the thickness of the polymer layer. The second process studied used exoelectrogenic bacteria in METs for the conversion of electrons to hydrogen via water electrolysis in a biocathodic system. Naturally occurring biocatalytic cell material on the cathode surface was used to lower the cathode overpotential. Different cell cultures ( G. sulfurreducens, Methanosarcina barkeri, and Escherichia coli) were tested for their effect on hydrogen formation using electrons supplied to an insoluble electrode. The mode of hydrogen production was investigated by monitoring hydrogen production over three to five months using G. sulfurreducens biofilms (pregrown under anodic conditions with acetate) that were: (1) not supplied with an organic carbon source for cell growth and maintenance, (2) killed with ethanol, or (3) supplied with lactate, an organic carbon source and electron donor for G. sulfurreducens. Hydrogen was produced at a rate 10--20 times higher over five months in reactors that were either not given organic carbon or killed with ethanol, compared to reactors with lactate added. The methanogen, M. barkeri, was also tested as a biocatalyst because it is able to grow autotrophically. However, M. barkeri cells did not grow in the reactor with the electrode potential poised, based on the lack of evidence for methane production. Despite the lack of cell activity, the rate of hydrogen production with M. barkeri was similar to the rate observed in killed G. sulfurreducens reactors. The addition of E. coli, a non-exoelectrogenic bacteria, resulted in an initial elevated hydrogen gas production, but hydrogen production rates similar to background levels after three months. No cells were detected on the electrode surfaces after five months using scanning electron microscopy and unique metals, such as iron, nickel, cobalt, and zinc, were detected on the electrode surfaces exposed to cells. The identifiable peptides extracted from the