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Sample records for acetogen moorella thermoacetica

  1. Anaerobic CO2 fixation by the acetogenic bacterium Moorella thermoacetica

    SciTech Connect

    Hu, P; Rismani-Yazdi, H; Stephanopoulos, G

    2013-05-16

    Anaerobic bacteria such as Moorella thermoacetica have the capacity of fixing carbon dioxide with carbon monoxide and hydrogen for the production of ethanol, acetic acid, and other useful chemicals. In this study, we evaluated the fixation of CO2 for the production of acetic acid, as a product in its own right but also as precursor for lipid synthesis by oleaginous organisms. We achieved maximum cell optical density of 11.3, acetic acid titer of 31 g/L, and productivity of 0.55 g/L-h at CO mass-transfer rate of 83 mM/h. We also showed electron availability by CO mass transfer limited the process at CO mass transfer rates lower than 30 mM/h. Further enhancement of mass-transfer rate removed such limitations in favor of biological kinetics as main limitation. This work underlines the potential of microbial processes for converting syngas to fuel and chemical products in processes suitable for distributed feedstock utilization. (c) 2013 American Institute of Chemical Engineers AIChE J, 59: 3176-3183, 2013

  2. THERMICANUS AEGYPTIUS GEN. NOV., SP. NOV., ISOLATED FROM OXIC SOIL, A FERMENTATIVE MICROAEROPHILE THAT GROWS COMMENSALLY WITH THE THERMOPHILIC ACETOGEN MOORELLA THERMOACETICA

    EPA Science Inventory

    A thermophilic, fermentative microaerophile (ET-5b) and a thermophilic acetogen (ET-5a) were coisolated from oxic soil obtained from Egypt. The 16S rRNA gene sequence of ET-5a was 99.8% identical to that of the classic acetogen Moorella thermoacetica. Further analyses confirmed t...

  3. THERMICANUS AEGYPTIUS GEN. NOV., SP. NOV., ISOLATED FROM OXIC SOIL, A FERMENTATIVE MICROAEROPHILE THAT GROWS COMMENSALLY WITH THE THERMOPHILIC ACETOGEN MOORELLA THERMOACETICA

    EPA Science Inventory

    A thermophilic, fermentative microaerophile (ET-5b) and a thermophilic acetogen (ET-5a) were coisolated from oxic soil obtained from Egypt. The 16S rRNA gene sequence of ET-5a was 99.8% identical to that of the classic acetogen Moorella thermoacetica. Further analyses confirmed t...

  4. Thermicanus aegyptius gen. nov., sp. nov., isolated from oxic soil, a fermentative microaerophile that grows commensally with the thermophilic acetogen Moorella thermoacetica.

    PubMed

    Gössner, A S; Devereux, R; Ohnemüller, N; Acker, G; Stackebrandt, E; Drake, H L

    1999-11-01

    A thermophilic, fermentative microaerophile (ET-5b) and a thermophilic acetogen (ET-5a) were coisolated from oxic soil obtained from Egypt. The 16S rRNA gene sequence of ET-5a was 99.8% similar to that of the classic acetogen Moorella thermoacetica. Further analyses confirmed that ET-5a was a new strain of M. thermoacetica. For ET-5b, the nearest 16S rRNA gene sequence similarity value to known genera was approximately 88%. ET-5b was found to be a motile rod with a genomic G+C content of 50.3 mol%. Cells were weakly gram positive and lacked spores. Growth was optimal at 55 to 60 degrees C and pH 6.5 to 7.0. ET-5b grew under both oxic and anoxic conditions, but growth was erratic under atmospheric concentrations of O(2). Utilizable substrates included oligosaccharides and monosaccharides. Acetate, formate, and succinate supported growth only under oxic conditions. Saccharides yielded succinate, lactate, ethanol, acetate, formate, and H(2) under anoxic conditions; fermentation products were also formed under oxic conditions. A new genus is proposed, the type strain being Thermicanus aegyptius ET-5b gen. nov., sp. nov. (DSMZ 12793). M. thermoacetica ET-5a (DSMZ 12797) grew commensally with T. aegyptius ET-5b on oligosaccharides via the interspecies transfer of H(2) formate, and lactate. In support of this interaction, uptake hydrogenase and formate dehydrogenase specific activities were fundamentally greater in M. thermoacetica ET-5a than in T. aegyptius ET-5b. These results demonstrate that (i) soils subject to high temperatures harbor uncharacterized thermophilic microaerophiles, (ii) the classic acetogen M. thermoacetica resides in such soils, and (iii) trophic links between such soil bacteria might contribute to their in situ activities.

  5. Electron bifurcation involved in the energy metabolism of the acetogenic bacterium Moorella thermoacetica growing on glucose or H2 plus CO2.

    PubMed

    Huang, Haiyan; Wang, Shuning; Moll, Johanna; Thauer, Rudolf K

    2012-07-01

    Moorella thermoacetica ferments glucose to three acetic acids. In the oxidative part of the fermentation, the hexose is converted to 2 acetic acids and 2 CO(2) molecules with the formation of 2 NADH and 2 reduced ferredoxin (Fd(red)(2-)) molecules. In the reductive part, 2 CO(2) molecules are reduced to acetic acid, consuming the 8 reducing equivalents generated in the oxidative part. An open question is how the two parts are electronically connected, since two of the four oxidoreductases involved in acetogenesis from CO(2) are NADP specific rather than NAD specific. We report here that the 2 NADPH molecules required for CO(2) reduction to acetic acid are generated by the reduction of 2 NADP(+) molecules with 1 NADH and 1 Fd(red)(2-) catalyzed by the electron-bifurcating NADH-dependent reduced ferredoxin:NADP(+) oxidoreductase (NfnAB). The cytoplasmic iron-sulfur flavoprotein was heterologously produced in Escherichia coli, purified, and characterized. The purified enzyme was composed of 30-kDa (NfnA) and 50-kDa (NfnB) subunits in a 1-to-1 stoichiometry. NfnA harbors a [2Fe2S] cluster and flavin adenine dinucleotide (FAD), and NfnB harbors two [4Fe4S] clusters and FAD. M. thermoacetica contains a second electron-bifurcating enzyme. Cell extracts catalyzed the coupled reduction of NAD(+) and Fd with 2 H(2) molecules. The specific activity of this cytoplasmic enzyme was 3-fold higher in H(2)-CO(2)-grown cells than in glucose-grown cells. The function of this electron-bifurcating hydrogenase is not yet clear, since H(2)-CO(2)-grown cells additionally contain high specific activities of an NADP(+)-dependent hydrogenase that catalyzes the reduction of NADP(+) with H(2). This activity is hardly detectable in glucose-grown cells.

  6. Electron Bifurcation Involved in the Energy Metabolism of the Acetogenic Bacterium Moorella thermoacetica Growing on Glucose or H2 plus CO2

    PubMed Central

    Huang, Haiyan; Wang, Shuning; Moll, Johanna

    2012-01-01

    Moorella thermoacetica ferments glucose to three acetic acids. In the oxidative part of the fermentation, the hexose is converted to 2 acetic acids and 2 CO2 molecules with the formation of 2 NADH and 2 reduced ferredoxin (Fdred2−) molecules. In the reductive part, 2 CO2 molecules are reduced to acetic acid, consuming the 8 reducing equivalents generated in the oxidative part. An open question is how the two parts are electronically connected, since two of the four oxidoreductases involved in acetogenesis from CO2 are NADP specific rather than NAD specific. We report here that the 2 NADPH molecules required for CO2 reduction to acetic acid are generated by the reduction of 2 NADP+ molecules with 1 NADH and 1 Fdred2− catalyzed by the electron-bifurcating NADH-dependent reduced ferredoxin:NADP+ oxidoreductase (NfnAB). The cytoplasmic iron-sulfur flavoprotein was heterologously produced in Escherichia coli, purified, and characterized. The purified enzyme was composed of 30-kDa (NfnA) and 50-kDa (NfnB) subunits in a 1-to-1 stoichiometry. NfnA harbors a [2Fe2S] cluster and flavin adenine dinucleotide (FAD), and NfnB harbors two [4Fe4S] clusters and FAD. M. thermoacetica contains a second electron-bifurcating enzyme. Cell extracts catalyzed the coupled reduction of NAD+ and Fd with 2 H2 molecules. The specific activity of this cytoplasmic enzyme was 3-fold higher in H2-CO2-grown cells than in glucose-grown cells. The function of this electron-bifurcating hydrogenase is not yet clear, since H2-CO2-grown cells additionally contain high specific activities of an NADP+-dependent hydrogenase that catalyzes the reduction of NADP+ with H2. This activity is hardly detectable in glucose-grown cells. PMID:22582275

  7. Two propanediol utilization-like proteins of Moorella thermoacetica with phosphotransacetylase activity.

    PubMed

    Breitkopf, Ronja; Uhlig, Ronny; Drenckhan, Tina; Fischer, Ralf-Jörg

    2016-09-01

    Moorella thermoacetica is one of the model acetogenic bacteria for the resolution of the Wood-Ljungdahl (acetyl-CoA) pathway in which CO2 is autotrophically assimilated yielding acetyl-CoA as central intermediate. Its further conversion into acetate relies on subsequent phosphotransacetylase (PTA) and acetate kinase reactions. However, the genome of M. thermoacetica contains no pta homologous gene. It has been speculated that the moth_0864 and moth_1181 gene products sharing similarities with an evolutionarily distinct phosphotransacylase involved in 1,2-propanediol utilization (PDUL) of Salmonella enterica act as PTAs in M. thermoacetica. Here, we demonstrate specific PTA activities with acetyl-CoA as substrate of 9.05 and 2.03 U/mg for the recombinant enzymes PDUL1 (Moth_1181) and PDUL2 (Moth_0864), respectively. Both showed maximal activity at 65 °C and pH 7.6. Native proteins (90 kDa) are homotetramers composed of four subunits with apparent molecular masses of about 23 kDa. Thus, one or both PDULs of M. thermoacetica might act as PTAs in vivo catalyzing the penultimate step of the Wood-Ljungdahl pathway toward the formation of acetate. In silico analysis underlined that up to now beside of M. thermoacetica, only Sporomusa ovata contains only PDUL like class(III)-PTAs but no other phosphotransacetylases or phosphotransbutyrylases (PTBs).

  8. The Complete Genome Sequence of Moorella thermoacetica (f. Clostridium thermoaceticum)

    PubMed Central

    Pierce, Elizabeth; Xie, Gary; Barabote, Ravi D.; Saunders, Elizabeth; Han, Cliff S.; Detter, John C.; Richardson, Paul; Brettin, Thomas S.; Das, Amaresh; Ljungdahl, Lars G.; Ragsdale, Stephen W.

    2008-01-01

    Summary This paper describes the genome sequence of M. thermoacetica (f. Clostridium thermoaceticum), which is the model acetogenic bacterium that has been widely used for elucidating the Wood-Ljungdahl pathway of CO and CO2 fixation. This pathway, which is also known as the reductive acetyl-CoA pathway, allows acetogenic (often called homoacetogenic) bacteria to convert glucose stoichiometrically into three mol of acetate and to grow autotrophically using H2 and CO as electron donors and CO2 as an electron acceptor. Methanogenic archaea use this pathway in reverse to grow by converting acetate into methane and CO2. Acetogenic bacteria also couple the Wood-Ljungdahl pathway to a variety of other pathways to allow the metabolism of a wide variety of carbon sources and electron donors (sugars, carboxylic acids, alcohols, and aromatic compounds) and electron acceptors (CO2, nitrate, nitrite, thiosulfate, dimethylsulfoxide, and aromatic carboxyl groups). The genome consists of a single circular 2628784 bp chromosome encoding 2615 open reading frames, which includes 2523 predicted protein-encoding genes. Of these, 1834 genes (70.13%) have been assigned tentative functions, 665 (25.43%) matched genes of unknown function, and the remaining 24 (0.92%) had no database match. Two thousand three hundred eighty-four (91.17%) of the ORFs in the M. thermoacetica genome can be grouped in ortholog clusters. This first genome sequence of an acetogenic bacterium provides important information related to how acetogens engage their extreme metabolic diversity by switching among different carbon substrates and electron donors/acceptors and how they conserve energy by anaerobic respiration. Our genome analysis indicates that the key genetic trait for homoacetogenesis is the core acs gene cluster of the Wood-Ljungdahl pathway. PMID:18631365

  9. Fermentation of lignocellulosic sugars to acetic acid by Moorella thermoacetica.

    PubMed

    Ehsanipour, Mandana; Suko, Azra Vajzovic; Bura, Renata

    2016-06-01

    A systematic study of bioconversion of lignocellulosic sugars to acetic acid by Moorella thermoacetica (strain ATCC 39073) was conducted. Four different water-soluble fractions (hydrolysates) obtained after steam pretreatment of lignocellulosic biomass were selected and fermented to acetic acid in batch fermentations. M. thermoacetica can effectively ferment xylose and glucose in hydrolysates from wheat straw, forest residues, switchgrass, and sugarcane straw to acetic acid. Xylose and glucose were completely utilized, with xylose being consumed first. M. thermoacetica consumed up to 62 % of arabinose, 49 % galactose and 66 % of mannose within 72 h of fermentation in the mixture of lignocellulosic sugars. The highest acetic acid yield was obtained from sugarcane straw hydrolysate, with 71 % of theoretical yield based on total sugars (17 g/L acetic acid from 24 g/L total sugars). The lowest acetic acid yield was observed in forest residues hydrolysate, with 39 % of theoretical yield based on total sugars (18 g/L acetic acid from 49 g/L total sugars). Process derived compounds from steam explosion pretreatment, including 5-hydroxymethylfurfural (0.4 g/L), furfural (0.1 g/L) and total phenolics (3 g/L), did not inhibit microbial growth and acetic acid production yield. This research identified two major factors that adversely affected acetic acid yield in all hydrolysates, especially in forest residues: (i) glucose to xylose ratio and (ii) incomplete consumption of arabinose, galactose and mannose. For efficient bioconversion of lignocellulosic sugars to acetic acid, it is imperative to have an appropriate balance of sugars in a hydrolysate. Hence, the choice of lignocellulosic biomass and steam pretreatment design are fundamental steps for the industrial application of this process.

  10. Evidence for a Hexaheteromeric Methylenetetrahydrofolate Reductase in Moorella thermoacetica

    PubMed Central

    Mock, Johanna; Wang, Shuning; Huang, Haiyan; Kahnt, Jörg

    2014-01-01

    Moorella thermoacetica can grow with H2 and CO2, forming acetic acid from 2 CO2 via the Wood-Ljungdahl pathway. All enzymes involved in this pathway have been characterized to date, except for methylenetetrahydrofolate reductase (MetF). We report here that the M. thermoacetica gene that putatively encodes this enzyme, metF, is part of a transcription unit also containing the genes hdrCBA, mvhD, and metV. MetF copurified with the other five proteins encoded in the unit in a hexaheteromeric complex with an apparent molecular mass in the 320-kDa range. The 40-fold-enriched preparation contained per mg protein 3.1 nmol flavin adenine dinucleotide (FAD), 3.4 nmol flavin mononucleotide (FMN), and 110 nmol iron, almost as predicted from the primary structure of the six subunits. It catalyzed the reduction of methylenetetrahydrofolate with reduced benzyl viologen but not with NAD(P)H in either the absence or presence of oxidized ferredoxin. It also catalyzed the reversible reduction of benzyl viologen with NADH (diaphorase activity). Heterologous expression of the metF gene in Escherichia coli revealed that the subunit MetF contains one FMN rather than FAD. MetF exhibited 70-fold-higher methylenetetrahydrofolate reductase activity with benzyl viologen when produced together with MetV, which in part shows sequence similarity to MetF. Heterologously produced HdrA contained 2 FADs and had NAD-specific diaphorase activity. Our results suggested that the physiological electron donor for methylenetetrahydrofolate reduction in M. thermoacetica is NADH and that the exergonic reduction of methylenetetrahydrofolate with NADH is coupled via flavin-based electron bifurcation with the endergonic reduction of an electron acceptor, whose identity remains unknown. PMID:25002540

  11. Glycerol acts as alternative electron sink during syngas fermentation by thermophilic anaerobe Moorella thermoacetica.

    PubMed

    Kimura, Zen-ichiro; Kita, Akihisa; Iwasaki, Yuki; Nakashimada, Yutaka; Hoshino, Tamotsu; Murakami, Katsuji

    2016-03-01

    Moorella thermoacetica is an anaerobic thermophilic acetogen that is capable of fermenting sugars, H(2)/CO(2) and syngas (H(2)/CO). For this reason, this bacterium is potentially useful for biotechnology applications, particularly the production of biofuel from CO(2). A soil isolate of M. thermoacetica, strain Y72, produces both ethanol and acetate from H(2)/CO(2); however, the maximum concentrations of these two products are too low to enable commercialization of the syngas fermentation process. In the present study, glycerol was identified as a novel electron sink among the fermentation products of strain Y72. Notably, a 1.5-fold increase in the production of ethanol (1.4 mM) was observed in cultures supplemented with glycerol during syngas fermentation. This discovery is expected to aid in the development of novel methods that allow for the regulation of metabolic pathways to direct and increase the production of desirable fermentative compounds. Copyright © 2015. Published by Elsevier B.V.

  12. Engineering of a functional thermostable kanamycin resistance marker for use in Moorella thermoacetica ATCC39073.

    PubMed

    Iwasaki, Yuki; Kita, Akihisa; Sakai, Shinsuke; Takaoka, Kazue; Yano, Shinichi; Tajima, Takahisa; Kato, Junichi; Nishio, Naomichi; Murakami, Katsuji; Nakashimada, Yutaka

    2013-06-01

    A transformation system for Moorella thermoacetica ATCC39073 was developed using thermostable kanamycin resistant gene (kanR) derived from the plasmid pJH1 that Streptococcus faecalis harbored. When kanR with its native promoter was introduced into uracil auxotrophic mutant of M. thermoacetica ATCC39073 together with a gene to complement the uracil auxotrophy as a selection marker, it did not give kanamycin resistance due to poor transcription level of kanR. However, the use of glyceraldehyde-3-phosphate dehydrogenase promoter cloned from M. thermoacetica ATCC39073 significantly improved transcription level of kanR and resulted in the cell growth in the presence of more than 150 μg mL(-1) kanamycin. It was also demonstrated that kanR with G3PD promoter can be used as a selection marker for transformation of wild-type strain of M. thermoacetica ATCC39073.

  13. Genome Sequence of the Acetogenic Bacterium Moorella mulderi DSM 14980T

    PubMed Central

    Castillo Villamizar, Genis Andrés

    2016-01-01

    Here, we report the draft genome sequence of Moorella mulderi DSM 14980T, a thermophilic acetogenic bacterium, which is able to grow autotrophically on H2 plus CO2 using the Wood-Ljungdahl pathway. The genome consists of a circular chromosome (2.99 Mb). PMID:27231372

  14. Influence of nitrate on oxalate- and glyoxylate-dependent growth and acetogenesis by Moorella thermoacetica.

    PubMed

    Seifritz, Corinna; Fröstl, Jürgen M; Drake, Harold L; Daniel, Steven L

    2002-12-01

    Oxalate and glyoxylate supported growth and acetate synthesis by Moorella thermoacetica in the presence of nitrate under basal (without yeast extract) culture conditions. In oxalate cultures, acetate formation occurred concomitant with growth and nitrate was reduced in the stationary phase. Growth in the presence of [(14)C]bicarbonate or [(14)C]oxalate showed that CO(2) reduction to acetate and biomass or oxalate oxidation to CO(2) was not affected by nitrate. However, cells engaged in oxalate-dependent acetogenesis in the presence of nitrate lacked a membranous b-type cytochrome, which was present in cells grown in the absence of nitrate. In glyoxylate cultures, growth was coupled to nitrate reduction and acetate was formed in the stationary phase after nitrate was totally consumed. In the absence of nitrate, glyoxylate-grown cells incorporated less CO(2) into biomass than oxalate-grown cells. CO(2) conversion to biomass by glyoxylate-grown cells decreased when cells were grown in the presence of nitrate. These results suggest that: (1) oxalate-grown cells prefer CO(2) as an electron sink and bypass the nitrate block on the acetyl-CoA pathway at the level of reductant flow and (2) glyoxylate-grown cells prefer nitrate as an electron sink and bypass the nitrate block of the acetyl-CoA pathway by assimilating carbon via an unknown process that supplements or replaces the acetyl-CoA pathway. In this regard, enzymes of known pathways for the assimilation of two-carbon compounds were not detected in glyoxylate- or oxalate-grown cells.

  15. Thermodynamics and economic feasibility of acetone production from syngas using the thermophilic production host Moorella thermoacetica.

    PubMed

    Redl, Stephanie; Sukumara, Sumesh; Ploeger, Tom; Wu, Liang; Ølshøj Jensen, Torbjørn; Nielsen, Alex Toftgaard; Noorman, Henk

    2017-01-01

    Syngas fermentation is a promising option for the production of biocommodities due to its abundance and compatibility with anaerobic fermentation. Using thermophilic production strains in a syngas fermentation process allows recovery of products with low boiling point from the off-gas via condensation. In this study we analyzed the production of acetone from syngas with the hypothetical production host derived from Moorella thermoacetica in a bubble column reactor at 60 °C with respect to thermodynamic and economic feasibility. We determined the cost of syngas production from basic oxygen furnace (BOF) process gas, from natural gas, and from corn stover and identified BOF gas as an economically interesting source for syngas. Taking gas-liquid mass transfer limitations into account, we applied a thermodynamics approach to derive the CO to acetone conversion rate under the process conditions. We estimated variable costs of production of 389 $/t acetone for a representative production scenario from BOF gas with costs for syngas as the main contributor. In comparison, the variable costs of production from natural gas- and corn stover-derived syngas were determined to be higher due to the higher feedstock costs (1724 and 2878 $/t acetone, respectively). We applied an approach of combining thermodynamic and economic assessment to analyze a hypothetical bioprocess in which the volatile product acetone is produced from syngas with a thermophilic microorganism. Our model allowed us to identify process metrics and quantify the variable production costs for different scenarios. Economical production of bulk chemicals is challenging, making rigorous thermodynamic/economic modeling critical before undertaking an experimental program and as an ongoing guide during the program. We intend this study to give an incentive to apply the demonstrated approach to other bioproduction processes.

  16. Crystallization and preliminary X-ray analysis of RsbS from Moorella thermoacetica at 2.5 A resolution.

    PubMed

    Quin, Maureen; Newman, Joseph; Firbank, Susan; Lewis, Richard J; Marles-Wright, Jon

    2008-03-01

    The thermophilic bacterium Moorella thermoacetica possesses an rsb operon that is related to the genetic locus common to many Gram-positive bacteria that regulates the activity of the stress-responsive sigma factor sigma(B). One of the gene products of this operon is RsbS, a single STAS-domain protein that is a component of higher order assemblies in Bacillus subtilis known as 'stressosomes'. It is expected that similar complexes are found in M. thermoacetica, but in this instance regulating the biosynthesis of cyclic di-GMP, a ubiquitous secondary messenger. Selenomethionine-labelled MtRsbS protein was crystallized at room temperature using the hanging-drop vapour-diffusion method. Crystals belonging to space group P2(1)2(1)2(1), with unit-cell parameters a = 51.07, b = 60.52, c = 89.28 A, diffracted to 2.5 A resolution on beamline I04 of the Diamond Light Source. The selenium substructure was solved using SHELX and it is believed that this represents the first reported ab initio crystal structure to be solved using diffraction data collected at DLS.

  17. Tolerance and metabolic response of acetogenic bacteria toward oxygen.

    PubMed

    Karnholz, Arno; Küsel, Kirsten; Gössner, Anita; Schramm, Andreas; Drake, Harold L

    2002-02-01

    The acetogens Sporomusa silvacetica, Moorella thermoacetica, Clostridium magnum, Acetobacterium woodii, and Thermoanaerobacter kivui (i) grew in both semisolid and liquid cultivation media containing O(2) and (ii) consumed small amounts of O(2). Low concentrations of O(2) caused a lag phase in growth but did not alter the ability of these acetogens to synthesize acetate via the acetyl coenzyme A pathway. Cell extracts of S. silvacetica, M. thermoacetica, and C. magnum contained peroxidase and NADH oxidase activities; catalase and superoxide dismutase activities were not detected.

  18. The three-dimensional structure of the Moorella thermoacetica selenocysteine insertion sequence RNA hairpin and its interaction with the elongation factor SelB

    PubMed Central

    Beribisky, Alexander V.; Tavares, Tony J.; Amborski, Andrew N.; Motamed, Mina; Johnson, Anne E.; Mark, Tobi L.; Johnson, Philip E.

    2007-01-01

    Incorporation of the amino acid selenocysteine into a growing protein chain involves the interaction between a hairpin in the mRNA termed the selenocysteine insertion sequence (SECIS) and the special elongation factor SelB. Here we present the structure of the SECIS from the thermophilic organism Moorella thermoacetica (SECIS-MT) determined using nuclear magnetic resonance (NMR) spectroscopy. The SECIS-MT hairpin structure contains a pentaloop with the first and fourth nucleotides of the loop forming a noncanonical GC base pair; the fifth loop nucleotide is bulged out and unstructured. The G and U in positions two and three are on opposite sides of the loop and solvent exposed. The backbone resonances of the SECIS-binding domain from the M. thermoacetica SelB protein were assigned, and the degree of chemical shift perturbations that occur upon SECIS binding were mapped onto the structure of the complex. We demonstrate that a region in the third winged-helix domain of SelB, not previously implicated in binding, is affected by SECIS binding. PMID:17901155

  19. Electrosynthesis of Organic Compounds from Carbon Dioxide Is Catalyzed by a Diversity of Acetogenic Microorganisms▿

    PubMed Central

    Nevin, Kelly P.; Hensley, Sarah A.; Franks, Ashley E.; Summers, Zarath M.; Ou, Jianhong; Woodard, Trevor L.; Snoeyenbos-West, Oona L.; Lovley, Derek R.

    2011-01-01

    Microbial electrosynthesis, a process in which microorganisms use electrons derived from electrodes to reduce carbon dioxide to multicarbon, extracellular organic compounds, is a potential strategy for capturing electrical energy in carbon-carbon bonds of readily stored and easily distributed products, such as transportation fuels. To date, only one organism, the acetogen Sporomusa ovata, has been shown to be capable of electrosynthesis. The purpose of this study was to determine if a wider range of microorganisms is capable of this process. Several other acetogenic bacteria, including two other Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, consumed current with the production of organic acids. In general acetate was the primary product, but 2-oxobutyrate and formate also were formed, with 2-oxobutyrate being the predominant identified product of electrosynthesis by C. aceticum. S. sphaeroides, C. ljungdahlii, and M. thermoacetica had high (>80%) efficiencies of electrons consumed and recovered in identified products. The acetogen Acetobacterium woodii was unable to consume current. These results expand the known range of microorganisms capable of electrosynthesis, providing multiple options for the further optimization of this process. PMID:21378039

  20. Electrosynthesis of Organic Compounds from Carbon Dioxide Is Catalyzed by a Diversity of Acetogenic Microorganisms

    SciTech Connect

    Nevin, KP; Hensley, SA; Franks, AE; Summers, ZM; Ou, JH; Woodard, TL; Snoeyenbos-West, OL; Lovley, DR

    2011-04-20

    Microbial electrosynthesis, a process in which microorganisms use electrons derived from electrodes to reduce carbon dioxide to multicarbon, extracellular organic compounds, is a potential strategy for capturing electrical energy in carbon-carbon bonds of readily stored and easily distributed products, such as transportation fuels. To date, only one organism, the acetogen Sporomusa ovata, has been shown to be capable of electrosynthesis. The purpose of this study was to determine if a wider range of microorganisms is capable of this process. Several other acetogenic bacteria, including two other Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, consumed current with the production of organic acids. In general acetate was the primary product, but 2-oxobutyrate and formate also were formed, with 2-oxobutyrate being the predominant identified product of electrosynthesis by C. aceticum. S. sphaeroides, C. ljungdahlii, and M. thermoacetica had high (> 80%) efficiencies of electrons consumed and recovered in identified products. The acetogen Acetobacterium woodii was unable to consume current. These results expand the known range of microorganisms capable of electrosynthesis, providing multiple options for the further optimization of this process.

  1. Electrosynthesis of organic compounds from carbon dioxide is catalyzed by a diversity of acetogenic microorganisms.

    PubMed

    Nevin, Kelly P; Hensley, Sarah A; Franks, Ashley E; Summers, Zarath M; Ou, Jianhong; Woodard, Trevor L; Snoeyenbos-West, Oona L; Lovley, Derek R

    2011-05-01

    Microbial electrosynthesis, a process in which microorganisms use electrons derived from electrodes to reduce carbon dioxide to multicarbon, extracellular organic compounds, is a potential strategy for capturing electrical energy in carbon-carbon bonds of readily stored and easily distributed products, such as transportation fuels. To date, only one organism, the acetogen Sporomusa ovata, has been shown to be capable of electrosynthesis. The purpose of this study was to determine if a wider range of microorganisms is capable of this process. Several other acetogenic bacteria, including two other Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, consumed current with the production of organic acids. In general acetate was the primary product, but 2-oxobutyrate and formate also were formed, with 2-oxobutyrate being the predominant identified product of electrosynthesis by C. aceticum. S. sphaeroides, C. ljungdahlii, and M. thermoacetica had high (>80%) efficiencies of electrons consumed and recovered in identified products. The acetogen Acetobacterium woodii was unable to consume current. These results expand the known range of microorganisms capable of electrosynthesis, providing multiple options for the further optimization of this process.

  2. New generation NMR bioreactor coupled with high-resolution NMR spectroscopy leads to novel discoveries in Moorella thermoaceticum metabolic profiles

    SciTech Connect

    Xue, Junfeng; Isern, Nancy G.; Ewing, R James; Liyu, Andrey V.; Sears, Jesse A.; Knapp, Harlan; Iversen, Jens; Sisk, Daniel R.; Ahring, Birgitte K.; Majors, Paul D.

    2014-06-20

    An in-situ nuclear magnetic resonance (NMR) bioreactor was developed and employed to monitor microbial metabolism under batch-growth conditions in real time. We selected Moorella thermoacetica ATCC 49707 as a test case. M. thermoacetica (formerly Clostridium thermoaceticum) is a strictly anaerobic, thermophilic, acetogenic, gram-positive bacterium with potential for industrial production of chemicals. The metabolic profiles of M. thermoacetica were characterized during growth in batch mode on xylose (a component of lignocellulosic biomass) using the new generation NMR bioreactor in combination with high-resolution, high sensitivity NMR (HR-NMR) spectroscopy. In-situ NMR measurements were performed using water-suppressed H-1 NMR spectroscopy at an NMR frequency of 500 MHz, and aliquots of the bioreactor contents were taken for 600 MHz HR-NMR spectroscopy at specific intervals to confirm metabolite identifications and expand metabolite coverage. M. thermoacetica demonstrated the metabolic potential to produce formate, ethanol and methanol from xylose, in addition to its known capability of producing acetic acid. Real-time monitoring of bioreactor conditions showed a temporary pH decrease, with a concomitant increase in formic acid during exponential growth. Fermentation experiments performed outside of the magnet showed that the strong magnetic field employed for NMR detection did not significantly affect cell metabolism. Use of the in-situ NMR bioreactor facilitated monitoring of the fermentation process in real time, enabling identification of intermediate and end-point metabolites and their correlation with pH and biomass produced during culture growth. Real-time monitoring of culture metabolism using the NMR bioreactor in combination with the HR-NMR spectroscopy will allow optimization of the metabolism of microorganisms producing valuable bioproducts.

  3. Phenotypic and genotypic characterization of some Moorella sp. strains isolated from canned foods.

    PubMed

    Carlier, Jean-Philippe; Bedora-Faure, Marie

    2006-11-01

    Six anaerobic thermophilic strains isolated from various spoiled cans including fish soups and cooked meats were characterized using a polyphasic approach. These strains were closely related to Moorella thermoacetica or Moorella thermoautotrophica species. Except the spacer region between the 16S and the 23S rRNA genes, which exhibited two PCR profiles distinguishing both species, the genotypic and phylogenetic analyses grouped these isolates, the type strains, and all sequences of Moorella thermoacetica and Moorella thermoautotrophica species contained in the GenBank database within a unique cluster. Moreover, all 16S rDNA sequences shared two characteristic DNA fragments, which were highly specific of Moorella thermoacetica/Moorella thermoautotrophica strains. However, taken together, the phenotypic, physiological and genotypic methods were conflicting, and did not enable affiliation of the isolates with one or the other species. To our knowledge, this study represents the first report of characterization of Moorella species isolated from spoiled cans. These results and previous work, very strongly argue in favor of questioning the taxonomic status of the two species.

  4. Carbon Isotope Fractionation during Catabolism and Anabolism in Acetogenic Bacteria Growing on Different Substrates

    PubMed Central

    Freude, Christoph

    2016-01-01

    Homoacetogenic bacteria are versatile microbes that use the acetyl coenzyme A (acetyl-CoA) pathway to synthesize acetate from CO2 and hydrogen. Likewise, the acetyl-CoA pathway may be used to incorporate other 1-carbon substrates (e.g., methanol or formate) into acetate or to homoferment monosaccharides completely to acetate. In this study, we analyzed the fractionation of pure acetogenic cultures grown on different carbon substrates. While the fractionation of Sporomusa sphaeroides grown on C1 compounds was strong (εC1, −49‰ to −64‰), the fractionation of Moorella thermoacetica and Thermoanaerobacter kivui using glucose (εGlu = −14.1‰) was roughly one-third as strong, suggesting a contribution of less-depleted acetate from fermentative processes. For M. thermoacetica, this could indeed be validated by the addition of nitrate, which inhibited the acetyl-CoA pathway, resulting in fractionation during fermentation (εferm = −0.4‰). In addition, we determined the fractionation into microbial biomass of T. kivui grown on H2/CO2 (εanabol. = −28.6‰) as well as on glucose (εanabol. = +2.9‰). PMID:26921422

  5. Carbon Isotope Fractionation during Catabolism and Anabolism in Acetogenic Bacteria Growing on Different Substrates.

    PubMed

    Freude, Christoph; Blaser, Martin

    2016-05-01

    Homoacetogenic bacteria are versatile microbes that use the acetyl coenzyme A (acetyl-CoA) pathway to synthesize acetate from CO2 and hydrogen. Likewise, the acetyl-CoA pathway may be used to incorporate other 1-carbon substrates (e.g., methanol or formate) into acetate or to homoferment monosaccharides completely to acetate. In this study, we analyzed the fractionation of pure acetogenic cultures grown on different carbon substrates. While the fractionation of Sporomusa sphaeroides grown on C1 compounds was strong (εC1, -49‰ to -64‰), the fractionation of Moorella thermoacetica and Thermoanaerobacter kivui using glucose (εGlu= -14.1‰) was roughly one-third as strong, suggesting a contribution of less-depleted acetate from fermentative processes. ForM. thermoacetica, this could indeed be validated by the addition of nitrate, which inhibited the acetyl-CoA pathway, resulting in fractionation during fermentation (εferm= -0.4‰). In addition, we determined the fractionation into microbial biomass of T. kivui grown on H2/CO2(εanabol.= -28.6‰) as well as on glucose (εanabol.= +2.9‰). Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  6. The Complete Genome Sequence of Clostridium aceticum: a Missing Link between Rnf- and Cytochrome-Containing Autotrophic Acetogens

    PubMed Central

    Poehlein, Anja; Cebulla, Martin; Ilg, Marcus M.; Bengelsdorf, Frank R.; Schiel-Bengelsdorf, Bettina; Whited, Gregg; Andreesen, Jan R.; Gottschalk, Gerhard; Daniel, Rolf

    2015-01-01

    ABSTRACT Clostridium aceticum was the first isolated autotrophic acetogen, converting CO2 plus H2 or syngas to acetate. Its genome has now been completely sequenced and consists of a 4.2-Mbp chromosome and a small circular plasmid of 5.7 kbp. Sequence analysis revealed major differences from other autotrophic acetogens. C. aceticum contains an Rnf complex for energy conservation (via pumping protons or sodium ions). Such systems have also been found in C. ljungdahlii and Acetobacterium woodii. However, C. aceticum also contains a cytochrome, as does Moorella thermoacetica, which has been proposed to be involved in the generation of a proton gradient. Thus, C. aceticum seems to represent a link between Rnf- and cytochrome-containing autotrophic acetogens. In C. aceticum, however, the cytochrome is probably not involved in an electron transport chain that leads to proton translocation, as no genes for quinone biosynthesis are present in the genome. PMID:26350967

  7. (Per)chlorate reduction by the thermophilic bacterium Moorella perchloratireducens sp. nov., isolated from underground gas storage.

    PubMed

    Balk, Melike; van Gelder, Ton; Weelink, Sander A; Stams, Alfons J M

    2008-01-01

    A thermophilic bacterium, strain An10, was isolated from underground gas storage with methanol as a substrate and perchlorate as an electron acceptor. Cells were gram-positive straight rods, 0.4 to 0.6 mum in diameter and 2 to 8 mum in length, growing as single cells or in pairs. Spores were terminal with a bulged sporangium. The temperature range for growth was 40 to 70 degrees C, with an optimum at 55 to 60 degrees C. The pH optimum was around 7. The salinity range for growth was between 0 and 40 g NaCl liter(-1) with an optimum at 10 g liter(-1). Strain An10 was able to grow on CO, methanol, pyruvate, glucose, fructose, cellobiose, mannose, xylose, and pectin. The isolate was able to respire with (per)chlorate, nitrate, thiosulfate, neutralized Fe(III) complexes, and anthraquinone-2,6-disulfonate. The G+C content of the DNA was 57.6 mol%. On the basis of 16S rRNA analysis, strain An10 was most closely related to Moorella thermoacetica and Moorella thermoautotrophica. The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell extracts. Strain An10 is the first thermophilic and gram-positive bacterium with the ability to use (per)chlorate as a terminal electron acceptor.

  8. Carbon Isotope Fractionation of 11 Acetogenic Strains Grown on H2 and CO2

    PubMed Central

    Dreisbach, Lisa K.; Conrad, Ralf

    2013-01-01

    Acetogenic bacteria are able to grow autotrophically on hydrogen and carbon dioxide by using the acetyl coenzyme A (acetyl-CoA) pathway. Acetate is the end product of this reaction. In contrast to the fermentative route of acetate production, which shows almost no fractionation of carbon isotopes, the acetyl-CoA pathway has been reported to exhibit a preference for light carbon. In Acetobacterium woodii the isotope fractionation factor (ε) for 13C and 12C has previously been reported to be ε = −58.6‰. To investigate whether such a strong fractionation is a general feature of acetogenic bacteria, we measured the stable carbon isotope fractionation factor of 10 acetogenic strains grown on H2 and CO2. The average fractionation factor was εTIC = −57.2‰ for utilization of total inorganic carbon and εacetate = −54.6‰ for the production of acetate. The strongest fractionation was found for Sporomusa sphaeroides (εTIC = −68.3‰), the lowest fractionation for Morella thermoacetica (εTIC = −38.2‰). To investigate the reproducibility of our measurements, we determined the fractionation factor of 21 biological replicates of Thermoanaerobacter kivui. In general, our study confirmed the strong fractionation of stable carbon during chemolithotrophic acetate formation in acetogenic bacteria. However, the specific characteristics of the bacterial strain, as well as the cultural conditions, may have a moderate influence on the overall fractionation. PMID:23275504

  9. Rumen fermentation and acetogen population changes in response to an exogenous acetogen TWA4 strain and Saccharomyces cerevisiae fermentation product.

    PubMed

    Yang, Chun-lei; Guan, Le-luo; Liu, Jian-xin; Wang, Jia-kun

    2015-08-01

    The presence of yeast cells could stimulate hydrogen utilization of acetogens and enhance acetogenesis. To understand the roles of acetogens in rumen fermentation, an in vitro rumen fermentation experiment was conducted with addition of acetogen strain (TWA4) and/or Saccharomyces cerevisiae fermentation product (XP). A 2×2 factorial design with two levels of TWA4 (0 or 2×10(7) cells/ml) and XP (0 or 2 g/L) was performed. Volatile fatty acids (VFAs) were increased (P<0.05) in XP and TWA4XP, while methane was increased only in TWA4XP (P<0.05). The increase rate of microorganisms with formyltetrahydrofolate synthetase, especially acetogens, was higher than that of methanogens under all treatments. Lachnospiraceae was predominant in all acetogen communities, but without close acetyl-CoA synthase (ACS) amino acid sequences from cultured isolates. Low-Acetitomaculum ruminis-like ACS was predominant in all acetogen communities, while four unique phylotypes in XP treatment were all amino acid identified low-Eubacterium limosum-like acetogens. It differs to XP treatment that more low-A. ruminis-like and less low-E. limosum-like sequences were identified in TWA4 and TWA4XP treatments. Enhancing acetogenesis by supplementation with an acetogen strain and/or yeast cells may be an approach to mitigate methane, by targeting proper acetogens such as uncultured low-E. limosum-like acetogens.

  10. Rumen fermentation and acetogen population changes in response to an exogenous acetogen TWA4 strain and Saccharomyces cerevisiae fermentation product*

    PubMed Central

    Yang, Chun-lei; Guan, Le-luo; Liu, Jian-xin; Wang, Jia-kun

    2015-01-01

    The presence of yeast cells could stimulate hydrogen utilization of acetogens and enhance acetogenesis. To understand the roles of acetogens in rumen fermentation, an in vitro rumen fermentation experiment was conducted with addition of acetogen strain (TWA4) and/or Saccharomyces cerevisiae fermentation product (XP). A 2×2 factorial design with two levels of TWA4 (0 or 2×107 cells/ml) and XP (0 or 2 g/L) was performed. Volatile fatty acids (VFAs) were increased (P<0.05) in XP and TWA4XP, while methane was increased only in TWA4XP (P<0.05). The increase rate of microorganisms with formyltetrahydrofolate synthetase, especially acetogens, was higher than that of methanogens under all treatments. Lachnospiraceae was predominant in all acetogen communities, but without close acetyl-CoA synthase (ACS) amino acid sequences from cultured isolates. Low-Acetitomaculum ruminis-like ACS was predominant in all acetogen communities, while four unique phylotypes in XP treatment were all amino acid identified low-Eubacterium limosum-like acetogens. It differs to XP treatment that more low-A. ruminis-like and less low-E. limosum-like sequences were identified in TWA4 and TWA4XP treatments. Enhancing acetogenesis by supplementation with an acetogen strain and/or yeast cells may be an approach to mitigate methane, by targeting proper acetogens such as uncultured low-E. limosum-like acetogens. PMID:26238546

  11. Energetics and Application of Heterotrophy in Acetogenic Bacteria

    PubMed Central

    Schuchmann, Kai

    2016-01-01

    Acetogenic bacteria are a diverse group of strictly anaerobic bacteria that utilize the Wood-Ljungdahl pathway for CO2 fixation and energy conservation. These microorganisms play an important part in the global carbon cycle and are a key component of the anaerobic food web. Their most prominent metabolic feature is autotrophic growth with molecular hydrogen and carbon dioxide as the substrates. However, most members also show an outstanding metabolic flexibility for utilizing a vast variety of different substrates. In contrast to autotrophic growth, which is hardly competitive, metabolic flexibility is seen as a key ability of acetogens to compete in ecosystems and might explain the almost-ubiquitous distribution of acetogenic bacteria in anoxic environments. This review covers the latest findings with respect to the heterotrophic metabolism of acetogenic bacteria, including utilization of carbohydrates, lactate, and different alcohols, especially in the model acetogen Acetobacterium woodii. Modularity of metabolism, a key concept of pathway design in synthetic biology, together with electron bifurcation, to overcome energetic barriers, appears to be the basis for the amazing substrate spectrum. At the same time, acetogens depend on only a relatively small number of enzymes to expand the substrate spectrum. We will discuss the energetic advantages of coupling CO2 reduction to fermentations that exploit otherwise-inaccessible substrates and the ecological advantages, as well as the biotechnological applications of the heterotrophic metabolism of acetogens. PMID:27208103

  12. Energetics and Application of Heterotrophy in Acetogenic Bacteria.

    PubMed

    Schuchmann, Kai; Müller, Volker

    2016-07-15

    Acetogenic bacteria are a diverse group of strictly anaerobic bacteria that utilize the Wood-Ljungdahl pathway for CO2 fixation and energy conservation. These microorganisms play an important part in the global carbon cycle and are a key component of the anaerobic food web. Their most prominent metabolic feature is autotrophic growth with molecular hydrogen and carbon dioxide as the substrates. However, most members also show an outstanding metabolic flexibility for utilizing a vast variety of different substrates. In contrast to autotrophic growth, which is hardly competitive, metabolic flexibility is seen as a key ability of acetogens to compete in ecosystems and might explain the almost-ubiquitous distribution of acetogenic bacteria in anoxic environments. This review covers the latest findings with respect to the heterotrophic metabolism of acetogenic bacteria, including utilization of carbohydrates, lactate, and different alcohols, especially in the model acetogen Acetobacterium woodii Modularity of metabolism, a key concept of pathway design in synthetic biology, together with electron bifurcation, to overcome energetic barriers, appears to be the basis for the amazing substrate spectrum. At the same time, acetogens depend on only a relatively small number of enzymes to expand the substrate spectrum. We will discuss the energetic advantages of coupling CO2 reduction to fermentations that exploit otherwise-inaccessible substrates and the ecological advantages, as well as the biotechnological applications of the heterotrophic metabolism of acetogens.

  13. Limitation of syntrophic coculture growth by the acetogen.

    PubMed

    Junicke, Helena; Feldman, Hannah; Van Loosdrecht, Mark C M; Kleerebezem, Robbert

    2016-03-01

    The syntrophic cooperation between hydrogen-producing acetogens and hydrogenotrophic methanogens relies on a critical balance between both partners. A recent study, provided several indications for the dependence of the biomass-specific growth rate of a methanogenic coculture on the acetogen. Nevertheless, final experimental proof was lacking since biomass-specific rates were obtained from a descriptive model, and not from direct measurement of individual biomass concentrations. In this study, a recently developed quantitative PCR approach was used to measure the individual biomass concentrations in the coculture of Desulfovibrio sp. G11 and Methanospirillum hungatei JF1 on lactate, formate or both. The model-derived growth yields and biomass-specific rates were successfully validated. Experimental findings identified the acetogen as the growth-limiting partner in the coculture on lactate. While the acetogen was operating at its maximum biomass-specific lactate consumption rate, the hydrogenotrophic methanogen showed a significant overcapacity. Furthermore, this study provides experimental evidence for different growth strategies followed by the syntrophic partners in order to maintain a common biomass-specific growth rate. During syntrophic lactate conversion, the biomass-specific electron transfer rate of Methanospirillum hungatei JF1 was three-fold higher compared to Desulfovibrio sp. G11. This is to compensate for the lower methanogenic biomass yield per electron-mole of substrate, which is dictated by the thermodynamics of the underlying reaction. © 2015 Wiley Periodicals, Inc.

  14. General medium for the autotrophic cultivation of acetogens.

    PubMed

    Groher, Anna; Weuster-Botz, Dirk

    2016-10-01

    Syngas fermentation, a microbial process in which synthesis gas serves as a substrate for acetogens, has attracted increasing interest in the last few years. For the purposeful selection of acetogens for various applications, it would be useful to characterize and compare the process performances of as many autotrophic strains as possible under identical process conditions. Unfortunately, all the media compositions so far recommended for syngas fermentation differ considerably with respect to each individual strain. Therefore, a general medium for syngas fermentation was designed. The suitability of this new general-acetogen medium (GA-medium) was proven based on the autotrophic batch cultivation of Acetobacterium fimetarium, Acetobacterium wieringae, Blautia hydrogenotrophica, Clostridium magnum, Eubacterium aggregans, Sporomusa acidovorans, Sporomusa ovata and Terrisporobacter mayombei in anaerobic flasks with an initial gas phase of H2:CO2 (66:34) (P = 200 kPa). A comparison of the autotrophic batch processes with this medium revealed T. mayombei as the bacterium with the highest maximum growth rate of 5.77 day(-1) which was more than 10 times higher than the lowest identified maximum growth rate of A. fimetarium. The maximum growth rates of A. wieringae, C. magnum and S. acidovorans were all in the same order of magnitude around 1.7 day(-1). The newly designed GA-medium offers the possibility to compare autotrophic process performances of different acetogens under similar conditions absent the effects of various media compositions.

  15. Ethylene Glycol Metabolism in the Acetogen Acetobacterium woodii

    PubMed Central

    Trifunović, Dragan; Schuchmann, Kai

    2016-01-01

    ABSTRACT The acetogenic bacterium Acetobacterium woodii is able to grow by the oxidation of diols, such as 1,2-propanediol, 2,3-butanediol, or ethylene glycol. Recent analyses demonstrated fundamentally different ways for oxidation of 1,2-propanediol and 2,3-butanediol. Here, we analyzed the metabolism of ethylene glycol. Our data demonstrate that ethylene glycol is dehydrated to acetaldehyde, which is then disproportionated to ethanol and acetyl coenzyme A (acetyl-CoA). The latter is further converted to acetate, and this pathway is coupled to ATP formation by substrate-level phosphorylation. Apparently, the product ethanol is in part further oxidized and the reducing equivalents are recycled by reduction of CO2 to acetate in the Wood-Ljungdahl pathway. Biochemical data as well as the results of protein synthesis analysis are consistent with the hypothesis that the propane diol dehydratase (PduCDE) and CoA-dependent propionaldehyde dehydrogenase (PduP) proteins, encoded by the pdu gene cluster, also catalyze ethylene glycol dehydration to acetaldehyde and its CoA-dependent oxidation to acetyl-CoA. Moreover, genes encoding bacterial microcompartments as part of the pdu gene cluster are also expressed during growth on ethylene glycol, arguing for a dual function of the Pdu microcompartment system. IMPORTANCE Acetogenic bacteria are characterized by their ability to use CO2 as a terminal electron acceptor by a specific pathway, the Wood-Ljungdahl pathway, enabling in most acetogens chemolithoautotrophic growth with H2 and CO2. However, acetogens are very versatile and can use a wide variety of different substrates for growth. Here we report on the elucidation of the pathway for utilization of ethylene glycol by the model acetogen Acetobacterium woodii. This diol is degraded by dehydration to acetaldehyde followed by a disproportionation to acetate and ethanol. We present evidence that this pathway is catalyzed by the same enzyme system recently described for the

  16. Ethylene Glycol Metabolism in the Acetogen Acetobacterium woodii.

    PubMed

    Trifunović, Dragan; Schuchmann, Kai; Müller, Volker

    2016-01-19

    The acetogenic bacterium Acetobacterium woodii is able to grow by the oxidation of diols, such as 1,2-propanediol, 2,3-butanediol, or ethylene glycol. Recent analyses demonstrated fundamentally different ways for oxidation of 1,2-propanediol and 2,3-butanediol. Here, we analyzed the metabolism of ethylene glycol. Our data demonstrate that ethylene glycol is dehydrated to acetaldehyde, which is then disproportionated to ethanol and acetyl coenzyme A (acetyl-CoA). The latter is further converted to acetate, and this pathway is coupled to ATP formation by substrate-level phosphorylation. Apparently, the product ethanol is in part further oxidized and the reducing equivalents are recycled by reduction of CO2 to acetate in the Wood-Ljungdahl pathway. Biochemical data as well as the results of protein synthesis analysis are consistent with the hypothesis that the propane diol dehydratase (PduCDE) and CoA-dependent propionaldehyde dehydrogenase (PduP) proteins, encoded by the pdu gene cluster, also catalyze ethylene glycol dehydration to acetaldehyde and its CoA-dependent oxidation to acetyl-CoA. Moreover, genes encoding bacterial microcompartments as part of the pdu gene cluster are also expressed during growth on ethylene glycol, arguing for a dual function of the Pdu microcompartment system. Acetogenic bacteria are characterized by their ability to use CO2 as a terminal electron acceptor by a specific pathway, the Wood-Ljungdahl pathway, enabling in most acetogens chemolithoautotrophic growth with H2 and CO2. However, acetogens are very versatile and can use a wide variety of different substrates for growth. Here we report on the elucidation of the pathway for utilization of ethylene glycol by the model acetogen Acetobacterium woodii. This diol is degraded by dehydration to acetaldehyde followed by a disproportionation to acetate and ethanol. We present evidence that this pathway is catalyzed by the same enzyme system recently described for the utilization of 1

  17. Investigation into Host Selection of the Cecal Acetogen Population in Rabbits after Weaning

    PubMed Central

    Yang, Chunlei; Mi, Lan; Hu, Xialu; Liu, Jianxin

    2016-01-01

    Homoacetogenic bacteria have received attention as a hydrogenotrophic population that offers a significant energetic advantage to the host animal. Reductive acetogenesis is likely an important hydrogen disposal mechanism in the cecum of rabbits. However, molecular ecology information about cecal acetogen candidates has rarely been reported. To better understand the effect of host selection in the rabbit cecal acetogen community with respect to growth, rabbits at four different age stages (30, 60, 120 and 180 days) with the same diet were studied. Although the abundance of potential acetogens and methanogens was high in the cecum of rabbits undergoing growth, many novel potential acetogen populations were observed in the cecum of rabbits across all age groups. Young and adult rabbits had their own distinct acetogen community although they received the same diet, which suggests that as the rabbit ages, acetogens in the cecum undergo developmental changes because of host selection that are independent of diet, and perhaps the different acetogen communities result in different hydrogenotrophic characteristics. The within-group similarity increased with age, indicating that the acetogen community converges to a more homogeneous and stable arrangement with aging. PMID:27379387

  18. Analysis of the Core Genome and Pan-Genome of Autotrophic Acetogenic Bacteria

    PubMed Central

    Shin, Jongoh; Song, Yoseb; Jeong, Yujin; Cho, Byung-Kwan

    2016-01-01

    Acetogens are obligate anaerobic bacteria capable of reducing carbon dioxide (CO2) to multicarbon compounds coupled to the oxidation of inorganic substrates, such as hydrogen (H2) or carbon monoxide (CO), via the Wood-Ljungdahl pathway. Owing to the metabolic capability of CO2 fixation, much attention has been focused on understanding the unique pathways associated with acetogens, particularly their metabolic coupling of CO2 fixation to energy conservation. Most known acetogens are phylogenetically and metabolically diverse bacteria present in 23 different bacterial genera. With the increased volume of available genome information, acetogenic bacterial genomes can be analyzed by comparative genome analysis. Even with the genetic diversity that exists among acetogens, the Wood-Ljungdahl pathway, a central metabolic pathway, and cofactor biosynthetic pathways are highly conserved for autotrophic growth. Additionally, comparative genome analysis revealed that most genes in the acetogen-specific core genome were associated with the Wood-Ljungdahl pathway. The conserved enzymes and those predicted as missing can provide insight into biological differences between acetogens and allow for the discovery of promising candidates for industrial applications. PMID:27733845

  19. Autotrophy at the thermodynamic limit of life: a model for energy conservation in acetogenic bacteria.

    PubMed

    Schuchmann, Kai; Müller, Volker

    2014-12-01

    Life on earth evolved in the absence of oxygen with inorganic gases as potential sources of carbon and energy. Among the alternative mechanisms for carbon dioxide (CO₂) fixation in the living world, only the reduction of CO₂ by the Wood-Ljungdahl pathway, which is used by acetogenic bacteria, complies with the two requirements to sustain life: conservation of energy and production of biomass. However, how energy is conserved in acetogenic bacteria has been an enigma since their discovery. In this Review, we discuss the latest progress on the biochemistry and genetics of the energy metabolism of model acetogens, elucidating how these bacteria couple CO₂ fixation to energy conservation.

  20. 2,3-Butanediol Metabolism in the Acetogen Acetobacterium woodii

    PubMed Central

    Hess, Verena; Oyrik, Olga; Trifunović, Dragan

    2015-01-01

    The acetogenic bacterium Acetobacterium woodii is able to reduce CO2 to acetate via the Wood-Ljungdahl pathway. Only recently we demonstrated that degradation of 1,2-propanediol by A. woodii was not dependent on acetogenesis, but that it is disproportionated to propanol and propionate. Here, we analyzed the metabolism of A. woodii on another diol, 2,3-butanediol. Experiments with growing and resting cells, metabolite analysis and enzymatic measurements revealed that 2,3-butanediol is oxidized in an NAD+-dependent manner to acetate via the intermediates acetoin, acetaldehyde, and acetyl coenzyme A. Ethanol was not detected as an end product, either in growing cultures or in cell suspensions. Apparently, all reducing equivalents originating from the oxidation of 2,3-butanediol were funneled into the Wood-Ljungdahl pathway to reduce CO2 to another acetate. Thus, the metabolism of 2,3-butanediol requires the Wood-Ljungdahl pathway. PMID:25934628

  1. Bioenergetic constraints for conversion of syngas to biofuels in acetogenic bacteria.

    PubMed

    Bertsch, Johannes; Müller, Volker

    2015-01-01

    Synthesis gas (syngas) is a gas mixture consisting mainly of H2, CO, and CO2 and can be derived from different sources, including renewable materials like lignocellulose. The fermentation of syngas to certain biofuels, using acetogenic bacteria, has attracted more and more interest over the last years. However, this technology is limited by two things: (1) the lack of complete knowledge of the energy metabolism of acetogenic bacteria, and (2) the lack of sophisticated genetic tools for the modification of acetogens. In this review, we discuss the bioenergetic constraints for the conversion of syngas to different biofuels. We will mainly focus on Acetobacterium woodii, which is the best understood acetogen in terms of energy conservation. Syngas fermentation with Clostridium autoethanogenum will also be discussed, since this organism is well suited to convert syngas to certain products and already used in large-scale industrial processes.

  2. Thermophilic Moorella thermoautotrophica-immobilized cathode enhanced microbial electrosynthesis of acetate and formate from CO2.

    PubMed

    Yu, Linpeng; Yuan, Yong; Tang, Jiahuan; Zhou, Shungui

    2017-10-01

    Microbial electrosynthesis (MES) is a promising technique that converts electricity and CO2 to biofuels using microbes as the catalysts. However, most of previous MES are conducted at mesophilic temperatures and challenged by low performances. Here we report a significant electrosynthesis performance enhancement via immobilization of a thermophilic microbe to cathodes. A temperature-dependent electron uptake rate of Moorella thermoautotrophica was observed at a cathode potential of -0.4V (vs. SHE), with a maximum current density of 63.47mAm(-2) at 55°C. Moreover, electrosynthesis rates of formate and acetate at 55°C were accelerated by 23.2 and 2.8 fold than those of 25°C, respectively. Compared with natural biofilms, immobilization of M. thermoautotrophica with carbon nanoparticles to electrodes further enhanced acetate and formate production rates (by 14 and 7.9 fold), reaching 58.2 and 63.2mmolm(-2)day(-1) at a coulombic efficiency of 65%, respectively. To our best knowledge, these are the highest electrosynthesis rates obtained thus far for pure cultures under the conditions of -0.4V (vs. SHE) and 55°C. This study, for the first time, demonstrates that embedding microbes to electrodes by carbon nanoparticles is a facile and efficient method of improving MES performance. Copyright © 2017. Published by Elsevier B.V.

  3. Industrial Acetogenic Biocatalysts: A Comparative Metabolic and Genomic Analysis

    PubMed Central

    Bengelsdorf, Frank R.; Poehlein, Anja; Linder, Sonja; Erz, Catarina; Hummel, Tim; Hoffmeister, Sabrina; Daniel, Rolf; Dürre, Peter

    2016-01-01

    Synthesis gas (syngas) fermentation by anaerobic acetogenic bacteria employing the Wood–Ljungdahl pathway is a bioprocess for production of biofuels and biocommodities. The major fermentation products of the most relevant biocatalytic strains (Clostridium ljungdahlii, C. autoethanogenum, C. ragsdalei, and C. coskatii) are acetic acid and ethanol. A comparative metabolic and genomic analysis using the mentioned biocatalysts might offer targets for metabolic engineering and thus improve the production of compounds apart from ethanol. Autotrophic growth and product formation of the four wild type (WT) strains were compared in uncontrolled batch experiments. The genomes of C. ragsdalei and C. coskatii were sequenced and the genome sequences of all four biocatalytic strains analyzed in comparative manner. Growth and product spectra (acetate, ethanol, 2,3-butanediol) of C. autoethanogenum, C. ljungdahlii, and C. ragsdalei were rather similar. In contrast, C. coskatii produced significantly less ethanol and its genome sequence lacks two genes encoding aldehyde:ferredoxin oxidoreductases (AOR). Comparative genome sequence analysis of the four WT strains revealed high average nucleotide identity (ANI) of C. ljungdahlii and C. autoethanogenum (99.3%) and C. coskatii (98.3%). In contrast, C. ljungdahlii WT and C. ragsdalei WT showed an ANI-based similarity of only 95.8%. Additionally, recombinant C. ljungdahlii strains were constructed that harbor an artificial acetone synthesis operon (ASO) consisting of the following genes: adc, ctfA, ctfB, and thlA (encoding acetoacetate decarboxylase, acetoacetyl-CoA:acetate/butyrate:CoA-transferase subunits A and B, and thiolase) under the control of thlA promoter (PthlA) from C. acetobutylicum or native pta-ack promoter (Ppta-ack) from C. ljungdahlii. Respective recombinant strains produced 2-propanol rather than acetone, due to the presence of a NADPH-dependent primary-secondary alcohol dehydrogenase that converts acetone to 2

  4. CO Metabolism in the Thermophilic Acetogen Thermoanaerobacter kivui

    PubMed Central

    Weghoff, Marie Charlotte

    2016-01-01

    The thermophilic acetogenic bacterium Thermoanaerobacter kivui, previously described not to use carbon monoxide as a carbon and energy source, was adapted to grow on CO. This was achieved by using a preculture grown on H2 plus CO2 and by increasing the CO concentration in small, 10% increments. T. kivui was finally able to grow within a 100% CO atmosphere. Growth on CO was found in complex and mineral media, and vitamins were not required. Carbon monoxide consumption was accompanied by acetate and hydrogen production. Cells also grew on synthesis gas (syngas) with the simultaneous use of CO and H2 coupled to acetate production. CO oxidation in resting cells was coupled to hydrogen and acetate production and accompanied by the synthesis of ATP. A protonophore abolished ATP synthesis but stimulated H2 production, which is consistent with a chemiosmotic mechanism of ATP synthesis. Hydrogenase activity was highest in crude extracts of CO-grown cells, and carbon monoxide dehydrogenase (CODH) activity was highest in H2-plus-CO2- or CO-grown cells. The genome of T. kivui harbors two CODH gene clusters, and both CODH proteins were present in crude extracts, but one CODH was more prevalent in crude extracts from CO-grown cells. PMID:26850300

  5. Hydrogen consumption in microbial electrochemical systems (MXCs): the role of homo-acetogenic bacteria.

    PubMed

    Parameswaran, Prathap; Torres, César I; Lee, Hyung-Sool; Rittmann, Bruce E; Krajmalnik-Brown, Rosa

    2011-01-01

    Homo-acetogens in the anode of a microbial electrolysis cell (MEC) fed with H(2) as sole electron donor allowed current densities similar to acetate-fed biofilm anodes (∼10 A/m(2)). Evidence for homo-acetogens included accumulation of acetate at high concentrations (up to 18 mM) in the anode compartment; detection of formate, a known intermediate during reductive acetogenesis by the acetyl-CoA pathway; and detection of formyl tetrahydrofolate synthetase (FTHFS) genes by quantitative real-time PCR. Current production and acetate accumulation increased in parallel in batch and continuous mode, while both values decreased simultaneously at short hydraulic retention times (1h) in the anode compartment, which limited suspended homo-acetogens. Acetate produced by homo-acetogens accounted for about 88% of the current density of 10A/m(2), but the current density was sustained at 4A/m(2) at short hydraulic retention time because of a robust partnership of homo-acetogens and anode respiring bacteria (ARB) in the biofilm anode.

  6. Enrichment of acetogenic bacteria in high rate anaerobic reactors under mesophilic and thermophilic conditions.

    PubMed

    Ryan, P; Forbes, C; McHugh, S; O'Reilly, C; Fleming, G T A; Colleran, E

    2010-07-01

    The objective of the current study was to expand the knowledge of the role of acetogenic Bacteria in high rate anaerobic digesters. To this end, acetogens were enriched by supplying a variety of acetogenic growth supportive substrates to two laboratory scale high rate upflow anaerobic sludge bed (UASB) reactors operated at 37 degrees C (R1) and 55 degrees C (R2). The reactors were initially fed a glucose/acetate influent. Having achieved high operational performance and granular sludge development and activity, both reactors were changed to homoacetogenic bacterial substrates on day 373 of the trial. The reactors were initially fed with sodium vanillate as a sole substrate. Although % COD removal indicated that the 55 degrees C reactor out performed the 37 degrees C reactor, effluent acetate levels from R2 were generally higher than from R1, reaching values as high as 5023 mg l(-1). Homoacetogenic activity in both reactors was confirmed on day 419 by specific acetogenic activity (SAA) measurement, with higher values obtained for R2 than R1. Sodium formate was introduced as sole substrate to both reactors on day 464. It was found that formate supported acetogenic activity at both temperatures. By the end of the trial, no specific methanogenic activity (SMA) was observed against acetate and propionate indicating that the methane produced was solely by hydrogenotrophic Archaea. Higher SMA and SAA values against H(2)/CO(2) suggested development of a formate utilising acetogenic population growing in syntrophy with hydrogenotrophic methanogens. Throughout the formate trial, the mesophilic reactor performed better overall than the thermophilic reactor.

  7. Isolation of Acetogenic Bacteria That Induce Biocorrosion by Utilizing Metallic Iron as the Sole Electron Donor

    PubMed Central

    Yumoto, Isao; Kamagata, Yoichi

    2014-01-01

    Corrosion of iron occurring under anoxic conditions, which is termed microbiologically influenced corrosion (MIC) or biocorrosion, is mostly caused by microbial activities. Microbial activity that enhances corrosion via uptake of electrons from metallic iron [Fe(0)] has been regarded as one of the major causative factors. In addition to sulfate-reducing bacteria and methanogenic archaea in marine environments, acetogenic bacteria in freshwater environments have recently been suggested to cause MIC under anoxic conditions. However, no microorganisms that perform acetogenesis-dependent MIC have been isolated or had their MIC-inducing mechanisms characterized. Here, we enriched and isolated acetogenic bacteria that induce iron corrosion by utilizing Fe(0) as the sole electron donor under freshwater, sulfate-free, and anoxic conditions. The enriched communities produced significantly larger amounts of Fe(II) than the abiotic controls and produced acetate coupled with Fe(0) oxidation prior to CH4 production. Microbial community analysis revealed that Sporomusa sp. and Desulfovibrio sp. dominated in the enrichments. Strain GT1, which is closely related to the acetogen Sporomusa sphaeroides, was eventually isolated from the enrichment. Strain GT1 grew acetogenetically with Fe(0) as the sole electron donor and enhanced iron corrosion, which is the first demonstration of MIC mediated by a pure culture of an acetogen. Other well-known acetogenic bacteria, including Sporomusa ovata and Acetobacterium spp., did not grow well on Fe(0). These results indicate that very few species of acetogens have specific mechanisms to efficiently utilize cathodic electrons derived from Fe(0) oxidation and induce iron corrosion. PMID:25304512

  8. Isolation of acetogenic bacteria that induce biocorrosion by utilizing metallic iron as the sole electron donor.

    PubMed

    Kato, Souichiro; Yumoto, Isao; Kamagata, Yoichi

    2015-01-01

    Corrosion of iron occurring under anoxic conditions, which is termed microbiologically influenced corrosion (MIC) or biocorrosion, is mostly caused by microbial activities. Microbial activity that enhances corrosion via uptake of electrons from metallic iron [Fe(0)] has been regarded as one of the major causative factors. In addition to sulfate-reducing bacteria and methanogenic archaea in marine environments, acetogenic bacteria in freshwater environments have recently been suggested to cause MIC under anoxic conditions. However, no microorganisms that perform acetogenesis-dependent MIC have been isolated or had their MIC-inducing mechanisms characterized. Here, we enriched and isolated acetogenic bacteria that induce iron corrosion by utilizing Fe(0) as the sole electron donor under freshwater, sulfate-free, and anoxic conditions. The enriched communities produced significantly larger amounts of Fe(II) than the abiotic controls and produced acetate coupled with Fe(0) oxidation prior to CH4 production. Microbial community analysis revealed that Sporomusa sp. and Desulfovibrio sp. dominated in the enrichments. Strain GT1, which is closely related to the acetogen Sporomusa sphaeroides, was eventually isolated from the enrichment. Strain GT1 grew acetogenetically with Fe(0) as the sole electron donor and enhanced iron corrosion, which is the first demonstration of MIC mediated by a pure culture of an acetogen. Other well-known acetogenic bacteria, including Sporomusa ovata and Acetobacterium spp., did not grow well on Fe(0). These results indicate that very few species of acetogens have specific mechanisms to efficiently utilize cathodic electrons derived from Fe(0) oxidation and induce iron corrosion.

  9. Isolation and characterization of the homoacetogenic thermophilic bacterium Moorella glycerini sp. nov.

    SciTech Connect

    Slobodkin, A.; Wiegel, J.; Reysenbach, A.L.

    1997-10-01

    A thermophilic, anaerobic, spore-forming bacterium (strain JW/AS-Y6) was isolated from a mixed sediment-water sample from a hot spring (Calcite Spring area) at Yellowstone National Park. The vegetative cells of this organism were straight rods, 0.5 to 0.6 by 3.0 to 6.5 {mu}m. Cells occurred singly and exhibited a slight tumbling motility. They formed round refractile endospores in terminal swollen sporangia. Cells stained gram positive. The temperature range for growth at pH 6.8 was 43 to 65{degrees}C, with optimum growth at 58{degrees}C. The range for growth at 60{degrees}C (pH{sup 60C}; with the pH meter calibrated at 60{degrees}C) was 5.9 to 7.8, with an optimum pH{sub 60C} of 6.3 to 6.5. The substrates utilized included glycerol, glucose, fructose, mannose, galactose, xylose, lactate, glycerate, pyruvate, and yeast extract. In the presence of CO{sub 2}, acetate was the only organic product from glyerol and carbohydrate fermentation. No H{sub 2} was produced during growth. The strain was not able to grow chemolithotrophically at the expense of H{sub 2}-CO{sub 2}; however, suspensions of cells in the exponential growth phase consumed H{sub 2}. The bacterium reduced fumarate to succinate and thiosulfate to elemental sulfur. Growth was exhibited by ampicillin, chloramphenicol, erythromycin, rifampin, and tetracycline, but not by streptomycin. The G+C content of the DNA was 54.5 mol% (as determined by high-performance liquid chromatography). The 16S ribosomal DNA sequence analysis placed the isolate in the Gram type-positive Bacillus-Clostridium subphylum. On the basis of physiological properties and phylogenetic analysis we propose that the isolated strain constitutes a new species, Moorella glycerini; the type strain is JW/AS-Y6 (= DSM 11254 = ATCC 700316).

  10. Complete genome sequence of a carbon monoxide-utilizing acetogen, Eubacterium limosum KIST612.

    PubMed

    Roh, Hanseong; Ko, Hyeok-Jin; Kim, Daehee; Choi, Dong Geon; Park, Shinyoung; Kim, Sujin; Chang, In Seop; Choi, In-Geol

    2011-01-01

    Eubacterium limosum KIST612 is an anaerobic acetogenic bacterium that uses CO as the sole carbon/energy source and produces acetate, butyrate, and ethanol. To evaluate its potential as a syngas microbial catalyst, we have sequenced the complete 4.3-Mb genome of E. limosum KIST612.

  11. ENUMERATION, ISOLATION, AND CHARACTERIZATION OF ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS (POSTER SESSION)

    EPA Science Inventory

    Seagrasses are rooted in anoxic, sulfate-reducing sediments. However, the seagrass root is oxygenated during the daytime, becoming anoxic at night. Root thin sections hydridized with 33P-labeled probes revealed the presence of acetogenic and sulfate-reducing bacteria in the rhizo...

  12. ENUMERATION, ISOLATION, AND CHARACTERIZATION OF ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS

    EPA Science Inventory

    Seagrasses are rooted in anoxic, sulfate-reducing sediments. However, the seagrass root is oxygenated during the daytime, becoming anoxic at night. Root thin sections hydridized with 33P-labeled probes revealed the presence of acetogenic and sulfate-reducing bacteria in the rhizo...

  13. Genome Sequence of the Acetogenic Bacterium Acetobacterium wieringae DSM 1911T

    PubMed Central

    Schiel-Bengelsdorf, Bettina; Daniel, Rolf

    2016-01-01

    Here, we report the draft genome sequence of Acetobacterium wieringae DSM 1911T, an anaerobic, autotrophic, acetogenic, d,l-lactate-utilizing bacterium. The genome consists of a chromosome (3.88 Mb) and 3,620 predicted protein-encoding genes. PMID:28007862

  14. Insights into the Genome of the Anaerobic Acetogen Sporomusa silvacetica DSM 10669

    PubMed Central

    Humphreys, Jonathan R.

    2017-01-01

    ABSTRACT Sporomusa silvacetica is a spore-forming, anaerobic acetogen isolated from soil derived from east central Germany. The genome contains genes of the Wood-Ljungdahl pathway required for carbon fixation and genes involved in the biosynthesis of the amino acid pyrrolysine. The genome (5.92 Mb) harbors 4,355 predicted protein-encoding genes. PMID:28935741

  15. ENUMERATION, ISOLATION, AND CHARACTERIZATION OF ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS (POSTER SESSION)

    EPA Science Inventory

    Seagrasses are rooted in anoxic, sulfate-reducing sediments. However, the seagrass root is oxygenated during the daytime, becoming anoxic at night. Root thin sections hydridized with 33P-labeled probes revealed the presence of acetogenic and sulfate-reducing bacteria in the rhizo...

  16. ENUMERATION, ISOLATION, AND CHARACTERIZATION OF ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS

    EPA Science Inventory

    Seagrasses are rooted in anoxic, sulfate-reducing sediments. However, the seagrass root is oxygenated during the daytime, becoming anoxic at night. Root thin sections hydridized with 33P-labeled probes revealed the presence of acetogenic and sulfate-reducing bacteria in the rhizo...

  17. Comparative reaction engineering analysis of different acetogenic bacteria for gas fermentation.

    PubMed

    Groher, Anna; Weuster-Botz, Dirk

    2016-06-20

    The production of chemicals by syngas fermentation is a promising alternative to heterotrophic fermentation processes. The autotrophic process performances of the so far not well studied acetogens Acetobacterium fimetarium, Acetobacterium wieringae, Blautia hydrogenotrophica, Clostridium magnum, Eubacterium aggregans, Sporomusa acidovorans, Sporomusa ovata and Terrisporobacter mayombei were characterized. Acetobacterium woodii was used as reference strain. Standardized batch experiments with continuous supply of the gaseous substrates CO2 and H2 were performed in fully controlled stirred-tank bioreactors. A. wieringae and S. ovata showed by far the highest growth rates and maximum biomass concentrations among the acetogens under study. Aside from the reference strain A. woodii, highest volumetric (17.96gL(-1)d(-1)) as well as cell specific acetate formation rates (21.03gg(-1)d(-1)) were observed with S. ovata resulting in a final acetate concentration of 32.2gL(-1). Accumulation of formate with up to 4.8gL(-1) was observed with all acetogens. Ethanol was produced autotrophically with up to 0.42gL(-1) by four of the acetogenic bacteria under study (A. wieringae, C. magnum, S. acidovorans and S. ovata) and also by A. woodii. Butyrate was formed with up to 0.14gL(-1) by three of the acetogenic bacteria under study (C. magnum, B. hydrogenotrophica and E. aggregans). Due to its superior process performances S. ovata may be a promising host for redirecting carbon fluxes by applying metabolic engineering and tools of synthetic biology to produce non-natural chemicals from syngas.

  18. Identification and characterization of oxalate oxidoreductase, a novel thiamine pyrophosphate-dependent 2-oxoacid oxidoreductase that enables anaerobic growth on oxalate.

    PubMed

    Pierce, Elizabeth; Becker, Donald F; Ragsdale, Stephen W

    2010-12-24

    Moorella thermoacetica is an anaerobic acetogen, a class of bacteria that is found in the soil, the animal gastrointestinal tract, and the rumen. This organism engages the Wood-Ljungdahl pathway of anaerobic CO(2) fixation for heterotrophic or autotrophic growth. This paper describes a novel enzyme, oxalate oxidoreductase (OOR), that enables M. thermoacetica to grow on oxalate, which is produced in soil and is a common component of kidney stones. Exposure to oxalate leads to the induction of three proteins that are subunits of OOR, which oxidizes oxalate coupled to the production of two electrons and CO(2) or bicarbonate. Like other members of the 2-oxoacid:ferredoxin oxidoreductase family, OOR contains thiamine pyrophosphate and three [Fe(4)S(4)] clusters. However, unlike previously characterized members of this family, OOR does not use coenzyme A as a substrate. Oxalate is oxidized with a k(cat) of 0.09 s(-1) and a K(m) of 58 μM at pH 8. OOR also oxidizes a few other 2-oxoacids (which do not induce OOR) also without any requirement for CoA. The enzyme transfers its reducing equivalents to a broad range of electron acceptors, including ferredoxin and the nickel-dependent carbon monoxide dehydrogenase. In conjunction with the well characterized Wood-Ljungdahl pathway, OOR should be sufficient for oxalate metabolism by M. thermoacetica, and it constitutes a novel pathway for oxalate metabolism.

  19. Trophic links between the acetogen Clostridium glycolicum KHa and the fermentative anaerobe Bacteroides xylanolyticus KHb, isolated from Hawaiian forest soil.

    PubMed

    Hunger, Sindy; Gössner, Anita S; Drake, Harold L

    2011-09-01

    Isolate KH was obtained from Hawaiian forest soil and found to be composed of two functionally linked anaerobes, KHa and KHb. Gene analyses (16S rRNA, fhs, cooS) identified KHa as an acetogenic strain of Clostridium glycolicum and KHb as Bacteroides xylanolyticus. KHb fermented xylan and other saccharides that KHa could not utilize and formed products (e.g., ethanol and H(2)) that supported the acetogenic growth of KHa.

  20. Design and testing of a functional group-specific DNA probe for the study of natural populations of acetogenic bacteria.

    PubMed Central

    Lovell, C R; Hui, Y

    1991-01-01

    The acetogens, although phylogenetically diverse, can be characterized by their possession of the acetyl coenzyme A (acetyl-CoA) pathway for autotrophic CO2 fixation. The gene encoding formyltetrahydrofolate synthetase, a key enzyme of the acetyl-CoA pathway, was previously cloned from the thermophilic acetogen Clostridium thermoaceticum and has now been tested as a group-specific probe for acetogens. Stable hybrids were formed between the probe and single DNA fragments from eight known acetogens representing six genera. A hybrid was also formed between the probe and a DNA fragment from one sulfate reducer known to be capable of both autotrophic CO2 fixation and acetate catabolism. No such hybrid was formed between the probe and DNA from a homoacetate fermenter not known to use the acetyl-CoA pathway, with two known formyltetrahydrofolate synthetase-producing purine fermenters, or with DNA from 27 other species representing 16 genera of organisms that do not use the acetyl-CoA pathway. DNA purified from cells extracted from horse manure was also screened with the acetogen probe. Six hybrids, indicating at least six detectable acetogen "strains," were observed. Images PMID:1768134

  1. Characterizing acetogenic metabolism using a genome-scale metabolic reconstruction of Clostridium ljungdahlii

    PubMed Central

    2013-01-01

    Background The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow autotrophically on H2/CO2, and more importantly on synthesis gas (H2/CO/CO2) make them very attractive candidates as production hosts for biofuels and biocommodities. Acetogenic metabolism is considered one of the earliest modes of bacterial metabolism. A thorough understanding of various factors governing the metabolism, in particular energy conservation mechanisms, is critical for metabolic engineering of acetogens for targeted production of desired chemicals. Results Here, we present the genome-scale metabolic network of Clostridium ljungdahlii, the first such model for an acetogen. This genome-scale model (iHN637) consisting of 637 genes, 785 reactions, and 698 metabolites captures all the major central metabolic and biosynthetic pathways, in particular pathways involved in carbon fixation and energy conservation. A combination of metabolic modeling, with physiological and transcriptomic data provided insights into autotrophic metabolism as well as aided the characterization of a nitrate reduction pathway in C. ljungdahlii. Analysis of the iHN637 metabolic model revealed that flavin based electron bifurcation played a key role in energy conservation during autotrophic growth and helped identify genes for some of the critical steps in this mechanism. Conclusions iHN637 represents a predictive model that recapitulates experimental data, and provides valuable insights into the metabolic response of C. ljungdahlii to genetic perturbations under various growth conditions. Thus, the model will be instrumental in guiding metabolic engineering of C. ljungdahlii for the industrial production of biocommodities and biofuels. PMID:24274140

  2. The complete genome sequence of Eubacterium limosum SA11, a metabolically versatile rumen acetogen.

    PubMed

    Kelly, William J; Henderson, Gemma; Pacheco, Diana M; Li, Dong; Reilly, Kerri; Naylor, Graham E; Janssen, Peter H; Attwood, Graeme T; Altermann, Eric; Leahy, Sinead C

    2016-01-01

    Acetogens are a specialized group of anaerobic bacteria able to produce acetate from CO2 and H2 via the Wood-Ljungdahl pathway. In some gut environments acetogens can compete with methanogens for H2, and as a result rumen acetogens are of interest in the development of microbial approaches for methane mitigation. The acetogen Eubacterium limosum SA11 was isolated from the rumen of a New Zealand sheep and its genome has been sequenced to examine its potential application in methane mitigation strategies, particularly in situations where hydrogenotrophic methanogens are inhibited resulting in increased H2 levels in the rumen. The 4.15 Mb chromosome of SA11 has an average G + C content of 47 %, and encodes 3805 protein-coding genes. There is a single prophage inserted in the chromosome, and several other gene clusters appear to have been acquired by horizontal transfer. These include genes for cell wall glycopolymers, a type VII secretion system, cell surface proteins and chemotaxis. SA11 is able to use a variety of organic substrates in addition to H2/CO2, with acetate and butyrate as the principal fermentation end-products, and genes involved in these metabolic pathways have been identified. An unusual feature is the presence of 39 genes encoding trimethylamine methyltransferase family proteins, more than any other bacterial genome. Overall, SA11 is a metabolically versatile organism, but its ability to grow on such a wide range of substrates suggests it may not be a suitable candidate to take the place of hydrogen-utilizing methanogens in the rumen.

  3. Characterizing acetogenic metabolism using a genome-scale metabolic reconstruction of Clostridium ljungdahlii

    SciTech Connect

    Nagarajan, H; Sahin, M; Nogales, J; Latif, H; Lovley, DR; Ebrahim, A; Zengler, K

    2013-11-25

    Background: The metabolic capabilities of acetogens to ferment a wide range of sugars, to grow autotrophically on H-2/CO2, and more importantly on synthesis gas (H-2/CO/CO2) make them very attractive candidates as production hosts for biofuels and biocommodities. Acetogenic metabolism is considered one of the earliest modes of bacterial metabolism. A thorough understanding of various factors governing the metabolism, in particular energy conservation mechanisms, is critical for metabolic engineering of acetogens for targeted production of desired chemicals. Results: Here, we present the genome-scale metabolic network of Clostridium ljungdahlii, the first such model for an acetogen. This genome-scale model (iHN637) consisting of 637 genes, 785 reactions, and 698 metabolites captures all the major central metabolic and biosynthetic pathways, in particular pathways involved in carbon fixation and energy conservation. A combination of metabolic modeling, with physiological and transcriptomic data provided insights into autotrophic metabolism as well as aided the characterization of a nitrate reduction pathway in C. ljungdahlii. Analysis of the iHN637 metabolic model revealed that flavin based electron bifurcation played a key role in energy conservation during autotrophic growth and helped identify genes for some of the critical steps in this mechanism. Conclusions: iHN637 represents a predictive model that recapitulates experimental data, and provides valuable insights into the metabolic response of C. ljungdahlii to genetic perturbations under various growth conditions. Thus, the model will be instrumental in guiding metabolic engineering of C. ljungdahlii for the industrial production of biocommodities and biofuels.

  4. Non-Growth-Associated Demethylation of Dimethylsulfoniopropionate by (Homo)acetogenic Bacteria

    PubMed Central

    Jansen, Michael; Hansen, Theo A.

    2001-01-01

    The demethylation of the algal osmolyte dimethylsulfoniopropionate (DMSP) to methylthiopropionate (MTPA) by (homo)acetogenic bacteria was studied. Five Eubacterium limosum strains (including the type strain), Sporomusa ovata DSM 2662T, Sporomusa sphaeroides DSM 2875T, and Acetobacterium woodii DSM 1030T were shown to demethylate DMSP stoichiometrically to MTPA. The (homo)acetogenic fermentation based on this demethylation did not result in any significant increase in biomass. The analogous demethylation of glycine betaine to dimethylglycine does support growth of acetogens. In batch cultures of E. limosum PM31 DMSP and glycine betaine were demethylated simultaneously. In mixed substrates experiments with fructose-DMSP or methanol-DMSP, DMSP was used rapidly but only after exhaustion of the fructose or the methanol. In steady-state fructose-limited chemostat cultures (at a dilution rate of 0.03 h−1) with DMSP as a second reservoir substrate, DMSP was biotransformed to MTPA but this did not result in higher biomass values than in cultures without DMSP; cells from such cultures demethylated DMSP at rates of approximately 50 nmol min−1 mg of protein−1, both after growth in the presence of DMSP and after growth in its absence. In cell extracts of glycine betaine-grown strain PM31, DMSP demethylation activities of 21 to 24 nmol min−1 mg of protein−1 were detected with tetrahydrofolate as a methyl acceptor; the activities seen with glycine betaine were approximately 10-fold lower. A speculative explanation for the demethylation of DMSP without an obvious benefit for the organism is that the DMSP-demethylating activity is catalyzed by the glycine betaine-demethylating enzyme and that a transport-related factor, in particular a higher energy demand for DMSP transport across the cytoplasmic membrane than for glycine betaine transport, may reduce the overall ATP yield of the fermentation to virtually zero. PMID:11133459

  5. Non-growth-associated demethylation of dimethylsulfoniopropionate by (homo)acetogenic bacteria.

    PubMed

    Jansen, M; Hansen, T A

    2001-01-01

    The demethylation of the algal osmolyte dimethylsulfoniopropionate (DMSP) to methylthiopropionate (MTPA) by (homo)acetogenic bacteria was studied. Five Eubacterium limosum strains (including the type strain), Sporomusa ovata DSM 2662(T), Sporomusa sphaeroides DSM 2875(T), and Acetobacterium woodii DSM 1030(T) were shown to demethylate DMSP stoichiometrically to MTPA. The (homo)acetogenic fermentation based on this demethylation did not result in any significant increase in biomass. The analogous demethylation of glycine betaine to dimethylglycine does support growth of acetogens. In batch cultures of E. limosum PM31 DMSP and glycine betaine were demethylated simultaneously. In mixed substrates experiments with fructose-DMSP or methanol-DMSP, DMSP was used rapidly but only after exhaustion of the fructose or the methanol. In steady-state fructose-limited chemostat cultures (at a dilution rate of 0.03 h(-1)) with DMSP as a second reservoir substrate, DMSP was biotransformed to MTPA but this did not result in higher biomass values than in cultures without DMSP; cells from such cultures demethylated DMSP at rates of approximately 50 nmol min(-1) mg of protein(-1), both after growth in the presence of DMSP and after growth in its absence. In cell extracts of glycine betaine-grown strain PM31, DMSP demethylation activities of 21 to 24 nmol min(-1) mg of protein(-1) were detected with tetrahydrofolate as a methyl acceptor; the activities seen with glycine betaine were approximately 10-fold lower. A speculative explanation for the demethylation of DMSP without an obvious benefit for the organism is that the DMSP-demethylating activity is catalyzed by the glycine betaine-demethylating enzyme and that a transport-related factor, in particular a higher energy demand for DMSP transport across the cytoplasmic membrane than for glycine betaine transport, may reduce the overall ATP yield of the fermentation to virtually zero.

  6. Acetate and ethanol production from H2 and CO2 by Moorella sp. using a repeated batch culture.

    PubMed

    Sakai, Shinsuke; Nakashimada, Yutaka; Inokuma, Kentaro; Kita, Masayuki; Okada, Hideki; Nishio, Naomichi

    2005-03-01

    The growth inhibition of Moorella sp. HUC22-1 by undissociated acetic acid was analyzed using a non-competitive inhibition model coupled with a pH inhibition model. In the cells grown on H2 and CO2, the inhibition constant, K(p) of the undissociated acetic acid was 6.2 mM (164 mM as the total acetate at pH 6.2, pKa = 4.795, 55 degrees C), which was 1.5-fold higher than that obtained in cells grown on fructose. When a pH-controlled batch culture was performed using a fermentor at pH 6.2 with H2 and CO2, a maximum of 0.92 g/l of dry cell weight and 339 mM of acetate were produced after 220 h, which were 4.4- and 6.8-fold higher than those produced in the pH-uncontrolled batch culture, respectively. In order to reduce acetate inhibition in the culture medium, a repeated batch culture with cell recycling was performed at a constant pH with H2 and CO2. At a pH of 6.2, the total acetate production reached 840 mmol/l-reactor with 4.7 mmol/l-reactor of total ethanol production after 420 h. When the culture pH was maintained at 5.8, which was the optimum for ethanol production, the total ethanol production reached 15.4 mmol/l-reactor after 430 h, although the total acetate production was decreased to 675 mmol/l-reactor.

  7. A new H2/CO2-using acetogenic bacterium from the rumen: description of Ruminococcus schinkii sp. nov.

    PubMed

    Rieu-Lesme, F; Morvan, B; Collins, M D; Fonty, G; Willems, A

    1996-07-01

    Two strains of H2/CO2-using acetogenic bacteria were isolated from the rumen of suckling lambs. Both strains displayed a coccobacillar morphology and possessed a Gram-positive type cell wall. Numerous organic substrates, including some O-methylated aromatic compounds, were used heterotrophically. 16S rRNA gene sequencing demonstrated that the two acetogenic isolates were phylogenetically identical and represent a new subline within Clostridium cluster XIVa. Based on phenotypic and phylogenetic considerations a new species, Ruminococcus schinkii sp. nov., is proposed.

  8. Physiology and biochemistry of single carbon catabolism by Butyribacterium methylotrophicum, and anaerobic acetogen

    SciTech Connect

    Kerby, R.

    1986-01-01

    The catabolism of methanol, formate, carbon monoxide, and carbon dioxide by the anaerobic acetogen Butyribacterium methylotrophicum was examined by several fermentation time course, /sup 13/C-NMR, /sup 14/C radioisotope tracer, and enzyme level analyses. During the simultaneous consumption of methanol and a more oxidized cosubstrate, methanol carbon was primarily funneled into the acetate methyl group with the co-substrate carbon predominantly incorporated into the acetate carboxyl. Formate and carbon monoxide were also simultaneously consumed and were preferentially distributed into the acetate methyl and carboxyl groups, respectively. These studies supported the function of a bifurcated single carbon catabolic pathway with carbonyl and methyl group synthesis routes jointed at acetyl-CoA, the primary reduced product. Isotope dilution experiments testified to the key role of carbon dioxide as the sole carbon unit which links the two halves of this catabolic mechanism. High levels of carbon monoxide and formate dehydrogenases and the tetrahydrofolate-requiring enzymes including methylene-tetrahydrofolate reductase correlated with the consumption of certain single carbon substrates. Implications of this catabolic scheme on acetogenic ATP synthesis via electron transport phosphorylation are discussed and the results of proton motive force analyses presented.

  9. Heterotrimeric NADH-oxidizing methylenetetrahydrofolate reductase from the acetogenic bacterium Acetobacterium woodii.

    PubMed

    Bertsch, Johannes; Öppinger, Christian; Hess, Verena; Langer, Julian D; Müller, Volker

    2015-05-01

    The methylenetetrahydrofolate reductase (MTHFR) of acetogenic bacteria catalyzes the reduction of methylene-THF, which is highly exergonic with NADH as the reductant. Therefore, the enzyme was suggested to be involved in energy conservation by reducing ferredoxin via electron bifurcation, followed by Na(+) translocation by the Rnf complex. The enzyme was purified from Acetobacterium woodii and shown to have an unprecedented subunit composition containing the three subunits RnfC2, MetF, and MetV. The stable complex contained 2 flavin mononucleotides (FMN), 23.5 ± 1.2 Fe and 24.5 ± 1.5 S, which fits well to the predicted six [4Fe4S] clusters in MetV and RnfC2. The enzyme catalyzed NADH:methylviologen and NADH:ferricyanide oxidoreductase activity but also methylene-tetrahydrofolate (THF) reduction with NADH as the reductant. The NADH:methylene-THF reductase activity was high (248 U/mg) and not stimulated by ferredoxin. Furthermore, reduction of ferredoxin, alone or in the presence of methylene-THF and NADH, was never observed. MetF or MetVF was not able to catalyze the methylene-THF-dependent oxidation of NADH, but MetVF could reduce methylene-THF using methyl viologen as the electron donor. The purified MTHFR complex did not catalyze the reverse reaction, the endergonic oxidation of methyl-THF with NAD(+) as the acceptor, and this reaction could not be driven by reduced ferredoxin. However, addition of protein fractions made the oxidation of methyl-THF to methylene-THF coupled to NAD(+) reduction possible. Our data demonstrate that the MTHFR of A. woodii catalyzes methylene-THF reduction according to the following reaction: NADH + methylene-THF → methyl-THF + NAD(+). The differences in the subunit compositions of MTHFRs of bacteria are discussed in the light of their different functions. Energy conservation in the acetogenic bacterium Acetobacterium woodii involves ferredoxin reduction followed by a chemiosmotic mechanism involving Na(+)-translocating ferredoxin

  10. Heterotrimeric NADH-Oxidizing Methylenetetrahydrofolate Reductase from the Acetogenic Bacterium Acetobacterium woodii

    PubMed Central

    Bertsch, Johannes; Öppinger, Christian; Hess, Verena; Langer, Julian D.

    2015-01-01

    ABSTRACT The methylenetetrahydrofolate reductase (MTHFR) of acetogenic bacteria catalyzes the reduction of methylene-THF, which is highly exergonic with NADH as the reductant. Therefore, the enzyme was suggested to be involved in energy conservation by reducing ferredoxin via electron bifurcation, followed by Na+ translocation by the Rnf complex. The enzyme was purified from Acetobacterium woodii and shown to have an unprecedented subunit composition containing the three subunits RnfC2, MetF, and MetV. The stable complex contained 2 flavin mononucleotides (FMN), 23.5 ± 1.2 Fe and 24.5 ± 1.5 S, which fits well to the predicted six [4Fe4S] clusters in MetV and RnfC2. The enzyme catalyzed NADH:methylviologen and NADH:ferricyanide oxidoreductase activity but also methylene-tetrahydrofolate (THF) reduction with NADH as the reductant. The NADH:methylene-THF reductase activity was high (248 U/mg) and not stimulated by ferredoxin. Furthermore, reduction of ferredoxin, alone or in the presence of methylene-THF and NADH, was never observed. MetF or MetVF was not able to catalyze the methylene-THF-dependent oxidation of NADH, but MetVF could reduce methylene-THF using methyl viologen as the electron donor. The purified MTHFR complex did not catalyze the reverse reaction, the endergonic oxidation of methyl-THF with NAD+ as the acceptor, and this reaction could not be driven by reduced ferredoxin. However, addition of protein fractions made the oxidation of methyl-THF to methylene-THF coupled to NAD+ reduction possible. Our data demonstrate that the MTHFR of A. woodii catalyzes methylene-THF reduction according to the following reaction: NADH + methylene-THF → methyl-THF + NAD+. The differences in the subunit compositions of MTHFRs of bacteria are discussed in the light of their different functions. IMPORTANCE Energy conservation in the acetogenic bacterium Acetobacterium woodii involves ferredoxin reduction followed by a chemiosmotic mechanism involving Na

  11. Elucidating acetogenic H2 consumption in dark fermentation using flux balance analysis.

    PubMed

    Lalman, Jerald A; Chaganti, Subba Rao; Moon, Chungman; Kim, Dong-Hoon

    2013-10-01

    In this study, a flux balance analysis (FBA) was adopted to estimate the activity of acetogenic H2-consuming reaction. Experimental data at different substrate concentrations of 10, 20, and 30 g COD/L showing the lowest, medium, and highest H2 yields, respectively, were used in the FBA to calculate the fluxes. It was interesting to note that the hydrogenase activity based on R12 (2Fd(+)+2H(+)→2Fd(2+)+H2, ferredoxin (Fd)) flux was most active at 10 g COD/L. The flux of R17 (4H2+2CO2→CH3COOH), a mechanism for reutilizing produced H2, increased in steps of 0.030, 0.119, and 0.467 as the substrate concentration decreased. Contradictory to our general understanding, acetate production found to have a negligible or even negative effect on the final H2 yield in dark fermentation.

  12. Acetogenic mixotrophy: novel options for yield improvement in biofuels and biochemicals production.

    PubMed

    Fast, Alan G; Schmidt, Ellinor D; Jones, Shawn W; Tracy, Bryan P

    2015-06-01

    Mass yields of biofuels and chemicals from sugar fermentations are limited by the decarboxylation reactions involved in Embden-Meyerhof-Parnas (EMP) glycolysis. This paper reviews one route to recapture evolved CO2 using the Wood-Ljungdahl carbon fixation pathway (WLP) in a process called anaerobic, non-photosynthetic (ANP) mixotrophic fermentation. In ANP mixotrophic fermentation, the two molecules of CO2 and eight electrons produced from glycolysis are used by the WLP to generate three molecules of acetyl-CoA from glucose, rather than the two molecules that are produced by typical fermentation processes. In this review, we define the bounds of ANP mixotrophy, calculate the potential metabolic advantages, and discuss the viability in a number of host organisms. Additionally, we highlight recent accomplishments in the field, including the recent discovery of electron bifurcation in acetogens, and close with recommendations to realize mixotrophic biofuel and biochemical production.

  13. Recovery and Analysis of Formyltetrahydrofolate Synthetase Gene Sequences from Natural Populations of Acetogenic Bacteria

    PubMed Central

    Leaphart, Adam B.; Lovell, Charles R.

    2001-01-01

    Primers for PCR amplification of partial (1,102 of 1,680 bp) formyltetrahydrofolate synthetase (FTHFS) gene sequences were developed and tested. Partial FTHFS sequences were successfully amplified from DNA from pure cultures of known acetogens, from other FTHFS-producing organisms, from the roots of the smooth cordgrass, Spartina alterniflora, and from fresh horse manure. The amplimers recovered were cloned, their nucleotide sequences were determined, and their translated amino acid sequences were used to construct phylogenetic trees. We found that FTHFS sequences from homoacetogens formed a monophyletic cluster that did not contain sequences from nonhomoacetogens and that FTHFS sequences appear to be informative regarding major physiological features of FTHFS-producing organisms. PMID:11229939

  14. Mevalonate production by engineered acetogen biocatalyst during continuous fermentation of syngas or CO₂/H₂ blend.

    PubMed

    Kiriukhin, Michael; Tyurin, Michael

    2014-02-01

    Naturally mevalonate-resistant acetogen Clostridium sp. MT1243 produced only 425 mM acetate during syngas fermentation. Using Clostridium sp. MT1243 we engineered biocatalyst selectively producing mevalonate from synthesis gas or CO₂/H₂ blend. Acetate production and spore formation were eliminated from Clostridium sp. MT1243 using Cre-lox66/lox71-system. Cell energy released via elimination of phosphotransacetylase, acetate kinase and early stage sporulation genes powered mevalonate accumulation in fermentation broth due to expression of synthetic thiolase, HMG-synthase, and HMG-reductase, three copies of each, integrated using Tn7-approach. Recombinants produced 145 mM mevalonate in five independent single-step fermentation runs 25 days each in five repeats using syngas blend 60% CO and 40% H₂ (v/v) (p < 0.005). Mevalonate production was 97 mM if only CO₂/H₂ blend was fed instead of syngas (p < 0.005). Mevalonate from CO₂/H₂ blend might serve as a commercial route to mitigate global warming in proportion to CO₂ fermentation scale worldwide.

  15. Acetogens and Acetoclastic Methanosarcinales Govern Methane Formation in Abandoned Coal Mines▿

    PubMed Central

    Beckmann, Sabrina; Lueders, Tillmann; Krüger, Martin; von Netzer, Frederick; Engelen, Bert; Cypionka, Heribert

    2011-01-01

    In abandoned coal mines, methanogenic archaea are responsible for the production of substantial amounts of methane. The present study aimed to directly unravel the active methanogens mediating methane release as well as active bacteria potentially involved in the trophic network. Therefore, the stable-isotope-labeled precursors of methane, [13C]acetate and H2-13CO2, were fed to liquid cultures from hard coal and mine timber from a coal mine in Germany. Guided by methane production rates, samples for DNA stable-isotope probing (SIP) with subsequent quantitative PCR and denaturing gradient gel electrophoretic (DGGE) analyses were taken over 6 months. Surprisingly, the formation of [13C]methane was linked to acetoclastic methanogenesis in both the [13C]acetate- and the H2-13CO2-amended cultures of coal and timber. H2-13CO2 was used mainly by acetogens related to Pelobacter acetylenicus and Clostridium species. Active methanogens, closely affiliated with Methanosarcina barkeri, utilized the readily available acetate rather than the thermodynamically more favorable hydrogen. Thus, the methanogenic microbial community appears to be highly adapted to the low-H2 conditions found in coal mines. PMID:21460109

  16. A Na+-translocating Pyrophosphatase in the Acetogenic Bacterium Acetobacterium woodii*

    PubMed Central

    Biegel, Eva; Müller, Volker

    2011-01-01

    The anaerobic acetogenic bacterium Acetobacterium woodii employs a novel type of Na+-motive anaerobic respiration, caffeate respiration. However, this respiration is at the thermodynamic limit of energy conservation, and even worse, in the first step, caffeate is activated by caffeyl-CoA synthetase, which hydrolyzes ATP to AMP and pyrophosphate. Here, we have addressed whether or not the energy stored in the anhydride bond of pyrophosphate is conserved by A. woodii. Inverted membrane vesicles of A. woodii have a membrane-bound pyrophosphatase that catalyzes pyrophosphate hydrolysis at a rate of 70–120 milliunits/mg of protein. Pyrophosphatase activity was dependent on the divalent cation Mg2+. In addition, activity was strictly dependent on Na+ with a Km of 1.1 mm. Hydrolysis of pyrophosphate was accompanied by 22Na+ transport into the lumen of the inverted membrane vesicles. Inhibitor studies revealed that 22Na+ transport was primary and electrogenic. Next to the Na+-motive ferredoxin:NAD+ oxidoreductase (Fno or Rnf), the Na+-pyrophosphatase is the second primary Na+-translocating enzyme in A. woodii. PMID:21173152

  17. Acetogenic and sulfate-reducing bacteria inhabiting the rhizoplane and deep cortex cells of the sea grass Halodule wrightii.

    PubMed

    Küsel, K; Pinkart, H C; Drake, H L; Devereux, R

    1999-11-01

    Recent declines in sea grass distribution underscore the importance of understanding microbial community structure-function relationships in sea grass rhizospheres that might affect the viability of these plants. Phospholipid fatty acid analyses showed that sulfate-reducing bacteria and clostridia were enriched in sediments colonized by the sea grasses Halodule wrightii and Thalassia testudinum compared to an adjacent unvegetated sediment. Most-probable-number analyses found that in contrast to butyrate-producing clostridia, acetogens and acetate-utilizing sulfate reducers were enriched by an order of magnitude in rhizosphere sediments. Although sea grass roots are oxygenated in the daytime, colorimetric root incubation studies demonstrated that acetogenic O-demethylation and sulfidogenic iron precipitation activities were tightly associated with washed, sediment-free H. wrightii roots. This suggests that the associated anaerobes are able to tolerate exposure to oxygen. To localize and quantify the anaerobic microbial colonization, root thin sections were hybridized with newly developed (33)P-labeled probes that targeted (i) low-G+C-content gram-positive bacteria, (ii) cluster I species of clostridia, (iii) species of Acetobacterium, and (iv) species of Desulfovibrio. Microautoradiography revealed intercellular colonization of the roots by Acetobacterium and Desulfovibrio species. Acetogenic bacteria occurred mostly in the rhizoplane and outermost cortex cell layers, and high numbers of sulfate reducers were detected on all epidermal cells and inward, colonizing some 60% of the deepest cortex cells. Approximately 30% of epidermal cells were colonized by bacteria that hybridized with an archaeal probe, strongly suggesting the presence of methanogens. Obligate anaerobes within the roots might contribute to the vitality of sea grasses and other aquatic plants and to the biogeochemistry of the surrounding sediment.

  18. Acetogenic and Sulfate-Reducing Bacteria Inhabiting the Rhizoplane and Deep Cortex Cells of the Sea Grass Halodule wrightii†

    PubMed Central

    Küsel, Kirsten; Pinkart, Holly C.; Drake, Harold L.; Devereux, Richard

    1999-01-01

    Recent declines in sea grass distribution underscore the importance of understanding microbial community structure-function relationships in sea grass rhizospheres that might affect the viability of these plants. Phospholipid fatty acid analyses showed that sulfate-reducing bacteria and clostridia were enriched in sediments colonized by the sea grasses Halodule wrightii and Thalassia testudinum compared to an adjacent unvegetated sediment. Most-probable-number analyses found that in contrast to butyrate-producing clostridia, acetogens and acetate-utilizing sulfate reducers were enriched by an order of magnitude in rhizosphere sediments. Although sea grass roots are oxygenated in the daytime, colorimetric root incubation studies demonstrated that acetogenic O-demethylation and sulfidogenic iron precipitation activities were tightly associated with washed, sediment-free H. wrightii roots. This suggests that the associated anaerobes are able to tolerate exposure to oxygen. To localize and quantify the anaerobic microbial colonization, root thin sections were hybridized with newly developed 33P-labeled probes that targeted (i) low-G+C-content gram-positive bacteria, (ii) cluster I species of clostridia, (iii) species of Acetobacterium, and (iv) species of Desulfovibrio. Microautoradiography revealed intercellular colonization of the roots by Acetobacterium and Desulfovibrio species. Acetogenic bacteria occurred mostly in the rhizoplane and outermost cortex cell layers, and high numbers of sulfate reducers were detected on all epidermal cells and inward, colonizing some 60% of the deepest cortex cells. Approximately 30% of epidermal cells were colonized by bacteria that hybridized with an archaeal probe, strongly suggesting the presence of methanogens. Obligate anaerobes within the roots might contribute to the vitality of sea grasses and other aquatic plants and to the biogeochemistry of the surrounding sediment. PMID:10543830

  19. Ruminococcus hydrogenotrophicus sp. nov., a new H2/CO2-utilizing acetogenic bacterium isolated from human feces.

    PubMed

    Bernalier, A; Willems, A; Leclerc, M; Rochet, V; Collins, M D

    1996-09-01

    A new H2/CO2-utilizing acetogenic bacterium was isolated from the feces of a non-methane-excreting human subject. The two strains S5a33 and S5a36 were strictly anaerobic, gram-positive, non-sporulating coccobacilli. The isolates grew autotrophically by metabolizing H2/CO2 to form acetate as sole metabolite and were also able to grow heterotrophically on a variety of organic compounds. The major end product of glucose and fructose fermentation was acetate; the strains also formed ethanol, lactate and, to a lesser extent, isobutyrate and isovalerate. The G+C content of DNA of strain S5a33 was 45.2 mol%. 16S rRNA gene sequencing demonstrated that the two acetogenic isolates were phylogenetically identical and represent a new subline within Clostridium cluster XIVa. Based on phenotypic and phylogenetic considerations, a new species, Ruminococcus hydrogenotrophicus, is proposed. The type strain of R. hydrogenotrophicus is S5a33 (DSM 10507). Furthermore, H2/CO2 acetogenesis appeared to be a common property of most of the species phylogenetically closely related to strain S5a33 (Clostridium coccoides, Ruminococcus hansenii, and Ruminococcus productus).

  20. Adaptation of the autotrophic acetogen Sporomusa ovata to methanol accelerates the conversion of CO2 to organic products.

    PubMed

    Tremblay, Pier-Luc; Höglund, Daniel; Koza, Anna; Bonde, Ida; Zhang, Tian

    2015-11-04

    Acetogens are efficient microbial catalysts for bioprocesses converting C1 compounds into organic products. Here, an adaptive laboratory evolution approach was implemented to adapt Sporomusa ovata for faster autotrophic metabolism and CO2 conversion to organic chemicals. S. ovata was first adapted to grow quicker autotrophically with methanol, a toxic C1 compound, as the sole substrate. Better growth on different concentrations of methanol and with H2-CO2 indicated the adapted strain had a more efficient autotrophic metabolism and a higher tolerance to solvent. The growth rate on methanol was increased 5-fold. Furthermore, acetate production rate from CO2 with an electrode serving as the electron donor was increased 6.5-fold confirming that the acceleration of the autotrophic metabolism of the adapted strain is independent of the electron donor provided. Whole-genome sequencing, transcriptomic, and biochemical studies revealed that the molecular mechanisms responsible for the novel characteristics of the adapted strain were associated with the methanol oxidation pathway and the Wood-Ljungdahl pathway of acetogens along with biosynthetic pathways, cell wall components, and protein chaperones. The results demonstrate that an efficient strategy to increase rates of CO2 conversion in bioprocesses like microbial electrosynthesis is to evolve the microbial catalyst by adaptive laboratory evolution to optimize its autotrophic metabolism.

  1. Unravelling the one-carbon metabolism of the acetogen Sporomusa strain An4 by genome and proteome analysis.

    PubMed

    Visser, Michael; Pieterse, Mervin M; Pinkse, Martijn W H; Nijsse, Bart; Verhaert, Peter D E M; de Vos, Willem M; Schaap, Peter J; Stams, Alfons J M

    2016-09-01

    The Sporomusa genus comprises anaerobic spore-forming acetogenic bacteria that stain Gram-negative. Sporomusa species typically grow with one-carbon substrates and N-methylated compounds. In the degradation of these compounds methyltransferases are involved. In addition, Sporomusa species can grow autotrophically with H2 and CO2 , and use a variety of sugars for acetogenic growth. Here we describe a genome analysis of Sporomusa strain An4 and a proteome analysis of cells grown under five different conditions. Comparison of the genomes of Sporomusa strain An4 and Sporomusa ovata strain H1 indicated that An4 is a S. ovata strain. Proteome analysis showed a high abundance of several methyltransferases, predominantly trimethylamine methyltransferases, during growth with betaine, whereas trimethylamine is one of the main end-products of betaine degradation. In methanol degradation methyltransferases are also involved. In methanol-utilizing methanogens, two methyltransferases catalyse methanol conversion, methyltransferase 1 composed of subunits MtaB and MtaC and methyltransferase 2, also called MtaA. The two methyltransferase 1 subunits MtaB and MtaC were highly abundant when strain An4 was grown with methanol. However, instead of MtaA a methyltetrahydrofolate methyltransferase was synthesized. We propose a novel methanol degradation pathway in Sporomusa strain An4 that uses a methyltetrahydrofolate methyltransferase instead of MtaA. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  2. Adaptation of the autotrophic acetogen Sporomusa ovata to methanol accelerates the conversion of CO2 to organic products

    PubMed Central

    Tremblay, Pier-Luc; Höglund, Daniel; Koza, Anna; Bonde, Ida; Zhang, Tian

    2015-01-01

    Acetogens are efficient microbial catalysts for bioprocesses converting C1 compounds into organic products. Here, an adaptive laboratory evolution approach was implemented to adapt Sporomusa ovata for faster autotrophic metabolism and CO2 conversion to organic chemicals. S. ovata was first adapted to grow quicker autotrophically with methanol, a toxic C1 compound, as the sole substrate. Better growth on different concentrations of methanol and with H2-CO2 indicated the adapted strain had a more efficient autotrophic metabolism and a higher tolerance to solvent. The growth rate on methanol was increased 5-fold. Furthermore, acetate production rate from CO2 with an electrode serving as the electron donor was increased 6.5-fold confirming that the acceleration of the autotrophic metabolism of the adapted strain is independent of the electron donor provided. Whole-genome sequencing, transcriptomic, and biochemical studies revealed that the molecular mechanisms responsible for the novel characteristics of the adapted strain were associated with the methanol oxidation pathway and the Wood-Ljungdahl pathway of acetogens along with biosynthetic pathways, cell wall components, and protein chaperones. The results demonstrate that an efficient strategy to increase rates of CO2 conversion in bioprocesses like microbial electrosynthesis is to evolve the microbial catalyst by adaptive laboratory evolution to optimize its autotrophic metabolism. PMID:26530351

  3. Clostridium huakuii sp. nov., an anaerobic, acetogenic bacterium isolated from methanogenic consortia.

    PubMed

    Ruan, Zhiyong; Wang, Yanwei; Zhang, Chi; Song, Jinlong; Zhai, Yi; Zhuang, Yan; Wang, Huimin; Chen, Xiaorong; Li, Yanting; Zhao, Bingqiang; Zhao, Bin

    2014-12-01

    A Gram-staining-positive, spore-forming, obligately anaerobic, acetogenic bacterium, designated LAM1030(T), was isolated from methanogenic consortia enriched from biogas slurry collected from the large-scale anaerobic digester of Modern Farming Corporation in Hebei Province, China. Cells of strain LAM1030(T) were motile, straight or spiral-rod-shaped. Strain LAM1030(T) could utilize glucose, fructose, maltose, galactose, lactose, sucrose, cellobiose, mannitol, pyruvate, succinic acid and tryptophan as the sole carbon source. Acetic acid, isovaleric acid and butanoic acid were the main products of glucose fermentation. Sodium sulfite was used as an electron acceptor. Growth of strain LAM1030(T) was completely inhibited by the addition of ampicillin, tetracycline, gentamicin or erythromycin at a concentration of 20 µg ml(-1). The main polar lipids of strain LAM1030(T) were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, 11 unknown glycolipids and two unknown phospholipids. No respiratory quinone was detected. The major fatty acids of strain LAM1030(T) were C16 : 0 (21.1 %), C14 : 0 (10.3 %), summed feature 9 (including C16:0 10-methyl and/or iso-C17:1 ω9c) (11.3% ), summed feature 3 (including C16:1 ω7c and/or C16:1 ω6c) (10.6% ) and iso-C15 : 0 (6.6 %). Analysis of the 16S rRNA gene sequence indicated that strain LAM1030(T) belonged to the genus Clostridium and was most closely related to Clostridium subterminale DSM 6970(T), Clostridium thiosulfatireducens DSM 13105(T) and Clostridium sulfidigenes DSM 18982(T), with 97.0, 96.9 and 96.8 % similarity, respectively. The G+C content of the genomic DNA of strain LAM1030(T) was 31.2±0.3 mol%. On the basis of its phenotypic, phylogenetic and chemotaxonomic characterization, strain LAM1030(T) is suggested to represent a novel species of the genus Clostridium, for which the name Clostridium huakuii sp. nov. is proposed. The type strain is LAM1030(T) ( = ACCC 00698(T

  4. Catabolite pathway for the production of skatole and indole acetic acid by the acetogen Clostridium drakei, Clostridium scatologenes, and swine manure

    USDA-ARS?s Scientific Manuscript database

    Skatole (3-methyl-indole) is a malodorous chemical in stored swine manure and is implicated as a component of foul tasting pork. Definitive evidence for the skatole pathway is lacking. Deuterium-labeled substrates were employed to resolve this pathway in the acetogenic bacterium Clostridium drakei...

  5. Genome tailoring powered production of isobutanol in continuous CO2/H2 blend fermentation using engineered acetogen biocatalyst.

    PubMed

    Gak, Eugene; Tyurin, Michael; Kiriukhin, Michael

    2014-05-01

    The cell energy fraction that powered maintenance and expression of genes encoding pro-phage elements, pta-ack cluster, early sporulation, sugar ABC transporter periplasmic proteins, 6-phosphofructokinase, pyruvate kinase, and fructose-1,6-disphosphatase in acetogen Clostridium sp. MT871 was re-directed to power synthetic operon encoding isobutanol biosynthesis at the expense of these genes achieved via their elimination. Genome tailoring decreased cell duplication time by 7.0 ± 0.1 min (p < 0.05) compared to the parental strain, with intact genome and cell duplication time of 68 ± 1 min (p < 0.05). Clostridium sp. MT871 with tailored genome was UVC-mutated to withstand 6.1 % isobutanol in fermentation broth to prevent product inhibition in an engineered commercial biocatalyst producing 5 % (674.5 mM) isobutanol during two-step continuous fermentation of CO2/H2 gas blend. Biocatalyst Clostridium sp. MT871RG- 11IBR6 was engineered to express six copies of synthetic operon comprising optimized synthetic format dehydrogenase, pyruvate formate lyase, acetolactate synthase, acetohydroxyacid reductoisomerase, 2,3-dihydroxy-isovalerate dehydratase, branched-chain alpha-ketoacid decarboxylase gene, aldehyde dehydrogenase, and alcohol dehydrogenase, regaining cell duplication time of 68 ± 1 min (p < 0.05) for the parental strain. This is the first report on isobutanol production by an engineered acetogen biocatalyst suitable for commercial manufacturing of this chemical/fuel using continuous fermentation of CO2/H2 blend thus contributing to the reversal of global warming.

  6. Reconstruction of an Acetogenic 2,3-Butanediol Pathway Involving a Novel NADPH-Dependent Primary-Secondary Alcohol Dehydrogenase

    PubMed Central

    Köpke, Michael; Gerth, Monica L.; Maddock, Danielle J.; Mueller, Alexander P.; Liew, FungMin

    2014-01-01

    Acetogenic bacteria use CO and/or CO2 plus H2 as their sole carbon and energy sources. Fermentation processes with these organisms hold promise for producing chemicals and biofuels from abundant waste gas feedstocks while simultaneously reducing industrial greenhouse gas emissions. The acetogen Clostridium autoethanogenum is known to synthesize the pyruvate-derived metabolites lactate and 2,3-butanediol during gas fermentation. Industrially, 2,3-butanediol is valuable for chemical production. Here we identify and characterize the C. autoethanogenum enzymes for lactate and 2,3-butanediol biosynthesis. The putative C. autoethanogenum lactate dehydrogenase was active when expressed in Escherichia coli. The 2,3-butanediol pathway was reconstituted in E. coli by cloning and expressing the candidate genes for acetolactate synthase, acetolactate decarboxylase, and 2,3-butanediol dehydrogenase. Under anaerobic conditions, the resulting E. coli strain produced 1.1 ± 0.2 mM 2R,3R-butanediol (23 μM h−1 optical density unit−1), which is comparable to the level produced by C. autoethanogenum during growth on CO-containing waste gases. In addition to the 2,3-butanediol dehydrogenase, we identified a strictly NADPH-dependent primary-secondary alcohol dehydrogenase (CaADH) that could reduce acetoin to 2,3-butanediol. Detailed kinetic analysis revealed that CaADH accepts a range of 2-, 3-, and 4-carbon substrates, including the nonphysiological ketones acetone and butanone. The high activity of CaADH toward acetone led us to predict, and confirm experimentally, that C. autoethanogenum can act as a whole-cell biocatalyst for converting exogenous acetone to isopropanol. Together, our results functionally validate the 2,3-butanediol pathway from C. autoethanogenum, identify CaADH as a target for further engineering, and demonstrate the potential of C. autoethanogenum as a platform for sustainable chemical production. PMID:24657865

  7. Rapid enrichment of (homo)acetogenic consortia from animal feces using a high mass-transfer gas-lift reactor fed with syngas.

    PubMed

    Park, Shinyoung; Yasin, Muhammad; Kim, Daehee; Park, Hee-Deung; Kang, Chang Min; Kim, Duk Jin; Chang, In Seop

    2013-09-01

    A gas-lift reactor having a high mass transfer coefficient (k(L)a = 80.28 h(-1)) for a relatively insoluble gas (carbon monoxide; CO) was used to enrich (homo)acetogens from animal feces. Samples of fecal matter from cow, rabbit, chicken, and goat were used as sources of inoculum for the enrichment of CO and H(2) utilizing microbial consortia. To confirm the successful enrichment, the Hungate roll tube technique was employed to count and then isolate putative CO utilizers. The results of this work showed that CO and H(2) utilizing consortia were established for each inoculum source after 8 days. The number of colony-forming units in cow, rabbit, chicken, and goat fecal samples were 3.83 × 10(9), 1.03 × 10(9), 8.3 × 10(8), and 3.25 × 10(8) cells/ml, respectively. Forty-two colonies from the animal fecal samples were screened for the ability to utilize CO/H(2). Ten of these 42 colonies were capable of utilizing CO/H(2). Five isolates from cow feces (samples 5, 6, 8, 16, and 22) were highly similar to previously unknown (homo)acetogen, while cow-7 has shown 99 % similarity with Acetobacterium sp. as acetogens. On the other hand, four isolates from chicken feces (samples 3, 8, 10, and 11) have also shown high CO/H(2) utilizing activity. Hence, it is expected that this research could be used as the basis for the rapid enrichment of (homo)acetogenic consortia from various environmental sources.

  8. Spectroscopic elucidation of energy transfer in hybrid inorganic-biological organisms for solar-to-chemical production.

    PubMed

    Kornienko, Nikolay; Sakimoto, Kelsey K; Herlihy, David M; Nguyen, Son C; Alivisatos, A Paul; Harris, Charles B; Schwartzberg, Adam; Yang, Peidong

    2016-10-18

    The rise of inorganic-biological hybrid organisms for solar-to-chemical production has spurred mechanistic investigations into the dynamics of the biotic-abiotic interface to drive the development of next-generation systems. The model system, Moorella thermoacetica-cadmium sulfide (CdS), combines an inorganic semiconductor nanoparticle light harvester with an acetogenic bacterium to drive the photosynthetic reduction of CO2 to acetic acid with high efficiency. In this work, we report insights into this unique electrotrophic behavior and propose a charge-transfer mechanism from CdS to M. thermoacetica Transient absorption (TA) spectroscopy revealed that photoexcited electron transfer rates increase with increasing hydrogenase (H2ase) enzyme activity. On the same time scale as the TA spectroscopy, time-resolved infrared (TRIR) spectroscopy showed spectral changes in the 1,700-1,900-cm(-1) spectral region. The quantum efficiency of this system for photosynthetic acetic acid generation also increased with increasing H2ase activity and shorter carrier lifetimes when averaged over the first 24 h of photosynthesis. However, within the initial 3 h of photosynthesis, the rate followed an opposite trend: The bacteria with the lowest H2ase activity photosynthesized acetic acid the fastest. These results suggest a two-pathway mechanism: a high quantum efficiency charge-transfer pathway to H2ase generating H2 as a molecular intermediate that dominates at long time scales (24 h), and a direct energy-transducing enzymatic pathway responsible for acetic acid production at short time scales (3 h). This work represents a promising platform to utilize conventional spectroscopic methodology to extract insights from more complex biotic-abiotic hybrid systems.

  9. Bioconversion of H2/CO 2 by acetogen enriched cultures for acetate and ethanol production: the impact of pH.

    PubMed

    Xu, Shuyun; Fu, Bo; Zhang, Lijuan; Liu, He

    2015-06-01

    Syngas fermentation into ethanol and other bioproducts by mixed cultures is considered a promising biotechnology. Effects of pH on product generation and microbial community during H2/CO2 utilization by acetogen enrichment cultures were investigated in this work. The maximum acetate concentration reached 95.41 mmol L(-1) at pH 7, which was 71.7, 21.8 and 50.9% higher than at pH 5, 9 and 11, respectively. The maximum ethanol concentration at pH 7 was 45.7, 50, 72% higher than that at pH 5, 9 and 11, respectively. The CO dehydrogenase (CODH) gene copy number was highest at pH 7, indicating that metabolically active acetogens reached their highest level at pH 7. The CODH gene copy number at pH 9 was lower than at pH 7, but higher than at pH 5 and 11. Correspondingly, the enrichment cultures at pH 7 had the highest species richness and diversity, while those at pH 9 had the second highest diversity, and those at pH 5 and 11 had the lowest diversity. The shift in microbial community structure and the different active acetogen contents resulting from different pHs were responsible for the differences in acetate and ethanol production.

  10. Improved conversion efficiencies for n-fatty acid reduction to primary alcohols by the solventogenic acetogen "Clostridium ragsdalei".

    PubMed

    Isom, Catherine E; Nanny, Mark A; Tanner, Ralph S

    2015-01-01

    "Clostridium ragsdalei" is an acetogen that ferments synthesis gas (syngas, predominantly H2:CO2:CO) to ethanol, acetate, and cell mass. Previous research showed that C. ragsdalei could also convert propionic acid to 1-propanol and butyric acid to 1-butanol at conversion efficiencies of 72.3 and 21.0 percent, respectively. Our research showed that C. ragsdalei can also reduce pentanoic and hexanoic acid to the corresponding primary alcohols. This reduction occurred independently of growth in an optimized medium with headspace gas exchange (vented and gassed with CO) every 48 h. Under these conditions, conversion efficiencies increased to 97 and 100 % for propionic and butyric acid, respectively. The conversion efficiencies for pentanoic and hexanoic acid to 1-pentanol and 1-hexanol, respectively, were 82 and 62 %. C. ragsdalei also reduced acetone to 2-propanol at a conversion efficiency of 100 %. Further, we showed that C. ragsdalei uses an aldehyde oxidoreductase-like enzyme to reduce n-fatty acids to the aldehyde intermediates in a reaction that requires ferredoxin and exogenous CO.

  11. Alkalibaculum bacchi gen. nov., sp. nov., a CO-oxidizing, ethanol-producing acetogen isolated from livestock-impacted soil.

    PubMed

    Allen, Toby D; Caldwell, Matthew E; Lawson, Paul A; Huhnke, Raymond L; Tanner, Ralph S

    2010-10-01

    Phenotypic and phylogenetic studies were performed on three strains of an acetogenic bacterium isolated from livestock-impacted soil. The bacterium stained Gram-negative and was a non-spore-forming rod that was motile by peritrichous flagella. The novel strains had an optimum pH for growth of 8.0-8.5 and utilized H₂ : CO₂, CO : CO₂, glucose, fructose, mannose, turanose, ribose, trimethylamine, pyruvate, methanol, ethanol, n-propanol and n-butanol as growth substrates. Acetate was produced from glucose. Acetate, CO₂ and ethanol were produced from CO : CO₂. 16S rRNA gene sequence analysis indicated that the novel strains formed a new subline in the family Eubacteriaceae (rRNA cluster XV) of the low G+C-containing Gram-positive bacteria of the class Clostridia. The DNA G+C base composition was 34 mol%. Cell wall analysis revealed the existence of a novel B-type peptidoglycan similar to the B2α-type (B4) configuration with a variation containing aspartic acid. Based on phylogenetic and phenotypic evidence, it is proposed that the new isolates represent a novel genus and species, for which the name Alkalibaculum bacchi gen. nov., sp. nov. is proposed. The type strain of the type species is CP11(T) (=ATCC BAA-1772(T)=DSM 22112(T)).

  12. A novel route for ethanol oxidation in the acetogenic bacterium Acetobacterium woodii: the acetaldehyde/ethanol dehydrogenase pathway.

    PubMed

    Bertsch, Johannes; Siemund, Anna Lena; Kremp, Florian; Müller, Volker

    2016-09-01

    Ethanol is a common substrate for anaerobic microorganisms despite its high redox potential (E0' etha- nol/acetaldehyde = -190mV), which does not allow for NAD(+) reduction. How this thermodynamic barrier is overcome is largely unknown. The acetogenic bacterium Acetobacterium woodii can also grow on ethanol. The genome harbours 11 genes encoding putative alcohol dehydrogenases, but only one, adhE, was upregulated during growth on ethanol. The bifunctional acetaldehyde/ethanol dehydrogenase (AdhE) was purified from ethanol-grown cells. It catalysed the NAD(+) - and CoA-dependent oxidation of ethanol via acetaldehyde to acetyl-CoA. The enzyme was regulated by free coenzyme A: in the absence of coenzyme A, the Km value for ethanol was shifted from 3.4 to 40 mM. The alcohol dehydrogenase domain could also oxidize 1-propanol and 1-butanol; however, the aldehyde dehydrogenase domain was highly specific for acetaldehyde as substrate. Apparently, the bifunctional AdhE allows for NAD(+) reduction by lowering the concentration of acetaldehyde, which makes the first oxidation reaction thermodynamically feasible. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  13. Visualizing molecular juggling within a B[subscript 12]-dependent methyltransferase complex

    SciTech Connect

    Kung, Yan; Ando, Nozomi; Doukov, Tzanko I.; Blasiak, Leah C.; Bender, Güne; #351; Seravalli, Javier; Ragsdale, Stephen W.; Drennan, Catherine L.

    2013-04-08

    Derivatives of vitamin B{sub 12} are used in methyl group transfer in biological processes as diverse as methionine synthesis in humans and CO{sub 2} fixation in acetogenic bacteria. This seemingly straightforward reaction requires large, multimodular enzyme complexes that adopt multiple conformations to alternately activate, protect and perform catalysis on the reactive B{sub 12} cofactor. Crystal structures determined thus far have provided structural information for only fragments of these complexes, inspiring speculation about the overall protein assembly and conformational movements inherent to activity. Here we present X-ray crystal structures of a complete 220 kDa complex that contains all enzymes responsible for B{sub 12}-dependent methyl transfer, namely the corrinoid iron-sulphur protein and its methyltransferase from the model acetogen Moorella thermoacetica. These structures provide the first three-dimensional depiction of all protein modules required for the activation, protection and catalytic steps of B{sub 12}-dependent methyl transfer. In addition, the structures capture B{sub 12} at multiple locations between its 'resting' and catalytic positions, allowing visualization of the dramatic protein rearrangements that enable methyl transfer and identification of the trajectory for B{sub 12} movement within the large enzyme scaffold. The structures are also presented alongside in crystallo spectroscopic data, which confirm enzymatic activity within crystals and demonstrate the largest known conformational movements of proteins in a crystalline state. Taken together, this work provides a model for the molecular juggling that accompanies turnover and helps explain why such an elaborate protein framework is required for such a simple, yet biologically essential reaction.

  14. Visualising molecular juggling within a B12-dependent methyltransferase complex

    PubMed Central

    Kung, Yan; Ando, Nozomi; Doukov, Tzanko I.; Blasiak, Leah C.; Bender, Güneş; Seravalli, Javier; Ragsdale, Stephen W.; Drennan, Catherine L.

    2012-01-01

    Derivatives of vitamin B12 are used in methyl group transfer in biological processes as diverse as methionine synthesis in humans and CO2 fixation in acetogenic bacteria1–3. This seemingly straightforward reaction requires large, multimodular enzyme complexes that adopt multiple conformations to alternately activate, protect, and perform catalysis on the reactive B12 cofactor. Crystal structures determined thus far have provided structural information for only fragments of these complexes4–12, inspiring speculation regarding the overall protein assembly and conformational movements inherent to activity. Here we present X-ray crystal structures of a complete ~220 kDa complex that contains all enzymes responsible for B12-dependent methyltransfer, namely the corrinoid iron-sulfur protein (CFeSP) and its methyltransferase (MeTr) from the model acetogen Moorella thermoacetica. These structures provide the first three-dimensional depiction of all protein modules required for the activation, protection, and catalytic steps of B12-dependent methyltransfer. In addition, the structures capture B12 at multiple locations between its “resting” and catalytic positions, allowing visualisation of the dramatic protein rearrangements that enable methyltransfer and identification of the trajectory for B12 movement within the large enzyme scaffold. The structures are also presented alongside in crystallo UV-vis spectroscopic data, which confirm enzymatic activity within crystals and demonstrate the largest known conformational movements of proteins in a crystalline state. Taken together, this work provides a model for the molecular juggling that accompanies turnover and helps explain why such an elaborate protein framework is required for such a simple, yet biologically essential reaction. PMID:22419154

  15. Tindallia californiensis sp. nov., a new anaerobic, haloalkaliphilic, spore-forming acetogen isolated from Mono Lake in California

    NASA Technical Reports Server (NTRS)

    Pikuta, E. V.; Hoover, R. B.; Bej, A. K.; Marsic, D.; Detkova, E. N.; Whitman, W. B.; Krader, P.

    2003-01-01

    A novel extremely haloalkaliphilic, strictly anaerobic, acetogenic bacterium strain APO was isolated from sediments of the athalassic, meromictic, alkaline Mono Lake in California. The Gram-positive, spore-forming, slightly curved rods with sizes 0.55- 0.7x1.7-3.0 microns were motile by a single laterally attached flagellum. Strain APO was mesophilic (range 10-48 C, optimum of 37 C); halophilic (NaCl range 1-20% (w/v) with optimum of 3-5% (w/v), and alkaliphilic (pH range 8.0-10.5, optimum 9.5). The novel isolate required sodium ions in the medium. Strain APO was an organotroph with a fermentative type of metabolism and used the substrates peptone, bacto-tryptone, casamino acid, yeast extract, L-serine, L-lysine, L-histidine, L-arginine, and pyruvate. The new isolate performed the Stickland reaction with the following amino acid pairs: proline + alanine, glycine + alanine, and tryptophan + valine. The main end product of growth was acetate. High activity of CO dehydrogenase and hydrogenase indicated the presence of a homoacetogenic, non-cycling acetyl-coA pathway. Strain APO was resistant to kanamycin but sensitive to chloramphenicol, tetracycline, and gentamycin. The G+C content of the genomic DNA was 44.4 mol% (by HPLC method). The sequence of the 16s rRNA gene of strain APO possessed 98.2% similarity with the sequence from Tindullia magadiensis Z-7934, but the DNA-DNA hybridization value between these organisms was only 55%. On the basis of these physiological and molecular properties, strain APO is proposed to be a novel species of the genus Tindallia with the name Tindallia californiensis sp. nov., (type strain APO = ATCC BAA-393 - DSM 14871).

  16. Tindallia californiensis sp. nov., a new anaerobic, haloalkaliphilic, spore-forming acetogen isolated from Mono Lake in California

    NASA Technical Reports Server (NTRS)

    Pikuta, E. V.; Hoover, R. B.; Bej, A. K.; Marsic, D.; Detkova, E. N.; Whitman, W. B.; Krader, P.

    2003-01-01

    A novel extremely haloalkaliphilic, strictly anaerobic, acetogenic bacterium strain APO was isolated from sediments of the athalassic, meromictic, alkaline Mono Lake in California. The Gram-positive, spore-forming, slightly curved rods with sizes 0.55- 0.7x1.7-3.0 microns were motile by a single laterally attached flagellum. Strain APO was mesophilic (range 10-48 C, optimum of 37 C); halophilic (NaCl range 1-20% (w/v) with optimum of 3-5% (w/v), and alkaliphilic (pH range 8.0-10.5, optimum 9.5). The novel isolate required sodium ions in the medium. Strain APO was an organotroph with a fermentative type of metabolism and used the substrates peptone, bacto-tryptone, casamino acid, yeast extract, L-serine, L-lysine, L-histidine, L-arginine, and pyruvate. The new isolate performed the Stickland reaction with the following amino acid pairs: proline + alanine, glycine + alanine, and tryptophan + valine. The main end product of growth was acetate. High activity of CO dehydrogenase and hydrogenase indicated the presence of a homoacetogenic, non-cycling acetyl-coA pathway. Strain APO was resistant to kanamycin but sensitive to chloramphenicol, tetracycline, and gentamycin. The G+C content of the genomic DNA was 44.4 mol% (by HPLC method). The sequence of the 16s rRNA gene of strain APO possessed 98.2% similarity with the sequence from Tindullia magadiensis Z-7934, but the DNA-DNA hybridization value between these organisms was only 55%. On the basis of these physiological and molecular properties, strain APO is proposed to be a novel species of the genus Tindallia with the name Tindallia californiensis sp. nov., (type strain APO = ATCC BAA-393 - DSM 14871).

  17. Tindallia californiensis sp. nov., a new anaerobic, haloalkaliphilic, spore-forming acetogen isolated from Mono Lake in California.

    PubMed

    Pikuta, Elena V; Hoover, Richard B; Bej, Asim K; Marsic, Damien; Detkova, Ekaterina N; Whitman, William B; Krader, Paul

    2003-08-01

    A novel extremely haloalkaliphilic, strictly anaerobic, acetogenic bacterium strain APO was isolated from sediments of the athalassic, meromictic, alkaline Mono Lake in California. The Gram-positive, spore-forming, slightly curved rods with sizes 0.55-0.7x1.7-3.0 microm were motile by a single laterally attached flagellum. Strain APO was mesophilic (range 10-48 degrees C, optimum of 37 degrees C); halophilic (NaCl range 1-20% (w/v) with optimum of 3-5% (w/v), and alkaliphilic (pH range 8.0-10.5, optimum 9.5). The novel isolate required sodium ions in the medium. Strain APO was an organotroph with a fermentative type of metabolism and used the substrates peptone, bacto-tryptone, casamino acid, yeast extract, l-serine, l-lysine, l-histidine, l-arginine, and pyruvate. The new isolate performed the Stickland reaction with the following amino acid pairs: proline + alanine, glycine + alanine, and tryptophan + valine. The main end product of growth was acetate. High activity of CO dehydrogenase and hydrogenase indicated the presence of a homoacetogenic, non-cycling acetyl-CoA pathway. Strain APO was resistant to kanamycin but sensitive to chloramphenicol, tetracycline, and gentamycin. The G+C content of the genomic DNA was 44.4 mol% (by HPLC method). The sequence of the 16S rRNA gene of strain APO possessed 98.2% similarity with the sequence from Tindallia magadiensis Z-7934, but the DNA-DNA hybridization value between these organisms was only 55%. On the basis of these physiological and molecular properties, strain APO is proposed to be a novel species of the genus Tindallia with the name Tindallia californiensis sp. nov., (type strain APO = ATCC BAA-393 = DSM 14871).

  18. Quantitative Analysis of Three Hydrogenotrophic Microbial Groups, Methanogenic Archaea, Sulfate-Reducing Bacteria, and Acetogenic Bacteria, within Plaque Biofilms Associated with Human Periodontal Disease▿

    PubMed Central

    Vianna, M. E.; Holtgraewe, S.; Seyfarth, I.; Conrads, G.; Horz, H. P.

    2008-01-01

    Human subgingival plaque biofilms are highly complex microbial ecosystems that may depend on H2-metabolizing processes. Here we investigated the ubiquity and proportions of methanogenic archaea, sulfate reducers, and acetogens in plaque samples from 102 periodontitis patients. In contrast to the case for 65 healthy control subjects, hydrogenotrophic groups were almost consistently detected in periodontal pockets, with the proportions of methanogens and sulfate reducers being significantly elevated in severe cases. In addition, antagonistic interactions among the three microbial groups indicated that they may function as alternative syntrophic partners of secondary fermenting periodontal pathogens. PMID:18326571

  19. Selective methanol or formate production during continuous CO₂ fermentation by the acetogen biocatalysts engineered via integration of synthetic pathways using Tn7-tool.

    PubMed

    Tyurin, Michael; Kiriukhin, Michael

    2013-09-01

    Methanol-resistant mutant acetogen Clostridium sp. MT1424 originally producing only 365 mM acetate from CO₂/CO was engineered to eliminate acetate production and spore formation using Cre-lox66/lox71-system to power subsequent methanol production via expressing synthetic methanol dehydrogenase, formaldehyde dehydrogenase and formate dehydrogenase, three copies of each, assembled in cluster and integrated to chromosome using Tn7-based approach. Production of 2.2 M methanol was steady (p < 0.005) in single step fermentations of 20 % CO₂ + 80 % H₂ blend (v/v) 25 day runs each in five independent repeats. If the integrated cluster comprised only three copies of formate dehydrogenase the respective recombinants produced 95 mM formate (p < 0.005) under the same conditions. For commercialization, the suggested source of inorganic carbon would be CO₂ waste of IGCC power plant. Hydrogen may be produced in situ via powered by solar panels electrolysis.

  20. Diversity of corrinoids in acetogenic bacteria. P-cresolylcobamide from Sporomusa ovata, 5-methoxy-6-methylbenzimidazolylcobamide from Clostridium formicoaceticum and vitamin B12 from Acetobacterium woodii.

    PubMed

    Stupperich, E; Eisinger, H J; Kräutler, B

    1988-03-01

    The Co beta-cyanocobamides obtained by cyanide extractions from several acetogenic bacteria were structurally characterized by ultraviolet/visible spectra, proton-nuclear-magnetic-resonance spectra and fast-atom-bombardment mass spectra. p-Cresolycobamide was detected as a major corrinoid from Sporomusa ovata. This 'complete' corrinoid was isolated from an organism for the first time. Instead of the common Co alpha bases of the known and biologically active cobamides, p-cresolylcobamide contained a glycosidically bound cresolyl function that was unable to coordinate to the cobalt of the corrin ring. An additional, previously unknown corrinoid from natural sources, Co alpha-[alpha-(5-methoxy-6-methylbenzimidazolyl)]-Co beta-cyanocobamide, was isolated along with vitamin B12 from Clostridium formicoaceticum. Both homoacetogenic eubacteria were grown on methanol and contained high amounts of corrinoids (greater than 950 nmol/g cell dry mass). Less corrinoid was isolated from Acetobacterium woodii and characterized as vitamin B12.

  1. Polycyclic Aromatic Hydrocarbon Affects Acetic Acid Production during Anaerobic Fermentation of Waste Activated Sludge by Altering Activity and Viability of Acetogen.

    PubMed

    Luo, Jingyang; Chen, Yinguang; Feng, Leiyu

    2016-07-05

    Till now, almost all the studies on anaerobic fermentation of waste activated sludge (WAS) for bioproducts generation focused on the influences of operating conditions, pretreatment methods and sludge characteristics, and few considered those of widespread persistent organic pollutants (POPs) in sludge, for example, polycyclic aromatic hydrocarbons (PAHs). Herein, phenanthrene, which was a typical PAH and widespread in WAS, was selected as a model compound to investigate its effect on WAS anaerobic fermentation for short-chain fatty acids (SCFAs) accumulation. Experimental results showed that the concentration of SCFAs derived from WAS was increased in the presence of phenanthrene during anaerobic fermentation. The yield of acetic acid which was the predominant SCFA in the fermentation reactor with the concentration of 100 mg/kg dry sludge was 1.8 fold of that in the control. Mechanism exploration revealed that the present phenanthrene mainly affected the acidification process of anaerobic fermentation and caused the shift of the microbial community to benefit the accumulation of acetic acid. Further investigation showed that both the activities of key enzymes (phosphotransacetylase and acetate kinase) involved in acetic acid production and the quantities of their corresponding encoding genes were enhanced in the presence of phenanthrene. Viability tests by determining the adenosine 5'-triphosphate content and membrane potential confirmed that the acetogens were more viable in anaerobic fermentation systems with phenanthrene, which resulted in the increased production of acetic acid.

  2. "Curing" of plasmid DNA in acetogen using microwave or applying an electric pulse improves cell growth and metabolite production as compared to the plasmid-harboring strain.

    PubMed

    Berzin, Vel; Kiriukhin, Michael; Tyurin, Michael

    2013-03-01

    Plasmid-free acetogen Clostridium sp. MT962 electrotransformed with a small cryptic plasmid pMT351 was used to develop time- and cost-effective methods for plasmid elimination. Elimination of pMT351 restored production of acetate and ethanol to the levels of the plasmid-free strain with no dry cell weight changes. Destabilizing cell membrane via microwave at 2.45 GHz, or exposure to a single 12 ms square electric pulse at 35 kV cm⁻¹, eliminated pMT351 in 42-47 % of cells. Plasmid elimination with a single square electric pulse required 10 versus 0.1 J needed to introduce the same 3,202-bp plasmid into the cells as calculated per cell sample of Clostridium sp. MT962. Microwave caused visible changes in repPCR pattern and increased ethanol production at the expense of acetate. This is the first report on microwave of microwave ovens, wireless routers, and mobile devices causing chromosomal DNA aberrations in microbes along with carbon flux change.

  3. Integrated bioprocess for conversion of gaseous substrates to liquids

    PubMed Central

    Hu, Peng; Chakraborty, Sagar; Kumar, Amit; Woolston, Benjamin; Liu, Hongjuan; Emerson, David; Stephanopoulos, Gregory

    2016-01-01

    In the quest for inexpensive feedstocks for the cost-effective production of liquid fuels, we have examined gaseous substrates that could be made available at low cost and sufficiently large scale for industrial fuel production. Here we introduce a new bioconversion scheme that effectively converts syngas, generated from gasification of coal, natural gas, or biomass, into lipids that can be used for biodiesel production. We present an integrated conversion method comprising a two-stage system. In the first stage, an anaerobic bioreactor converts mixtures of gases of CO2 and CO or H2 to acetic acid, using the anaerobic acetogen Moorella thermoacetica. The acetic acid product is fed as a substrate to a second bioreactor, where it is converted aerobically into lipids by an engineered oleaginous yeast, Yarrowia lipolytica. We first describe the process carried out in each reactor and then present an integrated system that produces microbial oil, using synthesis gas as input. The integrated continuous bench-scale reactor system produced 18 g/L of C16-C18 triacylglycerides directly from synthesis gas, with an overall productivity of 0.19 g⋅L−1⋅h−1 and a lipid content of 36%. Although suboptimal relative to the performance of the individual reactor components, the presented integrated system demonstrates the feasibility of substantial net fixation of carbon dioxide and conversion of gaseous feedstocks to lipids for biodiesel production. The system can be further optimized to approach the performance of its individual units so that it can be used for the economical conversion of waste gases from steel mills to valuable liquid fuels for transportation. PMID:26951649

  4. Spectroscopic elucidation of energy transfer in hybrid inorganic–biological organisms for solar-to-chemical production

    SciTech Connect

    Kornienko, Nikolay; Sakimoto, Kelsey K.; Herlihy, David M.; Nguyen, Son C.; Alivisatos, A. Paul; Harris, Charles. B.; Schwartzberg, Adam; Yang, Peidong

    2016-10-03

    We present that the rise of inorganic–biological hybrid organisms for solar-to-chemical production has spurred mechanistic investigations into the dynamics of the biotic–abiotic interface to drive the development of next-generation systems. The model system, Moorella thermoacetica–cadmium sulfide (CdS), combines an inorganic semiconductor nanoparticle light harvester with an acetogenic bacterium to drive the photosynthetic reduction of CO2 to acetic acid with high efficiency. In this work, we report insights into this unique electrotrophic behavior and propose a charge-transfer mechanism from CdS to M. thermoacetica. Transient absorption (TA) spectroscopy revealed that photoexcited electron transfer rates increase with increasing hydrogenase (H2ase) enzyme activity. On the same time scale as the TA spectroscopy, time-resolved infrared (TRIR) spectroscopy showed spectral changes in the 1,700–1,900-cm-1 spectral region. The quantum efficiency of this system for photosynthetic acetic acid generation also increased with increasing H2ase activity and shorter carrier lifetimes when averaged over the first 24 h of photosynthesis. However, within the initial 3 h of photosynthesis, the rate followed an opposite trend: The bacteria with the lowest H2ase activity photosynthesized acetic acid the fastest. These results suggest a two-pathway mechanism: a high quantum efficiency charge-transfer pathway to H2ase generating H2 as a molecular intermediate that dominates at long time scales (24 h), and a direct energy-transducing enzymatic pathway responsible for acetic acid production at short time scales (3 h). Lastly, this work represents a promising platform to utilize conventional spectroscopic methodology to extract insights from more complex biotic–abiotic hybrid systems.

  5. Spectroscopic elucidation of energy transfer in hybrid inorganic–biological organisms for solar-to-chemical production

    DOE PAGES

    Kornienko, Nikolay; Sakimoto, Kelsey K.; Herlihy, David M.; ...

    2016-10-03

    We present that the rise of inorganic–biological hybrid organisms for solar-to-chemical production has spurred mechanistic investigations into the dynamics of the biotic–abiotic interface to drive the development of next-generation systems. The model system, Moorella thermoacetica–cadmium sulfide (CdS), combines an inorganic semiconductor nanoparticle light harvester with an acetogenic bacterium to drive the photosynthetic reduction of CO2 to acetic acid with high efficiency. In this work, we report insights into this unique electrotrophic behavior and propose a charge-transfer mechanism from CdS to M. thermoacetica. Transient absorption (TA) spectroscopy revealed that photoexcited electron transfer rates increase with increasing hydrogenase (H2ase) enzyme activity. Onmore » the same time scale as the TA spectroscopy, time-resolved infrared (TRIR) spectroscopy showed spectral changes in the 1,700–1,900-cm-1 spectral region. The quantum efficiency of this system for photosynthetic acetic acid generation also increased with increasing H2ase activity and shorter carrier lifetimes when averaged over the first 24 h of photosynthesis. However, within the initial 3 h of photosynthesis, the rate followed an opposite trend: The bacteria with the lowest H2ase activity photosynthesized acetic acid the fastest. These results suggest a two-pathway mechanism: a high quantum efficiency charge-transfer pathway to H2ase generating H2 as a molecular intermediate that dominates at long time scales (24 h), and a direct energy-transducing enzymatic pathway responsible for acetic acid production at short time scales (3 h). Lastly, this work represents a promising platform to utilize conventional spectroscopic methodology to extract insights from more complex biotic–abiotic hybrid systems.« less

  6. Integrated bioprocess for conversion of gaseous substrates to liquids.

    PubMed

    Hu, Peng; Chakraborty, Sagar; Kumar, Amit; Woolston, Benjamin; Liu, Hongjuan; Emerson, David; Stephanopoulos, Gregory

    2016-04-05

    In the quest for inexpensive feedstocks for the cost-effective production of liquid fuels, we have examined gaseous substrates that could be made available at low cost and sufficiently large scale for industrial fuel production. Here we introduce a new bioconversion scheme that effectively converts syngas, generated from gasification of coal, natural gas, or biomass, into lipids that can be used for biodiesel production. We present an integrated conversion method comprising a two-stage system. In the first stage, an anaerobic bioreactor converts mixtures of gases of CO2 and CO or H2 to acetic acid, using the anaerobic acetogen Moorella thermoacetica The acetic acid product is fed as a substrate to a second bioreactor, where it is converted aerobically into lipids by an engineered oleaginous yeast, Yarrowia lipolytica We first describe the process carried out in each reactor and then present an integrated system that produces microbial oil, using synthesis gas as input. The integrated continuous bench-scale reactor system produced 18 g/L of C16-C18 triacylglycerides directly from synthesis gas, with an overall productivity of 0.19 g⋅L(-1)⋅h(-1) and a lipid content of 36%. Although suboptimal relative to the performance of the individual reactor components, the presented integrated system demonstrates the feasibility of substantial net fixation of carbon dioxide and conversion of gaseous feedstocks to lipids for biodiesel production. The system can be further optimized to approach the performance of its individual units so that it can be used for the economical conversion of waste gases from steel mills to valuable liquid fuels for transportation.

  7. Spectroscopic elucidation of energy transfer in hybrid inorganic–biological organisms for solar-to-chemical production

    SciTech Connect

    Kornienko, Nikolay; Sakimoto, Kelsey K.; Herlihy, David M.; Nguyen, Son C.; Alivisatos, A. Paul; Harris, Charles. B.; Schwartzberg, Adam; Yang, Peidong

    2016-10-03

    We present that the rise of inorganic–biological hybrid organisms for solar-to-chemical production has spurred mechanistic investigations into the dynamics of the biotic–abiotic interface to drive the development of next-generation systems. The model system, Moorella thermoacetica–cadmium sulfide (CdS), combines an inorganic semiconductor nanoparticle light harvester with an acetogenic bacterium to drive the photosynthetic reduction of CO2 to acetic acid with high efficiency. In this work, we report insights into this unique electrotrophic behavior and propose a charge-transfer mechanism from CdS to M. thermoacetica. Transient absorption (TA) spectroscopy revealed that photoexcited electron transfer rates increase with increasing hydrogenase (H2ase) enzyme activity. On the same time scale as the TA spectroscopy, time-resolved infrared (TRIR) spectroscopy showed spectral changes in the 1,700–1,900-cm-1 spectral region. The quantum efficiency of this system for photosynthetic acetic acid generation also increased with increasing H2ase activity and shorter carrier lifetimes when averaged over the first 24 h of photosynthesis. However, within the initial 3 h of photosynthesis, the rate followed an opposite trend: The bacteria with the lowest H2ase activity photosynthesized acetic acid the fastest. These results suggest a two-pathway mechanism: a high quantum efficiency charge-transfer pathway to H2ase generating H2 as a molecular intermediate that dominates at long time scales (24 h), and a direct energy-transducing enzymatic pathway responsible for acetic acid production at short time scales (3 h). Lastly, this work represents a promising platform to utilize conventional spectroscopic methodology to extract insights from more complex biotic–abiotic hybrid systems.

  8. Exploring biochemical pathways for mono-ethylene glycol (MEG) synthesis from synthesis gas.

    PubMed

    Islam, M Ahsanul; Hadadi, Noushin; Ataman, Meric; Hatzimanikatis, Vassily; Stephanopoulos, Gregory

    2017-05-01

    Mono-ethylene glycol (MEG) is an important petrochemical with widespread use in numerous consumer products. The current industrial MEG-production process relies on non-renewable fossil fuel-based feedstocks, such as petroleum, natural gas, and naphtha; hence, it is useful to explore alternative routes of MEG-synthesis from gases as they might provide a greener and more sustainable alternative to the current production methods. Technologies of synthetic biology and metabolic engineering of microorganisms can be deployed for the expression of new biochemical pathways for MEG-synthesis from gases, provided that such promising alternative routes are first identified. We used the BNICE.ch algorithm to develop novel and previously unknown biological pathways to MEG from synthesis gas by leveraging the Wood-Ljungdahl pathway of carbon fixation of acetogenic bacteria. We developed a set of useful pathway pruning and analysis criteria to systematically assess thousands of pathways generated by BNICE.ch. Published genome-scale models of Moorella thermoacetica and Clostridium ljungdahlii were used to perform the pathway yield calculations and in-depth analyses of seven (7) newly developed biological MEG-producing pathways from gases, including CO2, CO, and H2. These analyses helped identify not only better candidate pathways, but also superior chassis organisms that can be used for metabolic engineering of the candidate pathways. The pathway generation, pruning, and detailed analysis procedures described in this study can also be used to develop biochemical pathways for other commodity chemicals from gaseous substrates. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  9. Energy Conservation Model Based on Genomic and Experimental Analyses of a Carbon Monoxide-Utilizing, Butyrate-Forming Acetogen, Eubacterium limosum KIST612

    PubMed Central

    Jeong, Jiyeong; Bertsch, Johannes; Hess, Verena; Choi, Sunju; Choi, In-Geol

    2015-01-01

    Eubacterium limosum KIST612 is one of the few acetogens that can produce butyrate from carbon monoxide. We have used a genome-guided analysis to delineate the path of butyrate formation, the enzymes involved, and the potential coupling to ATP synthesis. Oxidation of CO is catalyzed by the acetyl-coenzyme A (CoA) synthase/CO dehydrogenase and coupled to the reduction of ferredoxin. Oxidation of reduced ferredoxin is catalyzed by the Rnf complex and Na+ dependent. Consistent with the finding of a Na+-dependent Rnf complex is the presence of a conserved Na+-binding motif in the c subunit of the ATP synthase. Butyrate formation is from acetyl-CoA via acetoacetyl-CoA, hydroxybutyryl-CoA, crotonyl-CoA, and butyryl-CoA and is consistent with the finding of a gene cluster that encodes the enzymes for this pathway. The activity of the butyryl-CoA dehydrogenase was demonstrated. Reduction of crotonyl-CoA to butyryl-CoA with NADH as the reductant was coupled to reduction of ferredoxin. We postulate that the butyryl-CoA dehydrogenase uses flavin-based electron bifurcation to reduce ferredoxin, which is consistent with the finding of etfA and etfB genes next to it. The overall ATP yield was calculated and is significantly higher than the one obtained with H2 + CO2. The energetic benefit may be one reason that butyrate is formed only from CO but not from H2 + CO2. PMID:25956767

  10. Energy Conservation Model Based on Genomic and Experimental Analyses of a Carbon Monoxide-Utilizing, Butyrate-Forming Acetogen, Eubacterium limosum KIST612.

    PubMed

    Jeong, Jiyeong; Bertsch, Johannes; Hess, Verena; Choi, Sunju; Choi, In-Geol; Chang, In Seop; Müller, Volker

    2015-07-01

    Eubacterium limosum KIST612 is one of the few acetogens that can produce butyrate from carbon monoxide. We have used a genome-guided analysis to delineate the path of butyrate formation, the enzymes involved, and the potential coupling to ATP synthesis. Oxidation of CO is catalyzed by the acetyl-coenzyme A (CoA) synthase/CO dehydrogenase and coupled to the reduction of ferredoxin. Oxidation of reduced ferredoxin is catalyzed by the Rnf complex and Na(+) dependent. Consistent with the finding of a Na(+)-dependent Rnf complex is the presence of a conserved Na(+)-binding motif in the c subunit of the ATP synthase. Butyrate formation is from acetyl-CoA via acetoacetyl-CoA, hydroxybutyryl-CoA, crotonyl-CoA, and butyryl-CoA and is consistent with the finding of a gene cluster that encodes the enzymes for this pathway. The activity of the butyryl-CoA dehydrogenase was demonstrated. Reduction of crotonyl-CoA to butyryl-CoA with NADH as the reductant was coupled to reduction of ferredoxin. We postulate that the butyryl-CoA dehydrogenase uses flavin-based electron bifurcation to reduce ferredoxin, which is consistent with the finding of etfA and etfB genes next to it. The overall ATP yield was calculated and is significantly higher than the one obtained with H2 + CO2. The energetic benefit may be one reason that butyrate is formed only from CO but not from H2 + CO2.

  11. Gene replacement and elimination using λRed- and FLP-based tool to re-direct carbon flux in acetogen biocatalyst during continuous CO₂/H₂ blend fermentation.

    PubMed

    Tyurin, Michael

    2013-07-01

    A time- and cost-efficient two-step gene elimination procedure was used for acetogen Clostridium sp. MT1834 capable of fermenting CO₂/H₂ blend to 245 mM acetate (p < 0.005). The first step rendered the targeted gene replacement without affecting the total genome size. We replaced the acetate pta-ack cluster with synthetic bi-functional acetaldehyde-alcohol dehydrogenase (al-adh). Replacement of pta-ack with al-adh rendered initiation of 243 mM ethanol accumulation at the expense of acetate production during CO₂/H₂ blend continuous fermentation (p < 0.005). At the second step, al-adh was eliminated to reduce the genome size. Resulting recombinants accumulated 25 mM mevalonate in fermentation broth (p < 0.005). Cell duplication time for recombinants with reduced genome size decreased by 9.5 % compared to Clostridium sp. MT1834 strain under the same fermentation conditions suggesting better cell energy pool management in the absence of the ack-pta gene cluster in the engineered biocatalyst. If the first gene elimination step was used alone for spo0A gene replacement with two copies of synthetic formate dehydrogenase in recombinants with a shortened genome, mevalonate production was replaced with 76.5 mM formate production in a single step continuous CO₂/H₂ blend fermentation (p < 0.005) with cell duplication time almost nearing that of the wild strain.

  12. Isolation and characterization of a thermophilic bacterium which oxidizes acetate in syntrophic association with a methanogen and which grows acetogenically on H/sub 2/-CO/sub 2/

    SciTech Connect

    Lee, M.J.; Zinder, S.H.

    1988-01-01

    The authors previously described a thermophilic (60/sup 0/C), syntrophic, two-membered culture which converted acetate to methane via a two-step mechanism in which acetate was oxidized to H/sub 2/ and CO/sub 2/. While the hydrogenotrophic methanogen Methanobacterium sp. strain THF in the biculture was readily isolated, we were unable to find a substrate that was suitable for isolation of the acetate-oxidizing member of the biculture. In this study, we found that the biculture grew on ethylene glycol, and an acetate-oxidizing, rod-shape bacterium (AOR) was isolated from the biculture by dilution into medium containing ethylene glycol as the growth substrate. When the axenic culture of the AOR was recombined with a pure culture of Methanobacterium sp. strain THF, the reconstituted biculture grew on acetate and converted it to CH/sub 4/. The AOR used ethylene glycol, 1,2-propanediol, formate, pyruvate, glycine-betaine, and H/sub 2/-CO/sub 2/ as growth substrates. Acetate was the major fermentation product detected from these substrates, except for 1,2-propanediol, which was converted to 1-propanol and propionate. N,N-Dimethylglycine was also formed from glycine-betaine. Acetate was formed in stoichiometric amounts during growth on H/sub 2/-CO/sub 2/, demonstrating that the AOR is an acetogen. This reaction, which was carried out by the pure culture of the AOR in the presence of high partial pressures of H/sub 2/, was the reverse of the acetate oxidation reaction carried out by the AOR when hydrogen partial pressures were kept low by coculturing it with Methanobacterium sp. strain THF. The DNA base composition of the AOR was 47 mol% guanine plus cytosine, and no cytochromes were detected.

  13. Advanced modelling, monitoring, and process control of bioconversion systems

    NASA Astrophysics Data System (ADS)

    Schmitt, Elliott C.

    Production of fuels and chemicals from lignocellulosic biomass is an increasingly important area of research and industrialization throughout the world. In order to be competitive with fossil-based fuels and chemicals, maintaining cost-effectiveness is critical. Advanced process control (APC) and optimization methods could significantly reduce operating costs in the biorefining industry. Two reasons APC has previously proven challenging to implement for bioprocesses include: lack of suitable online sensor technology of key system components, and strongly nonlinear first principal models required to predict bioconversion behavior. To overcome these challenges batch fermentations with the acetogen Moorella thermoacetica were monitored with Raman spectroscopy for the conversion of real lignocellulosic hydrolysates and a kinetic model for the conversion of synthetic sugars was developed. Raman spectroscopy was shown to be effective in monitoring the fermentation of sugarcane bagasse and sugarcane straw hydrolysate, where univariate models predicted acetate concentrations with a root mean square error of prediction (RMSEP) of 1.9 and 1.0 g L-1 for bagasse and straw, respectively. Multivariate partial least squares (PLS) models were employed to predict acetate, xylose, glucose, and total sugar concentrations for both hydrolysate fermentations. The PLS models were more robust than univariate models, and yielded a percent error of approximately 5% for both sugarcane bagasse and sugarcane straw. In addition, a screening technique was discussed for improving Raman spectra of hydrolysate samples prior to collecting fermentation data. Furthermore, a mechanistic model was developed to predict batch fermentation of synthetic glucose, xylose, and a mixture of the two sugars to acetate. The models accurately described the bioconversion process with an RMSEP of approximately 1 g L-1 for each model and provided insights into how kinetic parameters changed during dual substrate

  14. ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS

    EPA Science Inventory

    Seagrasses are adapted to being rooted in reduced, anoxic sediments with high rates of sulfate reduction. During the day, an oxygen gradient is generated around the roots, becoming anoxic at night. Thus, obligate anaerobic bacteria in the rhizosphere have to tolerate elevated oxy...

  15. ACETOGENIC BACTERIA ASSOCIATED WITH SEAGRASS ROOTS

    EPA Science Inventory

    Seagrasses are adapted to being rooted in reduced, anoxic sediments with high rates of sulfate reduction. During the day, an oxygen gradient is generated around the roots, becoming anoxic at night. Thus, obligate anaerobic bacteria in the rhizosphere have to tolerate elevated oxy...

  16. Acetogenic microbial degradation of vinyl chloride

    USGS Publications Warehouse

    Bradley, P.M.; Chapelle, F.H.

    2000-01-01

    Under methanogenic conditions, microbial degradation of [1,2-14C]vinyl chloride (VC) resulted in significant (14 ?? 3% maximum recovery) but transient recovery of radioactivity as 14C-acetate. Subsequently, 14C- acetate was degraded to 14CH4 and 14CO2 (18 ?? 2% and 54 ?? 3% final recoveries, respectively). In contrast, under 2-bromoethanesulfonic acid (BES) amended conditions, 14C-acetate recovery remained high (27 ?? 1% maximum recovery) throughout the study, no 14CH4 was produced, and the final recovery of 14CO2 was only 35 ?? 4%. These results demonstrate that oxidative acetogenesis may be an important mechanism for anaerobic VC biodegradation. Moreover, these results (1) demonstrate that microbial degradation of VC to CH4 and CO2 may involve oxidative acetogenesis followed by acetotrophic methanogenesis and (2) suggest that oxidative acetogenesis may be the initial step in the net oxidation of VC to CO2 reported previously under Fe(III)-reducing, SO4-reducing, and humic acids- reducing conditions.Under methanogenic conditions, microbial degradation of [1,2-14C]vinyl chloride (VC) resulted in significant (14 ?? 3% maximum recovery) but transient recovery of radioactivity as 14C-acetate. Subsequently, 14C-acetate was degraded to 14CH4 and 14CO2 (18 ?? 2% and 54 ?? 3% final recoveries respectively). In contrast, under 2-bromoethanesulfonic acid (BES) amended conditions, 14C-acetate recovery remained high (27 ?? 1% maximum recovery) throughout the study, no 14CH4 was produced, and the final recovery of 14CO2 was only 35 ?? 4%. These results demonstrate that oxidative acetogenesis may be an important mechanism for anaerobic VC biodegradation. Moreover, these results (1) demonstrate that microbial degradation of VC to CH4 and CO2 may involve oxidative acetogenesis followed by acetotrophic methanogenesis and (2) suggest that oxidative acetogenesis may be the initial step in the net oxidation of VC to CO2 reported previously under Fe(III)-reducing, SO4-reducing, and humic acids-reducing conditions.

  17. Electricity and H2 generation from hemicellulose by sequential fermentation and microbial fuel/electrolysis cell

    NASA Astrophysics Data System (ADS)

    Yan, Di; Yang, Xuewei; Yuan, Wenqiao

    2015-09-01

    Electricity and hydrogen generation by bacteria Geobacter sulfurreducens in a dual-chamber microbial fuel/electrolysis cell following the fermentation of hemicellulose by bacteria Moorella thermoacetica was investigated. Experimental results showed that 10 g l-1 xylose under 60 °C was appropriate for the fermentation of xylose by M. thermoacetica, yielding 0.87 g-acetic acid per gram of xylose consumed. Corncob hydrolysate could also be fermented to produce acetic acid, but with lower yield (0.74 g-acid per g-xylose). The broths of xylose and corncob hydrolysate fermented by M. thermoacetica containing acetic acid were fed to G. sulfurreducens in a dual-chamber microbial fuel/electrolysis cell for electricity and hydrogen generation. The highest open-circuit cell voltages generated were 802 and 745 mV, and hydrogen yields were 41.7 and 23.3 mmol per mol-acetate, in xylose and corncob hydrolysate fermentation broth media, respectively. The internal resistance of the microbial fuel/electrolysis cell fed with corncob hydrolysate fermentation broth (3472 Ω) was much higher than that with xylose fermentation broth (1993 Ω) or sodium acetate medium (467 Ω), which was believed to be the main cause of the variation in hydrogen yield of the three feeding media.

  18. Evaluation of peracetic acid sanitizers efficiency against spores isolated from spoiled cans in suspension and on stainless steel surfaces.

    PubMed

    André, S; Hédin, S; Remize, F; Zuber, F

    2012-02-01

    The aim of this study was to determine the inactivation effect of industrial formulations of peracetic acid biocides on bacterial spores adhering to stainless steel surfaces. A standardized protocol was used to validate biocide activity against spores in suspension. To validate sporicidal activity under practical conditions, we developed an additional protocol to simulate industrial sanitization of stainless steel surfaces with a foam sanitizer. Spores of three spore-forming bacteria, Clostridium sporogenes PA3679, Geobacillus stearothermophilus, and Moorella thermoacetica/thermoautotrophica, were sprayed onto stainless steel as bioaerosols. Sporicidal activity was high against the C. sporogenes spore suspension, with more than 5 log CFU ml(-1) destroyed at all liquid biocide contact times. Sporicidal activity also was high against G. stearothermophilus and M. thermoacetica/thermoautotrophica spores after 30 min of contact, but we found no population reduction at the 5-min contact time for the highest sporicide concentration tested. The foam biocide effectively inactivated C. sporogenes spores adhered to stainless steel but had a reduced decontamination effect on other species. For G. stearothermophilus spores, sanitization with the foam sporicide was more efficient on horizontal steel than on vertical steel, but foam sanitization was ineffective against M. thermoacetica/thermoautotrophica whatever the position. These results highlight that decontamination efficiency may differ depending on whether spores are suspended in an aqueous solution or adhered to a stainless steel surface. Biocide efficiency must be validated using relevant protocols and bacteria representative of the microbiological challenges and issues affecting each food industry.

  19. Self-photosensitization of nonphotosynthetic bacteria for solar-to-chemical production.

    PubMed

    Sakimoto, Kelsey K; Wong, Andrew Barnabas; Yang, Peidong

    2016-01-01

    Improving natural photosynthesis can enable the sustainable production of chemicals. However, neither purely artificial nor purely biological approaches seem poised to realize the potential of solar-to-chemical synthesis. We developed a hybrid approach, whereby we combined the highly efficient light harvesting of inorganic semiconductors with the high specificity, low cost, and self-replication and -repair of biocatalysts. We induced the self-photosensitization of a nonphotosynthetic bacterium, Moorella thermoacetica, with cadmium sulfide nanoparticles, enabling the photosynthesis of acetic acid from carbon dioxide. Biologically precipitated cadmium sulfide nanoparticles served as the light harvester to sustain cellular metabolism. This self-augmented biological system selectively produced acetic acid continuously over several days of light-dark cycles at relatively high quantum yields, demonstrating a self-replicating route toward solar-to-chemical carbon dioxide reduction. Copyright © 2016, American Association for the Advancement of Science.

  20. INTRACELLULAR COLONIZATION OF SEAGRASS ROOTS BY ACETOGENIC AND SULFIDOGENIC BACTERIA

    EPA Science Inventory

    The contribution of seagrasses to the stability and fertility of estuarine ecosystems is well established. Loss of seagrasses in recent years to disease and coastal development underscores the importance of understanding the microbial ecology of seagrasses, and the possible roles...

  1. The role of acetogens in microbially influenced corrosion of steel

    PubMed Central

    Mand, Jaspreet; Park, Hyung Soo; Jack, Thomas R.; Voordouw, Gerrit

    2014-01-01

    Microbially influenced corrosion (MIC) of iron (Fe0) by sulfate-reducing bacteria (SRB) has been studied extensively. Through a mechanism, that is still poorly understood, electrons or hydrogen (H2) molecules are removed from the metal surface and used as electron donor for sulfate reduction. The resulting ferrous ions precipitate in part with the sulfide produced, forming characteristic black iron sulfide. Hydrogenotrophic methanogens can also contribute to MIC. Incubation of pipeline water samples, containing bicarbonate and some sulfate, in serum bottles with steel coupons and a headspace of 10% (vol/vol) CO2 and 90% N2, indicated formation of acetate and methane. Incubation of these samples in serum bottles, containing medium with coupons and bicarbonate but no sulfate, also indicated that formation of acetate preceded the formation of methane. Microbial community analyses of these enrichments indicated the presence of Acetobacterium, as well as of hydrogenotrophic and acetotrophic methanogens. The formation of acetate by homoacetogens, such as Acetobacterium woodii from H2 (or Fe0) and CO2, is potentially important, because acetate is a required carbon source for many SRB growing with H2 and sulfate. A consortium of the SRB Desulfovibrio vulgaris Hildenborough and A. woodii was able to grow in defined medium with H2, CO2, and sulfate, because A. woodii provides the acetate, needed by D. vulgaris under these conditions. Likewise, general corrosion rates of metal coupons incubated with D. vulgaris in the presence of acetate or in the presence of A. woodii were higher than in the absence of acetate or A. woodii, respectively. An extended MIC model capturing these results is presented. PMID:24917861

  2. INTRACELLULAR COLONIZATION OF SEAGRASS ROOTS BY ACETOGENIC AND SULFIDOGENIC BACTERIA

    EPA Science Inventory

    The contribution of seagrasses to the stability and fertility of estuarine ecosystems is well established. Loss of seagrasses in recent years to disease and coastal development underscores the importance of understanding the microbial ecology of seagrasses, and the possible roles...

  3. Bacterial Cyanuric Acid Hydrolase for Water Treatment.

    PubMed

    Yeom, Sujin; Mutlu, Baris R; Aksan, Alptekin; Wackett, Lawrence P

    2015-10-01

    Di- and trichloroisocyanuric acids are widely used as water disinfection agents, but cyanuric acid accumulates with repeated additions and must be removed to maintain free hypochlorite for disinfection. This study describes the development of methods for using a cyanuric acid-degrading enzyme contained within nonliving cells that were encapsulated within a porous silica matrix. Initially, three different bacterial cyanuric acid hydrolases were compared: TrzD from Acidovorax citrulli strain 12227, AtzD from Pseudomonas sp. strain ADP, and CAH from Moorella thermoacetica ATCC 39073. Each enzyme was expressed recombinantly in Escherichia coli and tested for cyanuric acid hydrolase activity using freely suspended or encapsulated cell formats. Cyanuric acid hydrolase activities differed by only a 2-fold range when comparing across the different enzymes with a given format. A practical water filtration system is most likely to be used with nonviable cells, and all cells were rendered nonviable by heat treatment at 70°C for 1 h. Only the CAH enzyme from the thermophile M. thermoacetica retained significant activity under those conditions, and so it was tested in a flowthrough system simulating a bioreactive pool filter. Starting with a cyanuric acid concentration of 10,000 μM, more than 70% of the cyanuric acid was degraded in 24 h, it was completely removed in 72 h, and a respike of 10,000 μM cyanuric acid a week later showed identical biodegradation kinetics. An experiment conducted with water obtained from municipal swimming pools showed the efficacy of the process, although cyanuric acid degradation rates decreased by 50% in the presence of 4.5 ppm hypochlorite. In total, these experiments demonstrated significant robustness of cyanuric acid hydrolase and the silica bead materials in remediation. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  4. Evidence that Ferredoxin Interfaces with an Internal Redox Shuttle in Acetyl-CoA Synthase During Reductive Activation and Catalysis †

    PubMed Central

    Bender, Güneş; Ragsdale, Stephen W.

    2011-01-01

    Acetyl-CoA synthase (ACS), a subunit of the bifunctional CO dehydrogenase/acetyl-CoA synthase (CODH/ACS) complex of Moorella thermoacetica requires reductive activation in order to catalyze acetyl-CoA synthesis and related partial reactions, including the CO/[1-14C]-acetyl-CoA exchange reaction. We show that the M. thermoacetica ferredoxin(II) (Fd-II), which harbors two [4Fe-4S] clusters and is an electron acceptor for CODH, serves as a redox activator of ACS. The level of activation depends on the oxidation states of both ACS and Fd-II, which strongly suggests that Fd-II acts as a reducing agent. By the use of controlled potential enzymology, the midpoint reduction potential for the catalytic one-electron redox-active species in the CO/acetyl-CoA exchange reaction is −511 mV, which is similar to the midpoint reduction potential that was earlier measured for other reactions involving ACS. Incubation of ACS with Fd-II and CO leads to the formation of the NiFeC species, which also supports the role of Fd-II as a reductant for ACS. In addition to being a reductant, Fd-II can accept electrons from acetylated ACS, as observed by the increased intensity of the EPR spectrum of reduced Fd-II, indicating that there is a stored electron within an “electron shuttle” in the acetyl-Ni(II) form of ACS. This “shuttle” is proposed to serve as a redox mediator during activation and at different steps of the ACS catalytic cycle. PMID:21141812

  5. Bacterial Cyanuric Acid Hydrolase for Water Treatment

    PubMed Central

    Yeom, Sujin; Mutlu, Baris R.; Aksan, Alptekin

    2015-01-01

    Di- and trichloroisocyanuric acids are widely used as water disinfection agents, but cyanuric acid accumulates with repeated additions and must be removed to maintain free hypochlorite for disinfection. This study describes the development of methods for using a cyanuric acid-degrading enzyme contained within nonliving cells that were encapsulated within a porous silica matrix. Initially, three different bacterial cyanuric acid hydrolases were compared: TrzD from Acidovorax citrulli strain 12227, AtzD from Pseudomonas sp. strain ADP, and CAH from Moorella thermoacetica ATCC 39073. Each enzyme was expressed recombinantly in Escherichia coli and tested for cyanuric acid hydrolase activity using freely suspended or encapsulated cell formats. Cyanuric acid hydrolase activities differed by only a 2-fold range when comparing across the different enzymes with a given format. A practical water filtration system is most likely to be used with nonviable cells, and all cells were rendered nonviable by heat treatment at 70°C for 1 h. Only the CAH enzyme from the thermophile M. thermoacetica retained significant activity under those conditions, and so it was tested in a flowthrough system simulating a bioreactive pool filter. Starting with a cyanuric acid concentration of 10,000 μM, more than 70% of the cyanuric acid was degraded in 24 h, it was completely removed in 72 h, and a respike of 10,000 μM cyanuric acid a week later showed identical biodegradation kinetics. An experiment conducted with water obtained from municipal swimming pools showed the efficacy of the process, although cyanuric acid degradation rates decreased by 50% in the presence of 4.5 ppm hypochlorite. In total, these experiments demonstrated significant robustness of cyanuric acid hydrolase and the silica bead materials in remediation. PMID:26187963

  6. Gelria glutamica gen. nov., sp. nov., a thermophilic, obligately syntrophic, glutamate-degrading anaerobe.

    PubMed

    Plugge, Caroline M; Balk, Melike; Zoetendal, Erwin G; Stams, Alfons J M

    2002-03-01

    A novel anaerobic, gram-positive, thermophilic, spore-forming, obligately syntrophic, glutamate-degrading bacterium, strain TGO(T), was isolated from a propionate-oxidizing methanogenic enrichment culture. The axenic culture was obtained by growing the bacterium on pyruvate. Cells were rod-shaped and non-motile. The optimal temperature for growth was 50-55 degrees C and growth occurred between 37 and 60 degrees C. The pH range for growth was 5.5-8 with optimum growth at pH 7. In pure culture, strain TGO(T) could grow on pyruvate, lactate, glycerol and several sugars. In co-culture with the hydrogenotrophic methanogen Methanobacterium thermautotrophicum strain Z-245, strain TGO(T) could grow on glutamate, proline and Casamino acids. Glutamate was converted to H2, CO2, propionate and traces of succinate. Strain TGO(T) was not able to utilize sulphate, sulphite, thiosulphate, nitrate or fumarate as electron acceptors. The G+C content was 33.8 mol%. Sequence analysis of the 16S rDNA revealed that strain TGO(T) belongs to the thermophilic, endospore-forming anaerobes, though no close relations were found. Its closest relations were Moorella glycerini (92%) and Moorella thermoacetica (90%). Strain TGOT had an unusually long 16S rDNA of more than 1700 bp. The additional base pairs were found as long loops in the V1, V7 and V9 regions of the 16S rDNA. However, the loops were not found in the 16S rRNA. The name Gelria glutamica gen. nov., sp. nov. is proposed for strain TGO(T).

  7. High-Throughput Screening of Coenzyme Preference Change of Thermophilic 6-Phosphogluconate Dehydrogenase from NADP+ to NAD+

    PubMed Central

    Huang, Rui; Chen, Hui; Zhong, Chao; Kim, Jae Eung; Zhang, Yi-Heng Percival

    2016-01-01

    Coenzyme engineering that changes NAD(P) selectivity of redox enzymes is an important tool in metabolic engineering, synthetic biology, and biocatalysis. Here we developed a high throughput screening method to identify mutants of 6-phosphogluconate dehydrogenase (6PGDH) from a thermophilic bacterium Moorella thermoacetica with reversed coenzyme selectivity from NADP+ to NAD+. Colonies of a 6PGDH mutant library growing on the agar plates were treated by heat to minimize the background noise, that is, the deactivation of intracellular dehydrogenases, degradation of inherent NAD(P)H, and disruption of cell membrane. The melted agarose solution containing a redox dye tetranitroblue tetrazolium (TNBT), phenazine methosulfate (PMS), NAD+, and 6-phosphogluconate was carefully poured on colonies, forming a second semi-solid layer. More active 6PGDH mutants were examined via an enzyme-linked TNBT-PMS colorimetric assay. Positive mutants were recovered by direct extraction of plasmid from dead cell colonies followed by plasmid transformation into E. coli TOP10. By utilizing this double-layer screening method, six positive mutants were obtained from two-round saturation mutagenesis. The best mutant 6PGDH A30D/R31I/T32I exhibited a 4,278-fold reversal of coenzyme selectivity from NADP+ to NAD+. This screening method could be widely used to detect numerous redox enzymes, particularly for thermophilic ones, which can generate NAD(P)H reacted with the redox dye TNBT. PMID:27587230

  8. Ancient Evolution and Recent Evolution Converge for the Biodegradation of Cyanuric Acid and Related Triazines

    PubMed Central

    Seffernick, Jennifer L.

    2016-01-01

    Cyanuric acid was likely present on prebiotic Earth, may have been a component of early genetic materials, and is synthesized industrially today on a scale of more than one hundred million pounds per year in the United States. In light of this, it is not surprising that some bacteria and fungi have a metabolic pathway that sequentially hydrolyzes cyanuric acid and its metabolites to release the nitrogen atoms as ammonia to support growth. The initial reaction that opens the s-triazine ring is catalyzed by the unusual enzyme cyanuric acid hydrolase. This enzyme is in a rare protein family that consists of only cyanuric acid hydrolase (CAH) and barbiturase, with barbiturase participating in pyrimidine catabolism by some actinobacterial species. The X-ray structures of two cyanuric acid hydrolase proteins show that this family has a unique protein fold. Phylogenetic, bioinformatic, enzymological, and genetic studies are consistent with the idea that CAH has an ancient protein fold that was rare in microbial populations but is currently becoming more widespread in microbial populations in the wake of anthropogenic synthesis of cyanuric acid and other s-triazine compounds that are metabolized via a cyanuric acid intermediate. The need for the removal of cyanuric acid from swimming pools and spas, where it is used as a disinfectant stabilizer, can potentially be met using an enzyme filtration system. A stable thermophilic cyanuric acid hydrolase from Moorella thermoacetica is being tested for this purpose. PMID:26729715

  9. Five-Gene Cluster in Clostridium thermoaceticum Consisting of Two Divergent Operons Encoding Rubredoxin Oxidoreductase- Rubredoxin and Rubrerythrin–Type A Flavoprotein– High-Molecular-Weight Rubredoxin

    PubMed Central

    Das, Amaresh; Coulter, Eric D.; Kurtz, Donald M.; Ljungdahl, Lars G.

    2001-01-01

    A five-gene cluster encoding four nonheme iron proteins and a flavoprotein from the thermophilic anaerobic bacterium Clostridium thermoaceticum (Moorella thermoacetica) was cloned and sequenced. Based on analysis of deduced amino acid sequences, the genes were identified as rub (rubredoxin), rbo (rubredoxin oxidoreductase), rbr (rubrerythrin), fprA (type A flavoprotein), and a gene referred to as hrb (high-molecular-weight rubredoxin). Northern blot analysis demonstrated that the five-gene cluster is organized as two subclusters, consisting of two divergently transcribed operons, rbr-fprA-hrb and rbo-rub. The rbr, fprA, and rub genes were expressed in Escherichia coli, and their encoded recombinant proteins were purified. The molecular masses, UV-visible absorption spectra, and cofactor contents of the recombinant rubrerythrin, rubredoxin, and type A flavoprotein were similar to those of respective homologs from other microorganisms. Antibodies raised against Desulfovibrio vulgaris Rbr reacted with both native and recombinant Rbr from C. thermoaceticum, indicating that this protein was expressed in the native organism. Since Rbr and Rbo have been recently implicated in oxidative stress protection in several anaerobic bacteria and archaea, we suggest a similar function of these proteins in oxygen tolerance of C. thermoaceticum. PMID:11160086

  10. Haem-based sensors: a still growing old superfamily.

    PubMed

    Germani, Francesca; Moens, Luc; Dewilde, Sylvia

    2013-01-01

    The haem-based sensors are chimeric multi-domain proteins responsible for the cellular adaptive responses to environmental changes. The signal transduction is mediated by the sensing capability of the haem-binding domain, which transmits a usable signal to the cognate transmitter domain, responsible for providing the adequate answer. Four major families of haem-based sensors can be recognized, depending on the nature of the haem-binding domain: (i) the haem-binding PAS domain, (ii) the CO-sensitive carbon monoxide oxidation activator, (iii) the haem NO-binding domain, and (iv) the globin-coupled sensors. The functional classification of the haem-binding sensors is based on the activity of the transmitter domain and, traditionally, comprises: (i) sensors with aerotactic function; (ii) sensors with gene-regulating function; and (iii) sensors with unknown function. We have implemented this classification with newly identified proteins, that is, the Streptomyces avermitilis and Frankia sp. that present a C-terminal-truncated globin fused to an N-terminal cofactor-free monooxygenase, the structural-related class of non-haem globins in Bacillus subtilis, Moorella thermoacetica, and Bacillus anthracis, and a haemerythrin-coupled diguanylate cyclase in Vibrio cholerae. This review summarizes the structures, the functions, and the structure-function relationships known to date on this broad protein family. We also propose unresolved questions and new possible research approaches.

  11. EPR spectroscopic and computational characterization of the hydroxyethylidene-thiamine pyrophosphate radical intermediate of pyruvate:ferredoxin oxidoreductase.

    PubMed

    Mansoorabadi, Steven O; Seravalli, Javier; Furdui, Cristina; Krymov, Vladimir; Gerfen, Gary J; Begley, Tadhg P; Melnick, Jonathan; Ragsdale, Stephen W; Reed, George H

    2006-06-13

    The radical intermediate of pyruvate:ferredoxin oxidoreductase (PFOR) from Moorella thermoacetica was characterized using electron paramagnetic resonance (EPR) spectroscopy at X-band and D-band microwave frequencies. EPR spectra, obtained with various combinations of isotopically labeled substrate (pyruvate) and coenzyme (thiamine pyrophosphate (TPP)), were analyzed by spectral simulations. Parameters obtained from the simulations were compared with those predicted from electronic structure calculations on various radical structures. The g-values and 14N/15N-hyperfine splittings obtained from the spectra are consistent with a planar, hydroxyethylidene-thiamine pyrophosphate (HE-TPP) pi-radical, in which spin is delocalized onto the thiazolium sulfur and nitrogen atoms. The 1H-hyperfine splittings from the methyl group of pyruvate and the 13C-hyperfine splittings from C2 of both pyruvate and TPP are consistent with a model in which the pyruvate-derived oxygen atom of the HE-TPP radical forms a hydrogen bond. The hyperfine splitting constants and g-values are not compatible with those predicted for a nonplanar, sigma/n-type cation radical.

  12. High-throughput screening of coenzyme preference change of thermophilic 6-phosphogluconate dehydrogenase from NADP+ to NAD+

    DOE PAGES

    Huang, Rui; Chen, Hui; Zhong, Chao; ...

    2016-09-02

    Coenzyme engineering that changes NAD(P) selectivity of redox enzymes is an important tool in metabolic engineering, synthetic biology, and biocatalysis. Here we developed a high throughput screening method to identify mutants of 6-phosphogluconate dehydrogenase (6PGDH) from a thermophilic bacterium Moorella thermoacetica with reversed coenzyme selectivity from NADP+ to NAD+. Colonies of a 6PGDH mutant library growing on the agar plates were treated by heat to minimize the background noise, that is, the deactivation of intracellular dehydrogenases, degradation of inherent NAD(P)H, and disruption of cell membrane. The melted agarose solution containing a redox dye tetranitroblue tetrazolium (TNBT), phenazine methosulfate (PMS), NAD+,more » and 6-phosphogluconate was carefully poured on colonies, forming a second semi-solid layer. More active 6PGDH mutants were examined via an enzyme-linked TNBT-PMS colorimetric assay. Positive mutants were recovered by direct extraction of plasmid from dead cell colonies followed by plasmid transformation into E. coli TOP10. By utilizing this double-layer screening method, six positive mutants were obtained from two-round saturation mutagenesis. The best mutant 6PGDH A30D/R31I/T32I exhibited a 4,278-fold reversal of coenzyme selectivity from NADP+ to NAD+. Furthermore, this screening method could be widely used to detect numerous redox enzymes, particularly for thermophilic ones, which can generate NAD(P)H reacted with the redox dye TNBT.« less

  13. PCR detection of thermophilic spore-forming bacteria involved in canned food spoilage.

    PubMed

    Prevost, S; Andre, S; Remize, F

    2010-12-01

    Thermophilic bacteria that form highly heat-resistant spores constitute an important group of spoilage bacteria of low-acid canned food. A PCR assay was developed in order to rapidly trace these bacteria. Three PCR primer pairs were designed from rRNA gene sequences. These primers were evaluated for the specificity and the sensitivity of detection. Two primer pairs allowed detection at the species level of Geobacillus stearothermophilus and Moorella thermoacetica/thermoautrophica. The other pair allowed group-specific detection of anaerobic thermophilic bacteria of the genera Thermoanaerobacterium, Thermoanaerobacter, Caldanerobium and Caldanaerobacter. After a single enrichment step, these PCR assays allowed the detection of 28 thermophiles from 34 cans of spoiled low-acid food. In addition, 13 ingredients were screened for the presence of these bacteria. This PCR assay serves as a detection method for strains able to spoil low-acid canned food treated at 55°C. It will lead to better reactivity in the canning industry. Raw materials and ingredients might be qualified not only for quantitative spore contamination, but also for qualitative contamination by highly heat-resistant spores.

  14. Prevalence of Clostridium botulinum and thermophilic heat-resistant spores in raw carrots and green beans used in French canning industry.

    PubMed

    Sevenier, V; Delannoy, S; André, S; Fach, P; Remize, F

    2012-04-16

    Two categories of vegetables (carrots and green beans) that are widely used in the manufacture of canned food were surveyed for their spore contamination. Samples were recovered from 10 manufactures spread over all producing areas in France. Two samples over 316 raw vegetables collected were found positive for botulinum neurotoxin producing Clostridia spores as tested by PCR-based GeneDisc assay. Both positive samplestested positive for the type B neurotoxin gene (bont/B). In parallel, heat-resistant spores of thermophilic bacteria that are likely to be associated with canned food spoilage after prolonged incubation at 55 °C were surveyed after specific enrichment. Prevalence varied between 1.6% for Moorella thermoacetica/thermoautotrophica in green bean samples and 8.6% for either Geobacillus stearothermophilus or Thermoanaerobacterium spp. in carrot samples. Vegetable preparation, e.g. washing and edge cutting, considerably reduced spore contamination levels. These data constitute the first wide examination of vegetables specifically cultivated for industrialpurposes for their contamination by spores of thermophilic bacterial species.

  15. Reduction and Methyl Transfer Kinetics of the Alpha Subunit from Acetyl-Coenzyme A Synthase

    SciTech Connect

    Xiangshi Tan; Christopher Sewell; Qingwu Yang; Paul A. Lindahl

    2003-01-15

    OAK-B135 Stopped-flow was used to evaluate the methylation and reduction kinetics of the isolated alpha subunit of acetyl-Coenzyme A synthase from Moorella thermoacetica. This catalytically active subunit contains a novel Ni-X-Fe4S4 cluster and a putative unidentified n =2 redox site called D. The D-site must be reduced for a methyl group to transfer from a corrinoid-iron-sulfur protein, a key step in the catalytic synthesis of acetyl-CoA. The Fe4S4 component of this cluster is also redox active, raising the possibility that it is the D-site or a portion thereof. Results presented demonstrate that the D-site reduces far faster than the Fe4S4 component, effectively eliminating this possibility. Rather, this component may alter catalytically important properties of the Ni center. The D-site is reduced through a pathway that probably does not involve the Fe4S4 component of this active-site cluster.

  16. Clostridium ljungdahlii represents a microbial production platform based on syngas

    PubMed Central

    Köpke, Michael; Held, Claudia; Hujer, Sandra; Liesegang, Heiko; Wiezer, Arnim; Wollherr, Antje; Ehrenreich, Armin; Liebl, Wolfgang; Gottschalk, Gerhard; Dürre, Peter

    2010-01-01

    Clostridium ljungdahlii is an anaerobic homoacetogen, able to ferment sugars, other organic compounds, or CO2/H2 and synthesis gas (CO/H2). The latter feature makes it an interesting microbe for the biotech industry, as important bulk chemicals and proteins can be produced at the expense of CO2, thus combining industrial needs with sustained reduction of CO and CO2 in the atmosphere. Sequencing the complete genome of C. ljungdahlii revealed that it comprises 4,630,065 bp and is one of the largest clostridial genomes known to date. Experimental data and in silico comparisons revealed a third mode of anaerobic homoacetogenic metabolism. Unlike other organisms such as Moorella thermoacetica or Acetobacterium woodii, neither cytochromes nor sodium ions are involved in energy generation. Instead, an Rnf system is present, by which proton translocation can be performed. An electroporation procedure has been developed to transform the organism with plasmids bearing heterologous genes for butanol production. Successful expression of these genes could be demonstrated, leading to formation of the biofuel. Thus, C. ljungdahlii can be used as a unique microbial production platform based on synthesis gas and carbon dioxide/hydrogen mixtures. PMID:20616070

  17. Facile Construction of Random Gene Mutagenesis Library for Directed Evolution Without the Use of Restriction Enzyme in Escherichia coli.

    PubMed

    Kim, Jae-Eung; Huang, Rui; Chen, Hui; You, Chun; Zhang, Y-H Percival

    2016-09-01

    A foolproof protocol was developed for the construction of mutant DNA library for directed protein evolution. First, a library of linear mutant gene was generated by error-prone PCR or molecular shuffling, and a linear vector backbone was prepared by high-fidelity PCR. Second, the amplified insert and vector fragments were assembled by overlap-extension PCR with a pair of 5'-phosphorylated primers. Third, full-length linear plasmids with phosphorylated 5'-ends were self-ligated with T4 ligase, yielding circular plasmids encoding mutant variants suitable for high-efficiency transformation. Self-made competent Escherichia coli BL21(DE3) showed a transformation efficiency of 2.4 × 10(5) cfu/µg of the self-ligated circular plasmid. Using this method, three mutants of mCherry fluorescent protein were found to alter their colors and fluorescent intensities under visible and UV lights, respectively. Also, one mutant of 6-phosphorogluconate dehydrogenase from a thermophilic bacterium Moorella thermoacetica was found to show the 3.5-fold improved catalytic efficiency (kcat /Km ) on NAD(+) as compared to the wild-type. This protocol is DNA-sequence independent, and does not require restriction enzymes, special E. coli host, or labor-intensive optimization. In addition, this protocol can be used for subcloning the relatively long DNA sequences into any position of plasmids.

  18. (Per)chlorate reduction by an acetogenic bacterium, Sporomusa sp., isolated from an underground gas storage.

    PubMed

    Balk, Melike; Mehboob, Farrakh; van Gelder, Antonie H; Rijpstra, W Irene C; Damsté, Jaap S Sinninghe; Stams, Alfons J M

    2010-09-01

    A mesophilic bacterium, strain An4, was isolated from an underground gas storage reservoir with methanol as substrate and perchlorate as electron acceptor. Cells were Gram-negative, spore-forming, straight to curved rods, 0.5-0.8 microm in diameter, and 2-8 microm in length, growing as single cells or in pairs. The cells grew optimally at 37 degrees C, and the pH optimum was around 7. Strain An4 converted various alcohols, organic acids, fructose, acetoin, and H(2)/CO(2) to acetate, usually as the only product. Succinate was decarboxylated to propionate. The isolate was able to respire with (per)chlorate, nitrate, and CO(2). The G+C content of the DNA was 42.6 mol%. Based on the 16S rRNA gene sequence analysis, strain An4 was most closely related to Sporomusa ovata (98% similarity). The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell-free extracts.

  19. PHYSIOLOGICAL ECOLOGY OF CLOSTRIDIUM GLYCOLICUM RD-1, AN AEROTOLERANT ACETOGEN ISOLATED FROM SEA GRASS ROOTS

    EPA Science Inventory

    An anaerobic, H2-utilizing bacterium, strain RD-1, was isolated from the highest growth-positive dilution series of a root homogenate prepared from the sea grass Halodule wrightii. Cells of RD-1 were gram-positive, spore-forming, motile rods that were linked by connecting filamen...

  20. Physiological Ecology of Clostridium glycolicum RD-1, an Aerotolerant Acetogen Isolated from Sea Grass Roots

    PubMed Central

    Küsel, Kirsten; Karnholz, Arno; Trinkwalter, Tanja; Devereux, Richard; Acker, Georg; Drake, Harold L.

    2001-01-01

    An anaerobic, H2-utilizing bacterium, strain RD-1, was isolated from the highest growth-positive dilution series of a root homogenate prepared from the sea grass Halodule wrightii. Cells of RD-1 were gram-positive, spore-forming, motile rods that were linked by connecting filaments. Acetate was produced in stoichiometries indicative of an acetyl coenzyme A (acetyl-CoA) pathway-dependent metabolism when RD-1 utilized H2-CO2, formate, lactate, or pyruvate. Growth on sugars or ethylene glycol yielded acetate and ethanol as end products. RD-1 grew at the expense of glucose in the presence of low initial concentrations (up to 6% [vol/vol]) of O2 in the headspace of static, horizontally incubated culture tubes; the concentration of O2 decreased during growth in such cultures. Peroxidase, NADH oxidase, and superoxide dismutase activities were detected in the cytoplasmic fraction of cells grown in the presence of O2. In comparison to cultures incubated under strictly anoxic conditions, acetate production decreased, higher amounts of ethanol were produced, and lactate and H2 became significant end products when RD-1 was grown on glucose in the presence of O2. Similarly, when RD-1 was grown on fructose in the presence of elevated salt concentrations, lower amounts of acetate and higher amounts of ethanol and H2 were produced. When the concentration of O2 in the headspace exceeded 1% (vol/vol), supplemental H2 was not utilized. The 16S rRNA gene of RD-1 had a 99.7% sequence similarity to that of Clostridium glycolicum DSM 1288T, an organism characterized as a fermentative anaerobe. Comparative experiments with C. glycolicum DSM 1288T demonstrated that it had negligible H2- and formate-utilizing capacities. However, carbon monoxide dehydrogenase was detected in both RD-1 and C. glycolicum DSM 1288T. A 91.4% DNA-DNA hybridization between the genomic DNA of RD-1 and that of C. glycolicum DSM 1288T confirmed that RD-1 was a strain of C. glycolicum. These results indicate that (i) RD-1 metabolizes certain substrates via the acetyl-CoA pathway, (ii) RD-1 can tolerate and consume limited amounts of O2, (iii) oxic conditions favor the production of ethanol, lactate, and H2 by RD-1, and (iv) the ability of RD-1 to cope with limited amounts of O2 might contribute to its survival in a habitat subject to daily gradients of photosynthesis-derived O2. PMID:11571179

  1. PHYSIOLOGICAL ECOLOGY OF CLOSTRIDIUM GLYCOLICUM RD-1, AN AEROTOLERANT ACETOGEN ISOLATED FROM SEA GRASS ROOTS

    EPA Science Inventory

    An anaerobic, H2-utilizing bacterium, strain RD-1, was isolated from the highest growth-positive dilution series of a root homogenate prepared from the sea grass Halodule wrightii. Cells of RD-1 were gram-positive, spore-forming, motile rods that were linked by connecting filamen...

  2. (Per)chlorate reduction by an acetogenic bacterium, Sporomusa sp., isolated from an underground gas storage

    PubMed Central

    Mehboob, Farrakh; van Gelder, Antonie H.; Rijpstra, W. Irene C.; Damsté, Jaap S. Sinninghe; Stams, Alfons J. M.

    2010-01-01

    A mesophilic bacterium, strain An4, was isolated from an underground gas storage reservoir with methanol as substrate and perchlorate as electron acceptor. Cells were Gram-negative, spore-forming, straight to curved rods, 0.5–0.8 μm in diameter, and 2–8 μm in length, growing as single cells or in pairs. The cells grew optimally at 37°C, and the pH optimum was around 7. Strain An4 converted various alcohols, organic acids, fructose, acetoin, and H2/CO2 to acetate, usually as the only product. Succinate was decarboxylated to propionate. The isolate was able to respire with (per)chlorate, nitrate, and CO2. The G+C content of the DNA was 42.6 mol%. Based on the 16S rRNA gene sequence analysis, strain An4 was most closely related to Sporomusa ovata (98% similarity). The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell-free extracts. PMID:20680263

  3. Physiology and nutrition of Treponema primitia, an H2/CO2-acetogenic spirochete from termite hindguts.

    PubMed

    Graber, Joseph R; Breznak, John A

    2004-03-01

    Treponema primitia strains ZAS-1 and ZAS-2, the first spirochetes to be isolated from termite hindguts (J. R. Leadbetter, T. M. Schmidt, J. R. Graber, and J. A. Breznak, Science 283:686-689, 1999), were examined for nutritional, physiological, and biochemical properties relevant to growth and survival in their natural habitat. In addition to using H(2) plus CO(2) as substrates, these strains were capable of homoacetogenic growth on mono- and disaccharides and (in the case of ZAS-2) methoxylated benzenoids. Cells were also capable of mixotrophic growth (i.e., simultaneous utilization of H(2) and organic substrates). Cell extracts of T. primitia possessed enzyme activities of the Wood/Ljungdahl (acetyl coenzyme A) pathway of acetogenesis, including tetrahydrofolate-dependent enzymes of the methyl group-forming branch. However, a folate compound was required in the medium for growth. ZAS-1 and ZAS-2 growing on H(2) plus CO(2) displayed H(2) thresholds of 650 and 490 ppmv, respectively. Anoxic cultures of ZAS-1 and ZAS-2 maintained growth after the addition of as much as 0.5% (vol/vol) O(2) to the headspace atmosphere. Cell extracts exhibited NADH and NADPH peroxidase and NADH oxidase activities but neither catalase nor superoxide dismutase activity. Results indicate that (i) T. primitia is able to exploit a variety of substrates derived from the food of its termite hosts and in so doing contributes to termite nutrition via acetogenesis, (ii) in situ growth of T. primitia is likely dependent on secretion of a folate compound(s) by other members of the gut microbiota, and (iii) cells possess enzymatic adaptations to oxidative stress, which is likely to be encountered in peripheral regions of the termite hindgut.

  4. Natronoincola histidinovorans gen. nov., sp. nov., A new alkaliphilic acetogenic anaerobe.

    PubMed

    Zhilina, T N; Detkova, E N; Rainey, F A; Osipov, G A; Lysenko, A M; Kostrikina, N A; Zavarzin, G A

    1998-09-01

    Two strains, asporogenous Z-7940 and sporogenous Z-7939, of a moderately haloalkaliphilic, obligately anaerobic, fermentative bacteria, motile, with Gram-positive cell wall structure, were isolated from soda deposits in Lake Magadi, Kenya. Both strains are mesophilic and utilize only two amino acids, histidine and glutamate, with formation of acetate and ammonium as the main end products. Strain Z-7939 in addition is able to utilize pyruvate. DNA-DNA homology between strains Z-7940 and Z-7939 was 94%, indicating that in spite of phenotypic differences they belong to the same species. They are true alkaliphiles with a pH range for growth of the type strain Z-7940 from pH 8.0 to pH 10.5, optimum at pH 9.4. Both strains obligately depend on sodium and bicarbonate ions. The optimum salt concentration for growth of the type strain is 8-10% wt/vol and the range from 4% to 16%. The G+C content of strain Z-7940 is 31.9 mol% and the strain Z-7939 is 32.3 mol%. Analysis of 16S rDNA sequence of the type strain shows it to belong to cluster XI of the low G+C Gram-positive bacteria. On the basis of its distinct phylogenetic position and physiological properties, we propose a new genus and new species Natronoincola histidinovorans for these strains. The type strain is Z-7940 (=DSM 11416).

  5. Bio-electrochemical synthesis of commodity chemicals by autotrophic acetogens utilizing CO2 for environmental remediation.

    PubMed

    Jabeen, Gugan; Farooq, Robina

    2016-09-01

    Bio-electrochemical synthesis (BES) is a technique in which electro-autotrophic bacteria such as Clostridium ljungdahlii utilize electric currents as an electron source from the cathode to reduce CO2 to extracellular, multicarbon, exquisite products through autotrophic conversion. The BES of volatile fatty acids and alcohols directly from CO2 is a sustainable alternative for non-renewable, petroleum-based polymer production. This conversion of CO2 implies reduction of greenhouse gas emissions. The synthesis of heptanoic acid, heptanol, hexanoic acid and hexanol, for the first time, by Clostridium ljungdahlii was a remarkable achievement of BES. In our study, these microorganisms were cultivated on the cathode of a bio-electrochemical cell at -400 mV by a DC power supply at 37 degree Centrigrade, pH 6.8, and was studied for both batch and continuous systems. Pre-enrichment of bio-cathode enhanced the electroactivity of cells and resulted in maximizing extracellular products in less time. The main aim of the research was to investigate the impact of low-cost substrate CO2, and the longer cathode recovery range was due to bacterial reduction of CO2 to multicarbon chemical commodities with electrons driven from the cathode. Reactor design was simplified for cost-effectiveness and to enhance energy efficiencies. The Columbic recovery of ethanoic acid, ethanol, ethyl butyrate, hexanoic acid, heptanoic acid and hexanol being in excess of 80 percent proved that BES was a remarkable technology.

  6. Competition between Methanogens and Acetogens in Biocathodes: A Comparison between Potentiostatic and Galvanostatic Control

    PubMed Central

    Molenaar, Sam D.; Saha, Pradip; Mol, Annemerel R.; Sleutels, Tom H. J. A.; ter Heijne, Annemiek; Buisman, Cees J. N.

    2017-01-01

    Microbial electrosynthesis is a useful form of technology for the renewable production of organic commodities from biologically catalyzed reduction of CO2. However, for the technology to become applicable, process selectivity, stability and efficiency need strong improvement. Here we report on the effect of different electrochemical control modes (potentiostatic/galvanostatic) on both the start-up characteristics and steady-state performance of biocathodes using a non-enriched mixed-culture inoculum. Based on our results, it seems that kinetic differences exist between the two dominant functional microbial groups (i.e., homoacetogens and methanogens) and that by applying different current densities, these differences may be exploited to steer product selectivity and reactor performance. PMID:28106846

  7. Crystallographic Snapshots of Cyanide- and Water-Bound C-Clusters from Bifunctional Carbon Monoxide Dehydrogenase/Acetyl-CoA Synthase†,‡

    PubMed Central

    2009-01-01

    Nickel-containing carbon monoxide dehydrogenases (CODHs) reversibly catalyze the oxidation of carbon monoxide to carbon dioxide and are of vital importance in the global carbon cycle. The unusual catalytic CODH C-cluster has been crystallographically characterized as either a NiFe4S4 or a NiFe4S5 metal center, the latter containing a fifth, additional sulfide that bridges Ni and a unique Fe site. To determine whether this bridging sulfide is catalytically relevant and to further explore the mechanism of the C-cluster, we obtained crystal structures of the 310 kDa bifunctional CODH/acetyl-CoA synthase complex from Moorella thermoacetica bound both with a substrate H2O/OH− molecule and with a cyanide inhibitor. X-ray diffraction data were collected from native crystals and from identical crystals soaked in a solution containing potassium cyanide. In both structures, the substrate H2O/OH− molecule exhibits binding to the unique Fe site of the C-cluster. We also observe cyanide binding in a bent conformation to Ni of the C-cluster, adjacent the substrate H2O/OH− molecule. Importantly, the bridging sulfide is not present in either structure. As these forms of the C-cluster represent the coordination environment immediately before the reaction takes place, our findings do not support a fifth, bridging sulfide playing a catalytic role in the enzyme mechanism. The crystal structures presented here, along with recent structures of CODHs from other organisms, have led us toward a unified mechanism for CO oxidation by the C-cluster, the catalytic center of an environmentally important enzyme. PMID:19583207

  8. Xenon in And at the End of the Tunnel of Bifunctional Carbon Monoxide Dehydrogenase/Acetyl-CoA Synthase

    SciTech Connect

    Doukov, T.I.; Blasiak, L.C.; Seravalli, J.; Ragsdale, S.W.; Drennan, C.L.; /MIT /SLAC, SSRL /Nebraska U.

    2009-05-11

    A fascinating feature of some bifunctional enzymes is the presence of an internal channel or tunnel to connect the multiple active sites. A channel can allow for a reaction intermediate generated at one active site to be used as a substrate at a second active site, without the need for the intermediate to leave the safety of the protein matrix. One such bifunctional enzyme is carbon monoxide dehydrogenase/acetyl-CoA synthase from Moorella thermoacetica (mtCODH/ACS). A key player in the global carbon cycle, CODH/ACS uses a Ni-Fe-S center called the C-cluster to reduce carbon dioxide to carbon monoxide and uses a second Ni-Fe-S center, called the A-cluster, to assemble acetyl-CoA from a methyl group, coenzyme A, and C-cluster-generated CO. mtCODH/ACS has been proposed to contain one of the longest enzyme channels (138 A long) to allow for intermolecular CO transport. Here, we report a 2.5 A resolution structure of xenon-pressurized mtCODH/ACS and examine the nature of gaseous cavities within this enzyme. We find that the cavity calculation program CAVENV accurately predicts the channels connecting the C- and A-clusters, with 17 of 19 xenon binding sites within the predicted regions. Using this X-ray data, we analyze the amino acid composition surrounding the 19 Xe sites and consider how the protein fold is utilized to carve out such an impressive interior passageway. Finally, structural comparisons of Xe-pressurized mtCODH/ACS with related enzyme structures allow us to study channel design principles, as well as consider the conformational flexibility of an enzyme that contains a cavity through its center.

  9. Genome-guided analysis of physiological and morphological traits of the fermentative acetate oxidizer Thermacetogenium phaeum

    PubMed Central

    2012-01-01

    Background Thermacetogenium phaeum is a thermophilic strictly anaerobic bacterium oxidizing acetate to CO2 in syntrophic association with a methanogenic partner. It can also grow in pure culture, e.g., by fermentation of methanol to acetate. The key enzymes of homoacetate fermentation (Wood-Ljungdahl pathway) are used both in acetate oxidation and acetate formation. The obvious reversibility of this pathway in this organism is of specific interest since syntrophic acetate oxidation operates close to the energetic limitations of microbial life. Results The genome of Th. phaeum is organized on a single circular chromosome and has a total size of 2,939,057 bp. It comprises 3.215 open reading frames of which 75% could be assigned to a gene function. The G+C content is 53.88 mol%. Many CRISPR sequences were found, indicating heavy phage attack in the past. A complete gene set for a phage was found in the genome, and indications of phage action could also be observed in culture. The genome contained all genes required for CO2 reduction through the Wood-Ljungdahl pathway, including two formyl tetrahydrofolate ligases, three carbon monoxide dehydrogenases, one formate hydrogenlyase complex, three further formate dehydrogenases, and three further hydrogenases. The bacterium contains a menaquinone MQ-7. No indications of cytochromes or Rnf complexes could be found in the genome. Conclusions The information obtained from the genome sequence indicates that Th. phaeum differs basically from the three homoacetogenic bacteria sequenced so far, i.e., the sodium ion-dependent Acetobacterium woodii, the ethanol-producing Clostridium ljungdahlii, and the cytochrome-containing Moorella thermoacetica. The specific enzyme outfit of Th. phaeum obviously allows ATP formation both in acetate formation and acetate oxidation. PMID:23259483

  10. High-throughput screening of coenzyme preference change of thermophilic 6-phosphogluconate dehydrogenase from NADP+ to NAD+

    SciTech Connect

    Huang, Rui; Chen, Hui; Zhong, Chao; Kim, Jae Eung; Zhang, Yi-Heng Percival

    2016-09-02

    Coenzyme engineering that changes NAD(P) selectivity of redox enzymes is an important tool in metabolic engineering, synthetic biology, and biocatalysis. Here we developed a high throughput screening method to identify mutants of 6-phosphogluconate dehydrogenase (6PGDH) from a thermophilic bacterium Moorella thermoacetica with reversed coenzyme selectivity from NADP+ to NAD+. Colonies of a 6PGDH mutant library growing on the agar plates were treated by heat to minimize the background noise, that is, the deactivation of intracellular dehydrogenases, degradation of inherent NAD(P)H, and disruption of cell membrane. The melted agarose solution containing a redox dye tetranitroblue tetrazolium (TNBT), phenazine methosulfate (PMS), NAD+, and 6-phosphogluconate was carefully poured on colonies, forming a second semi-solid layer. More active 6PGDH mutants were examined via an enzyme-linked TNBT-PMS colorimetric assay. Positive mutants were recovered by direct extraction of plasmid from dead cell colonies followed by plasmid transformation into E. coli TOP10. By utilizing this double-layer screening method, six positive mutants were obtained from two-round saturation mutagenesis. The best mutant 6PGDH A30D/R31I/T32I exhibited a 4,278-fold reversal of coenzyme selectivity from NADP+ to NAD+. Furthermore, this screening method could be widely used to detect numerous redox enzymes, particularly for thermophilic ones, which can generate NAD(P)H reacted with the redox dye TNBT.

  11. Xenon in and at the end of the tunnel of bifunctional carbon monoxide dehydrogenase/acetyl-CoA synthase.

    PubMed

    Doukov, Tzanko I; Blasiak, Leah C; Seravalli, Javier; Ragsdale, Stephen W; Drennan, Catherine L

    2008-03-18

    A fascinating feature of some bifunctional enzymes is the presence of an internal channel or tunnel to connect the multiple active sites. A channel can allow for a reaction intermediate generated at one active site to be used as a substrate at a second active site, without the need for the intermediate to leave the safety of the protein matrix. One such bifunctional enzyme is carbon monoxide dehydrogenase/acetyl-CoA synthase from Moorella thermoacetica (mtCODH/ACS). A key player in the global carbon cycle, CODH/ACS uses a Ni-Fe-S center called the C-cluster to reduce carbon dioxide to carbon monoxide and uses a second Ni-Fe-S center, called the A-cluster, to assemble acetyl-CoA from a methyl group, coenzyme A, and C-cluster-generated CO. mtCODH/ACS has been proposed to contain one of the longest enzyme channels (138 A long) to allow for intermolecular CO transport. Here, we report a 2.5 A resolution structure of xenon-pressurized mtCODH/ACS and examine the nature of gaseous cavities within this enzyme. We find that the cavity calculation program CAVENV accurately predicts the channels connecting the C- and A-clusters, with 17 of 19 xenon binding sites within the predicted regions. Using this X-ray data, we analyze the amino acid composition surrounding the 19 Xe sites and consider how the protein fold is utilized to carve out such an impressive interior passageway. Finally, structural comparisons of Xe-pressurized mtCODH/ACS with related enzyme structures allow us to study channel design principles, as well as consider the conformational flexibility of an enzyme that contains a cavity through its center.

  12. Crystallographic snapshots of cyanide- and water-bound C-clusters from bifunctional carbon monoxide dehydrogenase/acetyl-CoA synthase.

    PubMed

    Kung, Yan; Doukov, Tzanko I; Seravalli, Javier; Ragsdale, Stephen W; Drennan, Catherine L

    2009-08-11

    Nickel-containing carbon monoxide dehydrogenases (CODHs) reversibly catalyze the oxidation of carbon monoxide to carbon dioxide and are of vital importance in the global carbon cycle. The unusual catalytic CODH C-cluster has been crystallographically characterized as either a NiFe(4)S(4) or a NiFe(4)S(5) metal center, the latter containing a fifth, additional sulfide that bridges Ni and a unique Fe site. To determine whether this bridging sulfide is catalytically relevant and to further explore the mechanism of the C-cluster, we obtained crystal structures of the 310 kDa bifunctional CODH/acetyl-CoA synthase complex from Moorella thermoacetica bound both with a substrate H(2)O/OH(-) molecule and with a cyanide inhibitor. X-ray diffraction data were collected from native crystals and from identical crystals soaked in a solution containing potassium cyanide. In both structures, the substrate H(2)O/OH(-) molecule exhibits binding to the unique Fe site of the C-cluster. We also observe cyanide binding in a bent conformation to Ni of the C-cluster, adjacent the substrate H(2)O/OH(-) molecule. Importantly, the bridging sulfide is not present in either structure. As these forms of the C-cluster represent the coordination environment immediately before the reaction takes place, our findings do not support a fifth, bridging sulfide playing a catalytic role in the enzyme mechanism. The crystal structures presented here, along with recent structures of CODHs from other organisms, have led us toward a unified mechanism for CO oxidation by the C-cluster, the catalytic center of an environmentally important enzyme.

  13. Silica Gel for Enhanced Activity and Hypochlorite Protection of Cyanuric Acid Hydrolase in Recombinant Escherichia coli

    PubMed Central

    Radian, Adi; Aukema, Kelly G.; Aksan, Alptekin

    2015-01-01

    ABSTRACT Chlorinated isocyanuric acids are widely used water disinfectants that generate hypochlorite, but with repeated application, they build up cyanuric acid (CYA) that must be removed to maintain disinfection. 3-Aminopropyltriethoxysilane (APTES)-treated Escherichia coli cells expressing cyanuric acid hydrolase (CAH) from Moorella thermoacetica exhibited significantly high CYA degradation rates and provided protection against enzyme inactivation by hypochlorite (chlorine). APTES coating or encapsulation of cells had two benefits: (i) overcoming diffusion limitations imposed by the cell wall and (ii) protecting against hypochlorite inactivation of CAH activity. Cells encapsulated in APTES gels degraded CYA three times faster than nonfunctionalized tetraethoxysilane (TEOS) gels, and cells coated with APTES degraded CYA at a rate of 29 µmol/min per mg of CAH protein, similar to the rate with purified enzyme. UV spectroscopy, fluorescence spectroscopy, and scanning electron microscopy showed that the higher rates were due to APTES increasing membrane permeability and enhancing cyanuric acid diffusion into the cytoplasm to reach the CAH enzyme. Purified CAH enzyme was shown to be rapidly inactivated by hypochlorite. APTES aggregates surrounding cells protected via the amine groups reacting with hypochlorite as shown by pH changes, zeta potential measurements, and infrared spectroscopy. APTES-encapsulated E. coli cells expressing CAH degraded cyanuric acid at high rates in the presence of 1 to 10 ppm hypochlorite, showing effectiveness under swimming pool conditions. In contrast, CAH activity in TEOS gels or free cells was completely inactivated by hypochlorite. These studies show that commercially available silica materials can selectively enhance, protect, and immobilize whole-cell biocatalysts for specialized applications. PMID:26530383

  14. Evidence for a Proton Transfer Network and a Required Persulfide-Bond-Forming Cysteine Residue in Ni-Containing Carbon Monoxide Dehydrogenases

    SciTech Connect

    Eun Jin Kim; Jian Feng; Matthew R. Bramlett; Paul A. Lindahl

    2004-05-18

    OAK-B135 Carbon monoxide dehydrogenase from Moorella thermoacetica catalyzes the reversible oxidation of CO to CO2 at a nickel-iron-sulfur active-site called the C-cluster. Mutants of a proposed proton transfer pathway and of a cysteine residue recently found to form a persulfide bond with the C-cluster were characterized. Four semi-conserved histidine residues were individually mutated to alanine. His116 and His122 were essential to catalysis, while His113 and His119 attenuated catalysis but were not essential. Significant activity was ''rescued'' by a double mutant where His116 was replaced by Ala and His was also introduced at position 115. Activity was also rescued in double mutants where His122 was replaced by Ala and His was simultaneously introduced at either position 121 or 123. Activity was also ''rescued'' by replacing His with Cys at position 116. Mutation of conserved Lys587 near the C-cluster attenuated activity but did not eliminate it. Activity was virtually abolished in a double mutant where Lys587 and His113 were both changed to Ala. Mutations of conserved Asn284 also attenuated activity. These effects suggest the presence of a network of amino acid residues responsible for proton transfer rather than a single linear pathway. The Ser mutant of the persulfide-forming Cys316 was essentially inactive and displayed no EPR signals originating from the C-cluster. Electronic absorption and metal analysis suggests that the C-cluster is absent in this mutant. The persulfide bond appears to be essential for either the assembly or stability of the C-cluster, and/or for eliciting the redox chemistry of the C-cluster required for catalytic activity.

  15. Effect of Sodium Sulfide on Ni-Containing Carbon Monoxide Dehydrogenases

    SciTech Connect

    Jian Feng; Paul A. Lindahl

    2004-07-28

    OAK-B135 The structure of the active-site C-cluster in CO dehydrogenase from Carboxythermus hydrogenoformans includes a {mu}{sup 2}-sulfide ion bridged to the Ni and unique Fe, while the same cluster in enzymes from Rhodospirillum rubrum (CODH{sub Rr}) and Moorella thermoacetica (CODH{sub Mt}) lack this ion. This difference was investigated by exploring the effects of sodium sulfide on activity and spectral properties. Sulfide partially inhibited the CO oxidation activity of CODH{sub Rr} and generated a lag prior to steady-state. CODH{sub Mt} was inhibited similarly but without a lag. Adding sulfide to CODH{sub Mt} in the C{sub red1} state caused the g{sub av} = 1.82 EPR signal to decline and new features to appear, including one with g = 1.95, 1.85 and (1.70 or 1.62). Removing sulfide caused the g{sub av} = 1.82 signal to reappear and activity to recover. Sulfide did not affect the g{sub av} = 1.86 signal from the C{sub red2} state. A model was developed in which sulfide binds reversibly to C{sub red1}, inhibiting catalysis. Reducing this adduct causes sulfide to dissociate, C{sub red2} to develop, and activity to recover. Using this model, apparent K{sub I} values are 40 {+-} 10 nM for CODH{sub Rr} and 60 {+-} 30 {micro}M for CODH{sub Mt}. Effects of sulfide are analogous to those of other anions, including the substrate hydroxyl group, suggesting that these ions also bridge the Ni and unique Fe. This proposed arrangement raises the possibility that CO binding labilizes the bridging hydroxyl and increases its nucleophilic tendency towards attacking Ni-bound carbonyl.

  16. Novel domain arrangement in the crystal structure of a truncated acetyl-CoA synthase fromMoorella thermoacetica†‡

    PubMed Central

    Volbeda, Anne; Darnault, Claudine; Tan, Xiangshi; Lindahl, Paul A.; Fontecilla-Camps, Juan C.

    2009-01-01

    Ni-dependent Acetyl-CoA synthase (ACS) and CO dehydrogenase (CODH) constitute the central enzyme complex of the Wood-Ljungdahl pathway of acetyl-CoA formation. The crystal structure of a recombinant bacterial ACS lacking the N-terminal domain that interacts with CODH shows a large reorganization of the remaining two globular domains, producing a narrow cleft of suitable size, shape and nature to bind CoA. Sequence comparisons with homologous archaeal enzymes that naturally lack the N-terminal domain show that many amino acids lining this cleft are conserved. Besides the typical [4Fe-4S] center, the A-cluster contains only one proximal metal ion that, according to anomalous scattering data, is most likely Cu or Zn. Incorporation of a functional Ni2Fe4S4 A-cluster would require only minor structural rearrangements. Using available structures, a plausible model of the interaction between CODH and the smaller ACS in archaeal multi-enzyme complexes is presented, along with a discussion of evolutionary relationships of the archaeal and bacterial enzymes. PMID:19650626

  17. Bacterial Community Profiling of H2/CO2 or Formate-Utilizing Acetogens Enriched from Diverse Ecosystems

    NASA Astrophysics Data System (ADS)

    Han, R.; Zhang, L.; Fu, B.; Liu, H.

    2014-12-01

    Synthetic gases are usually generated from either cellulosic agricultural waste combustion or industrial release and could be subsequently transformed into acetate, ethanol, and/or butyrate by homoacetogenic bacteria, which commonly possess reductive acetyl-CoA synthesis pathway. Homoacetogen-based syngas fermentation technology provides an alternative solution to link greenhouse gas emission control and cellulosic solid waste treatment with biofuels production. The objective of our current project is to hunt for homoacetogens with capabilities of highly efficiently converting syngases to chemical solvents. In this study, we evaluated homoacetogens population dynamics during enrichments and pinpointed dominant homoacetogens representing diverse ecosystems enriched by different substrates. We enriched homoacetogens from four different samples including waste activate sludge, freshwater sediment, anaerobic methanogenic sludge, and cow manure using H2/CO2 (4:1) or formate as substrate for homoacetogen enrichment. Along with the formyltetrahydrofolate synthetase (FTHFS) gene (fhs gene)-specific real time qPCR assay and Terminal Restriction Fragment Length Polymorphism (T-RFLP) analysis, 16S rRNA based 454 high-throughput pyrosequencing was applied to reveal the population dynamic and community structure during enrichment from different origins. Enrichment of homoacetogenic populations coincided with accumulations of short chain fatty acids such as acetate and butyrate. 454 high-throughput pyrosequencing revealed Firmicutes and Spirochaetes populations became dominant while the overall microbial diversity decreased after enrichment. The most abundant sequences among the four origins belonged to the following phyla: Firmicutes, Spirochaetes, Proteobacteria, and Bacteroidetes, accounting for 62.1%-99.1% of the total reads. The major putative homoacetogenic species enriched on H2/CO2 or formate belonged to Clostridium spp., Acetobacterium spp., Acetoanaerobium spp., Eubacterium spp., Sporomusa spp. This comprehensive molecular ecology study on homoacetogen enrichments provides molecular evidences for shaping homoacetogenic populations and targeting novel homoacetogenic species enriched from diverse ecosystems.

  18. ACETOGENIC AND SULPHATE-REDUCING BACTERIA INHABITING THE RHIZOPLANE AND DEEP CORTEX CELLS OF THE SEAGRASS HALODULE WRIGHTII

    EPA Science Inventory

    Recent declines in sea grass distribution underscore the importance of understanding microbial community structure-function relationships in sea grass rhizosphere that might affect the viability of these plants. Phospholipid fatty acid analyses showed that sulfate-reducing bacter...

  19. Bioconversion of Cellulose to Acetate with Pure Cultures of Ruminococcus albus and a Hydrogen-Using Acetogen.

    PubMed

    Miller, T L; Wolin, M J

    1995-11-01

    Bioconversion of cellulose to acetate was accomplished with cocultures of two organisms. One was the cellulolytic species Ruminococcus albus. It ferments crystalline cellulose (Avicel) to acetate, ethanol, CO(inf2), and H(inf2). The other organism (HA) obtains energy for growth by using H(inf2) to reduce CO(inf2) to acetate. HA is a gram-negative coccobacillus that was isolated from horse feces. Coculture of R. albus with HA in batch or continuous culture alters the fermentation products formed from crystalline cellulose by the ruminococcus via interspecies H(inf2) transfer. The major product of the fermentation by R. albus and HA coculture is acetate. High concentrations of acetate (333 mM) were obtained when batch cocultures grown on 5% cellulose were neutralized with Ca(OH)(inf2). Continuous cocultures grown at retention times of 2 and 3.1 days produced 109 and 102 mM acetate, respectively, when fed 1% cellulose with utilization of 84% of the substrate.

  20. Bioconversion of Cellulose to Acetate with Pure Cultures of Ruminococcus albus and a Hydrogen-Using Acetogen

    PubMed Central

    Miller, T. L.; Wolin, M. J.

    1995-01-01

    Bioconversion of cellulose to acetate was accomplished with cocultures of two organisms. One was the cellulolytic species Ruminococcus albus. It ferments crystalline cellulose (Avicel) to acetate, ethanol, CO(inf2), and H(inf2). The other organism (HA) obtains energy for growth by using H(inf2) to reduce CO(inf2) to acetate. HA is a gram-negative coccobacillus that was isolated from horse feces. Coculture of R. albus with HA in batch or continuous culture alters the fermentation products formed from crystalline cellulose by the ruminococcus via interspecies H(inf2) transfer. The major product of the fermentation by R. albus and HA coculture is acetate. High concentrations of acetate (333 mM) were obtained when batch cocultures grown on 5% cellulose were neutralized with Ca(OH)(inf2). Continuous cocultures grown at retention times of 2 and 3.1 days produced 109 and 102 mM acetate, respectively, when fed 1% cellulose with utilization of 84% of the substrate. PMID:16535158

  1. ACETOGENIC AND SULPHATE-REDUCING BACTERIA INHABITING THE RHIZOPLANE AND DEEP CORTEX CELLS OF THE SEAGRASS HALODULE WRIGHTII

    EPA Science Inventory

    Recent declines in sea grass distribution underscore the importance of understanding microbial community structure-function relationships in sea grass rhizosphere that might affect the viability of these plants. Phospholipid fatty acid analyses showed that sulfate-reducing bacter...

  2. Evidence for a super-reduced cobamide as the major corrinoid fraction in vivo and a histidine residue as a cobalt ligand of the p-cresolyl cobamide in the acetogenic bacterium Sporomusa ovata.

    PubMed

    Stupperich, E; Eisinger, H J; Albracht, S P

    1990-10-05

    The redox state of cobalt in p-cresolyl cobamide and one of its axial ligands were determined by EPR spectroscopy of Sporomusa ovata as harvested. The analyses revealed that less than 2% (less than 30 nmol/g dry cells) of the total corrinoids (greater than 2400 nmol/g dry cells) were in a low-spin Co(II) complex. The amount increased to about 15% (190-450 nmol/g dry cells) upon partial oxidation by air, indicating that the original valence state of cobalt was a Co(I) prior to this treatment. The cob(I)amide was quantified as Co(III)-CH3 after methylation by iodomethane. More than 45% (1100 nmol/g dry cells) of the extractable corrinoids were in the methylated form, whereas non-treated cells revealed less than 1% (less than 15 nmol g dry cells) of light-sensitive corrinoids. EPR spectra of the Co(II) complex exhibited a threefold N-hyperfine splitting in the gz region, which was similar to vitamin B12. Cells grown with [1.3-15N2]histidine showed a twofold N-hyperfine splitting, demonstrating that the axial N ligand of the corrinoid was derived from the imidazole group of histidine. It is concluded that the super-nucleophilic p-cresolyl cob(I)amide is the major corrinoid complex in vivo and that it is stabilized by its protein(s). The Co(II) ion of the prosthetic group was coordinated by one histidine residue of the apoprotein(s).

  3. A mathematical study of a syntrophic relationship of a model of anaerobic digestion process.

    PubMed

    El Hajji, Miled; Mazenc, Frédéric; Harmand, Jérôme

    2010-07-01

    A mathematical model involving the syntrophic relationship of two major populations of bacteria (acetogens and methanogens), each responsible for a stage of the methane fermentation process is proposed. A detailed qualitative analysis is carried out. The local and global stability analyses of the equilibria are performed. We demonstrate, under general assumptions of monotonicity, relevant from an applied point of view, the global asymptotic stability of a positive equilibrium point which corresponds to the coexistence of acetogenic and methanogenic bacteria.

  4. Importance of cobalt for individual trophic groups in an anaerobic methanol-degrading consortium

    SciTech Connect

    Florencio, L.; FIeld, J.A.; Lettinga, G. )

    1994-01-01

    Methanol is an important anaerobic substrate in industrial wastewater treatment and the natural environment. Previous studies indicate that cobalt greatly stimulates methane formation during anaerobic treatment of methanolic wastewaters. To evaluate the effect of cobalt in a mixed culture, a sludge with low background levels of cobalt was cultivated in an upflow anaerobic sludge blanket reactor. Specific inhibitors in batch assays were then utilized to study the effect of cobalt on the growth rate and activity of different microorganisms involved in the anaerboic degradation of methanol. Only methylotropic methanogens and acetogens were stimulated by cobalt additions, while the other trophic concentration of cobalt for the growth and activity of methanol-utilizing methanogens and acetogens were stimulated by cobalt additions, while the other trophic groups utilizing downstream intermediates, H[sub 2]-CO[sub 2] or acetate, were largely unaffected. The optimal concentration of cobalt for the growth and activity of methanol-utilizing methanogens and acetogens was 0.05 mg liter[sup [minus]1]. The higher requirement of cobalt is presumably due to the previously reported production of unique corrinoid-containing enzymes (or coenzymes) by direct utilizers of methanol. This distinctly high requirement of cobalt by methylotrophs should be considered during methanolic wastewater treatment. Methylotroph methanogens presented a 60-fold-higher affinity for methanol than acetogens. This result in combination with the fact that acetogens grow slightly faster than methanogens under optimal cobalt conditions indicates that acetogens can outcompete methanogens only when reactor methanol and cobalt concentrations are high, provided enough inorganic carbon is available.

  5. Genome Sequence of the Homoacetogenic, Gram-Negative, Endospore-Forming Bacterium Sporomusa acidovorans DSM 3132

    PubMed Central

    Humphreys, Jonathan R.

    2017-01-01

    ABSTRACT Sporomusa acidovorans DSM 3132 is a strictly anaerobic, spore-forming and acetogenic bacterium, which was isolated from effluent of an alcohol distillation fermenter. The genome harbors genes involved in the Wood-Ljungdahl pathway for carbon fixation and several genes for glycerol metabolism. The genome (6.06 Mb) contains 4,506 predicted protein-encoding genes. PMID:28935740

  6. Genome Sequence of the Solvent-Producing Bacterium Clostridium carboxidivorans Strain P7T▿

    PubMed Central

    Paul, Debarati; Austin, Frank W.; Arick, Tony; Bridges, Susan M.; Burgess, Shane C.; Dandass, Yoginder S.; Lawrence, Mark L.

    2010-01-01

    Clostridium carboxidivorans strain P7T is a strictly anaerobic acetogenic bacterium that produces acetate, ethanol, butanol, and butyrate. The C. carboxidivorans genome contains all the genes for the carbonyl branch of the Wood-Ljungdahl pathway for CO2 fixation, and it encodes enzymes for conversion of acetyl coenzyme A into butanol and butyrate. PMID:20729368

  7. Bacterial population development and chemical characteristics of refuse decomposition in a simulated sanitary landfill

    SciTech Connect

    Barlaz, M.A.; Schaefer, D.M.; Ham, R.K. )

    1989-01-01

    Population development of key groups of bacteria involved in municipal refuse conversion to methane was measured from the time of initial incubation through the onset of methane production. Hemicellulolytic bacteria, cellulolytic bacteria, hydrogen-producing acetogens, and acetate-and H{sub 2}-plus-CO{sub 2}-utilizing methanogens were enumerated by the most-probable-number technique with media containing oat spelt xylan, ball-milled cellulose, butyrate, acetate, and H{sub 2} plus CO{sub 2}, respectively. The methane concentration of the sampled containers increased to 64% by day 69, at which time the maximum methane production rate, 929 liters of CH{sub 4} per day kg-year, was measured. Population increases of 2, 4, 5, 5, and 6 orders of magnitude were measured between fresh refuse and the methane production phase for the hemicellulolytic bacteria, cellulolytic bacteria, butyrate-catabolizing acetogens, and acetate- and H{sub 2}-CO{sub 2}-utilizing methanogens, respectively. The cellulolytic bacteria and acetogens increased more slowly than the methanogens and only after the onset of methane production. The initial decrease in the pH of the refuse ecosystem from 7.5 to 5.7 was attributed to the accumulation of acidic end products of sugar fermentation, to the low acid-consuming activity of the acetogenic and methanogenic bacteria, and to levels of oxygen and nitrate in the fresh refuse sufficient for oxidation of only 8% of the sugars to carbon dioxide and water. Cellulose and hemicellulose decomposition was most rapid after establishment of the methanogenic and acetogenic populations and a reduction in the initial accumulation of carboxylic acids. Initially acetate utilization, but ultimately polymer hydrolysis, limited the rate of refuse conversion to methane.

  8. Tracking spore-forming bacteria in food: from natural biodiversity to selection by processes.

    PubMed

    Postollec, Florence; Mathot, Anne-Gabrielle; Bernard, Muriel; Divanac'h, Marie-Laure; Pavan, Sonia; Sohier, Danièle

    2012-08-01

    Sporeforming bacteria are ubiquitous in the environment and exhibit a wide range of diversity leading to their natural prevalence in foodstuff. The state of the art of sporeformer prevalence in ingredients and food was investigated using a multiparametric PCR-based tool that enables simultaneous detection and identification of various genera and species mostly encountered in food, i.e., Alicyclobacillus, Anoxybacillus flavithermus, Bacillus, B. cereus group, B. licheniformis, B. pumilus, B. sporothermodurans, B. subtilis, Brevibacillus laterosporus, Clostridium, Geobacillus stearothermophilus, Moorella and Paenibacillus species. In addition, 16S rDNA sequencing was used to extend identification to other possibly present contaminants. A total of 90 food products, with or without visible trace of spoilage were analysed, i.e., 30 egg-based products, 30 milk and dairy products and 30 canned food and ingredients. Results indicated that most samples contained one or several of the targeted genera and species. For all three tested food categories, 30 to 40% of products were contaminated with both Bacillus and Clostridium. The percentage of contaminations associated with Clostridium or Bacillus represented 100% in raw materials, 72% in dehydrated ingredients and 80% in processed foods. In the last two product types, additional thermophilic contaminants were identified (A. flavithermus, Geobacillus spp., Thermoanaerobacterium spp. and Moorella spp.). These results suggest that selection, and therefore the observed (re)-emergence of unexpected sporeforming contaminants in food might be favoured by the use of given food ingredients and food processing technologies.

  9. Syntrophic growth on formate: a new microbial niche in anoxic environments.

    PubMed

    Dolfing, Jan; Jiang, Bo; Henstra, Anne M; Stams, Alfons J M; Plugge, Caroline M

    2008-10-01

    Anaerobic syntrophic associations of fermentative bacteria and methanogenic archaea operate at the thermodynamic limits of life. The interspecies transfer of electrons from formate or hydrogen as a substrate for the methanogens is key. Contrary requirements of syntrophs and methanogens for growth-sustaining product and substrate concentrations keep the formate and hydrogen concentrations low and within a narrow range. Since formate is a direct substrate for methanogens, a niche for microorganisms that grow by the conversion of formate to hydrogen plus bicarbonate--or vice versa--may seem unlikely. Here we report experimental evidence for growth on formate by syntrophic communities of (i) Moorella sp. strain AMP in coculture with a thermophilic hydrogen-consuming Methanothermobacter species and of (ii) Desulfovibrio sp. strain G11 in coculture with a mesophilic hydrogen consumer, Methanobrevibacter arboriphilus AZ. In pure culture, neither Moorella sp. strain AMP, nor Desulfovibrio sp. strain G11, nor the methanogens grow on formate alone. These results imply the existence of a previously unrecognized microbial niche in anoxic environments.

  10. Bacterial community composition and fhs profiles of low- and high-ammonia biogas digesters reveal novel syntrophic acetate-oxidising bacteria.

    PubMed

    Müller, Bettina; Sun, Li; Westerholm, Maria; Schnürer, Anna

    2016-01-01

    Syntrophic acetate oxidation (SAO) is the predominant pathway for methane production in high ammonia anaerobic digestion processes. The bacteria (SAOB) occupying this niche and the metabolic pathway are poorly understood. Phylogenetic diversity and strict cultivation requirements hinder comprehensive research and discovery of novel SAOB. Most SAOB characterised to date are affiliated to the physiological group of acetogens. Formyltetrahydrofolate synthetase is a key enzyme of both acetogenic and SAO metabolism. The encoding fhs gene has therefore been identified as a suitable functional marker, using a newly designed primer pair. In this comparative study, we used a combination of terminal restriction fragment length polymorphism profiling, clone-based comparison, qPCR and Illumina amplicon sequencing to assess the bacterial community and acetogenic sub-community prevailing in high- and low-ammonia laboratory-scale digesters in order to delineate potential SAOB communities. Potential candidates identified were further tracked in a number of low-ammonia and high-ammonia laboratory-scale and large-scale digesters in order to reveal a potential function in SAO. All methodical approaches revealed significant changes in the bacterial community composition concurrently with increasing ammonia and predominance of SAO. The acetogenic community under high ammonia conditions was revealed to be generally heterogeneous, but formed distinct phylogenetic clusters. The clusters differed clearly from those found under low-ammonia conditions and represented an acetogenic assemblage unique for biogas processes and recurring in a number of high-ammonia processes, indicating potential involvement in SAO. The phylogenetic affiliation and population dynamics observed point to a key community, belonging mainly to the Clostridia class, in particular to the orders Clostridiales and Thermoanaerobacterales, which appear to specialise in SAO rather than being metabolically versatile. Overall

  11. Conversion of acids to alcohols by Clostridium ragsdalei strain P11: Process optimization and biochemistry

    NASA Astrophysics Data System (ADS)

    Isom, Catherine E.

    Research focus was directed toward the development of a biocatalyst that can be used to produce commodity chemicals and transportation fuels from volatile fatty acids ubiquitous in waste biomass. Clostridium ragsdalei was introduced to serve as an exemplar carboxidotrophic acetogen that reduces VFAs to alcohols of the same carbon structure with only acetate and ethanol as by-products of the fermentation. This dissertation developed a better understanding of this process in C. ragsdalei and, in turn, other similar bacteria and to supported previous discoveries as they relate to carboxylate reduction in acetogens. Additionally, pure culture studies allowed for a more detailed understanding of the biochemical behavior response to different compounds without skewing the results due to the influence of other species.

  12. Physiologically anaerobic microorganisms of the deep subsurface. Progress report, June 1, 1991--May 31, 1992

    SciTech Connect

    Stevens, S.E. Jr.; Chung, K.T.

    1992-06-01

    A variety of different media were used to isolate facultatively (FAB) and obligately anaerobic bacteria (OAB). These bacteria were isolated from core subsamples obtained from boreholes at the Idaho National Engineering Lab. (INEL) or at the Hanford Lab. (Yakima). Core material was sampled at various depths to 600 feet below the surface. All core samples with culturable bacteria contained at least FAB making thisthe most common physiological type of anaerobic bacteria present in the deep subsurface at these two sites. INEL core samples are characterized by isolates of both FAB and OAB. No isolates of acetogenic, methanogenic, or sulfate reducing bacteria were obtained. Yakima core samples are characterized by a marked predominance of FAB in comparison to OAB. In addition, isolates of acetogenic, methanogenic, and sulfate reducing bacteria were obtained. The Yakima site has the potential for complete anaerobic mineralization of organic compounds whereas this potential appears to be lacking at INEL.

  13. Physiologically anaerobic microorganisms of the deep subsurface

    SciTech Connect

    Stevens, S.E. Jr.; Chung, K.T.

    1992-06-01

    A variety of different media were used to isolate facultatively (FAB) and obligately anaerobic bacteria (OAB). These bacteria were isolated from core subsamples obtained from boreholes at the Idaho National Engineering Lab. (INEL) or at the Hanford Lab. (Yakima). Core material was sampled at various depths to 600 feet below the surface. All core samples with culturable bacteria contained at least FAB making thisthe most common physiological type of anaerobic bacteria present in the deep subsurface at these two sites. INEL core samples are characterized by isolates of both FAB and OAB. No isolates of acetogenic, methanogenic, or sulfate reducing bacteria were obtained. Yakima core samples are characterized by a marked predominance of FAB in comparison to OAB. In addition, isolates of acetogenic, methanogenic, and sulfate reducing bacteria were obtained. The Yakima site has the potential for complete anaerobic mineralization of organic compounds whereas this potential appears to be lacking at INEL.

  14. H2 and acetate transfers during xylan fermentation between a butyrate-producing xylanolytic species and hydrogenotrophic microorganisms from the human gut.

    PubMed

    Chassard, Christophe; Bernalier-Donadille, Annick

    2006-01-01

    The aim of this work was to investigate in vitro interrelationships during xylan fermentation between an H2 and butyrate-producing xylanolytic species recently isolated in our laboratory from human faeces and identified as Roseburia intestinalis and the H2-utilizing acetogen Ruminococcus hydrogenotrophicus or the methanogen Methanobrevibacter smithii. H2 transfer between M. smithii or Ru. hydrogenotrophicus and the xylanolytic species was evidenced, confirming the great potential of these H2-consuming microorganisms to reutilize fermentative H2 during fibre fermentation in the gut. In addition, acetate transfer was demonstrated between the xylanolytic Roseburia sp. and the acetogenic species, both metabolites transfers leading to butyric fermentation of oat xylan without production of H2.

  15. Enhancement in lipid content of Chlorella sp. MJ 11/11 from the spent medium of thermophilic biohydrogen production process.

    PubMed

    Ghosh, Supratim; Roy, Shantonu; Das, Debabrata

    2017-01-01

    The present study investigates the effect of spent media of acetogenic dark fermentation for mixotrophic algal cultivation for biodiesel production. Mixotrophic growth conditions were optimized in culture flask (250mL) using Chlorella sp. MJ 11/11. Maximum lipid accumulation (58% w/w) was observed under light intensity, pH, nitrate and phosphate concentration of 100μmolm(-2)s(-1), 7, 2.7mM and 1.8mM, respectively. Air lift (1.4L) and flat panel (1.4L) reactors were considered for algal cultivation. Air lift showed significant improvement in biomass and lipid production as compared to flat panel reactor. The results could help in development of sustainable technology involving acetogenic hydrogen production integrated with sequential mitigation of spent media by algal cultivation for improved energy recovery.

  16. Enzymology of the Wood–Ljungdahl Pathway of Acetogenesis

    PubMed Central

    Ragsdale, Stephen W.

    2011-01-01

    The biochemistry of acetogenesis is reviewed. The microbes that catalyze the reactions that are central to acetogenesis are described and the focus is on the enzymology of the process. These microbes play a key role in the global carbon cycle, producing over 10 trillion kilograms of acetic acid annually. Acetogens have the ability to anaerobically convert carbon dioxide and CO into acetyl-CoA by the Wood–Ljungdahl pathway, which is linked to energy conservation. They also can convert the six carbons of glucose stoichiometrically into 3 mol of acetate using this pathway. Acetogens and other anaerobic microbes (e.g., sulfate reducers and methanogens) use the Wood–Ljungdahl pathway for cell carbon synthesis. Important enzymes in this pathway that are covered in this review are pyruvate ferredoxin oxidoreductase, CO dehydrogenase/acetyl-CoA synthase, a corrinoid iron-sulfur protein, a methyltransferase, and the enzymes involved in the conversion of carbon dioxide to methyl-tetrahydrofolate. PMID:18378591

  17. Metagenomic analysis of the rumen microbial community following inhibition of methane formation by a halogenated methane analog

    PubMed Central

    Denman, Stuart E.; Martinez Fernandez, Gonzalo; Shinkai, Takumi; Mitsumori, Makoto; McSweeney, Christopher S.

    2015-01-01

    Japanese goats fed a diet of 50% Timothy grass and 50% concentrate with increasing levels of the anti-methanogenic compound, bromochloromethane (BCM) were investigated with respect to the microbial population and functional shifts in the rumen. Microbial ecology methods identified species that exhibited positive and negative responses to the increasing levels of BCM. The methane-inhibited rumen appeared to adapt to the higher H2 levels by shifting fermentation to propionate which was mediated by an increase in the population of H2-consuming Prevotella and Selenomonas spp. Metagenomic analysis of propionate production pathways was dominated by genomic content from these species. Reductive acetogenic marker gene libraries and metagenomics analysis indicate that reductive acetogenic species do not play a major role in the BCM treated rumen. PMID:26528253

  18. Bioelectrochemical Systems Workshop:Standardized Analyses, Design Benchmarks, and Reporting

    DTIC Science & Technology

    2012-01-01

    Producing Activities of Six Mesophilic Clostridium Species . Journal of Applied Microbiology, 2007. 103(6): p. 2258-2266. 27. Nevin, K.P., et al...lovleyi [30] and a comparative assessment of the exoelectrotrophic productivities of various acetogenic species [31]. There are two main strategies that...active species , such as mediators or biofilms. Steady-state kinetics should show an overlap in the forward and reverse CV curves. Unlike with

  19. 2012 Molecular Basis of Microbial One-Carbon Metabolism Gordon Research Conferences and Gordon Research Seminar, August 4-10,2012

    SciTech Connect

    Hanson, Thomas

    2012-08-10

    The 2012 Gordon Conference will present and discuss cutting-edge research in the field of microbial metabolism of C1 compounds. The conference will feature the roles and application of C1 metabolism in natural and synthetic systems at scales from molecules to ecosystems. The conference will stress molecular aspects of the unique metabolism exhibited by autotrophic bacteria, methanogens, methylotrophs, aerobic and anaerobic methanotrophs, and acetogens.

  20. Insights into CO2 Fixation Pathway of Clostridium autoethanogenum by Targeted Mutagenesis

    PubMed Central

    Liew, Fungmin; Henstra, Anne M.; Winzer, Klaus; Köpke, Michael; Simpson, Sean D.

    2016-01-01

    ABSTRACT The future sustainable production of chemicals and fuels from nonpetrochemical resources and reduction of greenhouse gas emissions are two of the greatest societal challenges. Gas fermentation, which utilizes the ability of acetogenic bacteria such as Clostridium autoethanogenum to grow and convert CO2 and CO into low-carbon fuels and chemicals, could potentially provide solutions to both. Acetogens fix these single-carbon gases via the Wood-Ljungdahl pathway. Two enzyme activities are predicted to be essential to the pathway: carbon monoxide dehydrogenase (CODH), which catalyzes the reversible oxidation of CO to CO2, and acetyl coenzyme A (acetyl-CoA) synthase (ACS), which combines with CODH to form a CODH/ACS complex for acetyl-CoA fixation. Despite their pivotal role in carbon fixation, their functions have not been confirmed in vivo. By genetically manipulating all three CODH isogenes (acsA, cooS1, and cooS2) of C. autoethanogenum, we highlighted the functional redundancies of CODH by demonstrating that cooS1 and cooS2 are dispensable for autotrophy. Unexpectedly, the cooS1 inactivation strain showed a significantly reduced lag phase and a higher growth rate than the wild type on H2 and CO2. During heterotrophic growth on fructose, the acsA inactivation strain exhibited 61% reduced biomass and the abolishment of acetate production (a hallmark of acetogens), in favor of ethanol, lactate, and 2,3-butanediol production. A translational readthrough event was discovered in the uniquely truncated (compared to those of other acetogens) C. autoethanogenum acsA gene. Insights gained from studying the function of CODH enhance the overall understanding of autotrophy and can be used for optimization of biotechnological production of ethanol and other commodities via gas fermentation. PMID:27222467

  1. A Deeply Branching Thermophilic Bacterium with an Ancient Acetyl-CoA Pathway Dominates a Subsurface Ecosystem

    PubMed Central

    Takami, Hideto; Noguchi, Hideki; Takaki, Yoshihiro; Uchiyama, Ikuo; Toyoda, Atsushi; Nishi, Shinro; Chee, Gab-Joo; Arai, Wataru; Nunoura, Takuro; Itoh, Takehiko; Hattori, Masahira; Takai, Ken

    2012-01-01

    A nearly complete genome sequence of Candidatus ‘Acetothermum autotrophicum’, a presently uncultivated bacterium in candidate division OP1, was revealed by metagenomic analysis of a subsurface thermophilic microbial mat community. Phylogenetic analysis based on the concatenated sequences of proteins common among 367 prokaryotes suggests that Ca. ‘A. autotrophicum’ is one of the earliest diverging bacterial lineages. It possesses a folate-dependent Wood-Ljungdahl (acetyl-CoA) pathway of CO2 fixation, is predicted to have an acetogenic lifestyle, and possesses the newly discovered archaeal-autotrophic type of bifunctional fructose 1,6-bisphosphate aldolase/phosphatase. A phylogenetic analysis of the core gene cluster of the acethyl-CoA pathway, shared by acetogens, methanogens, some sulfur- and iron-reducers and dechlorinators, supports the hypothesis that the core gene cluster of Ca. ‘A. autotrophicum’ is a particularly ancient bacterial pathway. The habitat, physiology and phylogenetic position of Ca. ‘A. autotrophicum’ support the view that the first bacterial and archaeal lineages were H2-dependent acetogens and methanogenes living in hydrothermal environments. PMID:22303444

  2. Syngas Biorefinery and Syngas Utilization.

    PubMed

    De Tissera, Sashini; Köpke, Michael; Simpson, Sean D; Humphreys, Christopher; Minton, Nigel P; Dürre, Peter

    2017-06-20

    Autotrophic acetogenic bacteria are able to capture carbon (CO or CO2) through gas fermentation, allowing them to grow on a spectrum of waste gases from industry (e.g., steel manufacture and oil refining, coal, and natural gas) and to produce ethanol. They can also consume syn(thesis) gas (CO and H2) made from the gasification of renewable/sustainable resources, such as biomass and domestic/agricultural waste. Acetogenic gas fermentation can, therefore, produce ethanol in any geographic region without competing for food or land. The commercialization of the process is now at an advanced stage. The real potential of acetogens, however, resides in their capacity to produce chemicals and fuels other than ethanol. This requires the redesign and implementation of more efficient metabolic pathways, adapting them to high performing manufacturing processes. Respective species, their bioenergetics, the genetic tools developed for their metabolic engineering, culture techniques and fermenter set-ups, as well as the commercialization, are comprehensively described and discussed in this chapter.

  3. Acetogenesis in the Energy-Starved Deep Biosphere – A Paradox?

    PubMed Central

    Lever, Mark Alexander

    2011-01-01

    Under anoxic conditions in sediments, acetogens are often thought to be outcompeted by microorganisms performing energetically more favorable metabolic pathways, such as sulfate reduction or methanogenesis. Recent evidence from deep subseafloor sediments suggesting acetogenesis in the presence of sulfate reduction and methanogenesis has called this notion into question, however. Here I argue that acetogens can successfully coexist with sulfate reducers and methanogens for multiple reasons. These include (1) substantial energy yields from most acetogenesis reactions across the wide range of conditions encountered in the subseafloor, (2) wide substrate spectra that enable niche differentiation by use of different substrates and/or pooling of energy from a broad range of energy substrates, (3) reduced energetic cost of biosynthesis among acetogens due to use of the reductive acetyl CoA pathway for both energy production and biosynthesis coupled with the ability to use many organic precursors to produce the key intermediate acetyl CoA. This leads to the general conclusion that, beside Gibbs free energy yields, variables such as metabolic strategy and energetic cost of biosynthesis need to be taken into account to understand microbial survival in the energy-depleted deep biosphere. PMID:22347874

  4. Acetogenesis in the energy-starved deep biosphere - a paradox?

    PubMed

    Lever, Mark Alexander

    2011-01-01

    Under anoxic conditions in sediments, acetogens are often thought to be outcompeted by microorganisms performing energetically more favorable metabolic pathways, such as sulfate reduction or methanogenesis. Recent evidence from deep subseafloor sediments suggesting acetogenesis in the presence of sulfate reduction and methanogenesis has called this notion into question, however. Here I argue that acetogens can successfully coexist with sulfate reducers and methanogens for multiple reasons. These include (1) substantial energy yields from most acetogenesis reactions across the wide range of conditions encountered in the subseafloor, (2) wide substrate spectra that enable niche differentiation by use of different substrates and/or pooling of energy from a broad range of energy substrates, (3) reduced energetic cost of biosynthesis among acetogens due to use of the reductive acetyl CoA pathway for both energy production and biosynthesis coupled with the ability to use many organic precursors to produce the key intermediate acetyl CoA. This leads to the general conclusion that, beside Gibbs free energy yields, variables such as metabolic strategy and energetic cost of biosynthesis need to be taken into account to understand microbial survival in the energy-depleted deep biosphere.

  5. Bacterial Population Development and Chemical Characteristics of Refuse Decomposition in a Simulated Sanitary Landfill

    PubMed Central

    Barlaz, M. A.; Schaefer, D. M.; Ham, R. K.

    1989-01-01

    Population development of key groups of bacteria involved in municipal refuse conversion to methane was measured from the time of initial incubation through the onset of methane production. Hemicellulolytic bacteria, cellulolytic bacteria, hydrogen-producing acetogens, and acetate- and H2-plus-CO2-utilizing methanogens were enumerated by the most-probable-number technique with media containing oat spelt xylan, ball-milled cellulose, butyrate, acetate, and H2 plus CO2, respectively. Refuse decomposition was monitored in multiple replicate laboratory-scale sanitary landfills. A laboratory-scale landfill was dismantled weekly for microbial and chemical analysis. Leachate was neutralized and recycled to ensure methanogenesis. The methane concentration of the sampled containers increased to 64% by day 69, at which time the maximum methane production rate, 929 liters of CH4 per dry kg-year, was measured. Population increases of 2, 4, 5, 5, and 6 orders of magnitude were measured between fresh refuse and the methane production phase for the hemicellulolytic bacteria, cellulolytic bacteria, butyrate-catabolizing acetogens, and acetate- and H2-CO2-utilizing methanogens, respectively. The cellulolytic bacteria and acetogens increased more slowly than the methanogens and only after the onset of methane production. The initial decrease in the pH of the refuse ecosystem from 7.5 to 5.7 was attributed to the accumulation of acidic end products of sugar fermentation, to the low acid-consuming activity of the acetogenic and methanogenic bacteria, and to levels of oxygen and nitrate in the fresh refuse sufficient for oxidation of only 8% of the sugars to carbon dioxide and water. Cellulose and hemicellulose decomposition was most rapid after establishment of the methanogenic and acetogenic populations and a reduction in the initial accumulation of carboxylic acids. A total of 72% of these carbohydrates were degraded in the container sampled after 111 days. Initially acetate

  6. Insights into CO2 Fixation Pathway of Clostridium autoethanogenum by Targeted Mutagenesis.

    PubMed

    Liew, Fungmin; Henstra, Anne M; Winzer, Klaus; Köpke, Michael; Simpson, Sean D; Minton, Nigel P

    2016-05-24

    The future sustainable production of chemicals and fuels from nonpetrochemical resources and reduction of greenhouse gas emissions are two of the greatest societal challenges. Gas fermentation, which utilizes the ability of acetogenic bacteria such as Clostridium autoethanogenum to grow and convert CO2 and CO into low-carbon fuels and chemicals, could potentially provide solutions to both. Acetogens fix these single-carbon gases via the Wood-Ljungdahl pathway. Two enzyme activities are predicted to be essential to the pathway: carbon monoxide dehydrogenase (CODH), which catalyzes the reversible oxidation of CO to CO2, and acetyl coenzyme A (acetyl-CoA) synthase (ACS), which combines with CODH to form a CODH/ACS complex for acetyl-CoA fixation. Despite their pivotal role in carbon fixation, their functions have not been confirmed in vivo By genetically manipulating all three CODH isogenes (acsA, cooS1, and cooS2) of C. autoethanogenum, we highlighted the functional redundancies of CODH by demonstrating that cooS1 and cooS2 are dispensable for autotrophy. Unexpectedly, the cooS1 inactivation strain showed a significantly reduced lag phase and a higher growth rate than the wild type on H2 and CO2 During heterotrophic growth on fructose, the acsA inactivation strain exhibited 61% reduced biomass and the abolishment of acetate production (a hallmark of acetogens), in favor of ethanol, lactate, and 2,3-butanediol production. A translational readthrough event was discovered in the uniquely truncated (compared to those of other acetogens) C. autoethanogenum acsA gene. Insights gained from studying the function of CODH enhance the overall understanding of autotrophy and can be used for optimization of biotechnological production of ethanol and other commodities via gas fermentation. Gas fermentation is an emerging technology that converts the greenhouse gases CO2 and CO in industrial waste gases and gasified biomass into fuels and chemical commodities. Acetogenic bacteria

  7. Attenuation of landfill leachate by UK Triassic sandstone aquifer materials. 2. Sorption and degradation of organic pollutants in laboratory columns

    NASA Astrophysics Data System (ADS)

    Thornton, Steven F.; Bright, Mildred I.; Lerner, David N.; Tellam, John H.

    2000-05-01

    The sorption and degradation of dissolved organic matter (DOM) and 13 organic micropollutants (BTEX, aromatic hydrocarbons, chloro-aromatic and -aliphatic compounds, and pesticides) in acetogenic and methanogenic landfill leachate was studied in laboratory columns containing Triassic sandstone aquifer materials from the English Midlands. Solute sorption and degradation relationships were evaluated using a simple transport model. Relative to predictions, micropollutant sorption was decreased up to eightfold in acetogenic leachate, but increased up to sixfold in methanogenic leachate. This behaviour reflects a combination of interactions between the micropollutants, leachate DOM and aquifer mineral fraction. Sorption of DOM was not significant. Degradation of organic fractions occurred under Mn-reducing and SO 4-reducing conditions. Degradation of some micropollutants occurred exclusively under Mn-reducing conditions. DOM and benzene were not significantly degraded under the conditions and time span (up to 280 days) of the experiments. Most micropollutants were degraded immediately or after a lag phase (32-115 days). Micropollutant degradation rates varied considerably (half-lives of 8 to >2000 days) for the same compounds (e.g., TeCE) in different experiments, and for compounds (e.g., naphthalene, DCB and TeCA) within the same experiment. Degradation of many micropollutants was both simultaneous and sequential, and inhibited by the utilisation of different substrates. This mechanism, in combination with lag phases, controls micropollutant degradation potential in these systems more than the degradation rate. These aquifer materials have a potentially large capacity for in situ bioremediation of organic pollutants in landfill leachate and significant degradation may occur in the Mn-reducing zones of leachate plumes. However, degradation of organic pollutants in acetogenic leachate may be limited in aquifers with low pH buffering capacity and reducible Mn oxides

  8. Competing Formate- and Carbon Dioxide-Utilizing Prokaryotes in an Anoxic Methane-Emitting Fen Soil▿†

    PubMed Central

    Hunger, Sindy; Schmidt, Oliver; Hilgarth, Maik; Horn, Marcus A.; Kolb, Steffen; Conrad, Ralf; Drake, Harold L.

    2011-01-01

    Methanogenesis in wetlands is dependent on intermediary substrates derived from the degradation of biopolymers. Formate is one such substrate and is stimulatory to methanogenesis and acetogenesis in anoxic microcosms of soil from the fen Schlöppnerbrunnen. Formate dissimilation also yields CO2 as a potential secondary substrate. The objective of this study was to resolve potential differences between anaerobic formate- and CO2-utilizing prokaryotes of this fen by stable isotope probing. Anoxic soil microcosms were pulsed daily with low concentrations of [13C]formate or 13CO2 (i.e., [13C]bicarbonate). Taxa were evaluated by assessment of 16S rRNA genes, mcrA (encoding the alpha-subunit of methyl-coenzyme M reductase), and fhs (encoding formyltetrahydrofolate synthetase). Methanogens, acetogens, and formate-hydrogen lyase-containing taxa appeared to compete for formate. Genes affiliated with Methanocellaceae, Methanobacteriaceae, Acetobacteraceae, and Rhodospirillaceae were 13C enriched (i.e., labeled) in [13C]formate treatments, whereas genes affiliated with Methanosarcinaceae, Conexibacteraceae, and Solirubrobacteraceae were labeled in 13CO2 treatments. [13C]acetate was enriched in [13C]formate treatments, but labeling of known acetogenic taxa was not detected. However, several phylotypes were affiliated with acetogen-containing taxa (e.g., Sporomusa). Methanosaetaceae-affiliated methanogens appeared to participate in the consumption of acetate. Twelve and 58 family-level archaeal and bacterial 16S rRNA phylotypes, respectively, were detected, approximately half of which had no isolated representatives. Crenarchaeota constituted half of the detected archaeal 16S rRNA phylotypes. The results highlight the unresolved microbial diversity of the fen Schlöppnerbrunnen, suggest that differing taxa competed for the same substrate, and indicate that Methanocellaceae, Methanobacteriaceae, Methanosarcinaceae, and Methanosaetaceae were linked to the production of methane

  9. Syntrophic Degradation of Lactate in Methanogenic Co-cultures

    SciTech Connect

    Meyer, Birte; Stahl, David

    2010-05-17

    In environments where the amount of the inorganic electron acceptors (oxygen, nitrate, sulfate, sulfur oroxidized metal ions (Fe3+;Mn4+) is insufficient for complete breakdown of organic matter, methane is formed as the major reduced end product. In such methanogenic environments organic acids are degraded by syntrophic associations of fermenting, acetogenic bacteria (e.g., sulfate-reducing bacteria (SRB) as"secondary fermenters") and methanogenic archaea. In these consortia, the conversion of lactate to acetate, CO2 and methane depends on the cooperating activities of both metabolically distinct microbial groups that are tightly linked by the need to maintain the exchanged metabolites (hydrogenandformate) at very low concentrations.

  10. Microbial Electrosynthesis: Feeding Microbes Electricity To Convert Carbon Dioxide and Water to Multicarbon Extracellular Organic Compounds

    PubMed Central

    Nevin, Kelly P.; Woodard, Trevor L.; Franks, Ashley E.; Summers, Zarath M.; Lovley, Derek R.

    2010-01-01

    The possibility of providing the acetogenic microorganism Sporomusa ovata with electrons delivered directly to the cells with a graphite electrode for the reduction of carbon dioxide to organic compounds was investigated. Biofilms of S. ovata growing on graphite cathode surfaces consumed electrons with the reduction of carbon dioxide to acetate and small amounts of 2-oxobutyrate. Electrons appearing in these products accounted for over 85% of the electrons consumed. These results demonstrate that microbial production of multicarbon organic compounds from carbon dioxide and water with electricity as the energy source is feasible. PMID:20714445

  11. Microbial electrosynthesis: feeding microbes electricity to convert carbon dioxide and water to multicarbon extracellular organic compounds.

    PubMed

    Nevin, Kelly P; Woodard, Trevor L; Franks, Ashley E; Summers, Zarath M; Lovley, Derek R

    2010-05-25

    The possibility of providing the acetogenic microorganism Sporomusa ovata with electrons delivered directly to the cells with a graphite electrode for the reduction of carbon dioxide to organic compounds was investigated. Biofilms of S. ovata growing on graphite cathode surfaces consumed electrons with the reduction of carbon dioxide to acetate and small amounts of 2-oxobutyrate. Electrons appearing in these products accounted for over 85% of the electrons consumed. These results demonstrate that microbial production of multicarbon organic compounds from carbon dioxide and water with electricity as the energy source is feasible.

  12. Recent Advances in Factors and Methods for Stimulation of Biomethane Production.

    PubMed

    Thakur, Neha; Khardenavis, Anshuman; Purohit, Hemant J

    2015-01-01

    The role of methanogenesis in the global carbon cycle is very important for recycle of renewable biomass which, has the potential for contribution to independence from fossil fuels. Anaerobic microbes comprised of fermentative and acetogenic species decompose the complex biomass to hydrogen, formate and, acetate that are further metabolized to methane by methanogens. A general review of biogenic production of methane and methanogenic diversity involved is presented. This review gives an overview of recent patents on methane production and focuses mainly on different methods, systems and, microbial methanogenic community involved in anaerobic digestion that can be used for improved understanding of the microbial community function and relationships in methanogenesis.

  13. (Fatty and aromatic acid catabolizing bacteria from methanogenic ecosystems). Annual technical progress report

    SciTech Connect

    Bryant, M.P.; Kammerer, J.J.

    1985-02-27

    A long-chain fatty acid degrading (beta oxidizing), obligate proton-reducing, acetogenic bacterium strain SD2 of the genus Syntrophomonas has been isolated in coculture with a hydrogen-using bacterium, Desulfovibrio strain G-11. The enzymology of fatty acid degradation is being studied to discover the differences of SD2 from S. wolfei which allow it to degrade long chain fatty acids. A new species, Clostridium pfennigii (V5-2) was isolated from the rumen. A new genus and species, Syntrophococcus sucromutans (S195) is present in relatively high numbers in rumen contents. Another new species is Eubacterium oxidoreducens. (ACR)

  14. Microbiology and physiology of anaerobic fermentation of cellulose. Annual report for 1990, 1992, 1993 and final report

    SciTech Connect

    Ljungdahl, L.G.; Wiegel, J.; Peck, H.D. Jr.; Mortenson, L.E.

    1993-08-31

    This report focuses on the bioconversion of cellulose to methane by various anaerobes. The structure and enzymatic activity of cellulosome and polycellulosome was studied in Clostridium thermocellum. The extracellular enzymes involved in the degradation of plant material and the physiology of fermentation was investigated in anaerobic fungi. Enzymes dealing with CO, CO{sub 2}, H{sub 2}, CH{sub 3}OH, as well as electron transport and energy generation coupled to the acetyl-CoA autotrophic pathway was studied in acetogenic clostridia.

  15. Anaerobic biodegradation of methyl esters by Acetobacterium woodii and Eubacterium limosum

    USGS Publications Warehouse

    Liu, Shi; Suflita, Joseph M.

    1994-01-01

    The ability ofAcetobacterium woodii andEubacterium limosum to degrade methyl esters of acetate, propionate, butyrate, and isobutyrate was examined under growing and resting-cell conditions. Both bacteria hydrolyzed the esters to the corresponding carboxylates and methanol under either condition. Methanol was further oxidized to formate under growing but not resting conditions. Unlike the metabolism of phenylmethylethers, no H2 requirement was evident for ester biotransformation. The hydrolysis of methyl carboxylates is thermodynamically favorable under standard conditions and the mixotrophic metabolism of ester/CO2 allowed for bacterial growth. These results suggest that the degradation of methyl carboxylates may be a heretofore unrecognized nutritional option for acetogenic bacteria.

  16. Population dynamics of anaerobic microbial consortia in thermophilic granular sludge in response to feed composition change.

    PubMed

    Syutsubo, K; Sinthurat, N; Ohashi, A; Harada, H

    2001-01-01

    A thermophilic UASB reactor was operated at 55 degrees C for greater than 470 days in order to investigate the effects of feed composition on the changes in microbial community structure where thermophilic granular sludge was used as the inoculum source. The feed compositions were changed with cultivation days; phase 1 (1-70 days), alcohol distillery wastewater; phase 2 (71-281 days), artificial acetate wastewater; phase 3 (282-474 days), artificial sucrose wastewater. During the first one month of each phase, the methanogenic activity and cell density of methanogens quantified by fluorescence in situ hybridization (FISH) drastically changed as a result of shift in feed composition. When artificial acetate wastewater was used as feed, retained granular sludge was partially disintegrated due to a decrease in the number of symbiotic bacterial community members: acetogens (acidogens) and hydrogenotrophic methanogens. In contrast, when the feed was shifted to sucrose (phase 3), granulation of biomass was promoted by a remarkable proliferation of the symbiotic community. The presence of hydrogen-utilizing methanogens and acetogens (acidogens) are shown to be effective for the enhancement of thermophilic granulation. The cell density of methanogens determined by FISH was strongly correlated with the methane-producing potential of the retained thermophilic granular sludge.

  17. Putting microbes to work in sequence: recent advances in temperature-phased anaerobic digestion processes.

    PubMed

    Lv, Wen; Schanbacher, Floyd L; Yu, Zhongtang

    2010-12-01

    Methane biogas production through anaerobic digestion (or biomethanation) is one of the few technologies that both produce bioenergy and protect the environment. When the focus of anaerobic digestion (AD) is shifted from primarily wastewater treatment to bioenergy production, efficiency and process stability become critical to the economic viability of AD technologies. Temperature-phased anaerobic digestion (TPAD) is a promising process that can significantly enhance both digestion efficiency and process robustness. A TPAD system separates the conventional AD process into two phases, so both phases can be optimized according to their individual functional needs. In the first, thermophilic phase, the often rate-limiting hydrolysis step of polymeric feedstock is accelerated by elevated temperatures, while in the second, mesophilic phase, the fastidious syntrophic acetogens and methanogens are provided with permissive conditions where inhibitions to key guilds (e.g., syntrophic acetogens and methanogens) are attenuated. Although large-scale TPAD systems have not been applied widely, researchers have demonstrated the potential superiority of TPAD systems over single-stage digesters and other AD processes with enhanced VS (volatile solids) and pathogen removal; increased methane yield, process stability, OLR (organic loading rate); shorter HRT (hydraulic retention time); decreased foaming and short-chain fatty acids in effluent. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  18. Application of real-time PCR to determination of combined effect of antibiotics on Bacteria, Methanogenic Archaea, Archaea in anaerobic sequencing batch reactors.

    PubMed

    Aydin, Sevcan; Ince, Bahar; Ince, Orhan

    2015-06-01

    This study evaluated the long-term effects of erythromycin-tetracycline-sulfamethoxazole (ETS) and sulfamethoxazole-tetracycline (ST) antibiotic combinations on the microbial community and examined the ways in which these antimicrobials impact the performance of anaerobic reactors. Quantitative real-time PCR was used to determine the effect that different antibiotic combinations had on the total and active Bacteria, Archae and Methanogenic Archae. Three primer sets that targeted metabolic genes encoding formylterahydrofolate synthetase, methyl-coenzyme M reductase and acetyl-coA synthetase were also used to determine the inhibition level on the mRNA expression of the homoacetogens, methanogens and specifically acetoclastic methanogens, respectively. These microorganisms play a vital role in the anaerobic degradation of organic waste and targeting these gene expressions offers operators or someone at a treatment plant the potential to control and the improve the anaerobic system. The results of the investigation revealed that acetogens have a competitive advantage over Archaea in the presence of ETS and ST combinations. Although the efficiency with which methane production takes place and the quantification of microbial populations in both the ETS and ST reactors decreased as antibiotic concentrations increased, the ETS batch reactor performed better than the ST batch reactor. According to the expression of genes results, the syntrophic interaction of acetogens and methanogens is critical to the performance of the ETS and ST reactors. Failure to maintain the stability of these microorganisms resulted in a decrease in the performance and stability of the anaerobic reactors.

  19. Microbial acetogenesis as a source of organic acids in ancient Atlantic Coastal Plain sediments

    USGS Publications Warehouse

    Chapelle, F.H.; Bradley, P.M.

    1996-01-01

    Field and laboratory evidence shows that deeply buried (90-888 m) fine-grained sediments of the Atlantic Coastal Plain contain viable acetogenic microorganisms, and that these microorganisms actively produce organic acids. Concentrations of formate, acetate, and propionate in pore waters extracted from fine-grained sediments ranged from 50 ??M to 5 mM and were much higher than in adjacent pore waters associated with sandy sediments (<2 ??M). Laboratory studies showed that asceptically cored fine-grained sediments incubated under a H2 atmosphere produced formate and acetate, and that H14CO-3 was converted to 14C-acetate and 14C-formate over time. An enrichment culture of these acetogenic microorganisms was recovered from one long-term incubation that showed the presence of several morphologically distinct gram-positive, rod-shaped bacteria. These microorganisms were capable of growth under autotrophic (H2 + CO2), heterotrophic (syringate), and mixotrophic (H2 + CO2 + syringate) conditions. These results suggest that microbial acetogenesis, rather than abiotic processes, is the most important organic acid-producing mechanism during low-temperature (???30 ??C) diagenesis of Atlantic Coastal Plain sediments.

  20. Shedding Light on the Microbial Community of the Macropod Foregut Using 454-Amplicon Pyrosequencing

    PubMed Central

    Gulino, Lisa-Maree; Ouwerkerk, Diane; Kang, Alicia Y. H.; Maguire, Anita J.; Kienzle, Marco; Klieve, Athol V.

    2013-01-01

    Twenty macropods from five locations in Queensland, Australia, grazing on a variety of native pastures were surveyed and the bacterial community of the foregut was examined using 454-amplicon pyrosequencing. Specifically, the V3/V4 region of 16S rRNA gene was examined. A total of 5040 OTUs were identified in the data set (post filtering). Thirty-two OTUs were identified as ‘shared’ OTUS (i.e. present in all samples) belonging to either Firmicutes or Bacteroidetes (Clostridiales/Bacteroidales). These phyla predominated the general microbial community in all macropods. Genera represented within the shared OTUs included: unclassified Ruminococcaceae, unclassified Lachnospiraceae, unclassified Clostridiales, Peptococcus sp. Coprococcus spp., Streptococcus spp., Blautia sp., Ruminoccocus sp., Eubacterium sp., Dorea sp., Oscillospira sp. and Butyrivibrio sp. The composition of the bacterial community of the foregut samples of each the host species (Macropus rufus, Macropus giganteus and Macropus robustus) was significantly different allowing differentiation between the host species based on alpha and beta diversity measures. Specifically, eleven dominant OTUs that separated the three host species were identified and classified as: unclassified Ruminococcaceae, unclassified Bacteroidales, Prevotella spp. and a Syntrophococcus sucromutans. Putative reductive acetogens and fibrolytic bacteria were also identified in samples. Future work will investigate the presence and role of fibrolytics and acetogens in these ecosystems. Ideally, the isolation and characterization of these organisms will be used for enhanced feed efficiency in cattle, methane mitigation and potentially for other industries such as the biofuel industry. PMID:23626688

  1. Genomic and enzymatic evidence for acetogenesis among multiple lineages of the archaeal phylum Bathyarchaeota widespread in marine sediments.

    PubMed

    He, Y; Li, M; Perumal, V; Feng, X; Fang, J; Xie, J; Sievert, S M; Wang, F

    2016-04-04

    Members of the archaeal phylum Bathyarchaeota are widespread and abundant in the energy-deficient marine subsurface sediments. However, their life strategies have remained largely elusive. Here, we provide genetic evidence that some lineages of Bathyarchaeota are acetogens, being capable of homoacetogenesis, a metabolism so far restricted to the domain Bacteria. Metabolic reconstruction based on genomic bins assembled from the metagenome of deep-sea subsurface sediments shows that the metabolism of some lineages of Bathyarchaeota is similar to that of bona fide bacterial homoacetogens, by having pathways for acetogenesis and for the fermentative utilization of a variety of organic substrates. Heterologous expression and activity assay of the acetate kinase gene ack from Bathyarchaeota, demonstrate further the capability of these Bathyarchaeota to grow as acetogens. The presence and expression of bathyarchaeotal genes indicative of active acetogenesis was also confirmed in Peru Margin subsurface sediments where Bathyarchaeota are abundant. The analyses reveal that this ubiquitous and abundant subsurface archaeal group has adopted a versatile life strategy to make a living under energy-limiting conditions. These findings further expand the metabolic potential of Archaea and argue for a revision of the role of Archaea in the carbon cycle of marine sediments.

  2. Bacterial growth with chlorinated methanes.

    PubMed Central

    Leisinger, T; Braus-Stromeyer, S A

    1995-01-01

    Chlorinated methanes are important industrial chemicals and significant environmental pollutants. While the highly chlorinated methanes, trichloromethane and tetrachloromethane, are not productively metabolized by bacteria, chloromethane and dichloromethane are used by both aerobic and anaerobic methylotrophic bacteria as carbon and energy sources. Some of the dehalogenation reactions involved in the utilization of the latter two compounds have been elucidated. In a strictly anaerobic acetogenic bacterium growing with chloromethane, an inducible enzyme forming methyltetrahydrofolate and chloride from chloromethane and tetrahydrofolate catalyzes dehalogenation of the growth substrate. A different mechanism for the nucleophilic displacement of chloride is observed in aerobic methylotrophic bacteria utilizing dichloromethane as the sole carbon and energy source. These organisms possess the enzyme dichloromethane dehalogenase which, in a glutathione-dependent reaction, converts dichloromethane to inorganic chloride and formaldehyde, a central metabolite of methylotrophic growth. Sequence comparisons have shown that bacterial dichloromethane dehalogenases belong to the glutathione S-transferase enzyme family, and within this family to class Theta. The dehalogenation reactions underlying aerobic utilization of chloromethane by a pure culture and anaerobic growth with dichloromethane by an acetogenic mixed culture are not known. It appears that they are based on mechanisms other than nucleophilic attack by tetrahydrofolate or glutathione. PMID:8565906

  3. Structure of the Alpha(2)F(2) Ni-Dependent CO Dehydrogenase Component of the Methanosarcina Barkeri Acetyl-CoA Decarbonylase/Synthase Complex

    SciTech Connect

    Gong, W.; Hao, B.; Wei, Z.; Ferguson, D.J.; Jr.; Tallant, T.; Krzycki, J.A.; Chang, M.K.

    2009-05-18

    Ni-dependent carbon monoxide dehydrogenases (Ni-CODHs) are a diverse family of enzymes that catalyze reversible CO:CO{sub 2} oxidoreductase activity in acetogens, methanogens, and some CO-using bacteria. Crystallography of Ni-CODHs from CO-using bacteria and acetogens has revealed the overall fold of the Ni-CODH core and has suggested structures for the C cluster that mediates CO:CO{sub 2} interconversion. Despite these advances, the mechanism of CO oxidation has remained elusive. Herein, we report the structure of a distinct class of Ni-CODH from methanogenic archaea: the {alpha}{sub 2}{epsilon}{sub 2} component from the {alpha}{sub 8}{beta}{sub 8}{gamma}{sub 8}{delta}{sub 8}{epsilon}{sub 8} CODH/acetyl-CoA decarbonylase/synthase complex, an enzyme responsible for the majority of biogenic methane production on Earth. The structure of this Ni-CODH component provides support for a hitherto unobserved state in which both CO and H{sub 2}O/OH{sup -} bind to the Ni and the exogenous FCII iron of the C cluster, respectively, and offers insight into the structures and functional roles of the {epsilon}-subunit and FeS domain not present in nonmethanogenic Ni-CODHs.

  4. Structure of the Alpha-2 Epsilon-2 Ni-dependent CO Dehydrogenase Component of the Methanosarcina Barkeri Acetyl-CoA Decarbonylase/Synthase Complex

    SciTech Connect

    Gong, W.; Hao, B; Wei, Z; Ferguson, Jr., D; Tallant, T; Krzycki, J; Chan, M

    2008-01-01

    Ni-dependent carbon monoxide dehydrogenases (Ni-CODHs) are a diverse family of enzymes that catalyze reversible CO:CO2 oxidoreductase activity in acetogens, methanogens, and some CO-using bacteria. Crystallography of Ni-CODHs from CO-using bacteria and acetogens has revealed the overall fold of the Ni-CODH core and has suggested structures for the C cluster that mediates CO:CO2 interconversion. Despite these advances, the mechanism of CO oxidation has remained elusive. Herein, we report the structure of a distinct class of Ni-CODH from methanogenic archaea: the ?2?2 component from the ?8?8?8?8?8 CODH/acetyl-CoA decarbonylase/synthase complex, an enzyme responsible for the majority of biogenic methane production on Earth. The structure of this Ni-CODH component provides support for a hitherto unobserved state in which both CO and H2O/OH- bind to the Ni and the exogenous FCII iron of the C cluster, respectively, and offers insight into the structures and functional roles of the ?-subunit and FeS domain not present in nonmethanogenic Ni-CODHs.

  5. How to sustainably feed a microbe: Strategies for biological production of carbon-based commodities with renewable electricity

    DOE PAGES

    Butler, Caitlyn S.; Lovley, Derek R.

    2016-11-28

    As interest and application of renewable energy grows, strategies are needed to align the asynchronous supply and demand. Microbial metabolisms are a potentially sustainable mechanism for transforming renewable electrical energy into biocommodities that are easily stored and transported. Acetogens and methanogens can reduce carbon dioxide to organic products including methane, acetic acid, and ethanol. The library of biocommodities is expanded when engineered metabolisms of acetogens are included. Typically, electrochemical systems are employed to integrate renewable energy sources with biological systems for production of carbon-based commodities. Within these systems, there are three prevailing mechanisms for delivering electrons to microorganisms for themore » conversion of carbon dioxide to reduce organic compounds: (1) electrons can be delivered to microorganisms via H2 produced separately in a electrolyzer, (2) H2 produced at a cathode can convey electrons to microorganisms supported on the cathode surface, and (3) a cathode can directly feed electrons to microorganisms. Each of these strategies has advantages and disadvantages that must be considered in designing full-scale processes. Furthermore, this review considers the evolving understanding of each of these approaches and the state of design for advancing these strategies toward viability.« less

  6. Metabolism of one-carbon compounds by the ruminal acetogan syntrophococcus sucromutans

    SciTech Connect

    Dore, J.; Bryant, M.P. )

    1990-04-01

    Syntrophococcus sucromutans is the predominant species capable of O demethylation of methoxylated lignin monoaromatic derivatives in the rumen. The enzymatic characterization of this acetogen indicated that it uses the acetyl coenzyme A (Woods) pathway. Cell extracts possess all the enzymes of the tetrahydrofolate pathway, as well as carbon monoxide dehydrogenase, at levels similar to those of other acetogens using this pathway. However, formate dehydrogenase could not be detected in cell extracts, whether formate or a methoxyaromatic was used as electron acceptor for growth of the cells on cellobiose. Labeled bicarbonate, formate, (1-{sup 14}C) pyruvate, and chemically synthesized O-(methyl-{sup 14}C) vanillate were used to further investigate the catabolism of one-carbon (C{sub 1}) compounds by using washed-cell preparations. The results were consistent with little or no contribution of formate dehydrogenase and pointed out some unique features. Conversion of formate to CO{sub 2} was detected, but labeled formate predominantly labeled the methyl group of acetate. Labeled CO{sub 2} readily exchanged with the carboxyl group of pyruvate but not with formate, and both labeled CO{sub 2} and pyruvate predominantly labeled the carboxyl group acetate. No CO{sub 2} was formed from O demethylation of vanillate, and the acetate produced was position labeled in the methyl group. The fermentation pattern and specific activities of products indicated a complete synthesis of acetate from pyruvate and the methoxyl group of vanillate.

  7. Enhanced anaerobic fermentation with azo dye as electron acceptor: simultaneous acceleration of organics decomposition and azo decolorization.

    PubMed

    Li, Yang; Zhang, Yaobin; Quan, Xie; Zhang, Jingxin; Chen, Shuo; Afzal, Shahzad

    2014-10-01

    Accumulation of hydrogen during anaerobic processes usually results in low decomposition of volatile organic acids (VFAs). On the other hand, hydrogen is a good electron donor for dye reduction, which would help the acetogenic conversion in keeping low hydrogen concentration. The main objective of the study was to accelerate VFA composition through using azo dye as electron acceptor. The results indicated that the azo dye serving as an electron acceptor could avoid H2 accumulation and accelerate anaerobic digestion of VFAs. After adding the azo dye, propionate decreased from 2400.0 to 689.5mg/L and acetate production increased from 180.0 to 519.5mg/L. It meant that the conversion of propionate into acetate was enhanced. Fluorescence in situ hybridization analysis showed that the abundance of propionate-utilizing acetogens with the presence of azo dye was greater than that in a reference without azo dye. The experiments via using glucose as the substrate further demonstrated that the VFA decomposition and the chemical oxygen demand (COD) removal increased by 319.7mg/L and 23.3% respectively after adding the azo dye. Therefore, adding moderate azo dye might be a way to recover anaerobic system from deterioration due to the accumulation of H2 or VFAs. Copyright © 2014. Published by Elsevier B.V.

  8. Methanogenic degradation of lignin-derived monoaromatic compounds by microbial enrichments from rice paddy field soil.

    PubMed

    Kato, Souichiro; Chino, Kanako; Kamimura, Naofumi; Masai, Eiji; Yumoto, Isao; Kamagata, Yoichi

    2015-09-24

    Anaerobic degradation of lignin-derived aromatics is an important metabolism for carbon and nutrient cycles in soil environments. Although there are some studies on degradation of lignin-derived aromatics by nitrate- and sulfate-reducing bacteria, knowledge on their degradation under methanogenic conditions are quite limited. In this study, methanogenic microbial communities were enriched from rice paddy field soil with lignin-derived methoxylated monoaromatics (vanillate and syringate) and their degradation intermediates (protocatechuate, catechol, and gallate) as the sole carbon and energy sources. Archaeal community analysis disclosed that both aceticlastic (Methanosarcina sp.) and hydrogenotrophic (Methanoculleus sp. and Methanocella sp.) methanogens dominated in all of the enrichments. Bacterial community analysis revealed the dominance of acetogenic bacteria (Sporomusa spp.) only in the enrichments on the methoxylated aromatics, suggesting that Sporomusa spp. initially convert vanillate and syringate into protocatechuate and gallate, respectively, with acetogenesis via O-demethylation. As the putative ring-cleavage microbes, bacteria within the phylum Firmicutes were dominantly detected from all of the enrichments, while the dominant phylotypes were not identical between enrichments on vanillate/protocatechuate/catechol (family Peptococcaceae bacteria) and on syringate/gallate (family Ruminococcaceae bacteria). This study demonstrates the importance of cooperation among acetogens, ring-cleaving fermenters/syntrophs and aceticlastic/hydrogenotrophic methanogens for degradation of lignin-derived aromatics under methanogenic conditions.

  9. Genome-Guided Analysis of Physiological Capacities of Tepidanaerobacter acetatoxydans Provides Insights into Environmental Adaptations and Syntrophic Acetate Oxidation

    PubMed Central

    Niazi, Adnan; Bongcam-Rudloff, Erik; Schnürer, Anna

    2015-01-01

    This paper describes the genome-based analysis of Tepidanaerobacter acetatoxydans strain Re1, a syntrophic acetate-oxidising bacterium (SAOB). Principal issues such as environmental adaptations, metabolic capacities, and energy conserving systems have been investigated and the potential consequences for syntrophic acetate oxidation discussed. Briefly, in pure culture, T. acetatoxydans grows with different organic compounds and produces acetate as the main product. In a syntrophic consortium with a hydrogenotrophic methanogen, it can also reverse its metabolism and instead convert acetate to formate/H2 and CO2. It can only proceed if the product formed is continuously removed. This process generates a very small amount of energy that is scarcely enough for growth, which makes this particular syntrophy of special interest. As a crucial member of the biogas-producing community in ammonium-rich engineered AD processes, genomic features conferring ammonium resistance, bacterial defense, oxygen and temperature tolerance were found, as well as attributes related to biofilm formation and flocculation. It is likely that T. acetatoxydans can form an electrochemical gradient by putative electron-bifurcating Rnf complex and [Fe-Fe] hydrogenases, as observed in other acetogens. However, genomic deficiencies related to acetogenic metabolism and anaerobic respiration were discovered, such as the lack of formate dehydrogenase and F1F0 ATP synthase. This has potential consequences for the metabolic pathways used under SAO and non-SAO conditions. The two complete sets of bacteriophage genomes, which were found to be encoded in the genome, are also worthy of mention. PMID:25811859

  10. Selective production of acetone during continuous synthesis gas fermentation by engineered biocatalyst Clostridium sp. MAceT113.

    PubMed

    Berzin, V; Kiriukhin, M; Tyurin, M

    2012-08-01

    To engineer acetogen biocatalyst capable of fermenting synthesis gas blend to acetone as the only liquid carbonaceous product. The metabolic engineering comprised inactivation of phosphotransacetylase via integration of a cassette comprising synthetic genes erm(B), thiolase and HMG-CoA synthase. Acetaldehyde dehydrogenase was inactivated via integration of a cassette consisting of synthetic genes cat, HMG-CoA lyase and acetoacetate decarboxylase. The engineered biocatalyst Clostridum sp. MAceT113 lost production of 253 mmol l(-1) ethanol and 296 mmol l(-1) acetate and started producing 1.8 mol l(-1) acetone in single-stage continuous syngas fermentation. The acetone concentration in culture broth is economical for bulk manufacture because it is about twenty times of that achieved with known acetone-butanol-ethanol fermentation of sugars. The process shows the opportunity to produce acetone from synthesis gas at concentrations comparable with production of acetone from products of petroleum cracking. This is the first report on elimination of acetate and acetaldehyde production and directing carbon flux from Acetyl-CoA to acetone via a non-naturally occurring in acetogen acetone biosynthesis pathway identified in eukaryotic organisms. © 2012 The Authors. Letters in Applied Microbiology © 2012 The Society for Applied Microbiology.

  11. Methanogenic degradation of lignin-derived monoaromatic compounds by microbial enrichments from rice paddy field soil

    PubMed Central

    Kato, Souichiro; Chino, Kanako; Kamimura, Naofumi; Masai, Eiji; Yumoto, Isao; Kamagata, Yoichi

    2015-01-01

    Anaerobic degradation of lignin-derived aromatics is an important metabolism for carbon and nutrient cycles in soil environments. Although there are some studies on degradation of lignin-derived aromatics by nitrate- and sulfate-reducing bacteria, knowledge on their degradation under methanogenic conditions are quite limited. In this study, methanogenic microbial communities were enriched from rice paddy field soil with lignin-derived methoxylated monoaromatics (vanillate and syringate) and their degradation intermediates (protocatechuate, catechol, and gallate) as the sole carbon and energy sources. Archaeal community analysis disclosed that both aceticlastic (Methanosarcina sp.) and hydrogenotrophic (Methanoculleus sp. and Methanocella sp.) methanogens dominated in all of the enrichments. Bacterial community analysis revealed the dominance of acetogenic bacteria (Sporomusa spp.) only in the enrichments on the methoxylated aromatics, suggesting that Sporomusa spp. initially convert vanillate and syringate into protocatechuate and gallate, respectively, with acetogenesis via O-demethylation. As the putative ring-cleavage microbes, bacteria within the phylum Firmicutes were dominantly detected from all of the enrichments, while the dominant phylotypes were not identical between enrichments on vanillate/protocatechuate/catechol (family Peptococcaceae bacteria) and on syringate/gallate (family Ruminococcaceae bacteria). This study demonstrates the importance of cooperation among acetogens, ring-cleaving fermenters/syntrophs and aceticlastic/hydrogenotrophic methanogens for degradation of lignin-derived aromatics under methanogenic conditions. PMID:26399549

  12. Energy Efficiency and Productivity Enhancement of Microbial Electrosynthesis of Acetate

    PubMed Central

    LaBelle, Edward V.; May, Harold D.

    2017-01-01

    It was hypothesized that a lack of acetogenic biomass (biocatalyst) at the cathode of a microbial electrosynthesis system, due to electron and nutrient limitations, has prevented further improvement in acetate productivity and efficiency. In order to increase the biomass at the cathode and thereby performance, a bioelectrochemical system with this acetogenic community was operated under galvanostatic control and continuous media flow through a reticulated vitreous carbon (RVC) foam cathode. The combination of galvanostatic control and the high surface area cathode reduced the electron limitation and the continuous flow overcame the nutrient limitation while avoiding the accumulation of products and potential inhibitors. These conditions were set with the intention of operating the biocathode through the production of H2. Biofilm growth occurred on and within the unmodified RVC foam regardless of vigorous H2 generation on the cathode surface. A maximum volumetric rate or space time yield for acetate production of 0.78 g/Lcatholyte/h was achieved with 8 A/Lcatholyte (83.3 A/m2projected surface area of cathode) supplied to the continuous flow/culture bioelectrochemical reactors. The total Coulombic efficiency in H2 and acetate ranged from approximately 80–100%, with a maximum of 35% in acetate. The overall energy efficiency ranged from approximately 35–42% with a maximum to acetate of 12%. PMID:28515713

  13. [Acidification and its effect on the population distributions of microorganisms in an anaerobic baffled reactor].

    PubMed

    Liu, Ran; Peng, Jian-feng; Song, Yong-hui; Wang, Yi-li; Yu, Lei; Yuan, Peng; Xie, Ming-shu

    2010-07-01

    The changes of pH, COD, volatile fatty acids (VFA) and microbial morphology of the acidification process in an anaerobic baffled reactor (ABR) were investigated. And the population succession process of the anaerobic microorganisms was quantitatively analyzed by using the Fluorescent In situ hybridization technology (FISH). The results show that the ABR reactor is acidified gradually from the front to the back. After the reactor is entirely acidified, the COD removal efficiency is only 30.9%, and the pH values are lowered by 1.0-2.2, while the VFA in effluent increases by 5.1 times. Additionally, the microbial morphology is significantly affected by the acidification process, in which not only the bacteria are deformed or died, but also the internal and external mass transfer of granular sludge becomes difficult. The quantitative analyses with FISH shows that in the acidification process the Archaea growth is inhibited but the Eubacteria growth is promoted, thus resulting in the sharp decrease of the three crucial microorganisms of the anaerobic digestion. The abundance of the butyrate-oxidizing acetogenic bacteria Syntrophomonas spp. reduces by 30.9%, the propionate-oxidizing acetogenic bacteria Syntrophobacter wolinii reduces by 85.5%, the homoacetogenic bacteria Acetobacterium species E. limosum reduces by 60.0%, and methanomicrobium Methanomicrobiales reduces by 54.3%. All these result in the upsetting of the mass transfer balances of different anaerobic microorganism populations.

  14. Impact of ethanol on the natural attenuation of MTBE in a normally sulfate-reducing aquifer.

    PubMed

    Mackay, Doug; de Sieyes, Nick; Einarson, Murray; Feris, Kevin; Pappas, Alex; Wood, Isaac; Jacobson, Lisa; Justice, Larry; Noske, Mark; Wilson, John; Adair, Cherri; Scow, Kate

    2007-03-15

    Side-by-side experiments were conducted in an aquifer contaminated with methyl-tert-butyl ether (MTBE) at a former fuel station to evaluate the effect of ethanol release on the fate of pre-existing MTBE contamination. On one side, for approximately 9 months we injected groundwater amended with 1-3 mg/L benzene, toluene, and o-xylene (BToX). On the other side, we injected the same, adding approximately 500 mg/L ethanol. The fates of BToX in both sides ("lanes") were addressed in a prior publication. No MTBE transformation was observed in the "No Ethanol Lane." In the "With Ethanol Lane", MTBE was transformed to tert-butyl alcohol (TBA) underthe methanogenic and/or acetogenic conditions induced by the in situ biodegradation of the ethanol downgradient of the injection wells. The lag time before onset of this transformation was less than 2 months and the pseudo-first-order reaction rate estimated after 7-8 months was 0.046 d(-1). Our results imply that rapid subsurface transformation of MTBE to TBA may be expected in situations where strongly anaerobic conditions are sustained and fluxes of requisite nutrients and electron donors allow development of an active acetogenic/methanogenic zone beyond the reach of inhibitory effects such as those caused by high concentrations of ethanol.

  15. Impact of ethanol on the natural attenuation of benzene, toluene, and o-xylene in a normally sulfate-reducing aquifer.

    PubMed

    Mackay, Douglas M; de Sieyes, Nicholas R; Einarson, Murray D; Feris, Kevin P; Pappas, Alexander A; Wood, Isaac A; Jacobson, Lisa; Justice, Larry G; Noske, Mark N; Scow, Kate M; Wilson, John T

    2006-10-01

    Side-by-side experiments were conducted in a sulfate-reducing aquifer at a former fuel station to evaluate the effect of ethanol on biodegradation of other gasoline constituents. On one side, for approximately 9 months we injected groundwater amended with 1-3 mg/L benzene, toluene, and o-xylene (BToX). On the other side, we injected the same, adding approximately 500 mg/L ethanol. Initially the BToX plumes on both sides ("lanes") extended approximately the same distance. Thereafter, the plumes in the "No Ethanol Lane" retracted significantly, which we hypothesize to be due to an initial acclimation period followed by improvement in efficiency of biodegradation under sulfate-reducing conditions. In the "With Ethanol Lane", the BToX plumes also retracted, but more slowly and not as far. The preferential biodegradation of ethanol depleted dissolved sulfate, leading to methanogenic/acetogenic conditions. We hypothesize that BToX in the ethanol-impacted lane were biodegraded in part within the methanogenic/acetogenic zone and, in part, within sulfate-reducing zones developing along the plume fringes due to mixing with sulfate-containing groundwater surrounding the plumes due to dispersion and/or shifts in flow direction. Overall, this research confirms that ethanol may reduce rates of biodegradation of aromatic fuel components in the subsurface, in both transient and near steady-state conditions.

  16. Shedding light on the microbial community of the macropod foregut using 454-amplicon pyrosequencing.

    PubMed

    Gulino, Lisa-Maree; Ouwerkerk, Diane; Kang, Alicia Y H; Maguire, Anita J; Kienzle, Marco; Klieve, Athol V

    2013-01-01

    Twenty macropods from five locations in Queensland, Australia, grazing on a variety of native pastures were surveyed and the bacterial community of the foregut was examined using 454-amplicon pyrosequencing. Specifically, the V3/V4 region of 16S rRNA gene was examined. A total of 5040 OTUs were identified in the data set (post filtering). Thirty-two OTUs were identified as 'shared' OTUS (i.e. present in all samples) belonging to either Firmicutes or Bacteroidetes (Clostridiales/Bacteroidales). These phyla predominated the general microbial community in all macropods. Genera represented within the shared OTUs included: unclassified Ruminococcaceae, unclassified Lachnospiraceae, unclassified Clostridiales, Peptococcus sp. Coprococcus spp., Streptococcus spp., Blautia sp., Ruminoccocus sp., Eubacterium sp., Dorea sp., Oscillospira sp. and Butyrivibrio sp. The composition of the bacterial community of the foregut samples of each the host species (Macropus rufus, Macropus giganteus and Macropus robustus) was significantly different allowing differentiation between the host species based on alpha and beta diversity measures. Specifically, eleven dominant OTUs that separated the three host species were identified and classified as: unclassified Ruminococcaceae, unclassified Bacteroidales, Prevotella spp. and a Syntrophococcus sucromutans. Putative reductive acetogens and fibrolytic bacteria were also identified in samples. Future work will investigate the presence and role of fibrolytics and acetogens in these ecosystems. Ideally, the isolation and characterization of these organisms will be used for enhanced feed efficiency in cattle, methane mitigation and potentially for other industries such as the biofuel industry.

  17. Investigation of the microbial metabolism of carbon dioxide and hydrogen in the kangaroo foregut by stable isotope probing

    PubMed Central

    Godwin, Scott; Kang, Alicia; Gulino, Lisa-Maree; Manefield, Mike; Gutierrez-Zamora, Maria-Luisa; Kienzle, Marco; Ouwerkerk, Diane; Dawson, Kerri; Klieve, Athol V

    2014-01-01

    Kangaroos ferment forage material in an enlarged forestomach analogous to the rumen, but in contrast to ruminants, they produce little or no methane. The objective of this study was to identify the dominant organisms and pathways involved in hydrogenotrophy in the kangaroo forestomach, with the broader aim of understanding how these processes are able to predominate over methanogenesis. Stable isotope analysis of fermentation end products and RNA stable isotope probing (RNA-SIP) were used to investigate the organisms and biochemical pathways involved in the metabolism of hydrogen and carbon dioxide in the kangaroo forestomach. Our results clearly demonstrate that the activity of bacterial reductive acetogens is a key factor in the reduced methane output of kangaroos. In in vitro fermentations, the microbial community of the kangaroo foregut produced very little methane, but produced a significantly greater proportion of acetate derived from carbon dioxide than the microbial community of the bovine rumen. A bacterial operational taxonomic unit closely related to the known reductive acetogen Blautia coccoides was found to be associated with carbon dioxide and hydrogen metabolism in the kangaroo foregut. Other bacterial taxa including members of the genera Prevotella, Oscillibacter and Streptococcus that have not previously been reported as containing hydrogenotrophic organisms were also significantly associated with metabolism of hydrogen and carbon dioxide in the kangaroo forestomach. PMID:24621520

  18. Elimination of acetate production to improve ethanol yield during continuous synthesis gas fermentation by engineered biocatalyst Clostridium sp. MTEtOH550.

    PubMed

    Berzin, Vel; Kiriukhin, Michael; Tyurin, Michael

    2012-05-01

    Acetogen strain Clostridum sp. MT653 produced acetate 273 mM (p < 0.005) and ethanol 250 mM (p < 0.005) from synthesis gas blend mixture of 64% CO and 36% H(2). Clostridum sp. MT653 was metabolically engineered to the biocatalyst strain Clostridium sp. MTEtOH550. The biocatalyst increased ethanol yield to 590 mM with no acetate production during single-stage continuous syngas fermentation due to expression of synthetic adh cloned in a multi-copy number expression vector. The acetate production was eliminated by inactivation of the pta gene in Clostridium sp. MTEtOH550. Gene introduction and gene elimination were achieved only using Syngas Biofuels Energy, Inc. electroporation generator. The electrotransformation efficiencies were 8.0 ± 0.2 × 10(6) per microgram of transforming DNA of the expression vector at cell viability ~15%. The frequency of suicidal vector integration to inactivate pta was ~10(-5) per the number of recipient cells. This is the first report on elimination of acetate production and overexpression of synthetic adh gene to engineer acetogen biocatalyst for selective biofuel ethanol production during continuous syngas fermentation.

  19. Selective n-butanol production by Clostridium sp. MTButOH1365 during continuous synthesis gas fermentation due to expression of synthetic thiolase, 3-hydroxy butyryl-CoA dehydrogenase, crotonase, butyryl-CoA dehydrogenase, butyraldehyde dehydrogenase, and NAD-dependent butanol dehydrogenase.

    PubMed

    Berzin, Vel; Tyurin, Michael; Kiriukhin, Michael

    2013-02-01

    Acetogen Clostridum sp. MT1962 produced 287 mM acetate (p < 0.005) and 293 mM ethanol (p < 0.005) fermenting synthesis gas blend 60% CO and 40% H₂ in single-stage continuous fermentation. This strain was metabolically engineered to the biocatalyst Clostridium sp. MTButOH1365. The engineered biocatalyst lost production of ethanol and acetate while initiated the production of 297 mM of n-butanol (p < 0.005). The metabolic engineering comprised Cre-lox66/lox71-based elimination of phosphotransacetylase and acetaldehyde dehydrogenase along with integration to chromosome synthetic thiolase, 3-hydroxy butyryl-CoA dehydrogenase, crotonase, butyryl-CoA dehydrogenase, butyraldehyde dehydrogenase, and NAD-dependent butanol dehydrogenase. This is the first report on elimination of acetate and ethanol production genes and expression of synthetic gene cluster encoding n-butanol biosynthesis pathway in acetogen biocatalyst for selective fuel n-butanol production with no antibiotic support for the introduced genes.

  20. Methanogenesis from methanol and methylamines and acetogenesis from hydrogen and carbon dioxide in the sediments of a eutrophic lake

    SciTech Connect

    Lovley, D.R.; Klug, M.J.

    1983-04-01

    /sup 14/C-tracer techniques were used to examine the metabolism of methanol and methylamines and acetogenesis from hydrogen and carbon dioxide in sediments from the profundal and littoral zones of eutrophic Wintergreen Lake, Michigan. Methanogens were primarily responsible for the metabolism of methanol, mono-methylammine, and trimethylamine and maintained the pool size of these substrates below 10 ..mu..M in both sediment types. Methanol and methylamines were the precursors for less than 5 and 1%, respectively, of the total methane produced. Methanol and methylamines continued to be metabolized to methane when the sulfate concentration in the sediment was increased to 20 mM. Less than 2% of the total acetate production was derived from carbon dioxide reduction. Hydrogen consumption by hydrogen-oxidizing acetogens was 5% or less of the total hydrogen uptake by acetogens and methanogens. These results, in conjunction with previous studies, emphasize that acetate and hydrogen are the major methane precursors and that methanogens are the predominant hydrogen consumers in the sediments of this eutrophic lake.

  1. Microbial ecophysiology of whey biomethanation

    SciTech Connect

    Chartrain, M.M.

    1986-01-01

    The biodegradation of lactose into methane was investigated in a chemostat ecosystem under steady state conditions in order to understand the intermediary metabolism and the responsible bacterial species; and, to model the anaerobic digestion of whey in a continuous contact process. Radioactive carbon tracer studies showed that lactose biomethanation occurred in three distinct but simultaneous metabolic steps with lactate, acetate and hydrogen/carbon dioxide as the major intermediary metabolites. Mixed culture studies on the ecosystem composition demonstrated that multiple species of well described anaerobic bacteria were participating in each of three trophic groups: hydrolytic, acetogenic, and methanogenic. Biomethanation performance studies analyzed the dynamics of bacterial species composition and competition in relation to dilution rate. These results demonstrated that the hydrolytic and acetogenic bacteria were coupled to the methanogenic bacteria by interspecies hydrogen transfer; that species competition and dominance for a given carbon metabolite or for hydrogen was related to specific substrate transformation kinetic properties; and that the data was useful for describing biomethanation with a mechanistic model. Starter cultures were developed by employing freeze drying techniques to preserve either a defined culture comprised of four prevalent digestor species or an adapted chemostat sludge. Both of these starter cultures were shown to effectively degrade lactose using either a defined medium or raw whey as biomethanation starting substrate.

  2. The effects of spatial and temporal variations in acetate and sulfate on methane cycling in two Michigan peatlands

    SciTech Connect

    Shannon, R.D.; White, J.R.

    1996-05-01

    In this 3-yr study, we investigated temporal and spatial changes in pools of pore-water methane, sulfate, and acetate (a methanogenic substrate), along with corresponding methane fluxes. Acetate accumulated to high concentrations (>500 {mu}M) in the shallow pore waters in winter and spring, indicating that acetogenic bacteria were metabolically active at 2-10{degree}C. High sulfate concentrations in the upper pore water of one site delayed onset of acetate accumulation, suggesting that acetate consumption by sulfate-reducing bacteria or competition between H{sub 2}-consuming sulfate-reducers and autotrophic acetogens caused the delay. Sites dominated by grasses and sedges has larger accumulations of acetate; pore-water pools and fluxes of methane increased rapidly following diminution of the acetate pools at these sites. At a site dominated by chamaedaphne calyculata, an ericaceous shrub, methane fluxes were lower by 2-3 orders of magnitude; little accumulation of acetate or methane was observed, except in the third year of the study when water levels were consistently high. The seasonal accumulation of acetate and subsequent methane production and flux seem to be dependent on long-term anaerobic conditions in the upper 20 cm of surface peat. 42 refs., 7 figs.

  3. How to sustainably feed a microbe: Strategies for biological production of carbon-based commodities with renewable electricity

    SciTech Connect

    Butler, Caitlyn S.; Lovley, Derek R.

    2016-11-28

    As interest and application of renewable energy grows, strategies are needed to align the asynchronous supply and demand. Microbial metabolisms are a potentially sustainable mechanism for transforming renewable electrical energy into biocommodities that are easily stored and transported. Acetogens and methanogens can reduce carbon dioxide to organic products including methane, acetic acid, and ethanol. The library of biocommodities is expanded when engineered metabolisms of acetogens are included. Typically, electrochemical systems are employed to integrate renewable energy sources with biological systems for production of carbon-based commodities. Within these systems, there are three prevailing mechanisms for delivering electrons to microorganisms for the conversion of carbon dioxide to reduce organic compounds: (1) electrons can be delivered to microorganisms via H2 produced separately in a electrolyzer, (2) H2 produced at a cathode can convey electrons to microorganisms supported on the cathode surface, and (3) a cathode can directly feed electrons to microorganisms. Each of these strategies has advantages and disadvantages that must be considered in designing full-scale processes. Furthermore, this review considers the evolving understanding of each of these approaches and the state of design for advancing these strategies toward viability.

  4. Microbial Methane Production Associated with Carbon Steel Corrosion in a Nigerian Oil Field.

    PubMed

    Mand, Jaspreet; Park, Hyung S; Okoro, Chuma; Lomans, Bart P; Smith, Seun; Chiejina, Leo; Voordouw, Gerrit

    2015-01-01

    Microbially influenced corrosion (MIC) in oil field pipeline systems can be attributed to many different types of hydrogenotrophic microorganisms including sulfate reducers, methanogens and acetogens. Samples from a low temperature oil reservoir in Nigeria were analyzed using DNA pyrotag sequencing. The microbial community compositions of these samples revealed an abundance of anaerobic methanogenic archaea. Activity of methanogens was demonstrated by incubating samples anaerobically in a basal salts medium, in the presence of carbon steel and carbon dioxide. Methane formation was measured in all enrichments and correlated with metal weight loss. Methanogens were prominently represented in pipeline solids samples, scraped from the inside of a pipeline, comprising over 85% of all pyrosequencing reads. Methane production was only witnessed when carbon steel beads were added to these pipeline solids samples, indicating that no methane was formed as a result of degradation of the oil organics present in these samples. These results were compared to those obtained for samples taken from a low temperature oil field in Canada, which had been incubated with oil, either in the presence or in the absence of carbon steel. Again, methanogens present in these samples catalyzed methane production only when carbon steel was present. Moreover, acetate production was also found in these enrichments only in the presence of carbon steel. From these studies it appears that carbon steel, not oil organics, was the predominant electron donor for acetate production and methane formation in these low temperature oil fields, indicating that the methanogens and acetogens found may contribute significantly to MIC.

  5. Microbial Methane Production Associated with Carbon Steel Corrosion in a Nigerian Oil Field

    PubMed Central

    Mand, Jaspreet; Park, Hyung S.; Okoro, Chuma; Lomans, Bart P.; Smith, Seun; Chiejina, Leo; Voordouw, Gerrit

    2016-01-01

    Microbially influenced corrosion (MIC) in oil field pipeline systems can be attributed to many different types of hydrogenotrophic microorganisms including sulfate reducers, methanogens and acetogens. Samples from a low temperature oil reservoir in Nigeria were analyzed using DNA pyrotag sequencing. The microbial community compositions of these samples revealed an abundance of anaerobic methanogenic archaea. Activity of methanogens was demonstrated by incubating samples anaerobically in a basal salts medium, in the presence of carbon steel and carbon dioxide. Methane formation was measured in all enrichments and correlated with metal weight loss. Methanogens were prominently represented in pipeline solids samples, scraped from the inside of a pipeline, comprising over 85% of all pyrosequencing reads. Methane production was only witnessed when carbon steel beads were added to these pipeline solids samples, indicating that no methane was formed as a result of degradation of the oil organics present in these samples. These results were compared to those obtained for samples taken from a low temperature oil field in Canada, which had been incubated with oil, either in the presence or in the absence of carbon steel. Again, methanogens present in these samples catalyzed methane production only when carbon steel was present. Moreover, acetate production was also found in these enrichments only in the presence of carbon steel. From these studies it appears that carbon steel, not oil organics, was the predominant electron donor for acetate production and methane formation in these low temperature oil fields, indicating that the methanogens and acetogens found may contribute significantly to MIC. PMID:26793176

  6. Growth dynamics of major microbial populations during biodegradation of o-phthalate in anaerobic sediment slurries under a CO2/H2 atmosphere.

    PubMed

    Liu, S-M; Lin, Y-L; Tsai, T-L

    2005-03-01

    o-Phthalate transformers increased about five orders of magnitude (to 1.6x10(11)cells g-1 sediment) just before the onset of fast biotransformation of o-phthalate (21.6 mg l-1) and then decreased sharply when the concentration of o-phthalate became low during biodegradation of o-phthalate in anaerobic sediment slurries under CO2/H2 (4:1, v/v). In contrast, the benzoate transformers increased about four orders of magnitude (to 1.6x10(11)cells g-1 sediment) in 48 days and then increased one more order (to 1.6x10(12)cells g-1 sediment) in 60 days and then remained at that high level in those sediment slurries. When making a comparison between the growth dynamics of o-phthalate transformers, acetogens, sulfate reducers, and methanogens and the time course of o-phthalate transformation, it appears that acetogens did not initiate biotransformation of o-phthalate, and that sulfate reducers and methanogens were not directly involved in o-phthalte degradation. o-Phthalate was not transformed in sediment slurries amended with BESA plus molybdate under CO2/H2.

  7. How to Sustainably Feed a Microbe: Strategies for Biological Production of Carbon-Based Commodities with Renewable Electricity

    PubMed Central

    Butler, Caitlyn S.; Lovley, Derek R.

    2016-01-01

    As interest and application of renewable energy grows, strategies are needed to align the asynchronous supply and demand. Microbial metabolisms are a potentially sustainable mechanism for transforming renewable electrical energy into biocommodities that are easily stored and transported. Acetogens and methanogens can reduce carbon dioxide to organic products including methane, acetic acid, and ethanol. The library of biocommodities is expanded when engineered metabolisms of acetogens are included. Typically, electrochemical systems are employed to integrate renewable energy sources with biological systems for production of carbon-based commodities. Within these systems, there are three prevailing mechanisms for delivering electrons to microorganisms for the conversion of carbon dioxide to reduce organic compounds: (1) electrons can be delivered to microorganisms via H2 produced separately in a electrolyzer, (2) H2 produced at a cathode can convey electrons to microorganisms supported on the cathode surface, and (3) a cathode can directly feed electrons to microorganisms. Each of these strategies has advantages and disadvantages that must be considered in designing full-scale processes. This review considers the evolving understanding of each of these approaches and the state of design for advancing these strategies toward viability. PMID:27965629

  8. How to Sustainably Feed a Microbe: Strategies for Biological Production of Carbon-Based Commodities with Renewable Electricity.

    PubMed

    Butler, Caitlyn S; Lovley, Derek R

    2016-01-01

    As interest and application of renewable energy grows, strategies are needed to align the asynchronous supply and demand. Microbial metabolisms are a potentially sustainable mechanism for transforming renewable electrical energy into biocommodities that are easily stored and transported. Acetogens and methanogens can reduce carbon dioxide to organic products including methane, acetic acid, and ethanol. The library of biocommodities is expanded when engineered metabolisms of acetogens are included. Typically, electrochemical systems are employed to integrate renewable energy sources with biological systems for production of carbon-based commodities. Within these systems, there are three prevailing mechanisms for delivering electrons to microorganisms for the conversion of carbon dioxide to reduce organic compounds: (1) electrons can be delivered to microorganisms via H2 produced separately in a electrolyzer, (2) H2 produced at a cathode can convey electrons to microorganisms supported on the cathode surface, and (3) a cathode can directly feed electrons to microorganisms. Each of these strategies has advantages and disadvantages that must be considered in designing full-scale processes. This review considers the evolving understanding of each of these approaches and the state of design for advancing these strategies toward viability.

  9. Effect of aluminium and sulphate on anaerobic digestion of sludge from wastewater enhanced primary treatment.

    PubMed

    Cabirol, N; Barragán, E J; Durán, A; Noyola, A

    2003-01-01

    The combined and individual effects of aluminium and sulphate at concentrations of 1,000 mg/l as Al(OH)3, and 150 mgSO4(2-)/L as K2SO4, respectively, on the anaerobic digestion of sludge from enhanced primary treatment (EPT) were evaluated in 1 L capacity semi continuous reactors. It was found that at 59 days, aluminium inhibits the specific methanogenic activity (SMA) of methanogenic and acetogenic bacteria resulting in a 50% to 72% decrease. Sulphate also inhibits (48% to 65%) the SMA of the same type of bacteria. Methanogenic and acetogenic bacteria were able to adapt, to a different extent, to the assayed concentrations of aluminium and sulphate. However, the combination of aluminium and sulphate resulted in a higher inhibition, especially of the hydrogenophilic methanogenic bacteria. Indeed, this effect remained during the time of the experiment, maintaining an inhibition of 44% at 114 days. Feeding with EPT sludge led to a bigger decrease in SMA of each bacterial group, with respect to the other treatments with time. It is concluded that the acidification of anaerobic reactors fed with EPT sludge is due, among other causes, to the concurrent presence of aluminium and sulphate.

  10. Investigation of the microbial metabolism of carbon dioxide and hydrogen in the kangaroo foregut by stable isotope probing.

    PubMed

    Godwin, Scott; Kang, Alicia; Gulino, Lisa-Maree; Manefield, Mike; Gutierrez-Zamora, Maria-Luisa; Kienzle, Marco; Ouwerkerk, Diane; Dawson, Kerri; Klieve, Athol V

    2014-09-01

    Kangaroos ferment forage material in an enlarged forestomach analogous to the rumen, but in contrast to ruminants, they produce little or no methane. The objective of this study was to identify the dominant organisms and pathways involved in hydrogenotrophy in the kangaroo forestomach, with the broader aim of understanding how these processes are able to predominate over methanogenesis. Stable isotope analysis of fermentation end products and RNA stable isotope probing (RNA-SIP) were used to investigate the organisms and biochemical pathways involved in the metabolism of hydrogen and carbon dioxide in the kangaroo forestomach. Our results clearly demonstrate that the activity of bacterial reductive acetogens is a key factor in the reduced methane output of kangaroos. In in vitro fermentations, the microbial community of the kangaroo foregut produced very little methane, but produced a significantly greater proportion of acetate derived from carbon dioxide than the microbial community of the bovine rumen. A bacterial operational taxonomic unit closely related to the known reductive acetogen Blautia coccoides was found to be associated with carbon dioxide and hydrogen metabolism in the kangaroo foregut. Other bacterial taxa including members of the genera Prevotella, Oscillibacter and Streptococcus that have not previously been reported as containing hydrogenotrophic organisms were also significantly associated with metabolism of hydrogen and carbon dioxide in the kangaroo forestomach.

  11. Perchlorate and chlorate reduction by the Crenarchaeon Aeropyrum pernix and two thermophilic Firmicutes.

    PubMed

    Liebensteiner, Martin G; Pinkse, Martijn W H; Nijsse, Bart; Verhaert, Peter D E M; Tsesmetzis, Nicolas; Stams, Alfons J M; Lomans, Bart P

    2015-12-01

    This study reports the ability of one hyperthermophilic and two thermophilic microorganisms to grow anaerobically by the reduction of chlorate and perchlorate. Physiological, genomic and proteome analyses suggest that the Crenarchaeon Aeropyrum pernix reduces perchlorate with a periplasmic enzyme related to nitrate reductases, but that it lacks a functional chlorite-disproportionating enzyme (Cld) to complete the pathway. Aeropyrum pernix, previously described as a strictly aerobic microorganism, seems to rely on the chemical reactivity of reduced sulfur compounds with chlorite, a mechanism previously reported for perchlorate-reducing Archaeoglobus fulgidus. The chemical oxidation of thiosulfate (in excessive amounts present in the medium) and the reduction of chlorite result in the release of sulfate and chloride, which are the products of a biotic-abiotic perchlorate reduction pathway in Ae. pernix. The apparent absence of Cld in two other perchlorate-reducing microorganisms, Carboxydothermus hydrogenoformans and Moorella glycerini strain NMP, and their dependence on sulfide for perchlorate reduction is consistent with the observations made on Ar. fulgidus. Our findings suggest that microbial perchlorate reduction at high temperature differs notably from the physiology of perchlorate- and chlorate-reducing mesophiles and that it is characterized by the lack of a chlorite dismutase and is enabled by a combination of biotic and abiotic reactions. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  12. The Heat Resistance of Microbial Cells Represented by D Values Can be Estimated by the Transition Temperature and the Coefficient of Linear Expansion.

    PubMed

    Nakanishi, Koichi; Kogure, Akinori; Deuchi, Keiji; Kuwana, Ritsuko; Takamatsu, Hiromu; Ito, Kiyoshi

    2015-01-01

    We previously developed a method for evaluating the heat resistance of microorganisms by measuring the transition temperature at which the coefficient of linear expansion of a cell changes. Here, we performed heat resistance measurements using a scanning probe microscope with a nano thermal analysis system. The microorganisms studied included six strains of the genus Bacillus or related genera, one strain each of the thermophilic obligate anaerobic bacterial genera Thermoanaerobacter and Moorella, two strains of heat-resistant mold, two strains of non-sporulating bacteria, and one strain of yeast. Both vegetative cells and spores were evaluated. The transition temperature at which the coefficient of linear expansion due to heating changed from a positive value to a negative value correlated strongly with the heat resistance of the microorganism as estimated from the D value. The microorganisms with greater heat resistance exhibited higher transition temperatures. There was also a strong negative correlation between the coefficient of linear expansion and heat resistance in bacteria and yeast, such that microorganisms with greater heat resistance showed lower coefficients of linear expansion. These findings suggest that our method could be useful for evaluating the heat resistance of microorganisms.

  13. Analysis of propionate-degrading consortia from agricultural biogas plants.

    PubMed

    Ahlert, Stephan; Zimmermann, Rita; Ebling, Johannes; König, Helmut

    2016-12-01

    In order to investigate the propionate-degrading community of agricultural biogas plants, four propionate-degrading consortia (Ap1a, N12, G12, and Wp2a) were established from different biogas plants which were fed with renewable resources. The consortia were cultivated in a batch for a period of 2-4 years and then analyzed in an 8-week batch experiment for microbial succession during propionate degradation. Community shifts showed considerable propagation of Syntrophobacter sulfatireducens, Cryptanaerobacter sp./Pelotomaculum sp., and "Candidatus Cloacamonas sp." in the course of decreasing propionate concentration. Methanogenic species belonged mainly to the genera Methanosarcina, Methanosaeta, and Methanoculleus. Due to the prevalent presence of the syntrophic acetate-oxidizing species Tepidanaerobacter acetatoxydans and potentially autotrophic homoacetogenic bacteria (Moorella sp., Thermacetogenium sp.), a theoretical involvement of syntrophic acetate oxidation and autotrophic homoacetogenesis in stable and efficient propionate degradation was indicated. Considering theoretical Gibbs free energy values at different hydrogen partial pressures, it is noticeable that syntrophic acetate oxidation and autotrophic homoacetogenesis have the potential to counterbalance adverse hydrogen partial pressure fluctuations, stabilizing most probably continuous and stable propionate degradation. © 2016 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  14. Deep Conversion of Carbon Monoxide to Hydrogen and Formation of Acetate by the Anaerobic Thermophile Carboxydothermus hydrogenoformans

    PubMed Central

    Henstra, Anne M.; Stams, Alfons J. M.

    2011-01-01

    Carboxydothermus hydrogenoformans is a thermophilic strictly anaerobic bacterium that catalyses the water gas shift reaction, the conversion of carbon monoxide with water to molecular hydrogen and carbon dioxide. The thermodynamically favorable growth temperature, compared to existing industrial catalytic processes, makes this organism an interesting alternative for production of cheap hydrogen gas suitable to fuel CO-sensitive fuel cells in a future hydrogen economy, provided sufficiently low levels of CO are reached. Here we study CO conversion and final CO levels in cultures of C. hydrogenoformans grown in batch cultures that were started with a 100% CO gas phase with and without removal of formed CO2. Final CO levels were 117 ppm without CO2 removal and below 2 ppm with CO2 removal. The Gibbs free energy change calculated with measured end concentrations and the detection of acetate suggest that C. hydrogenoformans shifted from a hydrogenogenic to an acetogenic metabolism. PMID:21754940

  15. Mathematical model of anaerobic digestion in a chemostat: effects of syntrophy and inhibition.

    PubMed

    Weedermann, Marion; Seo, Gunog; Wolkowicz, Gail S K

    2013-01-01

    Three of the four main stages of anaerobic digestion: acidogenesis, acetogenesis, and methanogenesis are described by a system of differential equations modelling the interaction of microbial populations in a chemostat. The microbes consume and/or produce simple substrates, alcohols and fatty acids, acetic acid, and hydrogen. Acetogenic bacteria and hydrogenotrophic methanogens interact through syntrophy. The model also includes the inhibition of acetoclastic and hydrogenotrophic methanogens due to sensitivity to varying pH-levels. To examine the effects of these interactions and inhibitions, we first study an inhibition-free model and obtain results for global stability using differential inequalities together with conservation laws. For the model with inhibition, we derive conditions for existence, local stability, and bistability of equilibria and present a global stability result. A case study illustrates the effects of inhibition on the regions of stability. Inhibition introduces regions of bistability and stabilizes some equilibria.

  16. Genesis of acetate and methane by gut bacteria of nutritionally diverse termites

    SciTech Connect

    Brauman, A.; Labat, M. ); Kane, M.D.; Breznak, J.A. )

    1992-09-04

    The evolution of different feeding guilds in termites is paralleled by differences in the activity of their gut microbiota. In wood-feeding termites, carbon dioxide-reducing acetogenic bacteria were found to generally outprocess carbon dioxide-reducing methanogenic bacteria for reductant (presumably hydrogen) generated during microbial fermentation in the hindgut. By contrast, acetogenesis from hydrogen and carbon dioxide was of little significance in fungus-growing and soil-feeding termites, which evolved more methane than their wood- and grass-feeding counterparts. Given the large biomass of termites on the earth and especially in the tropics, these findings should help refine global estimates of carbon dioxide reduction in anoxic habitats and the contribution of termite emissions to atmospheric methane concentrations.

  17. Development of biological coal gasification (MicGAS process). Final report, May 1, 1990--May 31, 1995

    SciTech Connect

    1998-12-31

    ARCTECH has developed a novel process (MicGAS) for direct, anaerobic biomethanation of coals. Biomethanation potential of coals of different ranks (Anthracite, bitumious, sub-bitumious, and lignites of different types), by various microbial consortia, was investigated. Studies on biogasification of Texas Lignite (TxL) were conducted with a proprietary microbial consortium, Mic-1, isolated from hind guts of soil eating termites (Zootermopsis and Nasutitermes sp.) and further improved at ARCTECH. Various microbial populations of the Mic-1 consortium carry out the multi-step MicGAS Process. First, the primary coal degraders, or hydrolytic microbes, degrade the coal to high molecular weight (MW) compounds. Then acedogens ferment the high MW compounds to low MW volatile fatty acids. The volatile fatty acids are converted to acetate by acetogens, and the methanogens complete the biomethanation by converting acetate and CO{sub 2} to methane.

  18. A coupled, pore-scale model for methanogenic microbial activity in underground hydrogen storage

    NASA Astrophysics Data System (ADS)

    Ebigbo, Anozie; Golfier, Fabrice; Quintard, Michel

    2013-11-01

    Underground hydrogen storage (UHS) as a means of energy storage is an efficient way of compensating for seasonal fluctuations in the availability of energy. One important factor which influences this technology is the activity of methanogenic microorganisms capable of utilising hydrogen and carbon dioxide for metabolism and leading to a change in the stored gas composition. A coupled, pore-scale model is presented which aids in the investigation of the mechanisms that govern the conversion of hydrogen to methane, i.e. advective hydrogen flow, its diffusion into microbial biofilms of multiple species, and its consumption within these biofilms. The model assumes that spherical grains are coated by a film of residual water and treats the biofilm development within each film in a quasi one-dimensional manner. A sample simulation using the presented model illustrates the biofilm growth process in these films as well as the competition between three different microbial species: methanogens, acetogens, and acetotrophs.

  19. Carbon recovery by fermentation of CO-rich off gases - Turning steel mills into biorefineries.

    PubMed

    Molitor, Bastian; Richter, Hanno; Martin, Michael E; Jensen, Rasmus O; Juminaga, Alex; Mihalcea, Christophe; Angenent, Largus T

    2016-09-01

    Technological solutions to reduce greenhouse gas (GHG) emissions from anthropogenic sources are required. Heavy industrial processes, such as steel making, contribute considerably to GHG emissions. Fermentation of carbon monoxide (CO)-rich off gases with wild-type acetogenic bacteria can be used to produce ethanol, acetate, and 2,3-butanediol, thereby, reducing the carbon footprint of heavy industries. Here, the processes for the production of ethanol from CO-rich off gases are discussed and a perspective on further routes towards an integrated biorefinery at a steel mill is given. Recent achievements in genetic engineering as well as integration of other biotechnology platforms to increase the product portfolio are summarized. Already, yields have been increased and the portfolio of products broadened. To develop a commercially viable process, however, the extraction from dilute product streams is a critical step and alternatives to distillation are discussed. Finally, another critical step is waste(water) treatment with the possibility to recover resources.

  20. My Lifelong Passion for Biochemistry and Anaerobic Microorganisms.

    PubMed

    Thauer, Rudolf Kurt

    2015-01-01

    Early parental influence led me first to medical school, but after developing a passion for biochemistry and sensing the need for a deeper foundation, I changed to chemistry. During breaks between semesters, I worked in various biochemistry labs to acquire a feeling for the different areas of investigation. The scientific puzzle that fascinated me most was the metabolism of the anaerobic bacterium Clostridium kluyveri, which I took on in 1965 in Karl Decker's lab in Freiburg, Germany. I quickly realized that little was known about the biochemistry of strict anaerobes such as clostridia, methanogens, acetogens, and sulfate-reducing bacteria and that these were ideal model organisms to study fundamental questions of energy conservation, CO2 fixation, and the evolution of metabolic pathways. My passion for anaerobes was born then and is unabated even after 50 years of study.

  1. Advanced studies of biological indirect liquefaction of coal: Topical report on Task 1: Culture identification

    SciTech Connect

    1989-01-01

    Culture identification and characterization studies carried out at the University of Arkansas and under contract to the University of Oklahoma, Department of Botany and Microbiology, have been essentially completed. The studies indicate that the organism is indeed a new clostridial strain, to be named Clostridium ljungdahlii, strain PETC, in honor of Dr. Lars G. Ljungdahl for his work on clostridia and acetogens. C. ljungdahlii is different from other clostridial strains and similar geni in its operating conditions and choice of substrates as sole carbon and energy sources. C. ljungdahlii, strain PETC, produces ethanol as a product only at low pH levels, with acetate the primary product at higher pH levels. 46 refs., 5 figs., 6 tabs.

  2. Gas fermentation: cellular engineering possibilities and scale up.

    PubMed

    Heijstra, Björn D; Leang, Ching; Juminaga, Alex

    2017-04-12

    Low carbon fuels and chemicals can be sourced from renewable materials such as biomass or from industrial and municipal waste streams. Gasification of these materials allows all of the carbon to become available for product generation, a clear advantage over partial biomass conversion into fermentable sugars. Gasification results into a synthesis stream (syngas) containing carbon monoxide (CO), carbon dioxide (CO2), hydrogen (H2) and nitrogen (N2). Autotrophy-the ability to fix carbon such as CO2 is present in all domains of life but photosynthesis alone is not keeping up with anthropogenic CO2 output. One strategy is to curtail the gaseous atmospheric release by developing waste and syngas conversion technologies. Historically microorganisms have contributed to major, albeit slow, atmospheric composition changes. The current status and future potential of anaerobic gas-fermenting bacteria with special focus on acetogens are the focus of this review.

  3. Occurrence of ferredoxin:NAD+ oxidoreductase activity and its ion specificity in several Gram-positive and Gram-negative bacteria

    PubMed Central

    Hess, Verena; Gallegos, Rene; Jones, J Andrew; Barquera, Blanca; Malamy, Michael H

    2016-01-01

    A ferredoxin:NAD+ oxidoreductase was recently discovered as a redox-driven ion pump in the anaerobic, acetogenic bacterium Acetobacterium woodii. The enzyme is assumed to be encoded by the rnf genes. Since these genes are present in the genomes of many bacteria, we tested for ferredoxin:NAD+ oxidoreductase activity in cytoplasmic membranes from several different Gram-positive and Gram-negative bacteria that have annotated rnf genes. We found this activity in Clostridium tetanomorphum, Clostridium ljungdahlii, Bacteroides fragilis, and Vibrio cholerae but not in Escherichia coli and Rhodobacter capsulatus. As in A. woodii, the activity was Na+-dependent in C. tetanomorphum and B. fragilis but Na+-independent in C. ljungdahlii and V. cholerae. We deleted the rnf genes from B. fragilis and demonstrated that the mutant has greatly reduced ferredoxin:NAD+ oxidoreductase activity. This is the first genetic proof that the rnf genes indeed encode the reduced ferredoxin:NAD+ oxidoreductase activity. PMID:26793417

  4. Genome sequence of Frateuria aurantia type strain (Kondo 67(T)), a xanthomonade isolated from Lilium auratium Lindl.

    SciTech Connect

    Anderson, Iain; Teshima, Hazuki; Nolan, Matt; Lapidus, Alla L.; Tice, Hope; Glavina Del Rio, Tijana; Cheng, Jan-Fang; Han, Cliff; Tapia, Roxanne; Goodwin, Lynne A.; Pitluck, Sam; Liolios, Konstantinos; Mavromatis, K; Pagani, Ioanna; Ivanova, N; Mikhailova, Natalia; Pati, Amrita; Chen, Amy; Palaniappan, Krishna; Land, Miriam L; Rohde, Manfred; Lang, Elke; Detter, J. Chris; Goker, Markus; Woyke, Tanja; Bristow, James; Eisen, Jonathan; Markowitz, Victor; Hugenholtz, Philip; Kyrpides, Nikos C; Klenk, Hans-Peter

    2013-01-01

    rateuria aurantia (ex Kondo and Ameyama 1958) Swings et al. 1980 is a member of the bispecific genus Frateuria in the family Xanthomonadaceae, which is already heavily targeted for non-type strain genome sequencing. Strain Kondo 67(T) was initially (1958) identified as a member of 'Acetobacter aurantius', a name that was not considered for the approved list. Kondo 67(T) was therefore later designated as the type strain of the newly proposed acetogenic species Frateuria aurantia. The strain is of interest because of its triterpenoids (hopane family). F. aurantia Kondo 67(T) is the first member of the genus Frateura whose genome sequence has been deciphered, and here we describe the features of this organism, together with the complete genome sequence and annotation. The 3,603,458-bp long chromosome with its 3,200 protein-coding and 88 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

  5. Electrobiocommodities: powering microbial production of fuels and commodity chemicals from carbon dioxide with electricity

    SciTech Connect

    Lovley, DR; Nevin, KP

    2013-06-01

    Electricity can be an energy source for microbially catalyzed production of fuels and other organic commodities from carbon dioxide. These electrobiocommodities (E-BCs) can be produced directly via electrode-to-microbe electron transfer or indirectly with electrochemically generated electron donors such as H-2 or formate. Producing E-BCs may be a more efficient and environmentally sustainable strategy for converting solar energy to biocommodities than approaches that rely on biological photosynthesis. A diversity of microbial physiologies could potentially be adapted for E-BC production, but to date acetogenic microorganisms are the only organisms shown to covert electrically generated low potential electrons and carbon dioxide into multi-carbon organic products with high recovery of electrons in product. Substantial research and development will be required for E-BC commercialization.

  6. Mathematical model of anaerobic digestion in a chemostat: effects of syntrophy and inhibition

    PubMed Central

    Weedermann, Marion; Seo, Gunog; Wolkowicz, Gail S.K.

    2013-01-01

    Three of the four main stages of anaerobic digestion: acidogenesis, acetogenesis, and methanogenesis are described by a system of differential equations modelling the interaction of microbial populations in a chemostat. The microbes consume and/or produce simple substrates, alcohols and fatty acids, acetic acid, and hydrogen. Acetogenic bacteria and hydrogenotrophic methanogens interact through syntrophy. The model also includes the inhibition of acetoclastic and hydrogenotrophic methanogens due to sensitivity to varying pH-levels. To examine the effects of these interactions and inhibitions, we first study an inhibition-free model and obtain results for global stability using differential inequalities together with conservation laws. For the model with inhibition, we derive conditions for existence, local stability, and bistability of equilibria and present a global stability result. A case study illustrates the effects of inhibition on the regions of stability. Inhibition introduces regions of bistability and stabilizes some equilibria. PMID:23336708

  7. Electrobiocommodities: powering microbial production of fuels and commodity chemicals from carbon dioxide with electricity.

    PubMed

    Lovley, Derek R; Nevin, Kelly P

    2013-06-01

    Electricity can be an energy source for microbially catalyzed production of fuels and other organic commodities from carbon dioxide. These electrobiocommodities (E-BCs) can be produced directly via electrode-to-microbe electron transfer or indirectly with electrochemically generated electron donors such as H2 or formate. Producing E-BCs may be a more efficient and environmentally sustainable strategy for converting solar energy to biocommodities than approaches that rely on biological photosynthesis. A diversity of microbial physiologies could potentially be adapted for E-BC production, but to date acetogenic microorganisms are the only organisms shown to covert electrically generated low potential electrons and carbon dioxide into multi-carbon organic products with high recovery of electrons in product. Substantial research and development will be required for E-BC commercialization. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Lipid growth requirement and influence of lipid supplement on fatty acid and aldehyde composition of Syntrophococcus sucromutans.

    PubMed Central

    Doré, J; Bryant, M P

    1989-01-01

    Results concerning the ruminal fluid growth requirement of the ruminal acetogen, Syntrophococcus sucromutans, indicate that octadecenoic acid isomers satisfy this essential requirement. Complex lipids, such as triglycerides and phospholipids, can also support growth. The cellular fatty acid and aldehyde composition closely reflects that of the lipid supplement provided to the cells. Up to 98% of the fatty acids and 80% of the fatty aldehydes are identical in chain length and degree of unsaturation to the octadecenoic acid supplement provided in the medium. S. sucromutans shows a tendency to have a greater proportion of the aldehyde form among its 18 carbon chains than it does with the shorter-chain simple lipids, which may be interpreted as a strategy to maintain membrane fluidity. 14C labeling showed that most of the oleic acid taken up from the medium was incorporated into the membrane fraction of the cells. PMID:2729991

  9. An Ancient Pathway Combining Carbon Dioxide Fixation with the Generation and Utilization of a Sodium Ion Gradient for ATP Synthesis

    PubMed Central

    Poehlein, Anja; Schmidt, Silke; Kaster, Anne-Kristin; Goenrich, Meike; Vollmers, John; Thürmer, Andrea; Bertsch, Johannes; Schuchmann, Kai; Voigt, Birgit; Hecker, Michael; Daniel, Rolf; Thauer, Rudolf K.; Gottschalk, Gerhard; Müller, Volker

    2012-01-01

    Synthesis of acetate from carbon dioxide and molecular hydrogen is considered to be the first carbon assimilation pathway on earth. It combines carbon dioxide fixation into acetyl-CoA with the production of ATP via an energized cell membrane. How the pathway is coupled with the net synthesis of ATP has been an enigma. The anaerobic, acetogenic bacterium Acetobacterium woodii uses an ancient version of this pathway without cytochromes and quinones. It generates a sodium ion potential across the cell membrane by the sodium-motive ferredoxin:NAD oxidoreductase (Rnf). The genome sequence of A. woodii solves the enigma: it uncovers Rnf as the only ion-motive enzyme coupled to the pathway and unravels a metabolism designed to produce reduced ferredoxin and overcome energetic barriers by virtue of electron-bifurcating, soluble enzymes. PMID:22479398

  10. A biological process effective for the conversion of CO-containing industrial waste gas to acetate.

    PubMed

    Kim, Tae Wan; Bae, Seung Seob; Lee, Jin Woo; Lee, Sung-Mok; Lee, Jung-Hyun; Lee, Hyun Sook; Kang, Sung Gyun

    2016-07-01

    Acetogens have often been observed to be inhibited by CO above an inhibition threshold concentration. In this study, a two-stage culture consisting of carboxydotrophic archaea and homoacetogenic bacteria is found to be effective in converting industrial waste gas derived from a steel mill process. In the first stage, Thermococcus onnurineus could grow on the Linz-Donawitz converter gas (LDG) containing ca. 56% CO as a sole energy source, converting the CO into H2 and CO2. Then, in the second stage, Thermoanaerobacter kivui could grow on the off-gas from the first stage culture, consuming the H2 and CO in the off-gas completely and producing acetate as a main product. T. kivui alone could not grow on the LDG gas. This work represents the first demonstration of acetate production using steel mill waste gas by a two-stage culture of carboxydotrophic hydrogenogenic microbes and homoacetogenic bacteria.

  11. Sulfate reduction at low pH to remediate acid mine drainage.

    PubMed

    Sánchez-Andrea, Irene; Sanz, Jose Luis; Bijmans, Martijn F M; Stams, Alfons J M

    2014-03-30

    Industrial activities and the natural oxidation of metallic sulfide-ores produce sulfate-rich waters with low pH and high heavy metals content, generally termed acid mine drainage (AMD). This is of great environmental concern as some heavy metals are highly toxic. Within a number of possibilities, biological treatment applying sulfate-reducing bacteria (SRB) is an attractive option to treat AMD and to recover metals. The process produces alkalinity, neutralizing the AMD simultaneously. The sulfide that is produced reacts with the metal in solution and precipitates them as metal sulfides. Here, important factors for biotechnological application of SRB such as the inocula, the pH of the process, the substrates and the reactor design are discussed. Microbial communities of sulfidogenic reactors treating AMD which comprise fermentative-, acetogenic- and SRB as well as methanogenic archaea are reviewed. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Metabolism of fatty acids by Syntrophomonas wolfei. Progress report, March 15, 1983-December 15, 1984

    SciTech Connect

    McInerney, M.J.

    1984-01-01

    The main objective in this research is to obtain basic information on the metabolism and physiology of the H/sub 2/-producing acetogenic bacteria. Syntrophomonas wolfei degrades butyrate, caproate and caprylate to acetate and H/sub 2/, valerate and heptanoate to acetate, propionate and H/sub 2/, and isoheptanoate to acetate, isovalerate and H/sub 2/. Since the degradation of these compounds with H/sub 2/ production is energetically favorable only when the partial pressure of H/sub 2/ is maintained at a very low level, S. wolfei can only be grown in coculture with H/sub 2/-using bacteria. The first objectives of this research were to determine if S. wolfei could be grown alone if the partial pressure of H/sub 2/ was maintained at a low level by sparging the culture with anaerobic gas.

  13. Supercooled water brines within permafrost-an unknown ecological niche for microorganisms: a model for astrobiology.

    PubMed

    Gilichinsky, D; Rivkina, E; Shcherbakova, V; Laurinavichuis, K; Tiedje, J

    2003-01-01

    This study describes brine lenses (cryopegs) found in Siberian permafrost derived from ancient marine sediment layers of the Arctic Ocean. The cryopegs were formed and isolated from sediment ~100,000-120,000 years ago. They remain liquid at the in situ temperature of -10 degrees C as a result of their high salt content (170-300 g/L). [(14)C] Glucose is taken up by the cryopeg biomass at -15 degrees C, indicating microbial metabolism at low temperatures in this habitat. Furthermore, aerobic, anaerobic heterotrophs, sulfate reducers, acetogens, and methanogens were detected by most probable number analysis. Two psychrophilic microbes were isolated from the cryopegs, a Clostridium and a Psychrobacter. The closest relatives of each were previously isolated from Antarctica. The cryopeg econiche might serve as a model for extraterrestrial life, and hence is of particular interest to astrobiology.

  14. C1-carbon sources for chemical and fuel production by microbial gas fermentation.

    PubMed

    Dürre, Peter; Eikmanns, Bernhard J

    2015-12-01

    Fossil resources for production of fuels and chemicals are finite and fuel use contributes to greenhouse gas emissions and global warming. Thus, sustainable fuel supply, security, and prices necessitate the implementation of alternative routes to the production of chemicals and fuels. Much attention has been focussed on use of cellulosic material, particularly through microbial-based processes. However, this is still costly and proving challenging, as are catalytic routes to biofuels from whole biomass. An alternative strategy is to directly capture carbon before incorporation into lignocellulosic biomass. Autotrophic acetogenic, carboxidotrophic, and methanotrophic bacteria are able to capture carbon as CO, CO2, or CH4, respectively, and reuse that carbon in products that displace their fossil-derived counterparts. Thus, gas fermentation represents a versatile industrial platform for the sustainable production of commodity chemicals and fuels from diverse gas resources derived from industrial processes, coal, biomass, municipal solid waste (MSW), and extracted natural gas.

  15. Deep Conversion of Carbon Monoxide to Hydrogen and Formation of Acetate by the Anaerobic Thermophile Carboxydothermus hydrogenoformans.

    PubMed

    Henstra, Anne M; Stams, Alfons J M

    2011-01-01

    Carboxydothermus hydrogenoformans is a thermophilic strictly anaerobic bacterium that catalyses the water gas shift reaction, the conversion of carbon monoxide with water to molecular hydrogen and carbon dioxide. The thermodynamically favorable growth temperature, compared to existing industrial catalytic processes, makes this organism an interesting alternative for production of cheap hydrogen gas suitable to fuel CO-sensitive fuel cells in a future hydrogen economy, provided sufficiently low levels of CO are reached. Here we study CO conversion and final CO levels in cultures of C. hydrogenoformans grown in batch cultures that were started with a 100% CO gas phase with and without removal of formed CO(2). Final CO levels were 117 ppm without CO(2) removal and below 2 ppm with CO(2) removal. The Gibbs free energy change calculated with measured end concentrations and the detection of acetate suggest that C. hydrogenoformans shifted from a hydrogenogenic to an acetogenic metabolism.

  16. Methods for increasing the production of ethanol from microbial fermentation

    DOEpatents

    Gaddy, James L.; Arora, Dinesh K.; Ko, Ching-Whan; Phillips, John Randall; Basu, Rahul; Wikstrom, Carl V.; Clausen, Edgar C.

    2007-10-23

    A stable continuous method for producing ethanol from the anaerobic bacterial fermentation of a gaseous substrate containing at least one reducing gas involves culturing a fermentation bioreactor anaerobic, acetogenic bacteria in a liquid nutrient medium; supplying the gaseous substrate to the bioreactor; and manipulating the bacteria in the bioreactor by reducing the redox potential, or increasing the NAD(P)H TO NAD(P) ratio, in the fermentation broth after the bacteria achieves a steady state and stable cell concentration in the bioreactor. The free acetic acid concentration in the bioreactor is maintained at less than 5 g/L free acid. This method allows ethanol to be produced in the fermentation broth in the bioreactor at a productivity greater than 10 g/L per day. Both ethanol and acetate are produced in a ratio of ethanol to acetate ranging from 1:1 to 20:1.

  17. Low-Carbon Fuel and Chemical Production by Anaerobic Gas Fermentation.

    PubMed

    Daniell, James; Nagaraju, Shilpa; Burton, Freya; Köpke, Michael; Simpson, Séan Dennis

    World energy demand is expected to increase by up to 40% by 2035. Over this period, the global population is also expected to increase by a billion people. A challenge facing the global community is not only to increase the supply of fuel, but also to minimize fossil carbon emissions to safeguard the environment, at the same time as ensuring that food production and supply is not detrimentally impacted. Gas fermentation is a rapidly maturing technology which allows low carbon fuel and commodity chemical synthesis. Unlike traditional biofuel technologies, gas fermentation avoids the use of sugars, relying instead on gas streams rich in carbon monoxide and/or hydrogen and carbon dioxide as sources of carbon and energy for product synthesis by specialized bacteria collectively known as acetogens. Thus, gas fermentation enables access to a diverse array of novel, large volume, and globally available feedstocks including industrial waste gases and syngas produced, for example, via the gasification of municipal waste and biomass. Through the efforts of academic labs and early stage ventures, process scale-up challenges have been surmounted through the development of specialized bioreactors. Furthermore, tools for the genetic improvement of the acetogenic bacteria have been reported, paving the way for the production of a spectrum of ever-more valuable products via this process. As a result of these developments, interest in gas fermentation among both researchers and legislators has grown significantly in the past 5 years to the point that this approach is now considered amongst the mainstream of emerging technology solutions for near-term low-carbon fuel and chemical synthesis.

  18. An Examination of the Carbon Isotope Effects Associated with Amino Acid Biosynthesis

    NASA Astrophysics Data System (ADS)

    Scott, James H.; O'Brien, Diane M.; Emerson, David; Sun, Henry; McDonald, Gene D.; Salgado, Antonio; Fogel, Marilyn L.

    2006-12-01

    Stable carbon isotope ratios (δ13C) were determined for alanine, proline, phenylalanine, valine, leucine, isoleucine, aspartate (aspartic acid and asparagine), glutamate (glutamic acid and glutamine), lysine, serine, glycine, and threonine from metabolically diverse microorganisms. The microorganisms examined included fermenting bacteria, organotrophic, chemolithotrophic, phototrophic, methylotrophic, methanogenic, acetogenic, acetotrophic, and naturally occurring cryptoendolithic communities from the Dry Valleys of Antarctica. Here we demonstrated that reactions involved in amino acid biosynthesis can be used to distinguish amino acids formed by life from those formed by nonbiological processes. The unique patterns of δ13C imprinted by life on amino acids produced a biological bias. We also showed that, by applying discriminant function analysis to the δ13C value of a pool of amino acids formed by biological activity, it was possible to identify key aspects of intermediary carbon metabolism in the microbial world. In fact, microorganisms examined in this study could be placed within one of three metabolic groups: (1) heterotrophs that grow by oxidizing compounds containing three or more carbon-to-carbon bonds (fermenters and organotrophs), (2) autotrophs that grow by taking up carbon dioxide (chemolitotrophs and phototrophs), and (3) acetoclastic microbes that grow by assimilation of formaldehyde or acetate (methylotrophs, methanogens, acetogens, and acetotrophs). Furthermore, we demonstrated that cryptoendolithic communities from Antarctica grouped most closely with the autotrophs, which indicates that the dominant metabolic pathways in these communities are likely those utilized for CO2 fixation. We propose that this technique can be used to determine the dominant metabolic types in a community and reveal the overall flow of carbon in a complex ecosystem.

  19. Nature's Helpers: Using Microorganisms to Remove Trichloroethene (TCE) from Groundwater

    NASA Astrophysics Data System (ADS)

    Delgado, A. G.; Krajmlanik-Brown, R.; Fajardo-Williams, D.; Halloum, I.

    2015-12-01

    Organic chlorinated solvents, such as perchloroethene (PCE) and trichloroethene (TCE), are toxic pollutants threatening ground water quality worldwide and present at many superfund sites. Bioremediation using microorganisms is a promising, green, efficient, and sustainable approach to remove PCE and TCE contamination from soil and groundwater. Under anaerobic conditions, specialized microorganisms (dechlorinators) can reduce these chlorinated ethenes to ethene, an innocuous product, and gain energy for growth by a process known as reductive dechlorination. Dechlorinators are most often present in the environment and in dechlorinating cultures alongside other microbes such as fermenters, methanogens, and acetogens. Fermenters, methanogens, and acetogens syntrophically provide essential nutrients and growth factors to dechlorinators, most specifically to the only members able to reduce TCE all the way to ethene: Dehalococcoides; unfortunately, they also compete with dechlorinators for electron donors. My laboratory devises reductive chlorination platforms to study competition and syntrophy among Dehalococcoides, and other microbes to optimize remediation reactions and transport in the subsurface. We look at competing processes present as part of the natural soil chemistry and microbiology and address these challenges through a combination of enrichment techniques, molecular microbial ecology (deep sequencing), water chemistry, and electron balances. We have applied knowledge gathered in my laboratory to: 1) enrich microbial dechlorinating cultures capable of some of the fastest rates of TCE to ethene dechlorination ever reported, and 2) successfully design and operate three different continuous dechlorinating reactor types. We attribute our successful reactor operations to our multidisciplinary approach which links microbiology and engineering. Our reactors produce robust dechlorinating cultures used for in-situ bioaugmentation of PCE and TCE at contaminated sites

  20. Hydrogen isotopic differences between C3 and C4 land plant lipids: consequences of compartmentation in C4 photosynthetic chemistry and C3 photorespiration.

    PubMed

    Zhou, Youping; Grice, Kliti; Stuart-Williams, Hilary; Hocart, Charles H; Gessler, Arthur; Farquhar, Graham D

    2016-12-01

    The (2) H/(1) H ratio of carbon-bound H in biolipids holds potential for probing plant lipid biosynthesis and metabolism. The biochemical mechanism underlying the isotopic differences between lipids from C3 and C4 plants is still poorly understood. GC-pyrolysis-IRMS (gas chromatography-pyrolysis-isotope ratio mass spectrometry) measurement of the (2) H/(1) H ratio of leaf lipids from controlled and field grown plants indicates that the biochemical isotopic fractionation (ε(2) Hlipid_biochem ) differed between C3 and C4 plants in a pathway-dependent manner: ε(2) HC4  > ε(2) HC3 for the acetogenic pathway, ε(2) HC4  < ε(2) HC3 for the mevalonic acid pathway and the 1-deoxy-D-xylulose 5-phosphate pathway across all species examined. It is proposed that compartmentation of photosynthetic CO2 fixation into C4 mesophyll (M) and bundle sheath (BS) cells and suppression of photorespiration in C4 M and BS cells both result in C4 M chloroplastic pyruvate - the precursor for acetogenic pathway - being more depleted in (2) H relative to pyruvate in C3 cells. In addition, compartmentation in C4 plants also results in (i) the transferable H of NADPH being enriched in (2) H in C4 M chloroplasts compared with that in C3 chloroplasts for the 1-deoxy-D-xylulose 5-phosphate pathway pathway and (ii) pyruvate relatively (2) H-enriched being used for the mevalonic acid pathway in the cytosol of BS cells in comparison with that in C3 cells.

  1. Methanogenic food web in the gut contents of methane-emitting earthworm Eudrilus eugeniae from Brazil

    PubMed Central

    Schulz, Kristin; Hunger, Sindy; Brown, George G; Tsai, Siu M; Cerri, Carlos C; Conrad, Ralf; Drake, Harold L

    2015-01-01

    The anoxic saccharide-rich conditions of the earthworm gut provide an ideal transient habitat for ingested microbes capable of anaerobiosis. It was recently discovered that the earthworm Eudrilus eugeniae from Brazil can emit methane (CH4) and that ingested methanogens might be associated with this emission. The objective of this study was to resolve trophic interactions of bacteria and methanogens in the methanogenic food web in the gut contents of E. eugeniae. RNA-based stable isotope probing of bacterial 16S rRNA as well as mcrA and mrtA (the alpha subunit of methyl-CoM reductase and its isoenzyme, respectively) of methanogens was performed with [13C]-glucose as a model saccharide in the gut contents. Concomitant fermentations were augmented by the rapid consumption of glucose, yielding numerous products, including molecular hydrogen (H2), carbon dioxide (CO2), formate, acetate, ethanol, lactate, succinate and propionate. Aeromonadaceae-affiliated facultative aerobes, and obligate anaerobes affiliated to Lachnospiraceae, Veillonellaceae and Ruminococcaceae were associated with the diverse fermentations. Methanogenesis was ongoing during incubations, and 13C-labeling of CH4 verified that supplemental [13C]-glucose derived carbon was dissimilated to CH4. Hydrogenotrophic methanogens affiliated with Methanobacteriaceae and Methanoregulaceae were linked to methanogenesis, and acetogens related to Peptostreptoccocaceae were likewise found to be participants in the methanogenic food web. H2 rather than acetate stimulated methanogenesis in the methanogenic gut content enrichments, and acetogens appeared to dissimilate supplemental H2 to acetate in methanogenic enrichments. These findings provide insight on the processes and associated taxa potentially linked to methanogenesis and the turnover of organic carbon in the alimentary canal of methane-emitting E. eugeniae. PMID:25615437

  2. Revisiting "You are what you eat, +1‰": Bacterial Trophic Structure and the Sedimentary Record

    NASA Astrophysics Data System (ADS)

    Pearson, A.; Tang, T.; Mohr, W.; Sattin, S.

    2015-12-01

    "You are what you eat, +1‰" is a central principle of carbon stable isotope (δ13C) distributions and is widely applied to understand the structure and ordering of macrobiotic ecosystems. Although based on observations from multicellular organisms that are able to ingest "food", this idea also has been applied to Precambrian ecosystems dominated by unicellular, microbial life, with the suggestion that such systems could sustain ordered trophic structures observable in their isotopes. However, using a new approach to community profiling known as protein stable isotope fingerprinting (P-SIF), we find that the carbon isotope ratios of whole proteins separated from environmental samples show differences only between metabolically-distinct autotrophs; heterotrophs are not 13C-enriched. In parallel, a survey of the relative distribution of 13C between biochemical classes - specifically acetogenic lipids, isoprenoid lipids, amino acids, and nucleic acids/sugars - across a variety of bacterial species appears to be a function of the main carbon metabolite, not an indicator of heterotrophy vs. autotrophy. Indeed, autotrophy, heterotrophy, and mixotrophy all are indistinguishable when the primary food source is fresh photosynthate, i.e., sugar. Significant assimilation of acetate is diagnosed by acetogenic lipids that are relatively 13C-enriched vs. isoprenoid lipids. Mixed-substrate heterotrophy, in contrast, satisfies the classic "…+1‰" rule for bulk biomass, yet simultaneously it collapses the biochemical patterns of 13C almost completely. Together these observations point to a paradigm shift for understanding the preservation of bulk organic and lipid δ13C signatures in the rock record, suggesting that patterns of δ13Corg must primarily reflect changing carbon inputs, not the extent or intensity of heterotrophy.

  3. Genes for selenium dependent and independent formate dehydrogenase in the gut microbial communities of three lower, wood-feeding termites and a wood-feeding roach.

    PubMed

    Zhang, Xinning; Matson, Eric G; Leadbetter, Jared R

    2011-02-01

    The bacterial Wood-Ljungdahl pathway for CO(2)-reductive acetogenesis is important for the nutritional mutualism occurring between wood-feeding insects and their hindgut microbiota. A key step in this pathway is the reduction of CO(2) to formate, catalysed by the enzyme formate dehydrogenase (FDH). Putative selenocysteine- (Sec) and cysteine- (Cys) containing paralogues of hydrogenase-linked FDH (FDH(H)) have been identified in the termite gut acetogenic spirochete, Treponema primitia, but knowledge of their relevance in the termite gut environment remains limited. In this study, we designed degenerate PCR primers for FDH(H) genes (fdhF) and assessed fdhF diversity in insect gut bacterial isolates and the gut microbial communities of termites and cockroaches. The insects examined herein represent three wood-feeding termite families, Termopsidae, Kalotermitidae and Rhinotermitidae (phylogenetically 'lower' termite taxa); the wood-feeding roach family Cryptocercidae (the sister taxon to termites); and the omnivorous roach family Blattidae. Sec and Cys FDH(H) variants were identified in every wood-feeding insect but not the omnivorous roach. Of 68 novel alleles obtained from inventories, 66 affiliated phylogenetically with enzymes from T. primitia. These formed two subclades (37 and 29 phylotypes) almost completely comprised of Sec-containing and Cys-containing enzymes respectively. A gut cDNA inventory showed transcription of both variants in the termite Zootermopsis nevadensis (family Termopsidae). The gene patterns suggest that FDH(H) enzymes are important for the CO(2)-reductive metabolism of uncultured acetogenic treponemes and imply that the availability of selenium, a trace element, shaped microbial gene content in the last common ancestor of dictyopteran, wood-feeding insects, and continues to shape it to this day.

  4. Establishment and Development of Ruminal Hydrogenotrophs in Methanogen-Free Lambs▿

    PubMed Central

    Fonty, Gérard; Joblin, Keith; Chavarot, Michel; Roux, Remy; Naylor, Graham; Michallon, Fabien

    2007-01-01

    The aim of this work was to determine whether reductive acetogenesis can provide an alternative to methanogenesis in the rumen. Gnotobiotic lambs were inoculated with a functional rumen microbiota lacking methanogens and reared to maturity on a fibrous diet. Lambs with a methanogen-free rumen grew well, and the feed intake and ruminal volatile fatty acid concentrations for lambs lacking ruminal methanogens were lower but not markedly dissimilar from those for conventional lambs reared on the same diet. A high population density (107 to 108 cells g−1) of ruminal acetogens slowly developed in methanogen-free lambs. Sulfate- and fumarate-reducing bacteria were present, but their population densities were highly variable. In methanogen-free lambs, the hydrogen capture from fermentation was low (28 to 46%) in comparison with that in lambs containing ruminal methanogens (>90%). Reductive acetogenesis was not a significant part of ruminal fermentation in conventional lambs but contributed 21 to 25% to the fermentation in methanogen-free meroxenic animals. Ruminal H2 utilization was lower in lambs lacking ruminal methanogens, but when a methanogen-free lamb was inoculated with a methanogen, the ruminal H2 utilization was similar to that in conventional lambs. H2 utilization in lambs containing a normal ruminal microflora was age dependent and increased with the animal age. The animal age effect was less marked in lambs lacking ruminal methanogens. Addition of fumarate to rumen contents from methanogen-free lambs increased H2 utilization. These findings provide the first evidence from animal studies that reductive acetogens can sustain a functional rumen and replace methanogens as a sink for H2 in the rumen. PMID:17675444

  5. Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity

    PubMed Central

    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

  6. Effect of tungstate on acetate and ethanol production by the electrosynthetic bacterium Sporomusa ovata.

    PubMed

    Ammam, Fariza; Tremblay, Pier-Luc; Lizak, Dawid M; Zhang, Tian

    2016-01-01

    Microbial electrosynthesis (MES) and gas fermentation are bioenergy technologies in which a microbial catalyst reduces CO2 into organic carbon molecules with electrons from the cathode of a bioelectrochemical system or from gases such as H2. The acetogen Sporomusa ovata has the capacity of reducing CO2 into commodity chemicals by both gas fermentation and MES. Acetate is often the only product generated by S. ovata during autotrophic growth. In this study, trace elements in S. ovata growth medium were optimized to improve MES and gas fermentation productivity. Augmenting tungstate concentration resulted in a 2.9-fold increase in ethanol production by S. ovata during H2:CO2-dependent growth. It also promoted electrosynthesis of ethanol in a S. ovata-driven MES reactor and increased acetate production 4.4-fold compared to unmodified medium. Furthermore, fatty acids propionate and butyrate were successfully converted to their corresponding alcohols 1-propanol and 1-butanol by S. ovata during gas fermentation. Increasing tungstate concentration enhanced conversion efficiency for both propionate and butyrate. Gene expression analysis suggested that tungsten-containing aldehyde ferredoxin oxidoreductases (AORs) and a tungsten-containing formate dehydrogenase (FDH) were involved in the improved biosynthesis of acetate, ethanol, 1-propanol, and 1-butanol. AORs and FDH contribute to the fatty acids re-assimilation pathway and the Wood-Ljungdahl pathway, respectively. This study presented here shows that optimization of microbial catalyst growth medium can improve productivity and lead to the biosynthesis of different products by gas fermentation and MES. It also provides insights on the metabolism of biofuels production in acetogens and demonstrates that S. ovata has an important untapped metabolic potential for the production of other chemicals than acetate via CO2-converting bioprocesses including MES.

  7. Biochemical fossils of the ancient transition from geoenergetics to bioenergetics in prokaryotic one carbon compound metabolism.

    PubMed

    Sousa, Filipa L; Martin, William F

    2014-07-01

    The deep dichotomy of archaea and bacteria is evident in many basic traits including ribosomal protein composition, membrane lipid synthesis, cell wall constituents, and flagellar composition. Here we explore that deep dichotomy further by examining the distribution of genes for the synthesis of the central carriers of one carbon units, tetrahydrofolate (H4F) and tetrahydromethanopterin (H4MPT), in bacteria and archaea. The enzymes underlying those distinct biosynthetic routes are broadly unrelated across the bacterial-archaeal divide, indicating that the corresponding pathways arose independently. That deep divergence in one carbon metabolism is mirrored in the structurally unrelated enzymes and different organic cofactors that methanogens (archaea) and acetogens (bacteria) use to perform methyl synthesis in their H4F- and H4MPT-dependent versions, respectively, of the acetyl-CoA pathway. By contrast, acetyl synthesis in the acetyl-CoA pathway - from a methyl group, CO2 and reduced ferredoxin - is simpler, uniform and conserved across acetogens and methanogens, and involves only transition metals as catalysts. The data suggest that the acetyl-CoA pathway, while being the most ancient of known CO2 assimilation pathways, reflects two phases in early evolution: an ancient phase in a geochemically confined and non-free-living universal common ancestor, in which acetyl thioester synthesis proceeded spontaneously with the help of geochemically supplied methyl groups, and a later phase that reflects the primordial divergence of the bacterial and archaeal stem groups, which independently invented genetically-encoded means to synthesize methyl groups via enzymatic reactions. This article is part of a Special Issue entitled: 18th European Bioenergetic Conference.

  8. Guided cobalamin biosynthesis supports Dehalococcoides mccartyi reductive dechlorination activity.

    PubMed

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

    2013-04-19

    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

  9. Acetate Synthesis from H2 plus CO2 by Termite Gut Microbes †

    PubMed Central

    Breznak, John A.; Switzer, Jodi M.

    1986-01-01

    Gut microbiota from Reticulitermes flavipes termites catalyzed an H2-dependent total synthesis of acetate from CO2. Rates of H2-CO2 acetogenesis in vitro were 1.11 ± 0.37 μmol of acetate g (fresh weight)−1 h−1 (equivalent to 4.44 ± 1.47 nmol termite−1 h−1) and could account for approximately 1/3 of all the acetate produced during the hindgut fermentation. Formate was also produced from H2 + CO2, as were small amounts of propionate, butyrate, and lactate-succinate. However, H2-CO2 formicogenesis seemed largely unrelated to acetogenesis and was believed not to be a significant reaction in situ. Little or no CH4 was formed from H2 + CO2 or from acetate. H2-CO2 acetogenesis was inhibited by O2, KCN, CHCl3, and iodopropane and could be abolished by prefeeding R. flavipes with antibacterial drugs. By contrast, prefeeding R. flavipes with starch resulted in almost complete defaunation but had little effect on H2-CO2 acetogenesis, suggesting that bacteria were the acetogenic agents in the gut. H2-CO2 acetogenesis was also observed with gut microbiota from Prorhinotermes simplex, Zootermopsis angusticollis, Nasutitermes costalis, and N. nigriceps; from the wood-eating cockroach Cryptocercus punctulatus; and from the American cockroach Periplaneta americana. Pure cultures of H2-CO2-acetogenic bacteria were isolated from N. nigriceps, and a preliminary account of their morphological and physiological properties is presented. Results indicate that in termites, CO2 reduction to acetate, rather than to CH4, represents the main electron sink reaction of the hindgut fermentation and can provide the insects with a significant fraction (ca. 1/3) of their principal oxidizable energy source, acetate. PMID:16347157

  10. Methanogenic food web in the gut contents of methane-emitting earthworm Eudrilus eugeniae from Brazil.

    PubMed

    Schulz, Kristin; Hunger, Sindy; Brown, George G; Tsai, Siu M; Cerri, Carlos C; Conrad, Ralf; Drake, Harold L

    2015-08-01

    The anoxic saccharide-rich conditions of the earthworm gut provide an ideal transient habitat for ingested microbes capable of anaerobiosis. It was recently discovered that the earthworm Eudrilus eugeniae from Brazil can emit methane (CH4) and that ingested methanogens might be associated with this emission. The objective of this study was to resolve trophic interactions of bacteria and methanogens in the methanogenic food web in the gut contents of E. eugeniae. RNA-based stable isotope probing of bacterial 16S rRNA as well as mcrA and mrtA (the alpha subunit of methyl-CoM reductase and its isoenzyme, respectively) of methanogens was performed with [(13)C]-glucose as a model saccharide in the gut contents. Concomitant fermentations were augmented by the rapid consumption of glucose, yielding numerous products, including molecular hydrogen (H2), carbon dioxide (CO2), formate, acetate, ethanol, lactate, succinate and propionate. Aeromonadaceae-affiliated facultative aerobes, and obligate anaerobes affiliated to Lachnospiraceae, Veillonellaceae and Ruminococcaceae were associated with the diverse fermentations. Methanogenesis was ongoing during incubations, and (13)C-labeling of CH4 verified that supplemental [(13)C]-glucose derived carbon was dissimilated to CH4. Hydrogenotrophic methanogens affiliated with Methanobacteriaceae and Methanoregulaceae were linked to methanogenesis, and acetogens related to Peptostreptoccocaceae were likewise found to be participants in the methanogenic food web. H2 rather than acetate stimulated methanogenesis in the methanogenic gut content enrichments, and acetogens appeared to dissimilate supplemental H2 to acetate in methanogenic enrichments. These findings provide insight on the processes and associated taxa potentially linked to methanogenesis and the turnover of organic carbon in the alimentary canal of methane-emitting E. eugeniae.

  11. Impact of formate on the growth and productivity of Clostridium ljungdahlii PETC and Clostridium carboxidivorans P7 grown on syngas.

    PubMed

    Ramió-Pujol, Sara; Ganigué, Ramon; Bañeras, Lluís; Colprim, Jesús

    2014-12-01

    The current energy model based on fossil fuels is coming to an end due to the increase in global energy demand. Biofuels such as ethanol and butanol can be produced through the syngas fermentation by acetogenic bacteria. The present work hypothesizes that formate addition would positively impact kinetic parameters for growth and alcohol production in Clostridium ljungdahlii PETC and Clostridium carboxidivorans P7 by diminishing the need for reducing equivalents. Fermentation experiments were conducted using completely anaerobic batch cultures at different pH values and formate concentrations. PETC cultures were more tolerant to formate concentrations than P7, specially at pH 5.0 and 6.0. Complete growth inhibition of PETC occurred at sodium formate concentrations of 30.0 mM; however, no differences in growth rates were observed at pH 7.0 for the two strains. Incubation at formate concentrations lower than 2.0 mM resulted in increased growth rates for both strains. The most recognizable effects of formate addition on the fermentation products were the increase in the total carbon fixed into acids and alcohols at pH 5.0 and pH 6.0, as well as, a higher ethanol to total products ratio at pH 7.0. Taken all together, these results show the ability of acetogens to use formate diminishing the energy demand for growth, and enhancing strain productivity. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

  12. A thermophilic, hydrogenogenic and carboxydotrophic bacterium, Calderihabitans maritimus gen. nov., sp. nov., from a marine sediment core of an undersea caldera.

    PubMed

    Yoneda, Yasuko; Yoshida, Takashi; Yasuda, Hisato; Imada, Chiaki; Sako, Yoshihiko

    2013-10-01

    A hydrogenogenic, carboxydotrophic marine bacterium, strain KKC1(T), was isolated from a sediment core sample taken from a submerged marine caldera. Cells were non-motile, Gram-stain-negative, 1.0-3.0 µm straight rods, often observed with round endospores. Strain KKC1(T) grew at 55-68 °C, pH 5.2-9.2 and 0.8-14 % (w/v) salinity. Optimum growth occurred at 65 °C, pH 7.0-7.5 and 2.46 % salinity with a doubling time of 3.7 h. The isolate grew chemolithotrophically, producing H2 from carbon monoxide (CO) oxidation with reduction of various electron acceptors, e.g. sulfite, thiosulfate, fumarate, ferric iron and AQDS (9,10-anthraquinone 2,6-disulfonate). KKC1(T) grew heterotrophically on pyruvate, lactate, fumarate, glucose, fructose and mannose with thiosulfate as an electron acceptor. When grown mixotrophically on CO and pyruvate, C16 : 0 constituted almost half of the total cellular fatty acids. The DNA G+C content was 50.6 mol%. The 16S rRNA gene sequence of KKC1(T) was most closely related to those of members of the genus Moorella with similarity ranging from 91 to 89 %. Based on physiological and phylogenetic novelty, we propose the isolate as a representative of a new genus and novel species with the name Calderihabitans maritimus gen. nov., sp. nov.; the type strain of the type species is KKC1(T) ( = DSM 26464(T) = NBRC 109353(T)).

  13. Reactive Oxygen Species on the Early Earth and Survival of Bacteria

    NASA Technical Reports Server (NTRS)

    Balk, Melikea; Mason, Paul; Stams, Alfons J. M.; Smidt, Hauke; Freund, Friedemann; Rothschild, Lynn

    2011-01-01

    An oxygen-rich atmosphere appears to have been a prerequisite for complex, multicellular life to evolve on Earth and possibly elsewhere in the Universe. However it remains unclear how free oxygen first became available on the early Earth. A potentially important, and as yet poorly constrained pathway, is the production of oxygen through the weathering of rocks and release into the near-surface environment. Reactive Oxygen Species (ROS), as precursors to molecular oxygen, are a key step in this process, and may have had a decisive impact on the evolution of life, present and past. ROS are generated from minerals in igneous rocks during hydrolysis of peroxy defects, which consist of pairs of oxygen anions oxidized to the valence state -1 and during (bio) transformations of iron sulphide minerals. ROS are produced and consumed by intracellular and extracellular reactions of Fe, Mn, C, N, and S species. We propose that, despite an overall reducing or neutral oxidation state of the macroenvironment and the absence of free O2 in the atmosphere, organisms on the early Earth had to cope with ROS in their microenvironments. They were thus under evolutionary pressure to develop enzymatic and other defences against the potentially dangerous, even lethal effects of oxygen and its derived ROS. Conversely it appears that microorganisms learned to take advantage of the enormous reactive potential and energy gain provided by nascent oxygen. We investigate how oxygen might be released through weathering. We test microorganisms in contact with rock surfaces and iron sulphides. We model bacteria such as Deionococcus radiodurans and Desulfotomaculum, Moorella and Bacillus species for their ability to grow or survive in the presence of ROS. We examine how early Life might have adapted to oxygen.

  14. The Rnf Complex of Clostridium ljungdahlii Is a Proton-Translocating Ferredoxin:NAD(+) Oxidoreductase Essential for Autotrophic Growth

    SciTech Connect

    Tremblay, PL; Zhang, T; Dar, SA; Leang, C; Lovley, DR

    2012-12-26

    It has been predicted that the Rnf complex of Clostridium ljungdahlii is a proton-translocating ferredoxin: NAD(+) oxidoreductase which contributes to ATP synthesis by an H+-translocating ATPase under both autotrophic and heterotrophic growth conditions. The recent development of methods for genetic manipulation of C. ljungdahlii made it possible to evaluate the possible role of the Rnf complex in energy conservation. Disruption of the C. ljungdahlii rnf operon inhibited autotrophic growth. ATP synthesis, proton gradient, membrane potential, and proton motive force collapsed in the Rnf-deficient mutant with H-2 as the electron source and CO2 as the electron acceptor. Heterotrophic growth was hindered in the absence of a functional Rnf complex, as ATP synthesis, proton gradient, and proton motive force were significantly reduced with fructose as the electron donor. Growth of the Rnf-deficient mutant was also inhibited when no source of fixed nitrogen was provided. These results demonstrate that the Rnf complex of C. ljungdahlii is responsible for translocation of protons across the membrane to elicit energy conservation during acetogenesis and is a multifunctional device also implicated in nitrogen fixation. IMPORTANCE Mechanisms for energy conservation in the acetogen Clostridium ljungdahlii are of interest because of its potential value as a chassis for the production of biocommodities with novel electron donors such as carbon monoxide, syngas, and electrons derived from electrodes. Characterizing the components implicated in the chemiosmotic ATP synthesis during acetogenesis by C. ljungdahlii is a prerequisite for the development of highly productive strains. The Rnf complex has been considered the prime candidate to be the pump responsible for the formation of an ion gradient coupled with ATP synthesis in multiple acetogens. However, experimental evidence for a proton-pumping Rnf complex has been lacking. This study establishes the C. ljungdahlii Rnf complex as

  15. The Rnf complex of Clostridium ljungdahlii is a proton-translocating ferredoxin:NAD+ oxidoreductase essential for autotrophic growth.

    PubMed

    Tremblay, Pier-Luc; Zhang, Tian; Dar, Shabir A; Leang, Ching; Lovley, Derek R

    2012-12-26

    It has been predicted that the Rnf complex of Clostridium ljungdahlii is a proton-translocating ferredoxin:NAD(+) oxidoreductase which contributes to ATP synthesis by an H(+)-translocating ATPase under both autotrophic and heterotrophic growth conditions. The recent development of methods for genetic manipulation of C. ljungdahlii made it possible to evaluate the possible role of the Rnf complex in energy conservation. Disruption of the C. ljungdahlii rnf operon inhibited autotrophic growth. ATP synthesis, proton gradient, membrane potential, and proton motive force collapsed in the Rnf-deficient mutant with H(2) as the electron source and CO(2) as the electron acceptor. Heterotrophic growth was hindered in the absence of a functional Rnf complex, as ATP synthesis, proton gradient, and proton motive force were significantly reduced with fructose as the electron donor. Growth of the Rnf-deficient mutant was also inhibited when no source of fixed nitrogen was provided. These results demonstrate that the Rnf complex of C. ljungdahlii is responsible for translocation of protons across the membrane to elicit energy conservation during acetogenesis and is a multifunctional device also implicated in nitrogen fixation. Mechanisms for energy conservation in the acetogen Clostridium ljungdahlii are of interest because of its potential value as a chassis for the production of biocommodities with novel electron donors such as carbon monoxide, syngas, and electrons derived from electrodes. Characterizing the components implicated in the chemiosmotic ATP synthesis during acetogenesis by C. ljungdahlii is a prerequisite for the development of highly productive strains. The Rnf complex has been considered the prime candidate to be the pump responsible for the formation of an ion gradient coupled with ATP synthesis in multiple acetogens. However, experimental evidence for a proton-pumping Rnf complex has been lacking. This study establishes the C. ljungdahlii Rnf complex as a

  16. Recent insights into intramolecular 13C isotope composition of biomolecules

    NASA Astrophysics Data System (ADS)

    Gilbert, A.; Yamada, K.; Julien, M.; Yoshida, N.; Remaud, G.; Robins, R.

    2016-12-01

    In 1961 Abelson & Hoering shown that the intramolecular 13C distribution in amino acids was not homogeneous, namely the carboxylic acid positions were 13C-enriched compared with the mean of the remaining C-atoms in the molecule [1]. Nearly 20 years later, Monson & Hayes were able to demonstrate that even and odd positions in acetogenic fatty acids also showed non-statistical 13C isotope distributions, and that the pattern varied depending on the organism [2]. It took a further decade for the intramolecular 13C distribution in the key metabolite, glucose, to be defined [3]. Although informative, much of this work was incomplete, a number of positions having to be deduced by difference. This limitation arose mainly due to the lack of techniques enabling the separation and quantification of 13C isotopomers of the target molecule. In the past decade, quantitative 13C NMR has been developed for the determination of the intramolecular isotope composition of a given molecule with a precision of 1‰ or better [4]. This breakthrough has made possible a comprehensive view of the determinants governing intramolecular isotope composition of biological molecules. In particular, it can be shown that intramolecular pattern in sugars is influenced by the C-assimilation pathway and by post-photosynthetic fractionation associated with carbohydrate metabolism [5]. In addition, analysis by NMR of the alkyl chain of acetogenic lipids (fatty acids, n-alkanes) shows an alternation between odd and even C-atom positions, as observed by Monson& Hayes [2], throughout the molecule [6]. Overall, it is becoming apparent that this pattern is influenced by two principal metabolic factors: (i) the 13C pattern extant in the starting compounds; (ii) isotope fractionation associated with the enzymes involved in the biosynthetic pathway. On the whole, the determination of intramolecular isotope patterns in biomolecules allows better insights into the conditions and pathways by which they are formed

  17. Towards a palaeosalinity proxy: hydrogen isotopic fractionation between source water and lipids produced via different biosynthetic pathways in haptophyte algae

    NASA Astrophysics Data System (ADS)

    Chivall, David; M'Boule, Daniela; Heinzelmann, Sandra M.; Kasper, Sebastian; Sinke-Schoen, Daniëlle; Sininnghe-Damsté, Jaap S.; Schouten, Stefan; van der Meer, Marcel T. J.

    2014-05-01

    Palaeosalinity is one of the most important oceanographic parameters that cannot currently be quantified with reasonable accuracy from sedimentary records. Hydrogen isotopic fractionation between water and alkenones is dependent, amongst other factors, upon the salinity in which alkenone-producing haptophyte algae grow and is represented by the fractionation factor, α, increasing with salinity.1 As such, the hydrogen isotopic composition of alkenones is emerging as a palaeosalinity proxy. Understanding the mechanism behind the sensitivity of fractionation to salinity is important for the correct application of the proxy, however this mechanism is currently unknown. Here we present hydrogen isotopic compositions of lipids produced via different biosynthetic pathways from batch cultures of Emiliania huxleyi CCMP 1516 and Isochrysis galbana CCMP 1323 grown over a range of salinities and discuss the possible sources of the sensitivity of hydrogen isotope fractionation to salinity. α for C37 alkenones (produced via an unknown biosynthetic pathway but assumed to be acetogenic; e.g.2) and that for C14:0, C16:0, and C18:1 fatty acids (acetogenic) from exponential growth phase I. galbana show a similar sensitivity to salinity, increasing at 0.0013-0.0019 per salinity unit (S-1). Meanwhile, in exponential growth phase E. huxleyi, α for C37 alkenones and α for brassicasterol (mevalonate pathway) increase at 0.0015-0.0022 S-1, but α for phytol (methylerythritol pathway) shows no significant relationship with salinity. These results suggest that fractionation is sensitive to salinity for lipids formed both in the chloroplast and cytosol. They also suggest that the sensitivity may either originate in glyceralde-3-phosphate or pyruvate but is then lost through hydrogen exchange with cell water during sugar rearrangements in the methylerythritol pathway or sensitivity originates with the production and consumption of acetate. References Schouten, S., Ossebaar, J., Schreiber

  18. Sequential Mixed Cultures: From Syngas to Malic Acid

    PubMed Central

    Oswald, Florian; Dörsam, Stefan; Veith, Nicolas; Zwick, Michaela; Neumann, Anke; Ochsenreither, Katrin; Syldatk, Christoph

    2016-01-01

    Synthesis gas (syngas) fermentation using acetogenic bacteria is an approach for production of bulk chemicals like acetate, ethanol, butanol, or 2,3-butandiol avoiding the fuel vs. food debate by using carbon monoxide, carbon dioxide, and hydrogen from gasification of biomass or industrial waste gases. Suffering from energetic limitations, yields of C4-molecules produced by syngas fermentation are quite low compared with ABE fermentation using sugars as a substrate. On the other hand, fungal production of malic acid has high yields of product per gram metabolized substrate but is currently limited to sugar containing substrates. In this study, it was possible to show that Aspergilus oryzae is able to produce malic acid using acetate as sole carbon source which is a main product of acetogenic syngas fermentation. Bioreactor cultivations were conducted in 2.5 L stirred tank reactors. During the syngas fermentation part of the sequential mixed culture, Clostridium ljungdahlii was grown in modified Tanner medium and sparged with 20 mL/min of artificial syngas mimicking a composition of clean syngas from entrained bed gasification of straw (32.5 vol-% CO, 32.5 vol-% H2, 16 vol-% CO2, and 19 vol-% N2) using a microsparger. Syngas consumption was monitored via automated gas chromatographic measurement of the off-gas. For the fungal fermentation part gas sparging was switched to 0.6 L/min of air and a standard sparger. Ammonia content of medium for syngas fermentation was reduced to 0.33 g/L NH4Cl to meet the requirements for fungal production of dicarboxylic acids. Malic acid production performance of A. oryzae in organic acid production medium and syngas medium with acetate as sole carbon source was verified and gave YP∕S values of 0.28 g/g and 0.37 g/g respectively. Growth and acetate formation of C. ljungdahlii during syngas fermentation were not affected by the reduced ammonia content and 66 % of the consumed syngas was converted to acetate. The overall conversion

  19. Sequential Mixed Cultures: From Syngas to Malic Acid.

    PubMed

    Oswald, Florian; Dörsam, Stefan; Veith, Nicolas; Zwick, Michaela; Neumann, Anke; Ochsenreither, Katrin; Syldatk, Christoph

    2016-01-01

    Synthesis gas (syngas) fermentation using acetogenic bacteria is an approach for production of bulk chemicals like acetate, ethanol, butanol, or 2,3-butandiol avoiding the fuel vs. food debate by using carbon monoxide, carbon dioxide, and hydrogen from gasification of biomass or industrial waste gases. Suffering from energetic limitations, yields of C4-molecules produced by syngas fermentation are quite low compared with ABE fermentation using sugars as a substrate. On the other hand, fungal production of malic acid has high yields of product per gram metabolized substrate but is currently limited to sugar containing substrates. In this study, it was possible to show that Aspergilus oryzae is able to produce malic acid using acetate as sole carbon source which is a main product of acetogenic syngas fermentation. Bioreactor cultivations were conducted in 2.5 L stirred tank reactors. During the syngas fermentation part of the sequential mixed culture, Clostridium ljungdahlii was grown in modified Tanner medium and sparged with 20 mL/min of artificial syngas mimicking a composition of clean syngas from entrained bed gasification of straw (32.5 vol-% CO, 32.5 vol-% H2, 16 vol-% CO2, and 19 vol-% N2) using a microsparger. Syngas consumption was monitored via automated gas chromatographic measurement of the off-gas. For the fungal fermentation part gas sparging was switched to 0.6 L/min of air and a standard sparger. Ammonia content of medium for syngas fermentation was reduced to 0.33 g/L NH4Cl to meet the requirements for fungal production of dicarboxylic acids. Malic acid production performance of A. oryzae in organic acid production medium and syngas medium with acetate as sole carbon source was verified and gave YP∕S values of 0.28 g/g and 0.37 g/g respectively. Growth and acetate formation of C. ljungdahlii during syngas fermentation were not affected by the reduced ammonia content and 66 % of the consumed syngas was converted to acetate. The overall conversion

  20. Low Fermentation pH Is a Trigger to Alcohol Production, but a Killer to Chain Elongation.

    PubMed

    Ganigué, Ramon; Sánchez-Paredes, Patricia; Bañeras, Lluis; Colprim, Jesús

    2016-01-01

    Gasification of organic wastes coupled to syngas fermentation allows the recovery of carbon in the form of commodity chemicals, such as carboxylates and biofuels. Acetogenic bacteria ferment syngas to mainly two-carbon compounds, although a few strains can also synthesize four-, and six-carbon molecules. In general, longer carbon chain products have a higher biotechnological (and commercial) value due to their higher energy content and their lower water solubility. However, de-novo synthesis of medium-chain products from syngas is quite uncommon in acetogenic bacteria. An alternative to de-novo synthesis is bioproduction of short-chain products (C2 and C4), and their subsequent elongation to C4, C6, or C8 through reversed β-oxidation metabolism. This two-step synergistic approach has been successfully applied for the production of up to C8 compounds, although the accumulation of alcohols in these mixed cultures remained below detection limits. The present work investigates the production of higher alcohols from syngas by open mixed cultures (OMC). A syngas-fermenting community was enriched from sludge of an anaerobic digester for a period of 109 days in a lab-scale reactor. At the end of this period, stable production of ethanol and butanol was obtained. C6 compounds were only transiently produced at the beginning of the enrichment phase, during which Clostridium kluyveri, a bacterium able to carry out carbon chain elongation, was detected in the community. Further experiments showed pH as a critical parameter to maintain chain elongation activity in the co-culture. Production of C6 compounds was recovered by preventing fermentation pH to decrease below pH 4.5-5. Finally, experiments showed maximal production of C6 compounds (0.8 g/L) and alcohols (1.7 g/L of ethanol, 1.1 g/L of butanol, and 0.6 g/L of hexanol) at pH 4.8. In conclusion, low fermentation pH is critical for the production of alcohols, although detrimental to C. kluyveri. Fine control of fermentation

  1. Low Fermentation pH Is a Trigger to Alcohol Production, but a Killer to Chain Elongation

    PubMed Central

    Ganigué, Ramon; Sánchez-Paredes, Patricia; Bañeras, Lluis; Colprim, Jesús

    2016-01-01

    Gasification of organic wastes coupled to syngas fermentation allows the recovery of carbon in the form of commodity chemicals, such as carboxylates and biofuels. Acetogenic bacteria ferment syngas to mainly two-carbon compounds, although a few strains can also synthesize four-, and six-carbon molecules. In general, longer carbon chain products have a higher biotechnological (and commercial) value due to their higher energy content and their lower water solubility. However, de-novo synthesis of medium-chain products from syngas is quite uncommon in acetogenic bacteria. An alternative to de-novo synthesis is bioproduction of short-chain products (C2 and C4), and their subsequent elongation to C4, C6, or C8 through reversed β-oxidation metabolism. This two-step synergistic approach has been successfully applied for the production of up to C8 compounds, although the accumulation of alcohols in these mixed cultures remained below detection limits. The present work investigates the production of higher alcohols from syngas by open mixed cultures (OMC). A syngas-fermenting community was enriched from sludge of an anaerobic digester for a period of 109 days in a lab-scale reactor. At the end of this period, stable production of ethanol and butanol was obtained. C6 compounds were only transiently produced at the beginning of the enrichment phase, during which Clostridium kluyveri, a bacterium able to carry out carbon chain elongation, was detected in the community. Further experiments showed pH as a critical parameter to maintain chain elongation activity in the co-culture. Production of C6 compounds was recovered by preventing fermentation pH to decrease below pH 4.5–5. Finally, experiments showed maximal production of C6 compounds (0.8 g/L) and alcohols (1.7 g/L of ethanol, 1.1 g/L of butanol, and 0.6 g/L of hexanol) at pH 4.8. In conclusion, low fermentation pH is critical for the production of alcohols, although detrimental to C. kluyveri. Fine control of

  2. Energy conservation via electron bifurcating ferredoxin reduction and proton/Na(+) translocating ferredoxin oxidation.

    PubMed

    Buckel, Wolfgang; Thauer, Rudolf K

    2013-02-01

    The review describes four flavin-containing cytoplasmatic multienzyme complexes from anaerobic bacteria and archaea that catalyze the reduction of the low potential ferredoxin by electron donors with higher potentials, such as NAD(P)H or H(2) at ≤ 100 kPa. These endergonic reactions are driven by concomitant oxidation of the same donor with higher potential acceptors such as crotonyl-CoA, NAD(+) or heterodisulfide (CoM-S-S-CoB). The process called flavin-based electron bifurcation (FBEB) can be regarded as a third mode of energy conservation in addition to substrate level phosphorylation (SLP) and electron transport phosphorylation (ETP). FBEB has been detected in the clostridial butyryl-CoA dehydrogenase/electron transferring flavoprotein complex (BcdA-EtfBC), the multisubunit [FeFe]hydrogenase from Thermotoga maritima (HydABC) and from acetogenic bacteria, the [NiFe]hydrogenase/heterodisulfide reductase (MvhADG-HdrABC) from methanogenic archaea, and the transhydrogenase (NfnAB) from many Gram positive and Gram negative bacteria and from anaerobic archaea. The Bcd/EtfBC complex that catalyzes electron bifurcation from NADH to the low potential ferredoxin and to the high potential crotonyl-CoA has already been studied in some detail. The bifurcating protein most likely is EtfBC, which in each subunit (βγ) contains one FAD. In analogy to the bifurcating complex III of the mitochondrial respiratory chain and with the help of the structure of the human ETF, we propose a conformational change by which γ-FADH(-) in EtfBC approaches β-FAD to enable the bifurcating one-electron transfer. The ferredoxin reduced in one of the four electron bifurcating reactions can regenerate H(2) or NADPH, reduce CO(2) in acetogenic bacteria and methanogenic archaea, or is converted to ΔμH(+)/Na(+) by the membrane-associated enzyme complexes Rnf and Ech, whereby NADH and H(2) are recycled, respectively. The mainly bacterial Rnf complexes couple ferredoxin oxidation by NAD(+) with

  3. Energy Conservation Associated with Ethanol Formation from H2 and CO2 in Clostridium autoethanogenum Involving Electron Bifurcation

    PubMed Central

    Mock, Johanna; Zheng, Yanning; Mueller, Alexander P.; Ly, San; Tran, Loan; Segovia, Simon; Nagaraju, Shilpa; Köpke, Michael; Dürre, Peter

    2015-01-01

    ABSTRACT Most acetogens can reduce CO2 with H2 to acetic acid via the Wood-Ljungdahl pathway, in which the ATP required for formate activation is regenerated in the acetate kinase reaction. However, a few acetogens, such as Clostridium autoethanogenum, Clostridium ljungdahlii, and Clostridium ragsdalei, also form large amounts of ethanol from CO2 and H2. How these anaerobes with a growth pH optimum near 5 conserve energy has remained elusive. We investigated this question by determining the specific activities and cofactor specificities of all relevant oxidoreductases in cell extracts of H2/CO2-grown C. autoethanogenum. The activity studies were backed up by transcriptional and mutational analyses. Most notably, despite the presence of six hydrogenase systems of various types encoded in the genome, the cells appear to contain only one active hydrogenase. The active [FeFe]-hydrogenase is electron bifurcating, with ferredoxin and NADP as the two electron acceptors. Consistently, most of the other active oxidoreductases rely on either reduced ferredoxin and/or NADPH as the electron donor. An exception is ethanol dehydrogenase, which was found to be NAD specific. Methylenetetrahydrofolate reductase activity could only be demonstrated with artificial electron donors. Key to the understanding of this energy metabolism is the presence of membrane-associated reduced ferredoxin:NAD+ oxidoreductase (Rnf), of electron-bifurcating and ferredoxin-dependent transhydrogenase (Nfn), and of acetaldehyde:ferredoxin oxidoreductase, which is present with very high specific activities in H2/CO2-grown cells. Based on these findings and on thermodynamic considerations, we propose metabolic schemes that allow, depending on the H2 partial pressure, the chemiosmotic synthesis of 0.14 to 1.5 mol ATP per mol ethanol synthesized from CO2 and H2. IMPORTANCE Ethanol formation from syngas (H2, CO, and CO2) and from H2 and CO2 that is catalyzed by bacteria is presently a much-discussed process for

  4. Energy Conservation Associated with Ethanol Formation from H2 and CO2 in Clostridium autoethanogenum Involving Electron Bifurcation.

    PubMed

    Mock, Johanna; Zheng, Yanning; Mueller, Alexander P; Ly, San; Tran, Loan; Segovia, Simon; Nagaraju, Shilpa; Köpke, Michael; Dürre, Peter; Thauer, Rudolf K

    2015-09-01

    Most acetogens can reduce CO2 with H2 to acetic acid via the Wood-Ljungdahl pathway, in which the ATP required for formate activation is regenerated in the acetate kinase reaction. However, a few acetogens, such as Clostridium autoethanogenum, Clostridium ljungdahlii, and Clostridium ragsdalei, also form large amounts of ethanol from CO2 and H2. How these anaerobes with a growth pH optimum near 5 conserve energy has remained elusive. We investigated this question by determining the specific activities and cofactor specificities of all relevant oxidoreductases in cell extracts of H2/CO2-grown C. autoethanogenum. The activity studies were backed up by transcriptional and mutational analyses. Most notably, despite the presence of six hydrogenase systems of various types encoded in the genome, the cells appear to contain only one active hydrogenase. The active [FeFe]-hydrogenase is electron bifurcating, with ferredoxin and NADP as the two electron acceptors. Consistently, most of the other active oxidoreductases rely on either reduced ferredoxin and/or NADPH as the electron donor. An exception is ethanol dehydrogenase, which was found to be NAD specific. Methylenetetrahydrofolate reductase activity could only be demonstrated with artificial electron donors. Key to the understanding of this energy metabolism is the presence of membrane-associated reduced ferredoxin:NAD(+) oxidoreductase (Rnf), of electron-bifurcating and ferredoxin-dependent transhydrogenase (Nfn), and of acetaldehyde:ferredoxin oxidoreductase, which is present with very high specific activities in H2/CO2-grown cells. Based on these findings and on thermodynamic considerations, we propose metabolic schemes that allow, depending on the H2 partial pressure, the chemiosmotic synthesis of 0.14 to 1.5 mol ATP per mol ethanol synthesized from CO2 and H2. Ethanol formation from syngas (H2, CO, and CO2) and from H2 and CO2 that is catalyzed by bacteria is presently a much-discussed process for sustainable

  5. Metagenomic Reconstruction of Key Anaerobic Digestion Pathways in Municipal Sludge and Industrial Wastewater Biogas-Producing Systems

    PubMed Central

    Cai, Mingwei; Wilkins, David; Chen, Jiapeng; Ng, Siu-Kin; Lu, Hongyuan; Jia, Yangyang; Lee, Patrick K. H.

    2016-01-01

    Anaerobic digestion (AD) is a microbial process widely used to treat organic wastes. While the microbes involved in digestion of municipal sludge are increasingly well characterized, the taxonomic and functional compositions of AD digesters treating industrial wastewater have been understudied. This study examined metagenomes from a biogas-producing digester treating municipal sludge in Shek Wu Hui (SWH), Hong Kong and an industrial wastewater digester in Guangzhou (GZ), China, and compared their taxonomic composition and reconstructed biochemical pathways. Genes encoding carbohydrate metabolism and protein metabolism functions were overrepresented in GZ, while genes encoding functions related to fatty acids, lipids and isoprenoids were overrepresented in SWH, reflecting the plants’ feedstocks. Mapping of genera to functions in each community indicated that both digesters had a high level of functional redundancy, and a more even distribution of genera in GZ suggested that it was more functionally stable. While fermentation in both samples was dominated by Clostridia, SWH had an overrepresentation of Proteobacteria, including syntrophic acetogens, reflecting its more complex substrate. Considering the growing importance of biogas as an alternative fuel source, a detailed mechanistic understanding of AD is important and this report will be a basis for further study of industrial wastewater AD. PMID:27252693

  6. Synergistic effects of the chitosan addition and polysaccharides-EPS on the formation of anaerobic granules.

    PubMed

    Hudayah, N; Suraraksa, B; Chaiprasert, P

    2016-11-01

    Concomitant early granulation with chitosan addition under a syntroph-specific substrate and enhancement of extracellular polymeric substances (EPS) production were aimed at to build anaerobic granules with high syntrophic activities in a short period. Two laboratory-scale upflow anaerobic sludge blanket reactors were operated as control (R1) and chitosan addition (R2) reactors during early granulation (phase 1). Chitosan decreased the negativity of microbial surface charges (zeta potential) to -10.5 mV on day 58 which led to increases in average diameter sizes, nuclei and granule ratio of approximately 115 µm, 55.1% and 8.2%, respectively. While zeta potential in R1 slightly changed, this resulted in less microbial aggregation. Although microbial aggregation in R2 was rapidly triggered by chitosan addition during phase 1, its structure was clumpy with rough surface due to lack of EPS. Substrate switching to glucose increased polysaccharides-EPS during phase 2 which was synergistically improved on the structural characteristics of microbial aggregate in R2, that is, more spherical and compact, with a smoother surface. Rapid-growth microorganism was also boosted, which then dominated the outer layer of the aggregate. The Archaea clumps were observed at a deeper layer and were surrounded by Eubacteria, presumably acetogens, indicating a syntrophic relationship due to substrate association between these microbial groups.

  7. Beating the acetyl coenzyme A-pathway to the origin of life.

    PubMed

    Nitschke, Wolfgang; Russell, Michael J

    2013-07-19

    Attempts to draft plausible scenarios for the origin of life have in the past mainly built upon palaeogeochemical boundary conditions while, as detailed in a companion article in this issue, frequently neglecting to comply with fundamental thermodynamic laws. Even if demands from both palaeogeochemistry and thermodynamics are respected, then a plethora of strongly differing models are still conceivable. Although we have no guarantee that life at its origin necessarily resembled biology in extant organisms, we consider that the only empirical way to deduce how life may have emerged is by taking the stance of assuming continuity of biology from its inception to the present day. Building upon this conviction, we have assessed extant types of energy and carbon metabolism for their appropriateness to conditions probably pertaining in those settings of the Hadean planet that fulfil the thermodynamic requirements for life to come into being. Wood-Ljungdahl (WL) pathways leading to acetyl CoA formation are excellent candidates for such primordial metabolism. Based on a review of our present understanding of the biochemistry and biophysics of acetogenic, methanogenic and methanotrophic pathways and on a phylogenetic analysis of involved enzymes, we propose that a variant of modern methanotrophy is more likely than traditional WL systems to date back to the origin of life. The proposed model furthermore better fits basic thermodynamic demands and palaeogeochemical conditions suggested by recent results from extant alkaline hydrothermal seeps.

  8. Stimulation of the hydrolytic stage for biogas production from cattle manure in an electrochemical bioreactor.

    PubMed

    Samani, Saeed; Abdoli, Mohammad Ali; Karbassi, Abdolreza; Amin, Mohammad Mehdi

    Electrical current in the hydrolytic phase of the biogas process might affect biogas yield. In this study, four 1,150 mL single membrane-less chamber electrochemical bioreactors, containing two parallel titanium plates were connected to the electrical source with voltages of 0, -0.5, -1 and -1.5 V, respectively. Reactor 1 with 0 V was considered as a control reactor. The trend of biogas production was precisely checked against pH, oxidation reduction potential and electrical power at a temperature of 37 ± 0.5°C amid cattle manure as substrate for 120 days. Biogas production increased by voltage applied to Reactors 2 and 3 when compared with the control reactor. In addition, the electricity in Reactors 2 and 3 caused more biogas production than Reactor 4. Acetogenic phase occurred more quickly in Reactor 3 than in the other reactors. The obtained results from Reactor 4 were indicative of acidogenic domination and its continuous behavior under electrical stimulation. The results of the present investigation clearly revealed that phasic electrical current could enhance the efficiency of biogas production.

  9. Metabolic engineering in chemolithoautotrophic hosts for the production of fuels and chemicals.

    PubMed

    Nybo, S Eric; Khan, Nymul E; Woolston, Benjamin M; Curtis, Wayne R

    2015-07-01

    The ability of autotrophic organisms to fix CO2 presents an opportunity to utilize this 'greenhouse gas' as an inexpensive substrate for biochemical production. Unlike conventional heterotrophic microorganisms that consume carbohydrates and amino acids, prokaryotic chemolithoautotrophs have evolved the capacity to utilize reduced chemical compounds to fix CO2 and drive metabolic processes. The use of chemolithoautotrophic hosts as production platforms has been renewed by the prospect of metabolically engineered commodity chemicals and fuels. Efforts such as the ARPA-E electrofuels program highlight both the potential and obstacles that chemolithoautotrophic biosynthetic platforms provide. This review surveys the numerous advances that have been made in chemolithoautotrophic metabolic engineering with a focus on hydrogen oxidizing bacteria such as the model chemolithoautotrophic organism (Ralstonia), the purple photosynthetic bacteria (Rhodobacter), and anaerobic acetogens. Two alternative strategies of microbial chassis development are considered: (1) introducing or enhancing autotrophic capabilities (carbon fixation, hydrogen utilization) in model heterotrophic organisms, or (2) improving tools for pathway engineering (transformation methods, promoters, vectors etc.) in native autotrophic organisms. Unique characteristics of autotrophic growth as they relate to bioreactor design and process development are also discussed in the context of challenges and opportunities for genetic manipulation of organisms as production platforms. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  10. Converting Carbon Dioxide to Butyrate with an Engineered Strain of Clostridium ljungdahlii

    PubMed Central

    Nevin, Kelly P.; Woodard, Trevor L.; Lovley, Derek R.

    2014-01-01

    ABSTRACT Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70% of the carbon and electron flow was diverted to the production of butyrate with H2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. PMID:25336453

  11. Methane production and consumption in grassland and boreal ecosystems

    NASA Technical Reports Server (NTRS)

    Schimel, David S.; Burke, Ingrid C.; Johnston, Carol; Pastor, John

    1994-01-01

    The objectives of the this project were to develop a mechanistic understanding of methane production and oxidation suitable for incorporation into spatially explicit models for spatial extrapolation. Field studies were undertaken in Minnesota, Canada, and Colorado to explore the process controls over the two microbial mediated methane transformations in a range of environments. Field measurements were done in conjunction with ongoing studies in Canada (the Canadian Northern Wetlands Projects: NOWES) and in Colorado (The Shortgrass Steppe Long Term Ecological Research Project: LTER). One of the central hypotheses of the proposal was that methane production should be substrate limited, as well as being controlled by physical variables influencing microbial activity (temperature, oxidation status, and pH). Laboratory studies of peats from Canada and Minnesota (Northern and Southern Boreal) were conducted with amendments of a methanogenic substrate at multiple temperatures and at multiple pHs (the latter by titrating samples). The studies showed control by substrate, pH, and temperature in order in anaerobic samples. Field and laboratory manipulations of natural plant litter, rather than an acetogenic substrate, showed similarly large effects. The studies concluded that substrate is an important control over methanogenesis, that substrate availability in the field is closely coupled to the chemistry of the dominant vegetation influencing its decomposition rate, that most methane is produced from recent plant litter, and that landscape changes in pH are an important control, highly correlated with vegetation.

  12. Formyltetrahydrofolate synthetase gene diversity in the guts of higher termites with different diets and lifestyles.

    PubMed

    Ottesen, Elizabeth A; Leadbetter, Jared R

    2011-05-01

    In this study, we examine gene diversity for formyl-tetrahydrofolate synthetase (FTHFS), a key enzyme in homoacetogenesis, recovered from the gut microbiota of six species of higher termites. The "higher" termites (family Termitidae), which represent the majority of extant termite species and genera, engage in a broader diversity of feeding and nesting styles than the "lower" termites. Previous studies of termite gut homoacetogenesis have focused on wood-feeding lower termites, from which the preponderance of FTHFS sequences recovered were related to those from acetogenic treponemes. While sequences belonging to this group were present in the guts of all six higher termites examined, treponeme-like FTHFS sequences represented the majority of recovered sequences in only two species (a wood-feeding Nasutitermes sp. and a palm-feeding Microcerotermes sp.). The remaining four termite species analyzed (a Gnathamitermes sp. and two Amitermes spp. that were recovered from subterranean nests with indeterminate feeding strategies and a litter-feeding Rhynchotermes sp.) yielded novel FTHFS clades not observed in lower termites. These termites yielded two distinct clusters of probable purinolytic Firmicutes and a large group of potential homoacetogens related to sequences previously recovered from the guts of omnivorous cockroaches. These findings suggest that the gut environments of different higher termite species may select for different groups of homoacetogens, with some species hosting treponeme-dominated homoacetogen populations similar to those of wood-feeding, lower termites while others host Firmicutes-dominated communities more similar to those of omnivorous cockroaches.

  13. Molecular analysis of deep subsurface Cretaceous rock indicates abundant Fe(III)- and S(zero)-reducing bacteria in a sulfate-rich environment.

    PubMed

    Kovacik, William P; Takai, Ken; Mormile, Melanie R; McKinley, James P; Brockman, Fred J; Fredrickson, James K; Holben, William E

    2006-01-01

    A multilevel sampler (MLS) was emplaced in a borehole straddling anaerobic, sulfate-rich Cretaceous-era shale and sandstone rock formations approximately 200 m below ground surface at Cerro Negro, New Mexico. Sterile quartzite sand contained in chambers in the sampler allowed in situ colonization and recovery of nucleic acids for molecular analyses. Denaturing gradient gel electrophoresis and 16S rRNA gene cloning results indicated a homogeneously distributed bacterial community across the shale-sandstone interface. delta-Proteobacteria sequences were common at all depths, and were dominated by members of the Geobacteraceae family (Pelobacter, Desulphuromonas and Geobacter). Other members of this group are capable of dissimilatory Fe(III) and/or S degrees reduction, but not sulfate reduction. RNA hybridization data also suggested that Fe(III)-/S degrees -reducing bacteria were predominant. These findings are striking considering the lack of significant concentrations of these electron acceptors in this environment. The next most abundant bacterial group indicated was the sulfate reducers, including Desulfobacterium, Desulfocapsa and Desulfobulbus. Sequences related to fermenters, denitrifiers and acetogens were also recovered. The presence of a phylogenetically and functionally diverse microbial community in this deep subsurface environment likely reflects the complex nature of the primary energy and carbon sources, kerogen associated with the shale.

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

    PubMed

    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.

  15. Cascade degradation of organic matters in brewery wastewater using a continuous stirred microbial electrochemical reactor and analysis of microbial communities

    PubMed Central

    Wang, Haiman; Qu, Youpeng; Li, Da; Ambuchi, John J.; He, Weihua; Zhou, Xiangtong; Liu, Jia; Feng, Yujie

    2016-01-01

    A continuous stirred microbial electrochemical reactor (CSMER), comprising of a complete mixing zone (CMZ) and microbial electrochemical zone (MEZ), was used for brewery wastewater treatment. The system realized 75.4 ± 5.7% of TCOD and 64.9 ± 4.9% of TSS when fed with brewery wastewater concomitantly achieving an average maximum power density of 304 ± 31 m W m−2. Cascade utilization of organic matters made the CSMER remove a wider range of substrates compared with a continuous stirred tank reactor (CSTR), in which process 79.1 ± 5.6% of soluble protein and 86.6 ± 2.2% of soluble carbohydrates were degraded by anaerobic digestion in the CMZ and short-chain volatile fatty acids were further decomposed and generated current in the MEZ. Co-existence of fermentative bacteria (Clostridium and Bacteroides, 19.7% and 5.0%), acetogenic bacteria (Syntrophobacter, 20.8%), methanogenic archaea (Methanosaeta and Methanobacterium, 40.3% and 38.4%) and exoelectrogens (Geobacter, 12.4%) as well as a clear spatial distribution and syntrophic interaction among them contributed to the cascade degradation process in CSMER. The CSMER shows great promise for practical wastewater treatment application due to high pre-hydrolysis and acidification rate, high energy recovery and low capital cost. PMID:27270788

  16. Mercury methylation by novel microorganisms from new environments

    SciTech Connect

    Gilmour, C C; Podar, Mircea; Bullock, Allyson L; Graham, Dr. Andrew M; Brown, Steven D; Somenahally, Anil C; Johs, Alexander; Hurt, Jr., Richard Ashley; Bailey, Kathryn L; Elias, Dwayne A

    2013-01-01

    Microbial mercury (Hg) methylation transforms a toxic trace metal into the highly bioaccumulated neurotoxin methylmercury (MeHg). The lack of a genetic marker for microbial MeHg production has prevented a clear understanding of Hg-methylating organism distribution in nature. Recently, a specific gene cluster (hgcAB) was linked to Hg methylation in two bacteria.1 Here we test if the presence of hgcAB orthologues is a reliable predictor of Hg methylation capability in microorganisms, a necessary confirmation for the development of molecular probes for Hg-methylation in nature. Although hgcAB orthologues are rare among all available microbial genomes, organisms are much more phylogenetically and environmentally diverse than previously thought. By directly measuring MeHg production in several bacterial and archaeal strains encoding hgcAB, we confirmed that possessing hgcAB predicts Hg methylation capability. For the first time, we demonstrated Hg methylation in a number of species other than sulfate- (SRB) and iron- (FeRB) reducing bacteria, including methanogens, and syntrophic, acetogenic, and fermentative Firmicutes. Several of these species occupy novel environmental niches for Hg methylation, including methanogenic habitats such as rice paddies, the animal gut, and extremes of pH and salinity. Identification of these organisms as Hg methylators now links methylation to discrete gene markers in microbial communities.

  17. Genomic evidence for distinct carbon substrate preferences and ecological niches of Bathyarchaeota in estuarine sediments.

    PubMed

    Lazar, Cassandre Sara; Baker, Brett J; Seitz, Kiley; Hyde, Andrew S; Dick, Gregory J; Hinrichs, Kai-Uwe; Teske, Andreas P

    2016-04-01

    Investigations of the biogeochemical roles of benthic Archaea in marine sediments are hampered by the scarcity of cultured representatives. In order to determine their metabolic capacity, we reconstructed the genomic content of four widespread uncultured benthic Archaea recovered from estuary sediments at 48% to 95% completeness. Four genomic bins were found to belong to different subgroups of the former Miscellaneous Crenarcheota Group (MCG) now called Bathyarchaeota: MCG-6, MCG-1, MCG-7/17 and MCG-15. Metabolic predictions based on gene content of the different genome bins indicate that subgroup 6 has the ability to hydrolyse extracellular plant-derived carbohydrates, and that all four subgroups can degrade detrital proteins. Genes encoding enzymes involved in acetate production as well as in the reductive acetyl-CoA pathway were detected in all four genomes inferring that these Archaea are organo-heterotrophic and autotrophic acetogens. Genes involved in nitrite reduction were detected in all Bathyarchaeota subgroups and indicate a potential for dissimilatory nitrite reduction to ammonium. Comparing the genome content of the different Bathyarchaeota subgroups indicated preferences for distinct types of carbohydrate substrates and implicitly, for different niches within the sedimentary environment.

  18. CO2 fixation by anaerobic non-photosynthetic mixotrophy for improved carbon conversion.

    PubMed

    Jones, Shawn W; Fast, Alan G; Carlson, Ellinor D; Wiedel, Carrissa A; Au, Jennifer; Antoniewicz, Maciek R; Papoutsakis, Eleftherios T; Tracy, Bryan P

    2016-09-30

    Maximizing the conversion of biogenic carbon feedstocks into chemicals and fuels is essential for fermentation processes as feedstock costs and processing is commonly the greatest operating expense. Unfortunately, for most fermentations, over one-third of sugar carbon is lost to CO2 due to the decarboxylation of pyruvate to acetyl-CoA and limitations in the reducing power of the bio-feedstock. Here we show that anaerobic, non-photosynthetic mixotrophy, defined as the concurrent utilization of organic (for example, sugars) and inorganic (for example, CO2) substrates in a single organism, can overcome these constraints to increase product yields and reduce overall CO2 emissions. As a proof-of-concept, Clostridium ljungdahlii was engineered to produce acetone and achieved a mass yield 138% of the previous theoretical maximum using a high cell density continuous fermentation process. In addition, when enough reductant (that is, H2) is provided, the fermentation emits no CO2. Finally, we show that mixotrophy is a general trait among acetogens.

  19. [A microbiological study of an underground gas storage in the process of gas extraction].

    PubMed

    Ivanova, A E; Borzenkov, I A; Tarasov, A L; Milekhina, E I; Beliaev, S S

    2007-01-01

    The numbers of microorganisms belonging to ecologically significant groups and the rates of terminal microbial processes of sulfate reduction and methanogenesis were determined in the liquid phase of an underground gas storage (UGS) in the period of gas extraction. The total number of microorganisms in water samples from the operation and injection wells reached 2.1 x 10(6) cells/ml. Aerobic organotrophs (including hydrocarbon- and oil-oxidizing ones) and various anaerobic microorganisms (fermenting bacteria, methanogens, acetogens, sulfate-, nitrate-, and iron-reducing bacteria) were constituent parts of the community. The radioisotopic method showed that, in all the UGS units, the terminal stages of organic matter decomposition included sulfate reduction and methanogenesis, with the maximal rate of these processes recorded in the aqueous phase of above-ground technological equipment which the gas enters from the operation wells. A comparative analysis by these parameters of different anaerobic ecotopes, including natural hydrocarbon fields, allows us to assess the rate of these processes in the UGS as high throughout the annual cycle of its operation. The data obtained indicate the existence in the UGS of a bacterial community that is unique in its diversity and metabolic capacities and able to make a certain contribution to the geochemistry of organic and inorganic compounds in the natural and technogenic ecosystem of the UGS and thus influence the industrial gas composition.

  20. Compositions of microbial communities associated with oil and water in a mesothermic oil field.

    PubMed

    Kryachko, Yuriy; Dong, Xiaoli; Sensen, Christoph W; Voordouw, Gerrit

    2012-03-01

    Samples of produced water and oil obtained from the Enermark field (near Medicine Hat, Alberta, Canada) were separated into oil and aqueous phases first gravitationally and then through centrifugation at 20°C in an atmosphere of 90% N(2) and 10% CO(2). Biomass that remained associated with oil after gravitational separation (1×g) was dislodged by centrifugation at 25,000×g. DNA was isolated from the aqueous and oil-associated biomass fractions and subjected to polymerase chain reaction amplification with primers targeting bacterial and archaeal 16S rRNA genes. DNA pyrosequencing and bioinformatics tools were used to characterize the resulting 16S rRNA gene amplicons. The oil-associated microbial community was less diverse than that of the aqueous phase and had consistently higher representation of hydrogenotrophs (methanogens of the genera Methanolobus and Methanobacterium and acetogens of the genus Acetobacterium), indicating the oil phase to be a primary source of hydrogen. Many known hydrocarbon degraders were also found to be oil-attached, e.g. representatives of the gammaproteobacterial genus Thalassolituus, the actinobacterial genus Rhodococcus and the alphaproteobacterial genera Sphingomonas, Brevundimonas and Stappia. In contrast, all eight representatives of genera of the Deltaproteobacteria identified were found to be associated with the aqueous phase, likely because their preferred growth substrates are mostly water-soluble. Hence, oil attachment was seen for genera acting on substrates found primarily in the oil phase.

  1. Effect of nonylphenol on volatile fatty acids accumulation during anaerobic fermentation of waste activated sludge.

    PubMed

    Duan, Xu; Wang, Xiao; Xie, Jing; Feng, Leiyu; Yan, Yuanyuan; Zhou, Qi

    2016-11-15

    Most of the reported studies on anaerobic fermentation of sludge focused on the influences of operating conditions, pretreatment methods, and its characteristics, and little attention was paid to those of persistent organic pollutants (POPs) which widespreadly appeared in sludge. In this study, the effect of nonylphenol, a typical POPs in waste activated sludge (WAS), on anaerobic fermentation for volatile fatty acids (VFAs) accumulation was investigated. The concentration of VFAs during WAS anaerobic fermentation was found to be affected positively from 2856 mg COD/L in the control (without NP) to 5620 mg COD/L with NP of 200 mg/kg dry sludge. Mechanism exploration exhibited that the main reason for the enhanced VFAs accumulation in the presence of NP was that more acetic acid was generated during the acidification of WAS, which was increased by almost three times (3790 versus 1310 mg COD/L). In WAS fermentation systems, the abundance of anaerobic functional microorganisms was advantageous to the accumulation of acetic acid. Further investigation by the pure acetogen revealed that both the viability and activity of Proteiniphilum acetatigenes were improved by NP during anaerobic fermentation, resulting in more production of acetic acid and showing good agreement with that in the real WAS fermentation systems. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. The role of homoacetogenic bacteria as efficient hydrogen scavengers in microbial electrochemical cells (MXCs).

    PubMed

    Parameswaran, Prathap; Torres, César I; Kang, Dae-Wook; Rittmann, Bruce E; Krajmalnik-Brown, Rosa

    2012-01-01

    We evaluated the consumption of hydrogen gas at the anode of a microbial electrolysis cell (MEC) and characterized the significance of new interactions between anode respiring bacteria (ARB) and homo-acetogens. We demonstrated the significance of biofilm limitation for direct consumption of H(2) over acetate by ARB, using the deep biofilm model. Selective inhibition of the major competing hydrogen sink at the biofilm anode, methanogenesis, resulted in significant increase in electron recovery as electric current (∼10-12 A/m(2)). The presence of acetate at high concentration in the anode compartment and detection of formate, a known intermediate of the acetyl-CoA pathway, provide evidence towards the role of homoacetogenic bacteria. We also assessed the activity of homoacetogens with reverse transcription quantitative PCR targeting formyltetrahydrofolate synthetase (FTHFS) transcripts, and observed a comparable decrease in the FTHFS transcript numbers with current density and acetate concentrations as we decreased the HRT below 4.5 h. The biofilm anode community was predominated by Deltaproteobacteria (70% of total readouts) along with a fraction of the homoacetogenic genus, Acetobacterium (4% of total readouts), established by pyrosequencing targeting the V6 region of the 16S rRNA. Homoacetogens seem to play a major role as syntrophic members of the biofilm anode community when electron recovery is high.

  3. Gas Fermentation—A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks

    PubMed Central

    Liew, FungMin; Martin, Michael E.; Tappel, Ryan C.; Heijstra, Björn D.; Mihalcea, Christophe; Köpke, Michael

    2016-01-01

    There is an immediate need to drastically reduce the emissions associated with global fossil fuel consumption in order to limit climate change. However, carbon-based materials, chemicals, and transportation fuels are predominantly made from fossil sources and currently there is no alternative source available to adequately displace them. Gas-fermenting microorganisms that fix carbon dioxide (CO2) and carbon monoxide (CO) can break this dependence as they are capable of converting gaseous carbon to fuels and chemicals. As such, the technology can utilize a wide range of feedstocks including gasified organic matter of any sort (e.g., municipal solid waste, industrial waste, biomass, and agricultural waste residues) or industrial off-gases (e.g., from steel mills or processing plants). Gas fermentation has matured to the point that large-scale production of ethanol from gas has been demonstrated by two companies. This review gives an overview of the gas fermentation process, focusing specifically on anaerobic acetogens. Applications of synthetic biology and coupling gas fermentation to additional processes are discussed in detail. Both of these strategies, demonstrated at bench-scale, have abundant potential to rapidly expand the commercial product spectrum of gas fermentation and further improve efficiencies and yields. PMID:27242719

  4. Methanol-driven enhanced biological phosphorus removal with a syntrophic consortium.

    PubMed

    Tayà, Carlota; Guerrero, Javier; Vanneste, Gianni; Guisasola, Albert; Baeza, Juan A

    2013-02-01

    The presence of suitable carbon sources for enhanced biological phosphorus removal (EBPR) plays a key role in phosphorus removal from wastewater in urban WWTP. For wastewaters with low volatile fatty acids (VFAs) content, an external carbon addition is necessary. As methanol is the most commonly external carbon source used for denitrification it could be a priori a promising alternative, but previous attempts to use it for EBPR have failed. This study is the first successful report of methanol utilization as external carbon source for EBPR. Since a direct replacement strategy (i.e., supply of methanol as a sole carbon source to a propionic-fed PAO-enriched sludge) failed, a novel process was designed and implemented successfully: development of a consortium with anaerobic biomass and polyphosphate accumulating organisms (PAOs). Methanol-degrading acetogens were (i) selected against other anaerobic methanol degraders from an anaerobic sludge; (ii) subjected to conventional EBPR conditions (anaerobic + aerobic); and (iii) bioaugmented with PAOs. EBPR with methanol as a sole carbon source was sustained in a mid-term basis with this procedure. Copyright © 2012 Wiley Periodicals, Inc.

  5. Microbial dark matter ecogenomics reveals complex synergistic networks in a methanogenic bioreactor

    PubMed Central

    Nobu, Masaru K; Narihiro, Takashi; Rinke, Christian; Kamagata, Yoichi; Tringe, Susannah G; Woyke, Tanja; Liu, Wen-Tso

    2015-01-01

    Ecogenomic investigation of a methanogenic bioreactor degrading terephthalate (TA) allowed elucidation of complex synergistic networks of uncultivated microorganisms, including those from candidate phyla with no cultivated representatives. Our previous metagenomic investigation proposed that Pelotomaculum and methanogens may interact with uncultivated organisms to degrade TA; however, many members of the community remained unaddressed because of past technological limitations. In further pursuit, this study employed state-of-the-art omics tools to generate draft genomes and transcriptomes for uncultivated organisms spanning 15 phyla and reports the first genomic insight into candidate phyla Atribacteria, Hydrogenedentes and Marinimicrobia in methanogenic environments. Metabolic reconstruction revealed that these organisms perform fermentative, syntrophic and acetogenic catabolism facilitated by energy conservation revolving around H2 metabolism. Several of these organisms could degrade TA catabolism by-products (acetate, butyrate and H2) and syntrophically support Pelotomaculum. Other taxa could scavenge anabolic products (protein and lipids) presumably derived from detrital biomass produced by the TA-degrading community. The protein scavengers expressed complementary metabolic pathways indicating syntrophic and fermentative step-wise protein degradation through amino acids, branched-chain fatty acids and propionate. Thus, the uncultivated organisms may interact to form an intricate syntrophy-supported food web with Pelotomaculum and methanogens to metabolize catabolic by-products and detritus, whereby facilitating holistic TA mineralization to CO2 and CH4. PMID:25615435

  6. Effect of 2-bromoethanesulfonic acid and Peptostreptococcus productus ATCC 35244 addition on stimulation of reductive acetogenesis in the ruminal ecosystem by selective inhibition of methanogenesis.

    PubMed Central

    Nollet, L; Demeyer, D; Verstraete, W

    1997-01-01

    Evidence is provided that reductive acetogenesis can be stimulated in ruminal samples during short-term (24-h) incubations when methanogenesis is inhibited selectively. While addition of the reductive acetogen Peptostreptococcus productus ATCC 35244 alone had no significant influence on CH4 and volatile fatty acid (VFA) production in ruminal samples, the addition of this strain together with 2-bromoethanesulfonic acid (BES) (final concentration, 0.01 or 0.03 mM) resulted in stimulation of acetic acid production and H2 consumption. Since acetate production exceeded amounts that could be attributed to reductive acetogenesis, as measured by H2 consumption, it was found that P. productus also fermented C6 units (glucose and fructose) heterotrophically to mainly acetate (> 99% of the total VFA). Using 14CH3COOH, we concluded that addition of BES and BES plus P. productus did not alter the consumption of acetate in ruminal samples. The addition of P. productus to BES-treated ruminal samples caused supplemental inhibition of CH4 production and stimulation of VFA production, representing a possible energy gain of about 13 to 15%. PMID:8979351

  7. Comparative mesophilic and thermophilic anaerobic digestion of palm oil mill effluent using upflow anaerobic sludge blanket.

    PubMed

    Khemkhao, Maneerat; Nuntakumjorn, Boonyarit; Techkarnjanaruk, Somkiet; Phalakornkule, Chantaraporn

    2012-07-01

    The effects of organic loading rate and operating temperature on the microbial diversity and performances of upflow anaerobic sludge blanket (UASB) reactors treating palm oil mill effluent (POME) were investigated. The following two UASB reactors were run in parallel for comparison: (1) under a mesophilic condition (37 degrees C) and (2) under a mesophilic condition in transition to a thermophilic condition (57 degrees C). A polymerase chain reaction (PCR)-based denaturing gradient gel electrophoresis (DGGE) analysis showed that the microbial population profiles significantly changed with the organic loading rate (OLR) and the temperature transition from the mesophilic to the thermophilic condition. Significant biomass washout was observed for the mesophilic UASB when operating at a high organic loading rate (OLR) of 9.5 g chemical oxygen demand (COD)/L.d. In contrast, the thermophilic UASB can be operated at this OLR and at a temperature of 57 degrees C with satisfactory COD removal and biogas production. The PCR-based DGGE analysis suggested that the thermophilic temperature of 57 degrees C was suitable for a number of hydrolytic, acidogenic, and acetogenic bacteria.

  8. UASB performance and microbial adaptation during a transition from mesophilic to thermophilic treatment of palm oil mill effluent.

    PubMed

    Khemkhao, Maneerat; Nuntakumjorn, Boonyarit; Techkarnjanaruk, Somkiet; Phalakornkule, Chantaraporn

    2012-07-30

    The treatment of palm oil mill effluent (POME) by an upflow anaerobic sludge bed (UASB) at organic loading rates (OLR) between 2.2 and 9.5 g COD l(-1) day(-1) was achieved by acclimatizing the mesophilic (37 °C) microbial seed to the thermophilic temperature (57 °C) by a series of stepwise temperature shifts. The UASB produced up to 13.2 l biogas d(-1) with methane content on an average of 76%. The COD removal efficiency ranged between 76 and 86%. Microbial diversity of granules from the UASB reactor was also investigated. The PCR-based DGGE analysis showed that the bacterial population profiles significantly changed with the temperature transition from mesophilic to thermophilic conditions. In addition, the results suggested that even though the thermophilic temperature of 57 °C was suitable for a number of hydrolytic, acidogenic and acetogenic bacteria, it may not be suitable for some Methanosaeta species acclimatized from 37 °C. Specifically, the bands associated with Methanosaeta thermophila PT and Methanosaeta harundinacea can be detected during the four consecutive operation phases of 37 °C, 42 °C, 47 °C and 52 °C, but their corresponding bands were found to fade out at 57 °C. The DGGE analysis predicted that the temperature transition can result in significant methanogenic biomass washout at 57 °C. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. Evolutionary ecology during the rise of dioxygen in the Earth's atmosphere.

    PubMed

    Sleep, Norman H; Bird, Dennis K

    2008-08-27

    Pre-photosynthetic niches were meagre with a productivity of much less than 10(-4) of modern photosynthesis. Serpentinization, arc volcanism and ridge-axis volcanism reliably provided H(2). Methanogens and acetogens reacted CO(2) with H(2) to obtain energy and make organic matter. These skills pre-adapted a bacterium for anoxygenic photosynthesis, probably starting with H(2) in lieu of an oxygen 'acceptor'. Use of ferrous iron and sulphide followed as abundant oxygen acceptors, allowing productivity to approach modern levels. The 'photobacterium' proliferated rooting much of the bacterial tree. Land photosynthetic microbes faced a dearth of oxygen acceptors and nutrients. A consortium of photosynthetic and soil bacteria aided weathering and access to ferrous iron. Biologically enhanced weathering led to the formation of shales and, ultimately, to granitic rocks. Already oxidized iron-poor sedimentary rocks and low-iron granites provided scant oxygen acceptors, as did freshwater in their drainages. Cyanobacteria evolved dioxygen production that relieved them of these vicissitudes. They did not immediately dominate the planet. Eventually, anoxygenic and oxygenic photosynthesis oxidized much of the Earth's crust and supplied sulphate to the ocean. Anoxygenic photosynthesis remained important until there was enough O(2) in downwelling seawater to quantitatively oxidize massive sulphides at mid-ocean ridge axes.

  10. Microbial activity of biofilm during start-up period of anaerobic hybrid reactor at low and high upflow feeding velocity.

    PubMed

    Suraruksa, B; Nopharatana, A; Chaiprasert, P; Tanticharoen, M; Bhumiratana, S

    2003-01-01

    With an aim to shorten start-up time of an Anaerobic Hybrid Reactor (AHR), initial biofilm development was studied, particularly at different upflow feeding velocities. At a low (0.01 m x h(-1)) upflow velocity, initial biofilm was found to develop via the attachment of suspended biomass in the packed zone, while microbial growth on the film was insignificant. Contrarily, with higher (1.0 m x h(-1)) upflow velocity, initial biofilm development was from both microbial attachment and growth on supporting media. Biofilm thickness was determined using confocal laser scanning microscopy (CLSM), which indicated that the biofilm developed faster with the higher velocity, due to the contribution of the microbial growth on supporting media. When operated beyond the initial biofilm development with the lower velocity, both the activity of acetogens and the methanogens increased, although there was a lower amount of attached biomass on the supporting media. Whereas, both groups were found to decrease with higher upflow velocity, but acidogenic activity increased. It can be concluded that higher upflow velocity positively affected the initial stage of biofilm development and has the potential to accelerate attached biomass on supporting media during the initial phase. Subsequently, the upflow velocity should be reduced to the normal rate to enhance the methanogenic activity.

  11. Continuous gas fermentation by Acetobacterium woodii in a submerged membrane reactor with full cell retention.

    PubMed

    Kantzow, Christina; Mayer, Alexander; Weuster-Botz, Dirk

    2015-10-20

    Acetogenic bacteria like Acetobacterium woodii represent an ancient group of anaerobic microorganisms which use hydrogen and carbon dioxide to produce acetate. Cell concentrations and space-time yields are usually low in gas fermentations. A standard stirred‑tank bioreactor with continuous gas supply was equipped with a customized submerged microfiltration unit. A. woodii showed similar growth behavior with an initial maximal growth rate of 1.2 d(-1) in continuous gas fermentations with full cell retention and varying dilution rates. A steady increase of cell mass concentrations was observed with the highest biomass formation at the highest dilution rate. By contrast the final acetate concentrations were lowest at the highest dilution rate. The highest final acetate space-time yield of 148 g l(-1) d(-1) was measured at the highest dilution rate (increase by factor 8 compared to a standard batch process or by factor 37 compared to published data). The highest reported cell concentration of A. woodii in gas fermentations of nearly 14 g l(-1) cell dry weight was achieved in the submerged membrane bioreactor with increased yeast extract concentrations in the feed medium. Product inhibition was observed when acetate concentrations exceeded 8-12 g l(-1) causing a steady decrease in cell mass specific acetate production rates. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Expression of amplified synthetic ethanol pathway integrated using Tn7-tool and powered at the expense of eliminated pta, ack, spo0A and spo0J during continuous syngas or CO2 /H2 blend fermentation.

    PubMed

    Kiriukhin, M; Tyurin, M

    2013-04-01

    To engineer acetogen biocatalyst selectively overproducing ethanol from synthesis gas or CO2 /H2 as the only liquid carbonaceous product. Ethanol-resistant mutant originally capable of producing only acetate from CO2 /CO was engineered to eliminate acetate production and spore formation using our proprietary Cre-lox66/lox71-system. Bi-functional aldehyde/alcohol dehydrogenase was inserted into the chromosome of the engineered mutant using Tn7-based approach. Recombinants with three or six copies of the inserted gene produced 525 mmol l(-1) and 1018 mmol l(-1) of ethanol, respectively, in five independent single-step fermentation runs 25 days each (P < 0.005) in five independent repeats using syngas blend 60% CO and 40% H2 . Ethanol production was 64% if only CO2 + H2 blend was used compared with syngas blend (P < 0.005). Elimination of genes unnecessary for syngas fermentation can boost artificial integrated pathway performance. Cell energy released via elimination of phosphotransacetylase, acetate kinase and early-stage sporulation genes boosted ethanol production. Deletion of sporulation genes added theft-proof feature to the engineered biocatalyst. Production of ethanol from CO2 /H2 blend might be utilized as a tool to mitigate global warming proportional to CO2 fermentation scale. © 2013 The Society for Applied Microbiology.

  13. Influence of the composition of the cellulolytic flora on the development of hydrogenotrophic microorganisms, hydrogen utilization, and methane production in the rumens of gnotobiotically reared lambs.

    PubMed

    Chaucheyras-Durand, Frédérique; Masséglia, Sébastien; Fonty, Gérard; Forano, Evelyne

    2010-12-01

    We investigated the influence of the composition of the fibrolytic microbial community on the development and activities of hydrogen-utilizing microorganisms in the rumens of gnotobiotically reared lambs. Two groups of lambs were reared. The first group was inoculated with Fibrobacter succinogenes, a non-H(2)-producing species, as the main cellulolytic organism, and the second group was inoculated with Ruminococcus albus, Ruminococcus flavefaciens, and anaerobic fungi that produce hydrogen. The development of hydrogenotrophic bacterial communities, i.e., acetogens, fumarate and sulfate reducers, was monitored in the absence of methanogens and after inoculation of methanogens. Hydrogen production and utilization and methane production were measured in rumen content samples incubated in vitro in the presence of exogenous hydrogen (supplemented with fumarate or not supplemented with fumarate) or in the presence of ground alfalfa hay as a degradable substrate. Our results show that methane production was clearly reduced when the dominant fibrolytic species was a non-H(2)-producing species, such as Fibrobacter succinogenes, without significantly impairing fiber degradation and fermentations in the rumen. The addition of fumarate to the rumen contents stimulated H(2) utilization only by the ruminal microbiota inoculated with F. succinogenes, suggesting that these communities could play an important role in fumarate reduction in vivo.

  14. Performance and spatial community succession of an anaerobic baffled reactor treating acetone-butanol-ethanol fermentation wastewater.

    PubMed

    Zhang, Jun; Wei, Yongjun; Xiao, Wei; Zhou, Zhihua; Yan, Xing

    2011-08-01

    An anaerobic baffled reactor with four compartments (C1-C4) was successfully used for treatment of acetone-butanol-ethanol fermentation wastewater and methane production. The chemical oxygen demand (COD) removal efficiency was 88.2% with a CH(4) yield of 0.25L/(g COD(removed)) when organic loading rate (OLR) was 5.4kg CODm(-3)d(-1). C1 played the most important role in solvents (acetone, butanol and ethanol) and COD removal. Community structure of C2 was similar to that in C1 at stage 3 with higher OLR, but was similar to those in C3 and C4 at stages 1-2 with lower OLR. This community variation in C2 was consistent with its increased role in COD and solvent removal at stage 3. During community succession from C1 to C4 at stage 3, abundance of Firmicutes (especially OTUs ABRB07 and ABRB10) and Methanoculleus decreased, while Bacteroidetes and Methanocorpusculum became dominant. Thus, ABRB07 coupled with Methanoculleus and/or acetogen (ABRB10) may be key species for solvents degradation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  15. Microbial Anaerobic Digestion (Bio-Digesters) as an Approach to the Decontamination of Animal Wastes in Pollution Control and the Generation of Renewable Energy

    PubMed Central

    Manyi-Loh, Christy E.; Mamphweli, Sampson N.; Meyer, Edson L.; Okoh, Anthony I.; Makaka, Golden; Simon, Michael

    2013-01-01

    With an ever increasing population rate; a vast array of biomass wastes rich in organic and inorganic nutrients as well as pathogenic microorganisms will result from the diversified human, industrial and agricultural activities. Anaerobic digestion is applauded as one of the best ways to properly handle and manage these wastes. Animal wastes have been recognized as suitable substrates for anaerobic digestion process, a natural biological process in which complex organic materials are broken down into simpler molecules in the absence of oxygen by the concerted activities of four sets of metabolically linked microorganisms. This process occurs in an airtight chamber (biodigester) via four stages represented by hydrolytic, acidogenic, acetogenic and methanogenic microorganisms. The microbial population and structure can be identified by the combined use of culture-based, microscopic and molecular techniques. Overall, the process is affected by bio-digester design, operational factors and manure characteristics. The purpose of anaerobic digestion is the production of a renewable energy source (biogas) and an odor free nutrient-rich fertilizer. Conversely, if animal wastes are accidentally found in the environment, it can cause a drastic chain of environmental and public health complications. PMID:24048207

  16. Effect of carbon monoxide, hydrogen and sulfate on thermophilic (55 degrees C) hydrogenogenic carbon monoxide conversion in two anaerobic bioreactor sludges.

    PubMed

    Sipma, J; Meulepas, R J W; Parshina, S N; Stams, A J M; Lettinga, G; Lens, P N L

    2004-04-01

    The conversion routes of carbon monoxide (CO) at 55 degrees C by full-scale grown anaerobic sludges treating paper mill and distillery wastewater were elucidated. Inhibition experiments with 2-bromoethanesulfonate (BES) and vancomycin showed that CO conversion was performed by a hydrogenogenic population and that its products, i.e. hydrogen and CO2, were subsequently used by methanogens, homo-acetogens or sulfate reducers depending on the sludge source and inhibitors supplied. Direct methanogenic CO conversion occurred only at low CO concentrations [partial pressure of CO (PCO) <0.5 bar (1 bar=10(5) Pa)] with the paper mill sludge. The presence of hydrogen decreased the CO conversion rates, but did not prevent the depletion of CO to undetectable levels (<400 ppm). Both sludges showed interesting potential for hydrogen production from CO, especially since after 30 min exposure to 95 degrees C, the production of CH4 at 55 degrees C was negligible. The paper mill sludge was capable of sulfate reduction with hydrogen, tolerating and using high CO concentrations (PCO>1.6 bar), indicating that CO-rich synthesis gas can be used efficiently as an electron donor for biological sulfate reduction.

  17. Gas Fermentation-A Flexible Platform for Commercial Scale Production of Low-Carbon-Fuels and Chemicals from Waste and Renewable Feedstocks.

    PubMed

    Liew, FungMin; Martin, Michael E; Tappel, Ryan C; Heijstra, Björn D; Mihalcea, Christophe; Köpke, Michael

    2016-01-01

    There is an immediate need to drastically reduce the emissions associated with global fossil fuel consumption in order to limit climate change. However, carbon-based materials, chemicals, and transportation fuels are predominantly made from fossil sources and currently there is no alternative source available to adequately displace them. Gas-fermenting microorganisms that fix carbon dioxide (CO2) and carbon monoxide (CO) can break this dependence as they are capable of converting gaseous carbon to fuels and chemicals. As such, the technology can utilize a wide range of feedstocks including gasified organic matter of any sort (e.g., municipal solid waste, industrial waste, biomass, and agricultural waste residues) or industrial off-gases (e.g., from steel mills or processing plants). Gas fermentation has matured to the point that large-scale production of ethanol from gas has been demonstrated by two companies. This review gives an overview of the gas fermentation process, focusing specifically on anaerobic acetogens. Applications of synthetic biology and coupling gas fermentation to additional processes are discussed in detail. Both of these strategies, demonstrated at bench-scale, have abundant potential to rapidly expand the commercial product spectrum of gas fermentation and further improve efficiencies and yields.

  18. Formyltetrahydrofolate Synthetase Gene Diversity in the Guts of Higher Termites with Different Diets and Lifestyles ▿ †

    PubMed Central

    Ottesen, Elizabeth A.; Leadbetter, Jared R.

    2011-01-01

    In this study, we examine gene diversity for formyl-tetrahydrofolate synthetase (FTHFS), a key enzyme in homoacetogenesis, recovered from the gut microbiota of six species of higher termites. The “higher” termites (family Termitidae), which represent the majority of extant termite species and genera, engage in a broader diversity of feeding and nesting styles than the “lower” termites. Previous studies of termite gut homoacetogenesis have focused on wood-feeding lower termites, from which the preponderance of FTHFS sequences recovered were related to those from acetogenic treponemes. While sequences belonging to this group were present in the guts of all six higher termites examined, treponeme-like FTHFS sequences represented the majority of recovered sequences in only two species (a wood-feeding Nasutitermes sp. and a palm-feeding Microcerotermes sp.). The remaining four termite species analyzed (a Gnathamitermes sp. and two Amitermes spp. that were recovered from subterranean nests with indeterminate feeding strategies and a litter-feeding Rhynchotermes sp.) yielded novel FTHFS clades not observed in lower termites. These termites yielded two distinct clusters of probable purinolytic Firmicutes and a large group of potential homoacetogens related to sequences previously recovered from the guts of omnivorous cockroaches. These findings suggest that the gut environments of different higher termite species may select for different groups of homoacetogens, with some species hosting treponeme-dominated homoacetogen populations similar to those of wood-feeding, lower termites while others host Firmicutes-dominated communities more similar to those of omnivorous cockroaches. PMID:21441328

  19. Microbial methane formation in deep aquifers of a coal-bearing sedimentary basin, Germany

    PubMed Central

    Gründger, Friederike; Jiménez, Núria; Thielemann, Thomas; Straaten, Nontje; Lüders, Tillmann; Richnow, Hans-Hermann; Krüger, Martin

    2015-01-01

    Coal-bearing sediments are major reservoirs of organic matter potentially available for methanogenic subsurface microbial communities. In this study the specific microbial community inside lignite-bearing sedimentary basin in Germany and its contribution to methanogenic hydrocarbon degradation processes was investigated. The stable isotope signature of methane measured in groundwater and coal-rich sediment samples indicated methanogenic activity. Analysis of 16S rRNA gene sequences showed the presence of methanogenic Archaea, predominantly belonging to the orders Methanosarcinales and Methanomicrobiales, capable of acetoclastic or hydrogenotrophic methanogenesis. Furthermore, we identified fermenting, sulfate-, nitrate-, and metal-reducing, or acetogenic Bacteria clustering within the phyla Proteobacteria, complemented by members of the classes Actinobacteria, and Clostridia. The indigenous microbial communities found in the groundwater as well as in the coal-rich sediments are able to degrade coal-derived organic components and to produce methane as the final product. Lignite-bearing sediments may be an important nutrient and energy source influencing larger compartments via groundwater transport. PMID:25852663

  20. Interaction of organic contaminants with natural clay type geosorbents: potential use as geologic barrier in urban landfill.

    PubMed

    Sánchez-Jiménez, N; Sevilla, M T; Cuevas, J; Rodríguez, M; Procopio, J R

    2012-03-01

    The aim of this work is to characterize the capability of several clay materials as preservative of organic pollution for use as landfill barrier. Interaction of representative organic pollutants with different polarity and water solubility (atrazine, benzamide, methomyl, paraquat and toluene) with several clay materials coming from several locations of Spain were studied. Batch suspension method was used to study the pesticide adsorption onto the clay sorbents in solution conditions that simulate the composition of a young leachate in its aerobic acetogenic stage (pH=5 and I=0.15) The obtained data of the analytes sorption were modelized by several sorption isotherm models, and the best fitted data were got with a generalized Langmuir adsorption isotherm. The higher maxima adsorptions were observed for paraquat (50-62 mmol kg(-1)) and toluene (19-34 mmol kg(-1)) whereas more hydrophobic compounds present lower adsorption (0.7-2.5 mmol kg(-1)). Paraquat is the compound that presents the higher bonding coefficients. Therefore these clays could be used as components of the multibarriers in controlled urban landfill. Copyright © 2011 Elsevier Ltd. All rights reserved.

  1. Control of amphibious weed ipomoea (Ipomoea carnea) by utilizing it for the extraction of volatile fatty acids as energy precursors

    PubMed Central

    Rafiq Kumar, M.; Tauseef, S.M.; Abbasi, Tasneem; Abbasi, S.A.

    2014-01-01

    Volatile fatty acids (VFAs), comprising mainly of acetic acid and lesser quantities of propionic and butyric acids, are generated when zoomass or phytomass is acted upon by acidogenic and acetogenic microorganisms. VFAs can be utilized by methanogens under anaerobic conditions to generate flammable methane–carbon dioxide mixtures known as ‘biogas’. Acting on the premise that this manner of VFA utilization for generating relatively clean energy can be easily accomplished in a controlled fashion in conventional biogas plants as well as higher-rate anaerobic digesters, we have carried out studies aimed to generate VFAs from the pernicious weed ipomoea (Ipomoea carnea). The VFA extraction was accomplished by a simple yet effective technology, appropriate for use even by laypersons. For this acid-phase reactors were set, to which measured quantities of ipomoea leaves were charged along with water inoculated with cow dung. The reactors were stirred intermittently. It was found that VFA production started within hours of the mixing of the reactants and peaked by the 10th or 11th day in all the reactors, effecting a conversion of over 10% of the biomass into VFAs. The reactor performance had good reproducibility and the process appeared easily controllable, frugal and robust. PMID:25685545

  2. Biomass adaptation over anaerobic co-digestion of sewage sludge and trapped grease waste.

    PubMed

    Silvestre, G; Rodríguez-Abalde, A; Fernández, B; Flotats, X; Bonmatí, A

    2011-07-01

    The feasibility of sewage sludge co-digestion using intermediate waste generated inside a wastewater treatment plant, i.e. trapped grease waste from the dissolved air flotation unit, has been assessed in a continuous stirred lab reactor operating at 35°C with a hydraulic retention time of 20 days. Three different periods of co-digestion were carried out as the grease waste dose was increased. When the grease waste addition was 23% of the volatile solids fed (organic loading rate 3.0 kg(COD)m(-3)d(-1)), an increase in methane yield of 138% was reported. Specific activity tests suggested that anaerobic biomass had adapted to the co-substrate. The adapted inoculum showed higher acetoclastic methanogenic and β-oxidation synthrophic acetogenic activities but lower hydrogenotrophic methanogenic activity. The results indicate that a slow increase in the grease waste dose could be a strategy that favours biomass acclimation to fat-rich co-substrate, increases long chain fatty acid degradation and reduces the latter's inhibitory effect.

  3. Microbial anaerobic digestion (bio-digesters) as an approach to the decontamination of animal wastes in pollution control and the generation of renewable energy.

    PubMed

    Manyi-Loh, Christy E; Mamphweli, Sampson N; Meyer, Edson L; Okoh, Anthony I; Makaka, Golden; Simon, Michael

    2013-09-17

    With an ever increasing population rate; a vast array of biomass wastes rich in organic and inorganic nutrients as well as pathogenic microorganisms will result from the diversified human, industrial and agricultural activities. Anaerobic digestion is applauded as one of the best ways to properly handle and manage these wastes. Animal wastes have been recognized as suitable substrates for anaerobic digestion process, a natural biological process in which complex organic materials are broken down into simpler molecules in the absence of oxygen by the concerted activities of four sets of metabolically linked microorganisms. This process occurs in an airtight chamber (biodigester) via four stages represented by hydrolytic, acidogenic, acetogenic and methanogenic microorganisms. The microbial population and structure can be identified by the combined use of culture-based, microscopic and molecular techniques. Overall, the process is affected by bio-digester design, operational factors and manure characteristics. The purpose of anaerobic digestion is the production of a renewable energy source (biogas) and an odor free nutrient-rich fertilizer. Conversely, if animal wastes are accidentally found in the environment, it can cause a drastic chain of environmental and public health complications.

  4. Contributions of available substrates and activities of trophic microbial community to methanogenesis in vegetative and reproductive rice rhizospheric soil.

    PubMed

    Chawanakul, Sansanee; Chaiprasert, Pawinee; Towprayoon, Sirintornthep; Tanticharoen, Morakot

    2009-01-01

    Potential of methane production and trophic microbial activities at rhizospheric soil during rice cv. Supanbunri 1 cultivation were determined by laboratory anaerobic diluents vials. The methane production was higher from rhizospheric than non-rhizospheric soil, with the noticeable peaks during reproductive phase (RP) than vegetative phase (VP). Glucose, ethanol and acetate were the dominant available substrates found in rhizospheric soil during methane production at both phases. The predominance activities of trophic microbial consortium in methanogenesis, namely fermentative bacteria (FB), acetogenic bacteria (AGB), acetate utilizing bacteria (AB) and acetoclastic methanogens (AM) were also determined. At RP, these microbial groups were enhanced in the higher of methane production than VP. This correlates with our finding that methane production was greater at the rhizospheric soil with the noticeable peaks during RP (1,150 +/- 60 nmol g dw(-1) d(-1)) compared with VP (510 +/- 30 nmol g dw(-1) d(-1)). The high number of AM showed the abundant (1.1x10(4) cell g dw(-1)) with its high activity at RP, compared to the less activity with AM number at VP (9.8x10(2) cell g dw(-1)). Levels of AM are low in the total microbial population, being less than 1% of AB. These evidences revealed that the microbial consortium of these two phases were different.

  5. Molecular monitoring of culturable bacteria from deep-sea sediment of the Nankai Trough, Leg 190 Ocean Drilling Program.

    PubMed

    Toffin, Laurent; Webster, Gordon; Weightman, Andrew J; Fry, John C; Prieur, Daniel

    2004-06-01

    Culturable bacteria were detected in deep-sea sediment samples collected from the Nankai Trough site 1173 (Ocean Drilling Program, ODP, Leg 190) at 4.15 m below the seafloor with 4791 m of overlying water. In this deep ocean near surface sediment, mainly fermentative heterotrophs, autotrophic acetogens and sulfate-reducing bacteria were enriched by using two different non-selective enrichment culture media. Culturable bacterial population shifts within the deep marine sediment enrichments were monitored by using denaturating gradient gel electrophoresis (DGGE). DGGE analysis revealed a decrease in the number of 16S rRNA gene fragments from high to low carbon concentrations, and from low to high dilution of inoculum, suggesting that fast-growing bacteria were numerically dominant in enrichment culture samples. The dominant 16S rRNA fragments observed in DGGE gels were assigned to the Firmicutes, Proteobacteria (gamma and delta subgroups) and Spirochaeta phyla. Continual sub-culture and purification resulted in two isolates which were phylogenetically identified as members of the genera Acetobacterium and Marinilactibacillus. Our results, which combine enrichment culturing with DGGE analysis, indicated that enrichment cultures derived from inoculum dilution and media with various concentrations of carbon could facilitate the detection and isolation of a greater number of environmentally relevant bacterial species than when using traditional enrichment techniques alone.

  6. CO2 fixation by anaerobic non-photosynthetic mixotrophy for improved carbon conversion

    PubMed Central

    Jones, Shawn W.; Fast, Alan G.; Carlson, Ellinor D.; Wiedel, Carrissa A.; Au, Jennifer; Antoniewicz, Maciek R.; Papoutsakis, Eleftherios T.; Tracy, Bryan P.

    2016-01-01

    Maximizing the conversion of biogenic carbon feedstocks into chemicals and fuels is essential for fermentation processes as feedstock costs and processing is commonly the greatest operating expense. Unfortunately, for most fermentations, over one-third of sugar carbon is lost to CO2 due to the decarboxylation of pyruvate to acetyl-CoA and limitations in the reducing power of the bio-feedstock. Here we show that anaerobic, non-photosynthetic mixotrophy, defined as the concurrent utilization of organic (for example, sugars) and inorganic (for example, CO2) substrates in a single organism, can overcome these constraints to increase product yields and reduce overall CO2 emissions. As a proof-of-concept, Clostridium ljungdahlii was engineered to produce acetone and achieved a mass yield 138% of the previous theoretical maximum using a high cell density continuous fermentation process. In addition, when enough reductant (that is, H2) is provided, the fermentation emits no CO2. Finally, we show that mixotrophy is a general trait among acetogens. PMID:27687501

  7. Genome analysis of Desulfotomaculum gibsoniae strain Groll(T) a highly versatile Gram-positive sulfate-reducing bacterium.

    PubMed

    Kuever, Jan; Visser, Michael; Loeffler, Claudia; Boll, Matthias; Worm, Petra; Sousa, Diana Z; Plugge, Caroline M; Schaap, Peter J; Muyzer, Gerard; Pereira, Ines A C; Parshina, Sofiya N; Goodwin, Lynne A; Kyrpides, Nikos C; Detter, Janine; Woyke, Tanja; Chain, Patrick; Davenport, Karen W; Rohde, Manfred; Spring, Stefan; Klenk, Hans-Peter; Stams, Alfons J M

    2014-06-15

    Desulfotomaculum gibsoniae is a mesophilic member of the polyphyletic spore-forming genus Desulfotomaculum within the family Peptococcaceae. This bacterium was isolated from a freshwater ditch and is of interest because it can grow with a large variety of organic substrates, in particular several aromatic compounds, short-chain and medium-chain fatty acids, which are degraded completely to carbon dioxide coupled to the reduction of sulfate. It can grow autotrophically with H2 + CO2 and sulfate and slowly acetogenically with H2 + CO2, formate or methoxylated aromatic compounds in the absence of sulfate. It does not require any vitamins for growth. Here, we describe the features of D. gibsoniae strain Groll(T) together with the genome sequence and annotation. The chromosome has 4,855,529 bp organized in one circular contig and is the largest genome of all sequenced Desulfotomaculum spp. to date. A total of 4,666 candidate protein-encoding genes and 96 RNA genes were identified. Genes of the acetyl-CoA pathway, possibly involved in heterotrophic growth and in CO2 fixation during autotrophic growth, are present. The genome contains a large set of genes for the anaerobic transformation and degradation of aromatic compounds, which are lacking in the other sequenced Desulfotomaculum genomes.

  8. Effects of octahedral molecular sieve on treatment performance, microbial metabolism, and microbial community in expanded granular sludge bed reactor.

    PubMed

    Pan, Fei; Xu, Aihua; Xia, Dongsheng; Yu, Yang; Chen, Guo; Meyer, Melissa; Zhao, Dongye; Huang, Ching-Hua; Wu, Qihang; Fu, Jie

    2015-12-15

    This study evaluated the effects of synthesized octahedral molecular sieve (OMS-2) nanoparticles on the anaerobic microbial community in a model digester, expanded granular sludge bed (EGSB) reactor. The addition of OMS-2 (0.025 g/L) in the EGSB reactors resulted in an enhanced operational performance, i.e., COD removal and biogas production increased by 4% and 11% respectively, and effluent volatile fatty acid (VFA) decreased by 11% relative to the control group. The Biolog EcoPlate™ test was employed to investigate microbial metabolism in the EGSB reactors. Results showed that OMS-2 not only increased the microbial metabolic level but also significantly changed the community level physiological profiling of the microorganisms. The Illumina MiSeq high-throughput sequencing of 16S rRNA gene indicated OMS-2 enhanced the microbial diversity and altered the community structure. The largest bacterial genus Lactococcus, a lactic acid bacterium, reduced from 29.3% to 20.4% by abundance in the presence of 0.25 g/L OMS-2, which may be conducive to decreasing the VFA production and increasing the microbial diversity. OMS-2 also increased the quantities of acetogenic bacteria and Archaea, and promoted the acetogenesis and methanogenesis. The X-ray photoelectron spectroscopy illustrated that Mn(IV)/Mn(III) with high redox potential in OMS-2 were reduced to Mn(II) in the EGSB reactors; this in turn affected the microbial community. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Ammonia threshold for inhibition of anaerobic digestion of thin stillage and the importance of organic loading rate.

    PubMed

    Moestedt, Jan; Müller, Bettina; Westerholm, Maria; Schnürer, Anna

    2016-03-01

    Biogas production from nitrogen-rich feedstock results in release of ammonia (NH3), causing inhibition of the microbial process. The reported threshold ammonia value for stable biogas production varies greatly between studies, probably because of differences in operating conditions. Moreover, it is often difficult to separate the effect of ammonia inhibition from that of organic loading rate (OLR), as these two factors are often interrelated. This study attempted to distinguish the effects of ammonia and OLR by analysis of two laboratory-scale biogas reactors operating with thin stillage and subjected to an increase in free ammonia (from 0.30 to 1.1 g L(-1)) either by addition of an external nitrogen source (urea) or by increasing the OLR (3.2-6.0 g volatile solids L(-1) d(-1)). The results showed that ammonia concentration was detrimental for process performance, with the threshold for stability in both processes identified as being about 1 g NH3-N L(-1), irrespective of OLR. Analysis of the methanogenic community showed limited differences between the two reactors on order level and a clear increase in the abundance of Methanomicrobiales, particularly Methanoculleus sp., in response to increasing ammonia concentration. Further comprehensive molecular analysis revealed that diverse Methanoculleus species dominated in the reactors at a given ammonia level at different OLR. The acetogenic community was clearly affected by both ammonia concentration and OLR, suggesting that the volatile fatty acid load in relation to the higher OLR was important for the dynamics of this community.

  10. Changes in microbial community during hydrogen and methane production in two-stage thermophilic anaerobic co-digestion process from biowaste.

    PubMed

    Zahedi, S; Solera, R; Micolucci, F; Cavinato, C; Bolzonella, D

    2016-03-01

    In this paper, the microbial community in a two-phase thermophilic anaerobic co-digestion process was investigated for its role in hydrogen and methane production, treating waste activated sludge and treating the organic fraction of municipal solid waste. In the acidogenic phase, in which hydrogen is produced, Clostridium sp. clusters represented 76% of total Firmicutes. When feeding the acidogenic effluent into the methanogenic reactors, these acidic conditions negatively influenced methanogenic microorganisms: Methanosaeta sp., (Methanobacteriales, Methanomicrobiales, Methanococcales) decreased by 75%, 50%, 38% and 52%, respectively. At the same time, methanogenic digestion lowered the numbers of Clostridium sp. clusters due to both pH increasing and substrate reduction, and an increase in both Firmicutes genera (non Clostridium) and methanogenic microorganisms, especially Methanosaeta sp. (208%). This was in accordance with the observed decrease in acetic (98%) and butyric (100%) acid contents. To ensure the activity of the acetate-utilizing methanogens (AUM) and the acetogens, high ratios of H2-utilizing methanogens (HUM)/AUM (3.6) were required.

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

    PubMed Central

    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

  12. Analysis of the Microbial Community in an Acidic Hollow-Fiber Membrane Biofilm Reactor (Hf-MBfR) Used for the Biological Conversion of Carbon Dioxide to Methane

    PubMed Central

    Jeon, Byoung Seung; Choi, Okkyoung; Kim, Hyun Wook; Um, Youngsoon; Lee, Dong-Hoon; Sang, Byoung-In

    2015-01-01

    Hydrogenotrophic methanogens can use gaseous substrates, such as H2 and CO2, in CH4 production. H2 gas is used to reduce CO2. We have successfully operated a hollow-fiber membrane biofilm reactor (Hf-MBfR) for stable and continuous CH4 production from CO2 and H2. CO2 and H2 were diffused into the culture medium through the membrane without bubble formation in the Hf-MBfR, which was operated at pH 4.5–5.5 over 70 days. Focusing on the presence of hydrogenotrophic methanogens, we analyzed the structure of the microbial community in the reactor. Denaturing gradient gel electrophoresis (DGGE) was conducted with bacterial and archaeal 16S rDNA primers. Real-time qPCR was used to track changes in the community composition of methanogens over the course of operation. Finally, the microbial community and its diversity at the time of maximum CH4 production were analyzed by pyrosequencing methods. Genus Methanobacterium, related to hydrogenotrophic methanogens, dominated the microbial community, but acetate consumption by bacteria, such as unclassified Clostridium sp., restricted the development of acetoclastic methanogens in the acidic CH4 production process. The results show that acidic operation of a CH4 production reactor without any pH adjustment inhibited acetogenic growth and enriched the hydrogenotrophic methanogens, decreasing the growth of acetoclastic methanogens. PMID:26694756

  13. Microbial dark matter ecogenomics reveals complex synergistic networks in a methanogenic bioreactor.

    PubMed

    Nobu, Masaru K; Narihiro, Takashi; Rinke, Christian; Kamagata, Yoichi; Tringe, Susannah G; Woyke, Tanja; Liu, Wen-Tso

    2015-08-01

    Ecogenomic investigation of a methanogenic bioreactor degrading terephthalate (TA) allowed elucidation of complex synergistic networks of uncultivated microorganisms, including those from candidate phyla with no cultivated representatives. Our previous metagenomic investigation proposed that Pelotomaculum and methanogens may interact with uncultivated organisms to degrade TA; however, many members of the community remained unaddressed because of past technological limitations. In further pursuit, this study employed state-of-the-art omics tools to generate draft genomes and transcriptomes for uncultivated organisms spanning 15 phyla and reports the first genomic insight into candidate phyla Atribacteria, Hydrogenedentes and Marinimicrobia in methanogenic environments. Metabolic reconstruction revealed that these organisms perform fermentative, syntrophic and acetogenic catabolism facilitated by energy conservation revolving around H2 metabolism. Several of these organisms could degrade TA catabolism by-products (acetate, butyrate and H2) and syntrophically support Pelotomaculum. Other taxa could scavenge anabolic products (protein and lipids) presumably derived from detrital biomass produced by the TA-degrading community. The protein scavengers expressed complementary metabolic pathways indicating syntrophic and fermentative step-wise protein degradation through amino acids, branched-chain fatty acids and propionate. Thus, the uncultivated organisms may interact to form an intricate syntrophy-supported food web with Pelotomaculum and methanogens to metabolize catabolic by-products and detritus, whereby facilitating holistic TA mineralization to CO2 and CH4.

  14. [Modern Approaches to the Creation of Industrial Microorganism Strains].

    PubMed

    Debabov, V G

    2015-04-01

    Microorganism producer strains are the basis of industrial biotechnology. Their properties determine the economical parameters of the production. Methods of rational design (metabolic engineering) and combinatorial methods of mutagenesis and selection (laboratory evolution, adaptive evolution, protein and genomic shuffling) are used for the construction of microorganism strains. Combination of these methods is frequently used. Modern strains usually do not contain plasmids and markers of drug resistance. All changes are introduced into the chromosome by the methods of homologous and site-specific recombination. The sum of such approaches is called recombineering. Gene expression is carried out at the optimal level under the control of promoters of a certain power (frequently regulated). Knowledge of a complete genomic sequence is almost a mandatory condition for the use of methods of metabolic engineering. Bioinformatics significantly assists in the selection of enzymes and the search for necessary genes and metabolic reactions. Measurement of metabolic fluxes largely assists in the construction of strains. The current level of science makes it possible to construct metabolic pathways de novo in strains for the production of chemicals and biofuel. Carbon dioxide has potential as a raw material for microbiological industry; therefore, the study of CO2 fixation by acetogens and electrogens is a promising direction of studies.

  15. A Bacterial Electron-bifurcating Hydrogenase*

    PubMed Central

    Schuchmann, Kai; Müller, Volker

    2012-01-01

    The Wood-Ljungdahl pathway of anaerobic CO2 fixation with hydrogen as reductant is considered a candidate for the first life-sustaining pathway on earth because it combines carbon dioxide fixation with the synthesis of ATP via a chemiosmotic mechanism. The acetogenic bacterium Acetobacterium woodii uses an ancient version of the pathway that has only one site to generate the electrochemical ion potential used to drive ATP synthesis, the ferredoxin-fueled, sodium-motive Rnf complex. However, hydrogen-based ferredoxin reduction is endergonic, and how the steep energy barrier is overcome has been an enigma for a long time. We have purified a multimeric [FeFe]-hydrogenase from A. woodii containing four subunits (HydABCD) which is predicted to have one [H]-cluster, three [2Fe2S]-, and six [4Fe4S]-clusters consistent with the experimental determination of 32 mol of Fe and 30 mol of acid-labile sulfur. The enzyme indeed catalyzed hydrogen-based ferredoxin reduction, but required NAD+ for this reaction. NAD+ was also reduced but only in the presence of ferredoxin. NAD+ and ferredoxin reduction both required flavin. Spectroscopic analyses revealed that NAD+ and ferredoxin reduction are strictly coupled and that they are reduced in a 1:1 stoichiometry. Apparently, the multimeric hydrogenase of A. woodii is a soluble energy-converting hydrogenase that uses electron bifurcation to drive the endergonic ferredoxin reduction by coupling it to the exergonic NAD+ reduction. PMID:22810230

  16. First insights into the syntrophic acetate-oxidizing bacteria – a genetic study

    PubMed Central

    Müller, Bettina; Sun, Li; Schnürer, Anna

    2013-01-01

    Syntrophic acetate-oxidizing bacteria have been identified as key organisms for efficient biogas production from protein-rich materials. They normally grow as lithotrophs or heterotrophs, producing acetate through the Wood–Ljungdahl pathway, but when growing in syntrophy with methanogens, they reportedly reverse this pathway and oxidize acetate to hydrogen and carbon dioxide. However, the biochemical and regulatory mechanisms behind the shift and the way in which the bacteria regain energy remain unknown. In a genome-walking approach, starting with degenerated primers, we identified those gene clusters in Syntrophaceticus schinkii, Clostridium ultunense, and Tepidanaerobacter acetatoxydans that comprise the formyltetrahydrofolate synthetase gene (fhs), encoding a key enzyme of the Wood–Ljungdahl pathway. We also discovered that the latter two harbor two fhs alleles. The fhs genes are phylogenetically separated and in the case of S. schinkii functionally linked to sulfate reducers. The T. acetatoxydans fhs1 cluster combines features of acetogens, sulfate reducers, and carbon monoxide oxidizers and is organized as a putative operon. The T. acetatoxydans fhs2 cluster encodes Wood–Ljungdahl pathway enzymes, which are also known to be involved in C1 carbon metabolism. Isolation of the enzymes illustrated that both formyltetrahydrofolate synthetases of T. acetatoxydans were functionally active. However, only fhs1 was expressed, confirming bidirectional usage of the pathway. PMID:23239474

  17. Microbiology and physiology of anaerobic fermentations of cellulose. Progress report

    SciTech Connect

    Peck, H.D. Jr.; Ljungdahl, L.G.

    1986-01-01

    Investigations into the biochemistry and physiology of the four major groups of microorganisms (primary, ancillary, secondary and methane bacteria) involved in the anaerobic conversion of cellulose to methane and carbon dioxide are presented. The investigations of the ancillary bacteria emphasize the isolation of new strains and increasing ethanol production with T. ethanolicus. These studies involve genetic modifications, enzymological studies on the regulation of appropriate enzymes and a study of the effect of inorganic pyrophosphate on growth and fermentation patterns. The acetogenic bacteria forming acetate from carbon dioxide were studied from the aspects of the enzymology of acetate from the standpoint from one carbon compound, bioenergetics emphasizing hydrogen metabolism and energy coupling H/sub 2/ cycling and the structure and function of electron transfer components. Research on secondary bacteria emphasizes the sulfate reducing bacteria from the aspects of H/sub 2/ cycling, specificities of electron transfer proteins and enzymes, the mechanism of bisulfite reductase and the enzymology and physiology of new genera of sulfate reducing bacteria. The biochemistry and physiology of both H/sub 2/-utilizing and acetate utilizing methanogenic are reported. The studies with H/sub 2/-utilizing methanogens stress the hydrogenase and the effect of inorganic pyrophosphate on growth. The research on the acetate-utilizing methanogens involve the bioenergetics of sulfite reduction and the mechanism of acetate formation induced by pyrophosphate. 143 refs., 15 figs., 10 tabs.

  18. Growth Inhibition of Sporomusa ovata by Incorporation of Benzimidazole Bases into Cobamides

    PubMed Central

    Mok, Kenny C.

    2013-01-01

    Phenolyl cobamides are unique members of a class of cobalt-containing cofactors that includes vitamin B12 (cobalamin). Cobamide cofactors facilitate diverse reactions in prokaryotes and eukaryotes. Phenolyl cobamides are structurally and chemically distinct from the more commonly used benzimidazolyl cobamides such as cobalamin, as the lower axial ligand is a phenolic group rather than a benzimidazole. The functional significance of this difference is not well understood. Here we show that in the bacterium Sporomusa ovata, the only organism known to synthesize phenolyl cobamides, several cobamide-dependent acetogenic metabolisms have a requirement or preference for phenolyl cobamides. The addition of benzimidazoles to S. ovata cultures results in a decrease in growth rate when grown on methanol, 3,4-dimethoxybenzoate, H2 plus CO2, or betaine. Suppression of native p-cresolyl cobamide synthesis and production of benzimidazolyl cobamides occur upon the addition of benzimidazoles, indicating that benzimidazolyl cobamides are not functionally equivalent to the phenolyl cobamide cofactors produced by S. ovata. We further show that S. ovata is capable of incorporating other phenolic compounds into cobamides that function in methanol metabolism. These results demonstrate that S. ovata can incorporate a wide range of compounds as cobamide lower ligands, despite its preference for phenolyl cobamides in the metabolism of certain energy substrates. To our knowledge, S. ovata is unique among cobamide-dependent organisms in its preferential utilization of phenolyl cobamides. PMID:23417488

  19. History of adaptation determines short-term shifts in performance and community structure of hydrogen-producing microbial communities degrading wheat straw.

    PubMed

    Valdez-Vazquez, Idania; Morales, Ana L; Escalante, Ana E

    2017-03-14

    This study addresses the question of ecological interest for the determination of structure and diversity of microbial communities that degrade lignocellulosic biomasses to produce biofuels. Two microbial consortia with different history, native of wheat straw (NWS) and from a methanogenic digester (MD) fed with cow manure, were contrasted in terms of hydrogen performance, substrate disintegration and microbial diversity. NWS outperformed the hydrogen production rate of MD. Microscopic images revealed that NWS acted on the cuticle and epidermis, generating cellulose strands with high crystallinity, while MD degraded deeper layers, equally affecting all polysaccharides. The bacterial composition markedly differed according to the inocula origin. NWS almost solely comprised hydrogen producers of the phyla Firmicutes and Proteobacteria, with 38% members of Enterococcus. After hydrogen fermentation, NWS comprised 8% Syntrophococcus, an acetogen that cleaves aryl ethers of constituent groups on the aromatic components of lignin. Conversely, MD comprised thirteen phyla, primarily including Firmicutes with H2 -producing members, and Bacteroidetes with non-H2 -producing members, which reduced the hydrogen performance. Overall, the results of this study provide clear evidence that the history of adaptation of NWS enhanced the hydrogen performance from untreated wheat straw. Further, native wheat straw communities have the potential to refine cellulose fibers and produce biofuels simultaneously.

  20. Lactococcus lactis catalyses electricity generation at microbial fuel cell anodes via excretion of a soluble quinone.

    PubMed

    Freguia, Stefano; Masuda, Masaki; Tsujimura, Seiya; Kano, Kenji

    2009-09-01

    Lactococcus lactis is a gram-positive, normally homolactic fermenter that is known to produce several kinds of membrane associated quinones, which are able to mediate electron transfer to extracellular electron acceptors such as Fe(3+), Cu(2+) and hexacyanoferrate. Here we show that this bacterium is also capable of performing extracellular electron transfer to anodes by utilizing at least two soluble redox mediators, as suggested by the two-step catalytic current developed. One of these two mediators was herein suggested to be 2-amino-3-dicarboxy-1,4-naphthoquinone (ACNQ), via evaluation of standard redox potential, ability of the bacterium to exploit the quinone when exogenously provided, as well as by high performance liquid chromatography coupled with UV spectrum analysis. During electricity generation, L. lactis slightly deviated from its normal homolactic metabolism by excreting acetate and pyruvate in stoichiometric amounts with respect to the electrical current. In this metabolism, the anode takes on the role of electron sink for acetogenic fermentation. The finding that L. lactis self-catalyses anodic electron transfer by excretion of redox mediators is remarkable as the mechanisms of extracellular electron transfer by pure cultures of gram-positive bacteria had previously never been elucidated.

  1. Compaction of forest soil by logging machinery favours occurrence of prokaryotes.

    PubMed

    Schnurr-Pütz, Silvia; Bååth, Erland; Guggenberger, Georg; Drake, Harold L; Küsel, Kirsten

    2006-12-01

    Soil compaction caused by passage of logging machinery reduces the soil air capacity. Changed abiotic factors might induce a change in the soil microbial community and favour organisms capable of tolerating anoxic conditions. The goals of this study were to resolve differences between soil microbial communities obtained from wheel-tracks (i.e. compacted) and their adjacent undisturbed sites, and to evaluate differences in potential anaerobic microbial activities of these contrasting soils. Soil samples obtained from compacted soil had a greater bulk density and a higher pH than uncompacted soil. Analyses of phospholipid fatty acids demonstrated that the eukaryotic/prokaryotic ratio in compacted soils was lower than that of uncompacted soils, suggesting that fungi were not favoured by the in situ conditions produced by compaction. Indeed, most-probable-number (MPN) estimates of nitrous oxide-producing denitrifiers, acetate- and lactate-utilizing iron and sulfate reducers, and methanogens were higher in compacted than in uncompacted soils obtained from one site that had large differences in bulk density. Compacted soils from this site yielded higher iron-reducing, sulfate-reducing and methanogenic potentials than did uncompacted soils. MPN estimates of H2-utilizing acetogens in compacted and uncompacted soils were similar. These results indicate that compaction of forest soil alters the structure and function of the soil microbial community and favours occurrence of prokaryotes.

  2. Beating the acetyl coenzyme A-pathway to the origin of life

    PubMed Central

    Nitschke, Wolfgang; Russell, Michael J.

    2013-01-01

    Attempts to draft plausible scenarios for the origin of life have in the past mainly built upon palaeogeochemical boundary conditions while, as detailed in a companion article in this issue, frequently neglecting to comply with fundamental thermodynamic laws. Even if demands from both palaeogeochemistry and thermodynamics are respected, then a plethora of strongly differing models are still conceivable. Although we have no guarantee that life at its origin necessarily resembled biology in extant organisms, we consider that the only empirical way to deduce how life may have emerged is by taking the stance of assuming continuity of biology from its inception to the present day. Building upon this conviction, we have assessed extant types of energy and carbon metabolism for their appropriateness to conditions probably pertaining in those settings of the Hadean planet that fulfil the thermodynamic requirements for life to come into being. Wood–Ljungdahl (WL) pathways leading to acetyl CoA formation are excellent candidates for such primordial metabolism. Based on a review of our present understanding of the biochemistry and biophysics of acetogenic, methanogenic and methanotrophic pathways and on a phylogenetic analysis of involved enzymes, we propose that a variant of modern methanotrophy is more likely than traditional WL systems to date back to the origin of life. The proposed model furthermore better fits basic thermodynamic demands and palaeogeochemical conditions suggested by recent results from extant alkaline hydrothermal seeps. PMID:23754811

  3. The Acimet{reg_sign} Process: An innovative approach to biogasification of municipal sludge

    SciTech Connect

    Ghosh, S.; Buoy, K.

    1993-12-31

    This paper reports the results of successful completion of an advanced anaerobic-digestion-process commercialization program supported by the County of DuPage, IL and the State of Illinois Department of Energy and Natural Resources, Springfield, IL. The project entailed anaerobic digestion of municipal sludge by pilot- and full-scale Acimet Process, which replaced an existing high-rate digestion system. The project was conducted at the Woodridge-Greenevalley wastewater treatment plant of DuPage County, IL. The Acimet Process relies on the application of two-phase anaerobic digestion for enhanced methane production and stabilization of municipal sludges, the disposal of which poses intractable problems in many municipal wastewater treatment plants. Unlike conventional anaerobic digestion processes, the Acimet System optimizes the liquefaction- acidification and acetogenic-methanogenic fermentations in separate acid- and methane-phase digesters operated at different hydraulic residence times (HRTs) to maximize feed hydrolysis and acidification, as well as biomethanation of the acidic intermediates. The Acimet System installed at the Woodridge Plant utilizes two mesophilic upflow digesters of novel design exhibiting unusually high product-formation efficiencies achieved without any mechanical mixing of the reactor contents.

  4. Biotransformation of nitrocellulose under methanogenic conditions

    SciTech Connect

    Caenepeel, B.M.; Freedman, D.; Kim, B.

    1996-11-01

    Treatment/disposal of nitrocellulose (NC) waste fines is a serious problem currently facing the US Army. For example, over 500,000 pounds of NC waste fines are generated annually at the Radford Army Ammunition Plant. While NC does not pose any human health problems, it is classified as a KO44 hazardous material because it is a highly energetic compound. Biological treatment of NC has been proposed as an economical alternative. However, evaluating the feasibility of biological treatment has been complicated by the lack of analytical methods for directly measuring NC. The purpose of this research was to adapt a cold acid digestion method for measuring the nitrogen content of NC when exposed to methanogenic and acetogenic conditions. Previous studies have indicated that NC inhibits methanogenesis and concluded that NC is not biodegradable under anaerobic conditions. The authors are examining the possibility that NC acts as a fortuitous electron acceptor. If so, the energetic content of NC will be reduced, resulting in a nonhazardous material. Results will be presented for the reduction of NC in methanol enrichment cultures developed from sewage sludge inoculum.

  5. Early Microbial Evolution: The Age of Anaerobes.

    PubMed

    Martin, William F; Sousa, Filipa L

    2015-12-18

    In this article, the term "early microbial evolution" refers to the phase of biological history from the emergence of life to the diversification of the first microbial lineages. In the modern era (since we knew about archaea), three debates have emerged on the subject that deserve discussion: (1) thermophilic origins versus mesophilic origins, (2) autotrophic origins versus heterotrophic origins, and (3) how do eukaryotes figure into early evolution. Here, we revisit those debates from the standpoint of newer data. We also consider the perhaps more pressing issue that molecular phylogenies need to recover anaerobic lineages at the base of prokaryotic trees, because O2 is a product of biological evolution; hence, the first microbes had to be anaerobes. If molecular phylogenies do not recover anaerobes basal, something is wrong. Among the anaerobes, hydrogen-dependent autotrophs--acetogens and methanogens--look like good candidates for the ancestral state of physiology in the bacteria and archaea, respectively. New trees tend to indicate that eukaryote cytosolic ribosomes branch within their archaeal homologs, not as sisters to them and, furthermore tend to root archaea within the methanogens. These are major changes in the tree of life, and open up new avenues of thought. Geochemical methane synthesis occurs as a spontaneous, abiotic exergonic reaction at hydrothermal vents. The overall similarity between that reaction and biological methanogenesis fits well with the concept of a methanogenic root for archaea and an autotrophic origin of microbial physiology. Copyright © 2016 Cold Spring Harbor Laboratory Press; all rights reserved.

  6. Control of interspecies electron flow during anaerobic digestion: significance of formate transfer versus hydrogen transfer during syntrophic methanogenesis in flocs. [Methanobacterium formicicum; Desulfovibrio vulgaris

    SciTech Connect

    Thiele, J.H.; Zeikus, J.G.

    1988-01-01

    Microbial formate production and consumption during syntrophic conversion of ethanol or lactate to methane was examined in purified flocs and digestor contents obtained from a whey-processing digestor. Formate production by digestor contents or purified digestor flocs was dependent on CO/sub 2/ and either ethanol or lactate but not H/sub 2/ gas as an electron donor. Floc preparations accumulated fourfold-higher levels of formate (40 ..mu..M) than digestor contents, and the free flora was the primary site for formate cleavage to CO/sub 2/ and H/sub 2/ (90 ..mu..M formate per h). Inhibition of methanogenesis by CHCl/sub 3/ resulted in formate accumulation and suppression of syntrophic ethanol oxidation. H/sub 2/ gas was an insignificant intermediary metabolite of syntrophic ethanol conversion by flocs, and it exogenous addition neither stimulated methanogenes nor inhibited the initial rate of ethanol oxidation. These results demonstrated that >90% of the syntrophic ethanol conversion to methane by mixed cultures containing primarily Desulfovibrio vulgaris and Methanobacterium formicicum was mediated via interspecies formate transfer and the <10% was mediated via interspecies H/sub 2/ transfer. The results are discussed in relation to biochemical thermodynamics. A model is presented which describes the dynamics of a bicarbonate-formate electron shuttle mechanism for control of carbon and electron flow during syntrophic methanogenesis and provides a novel mechanism for energy conservation by syntrophic acetogens.

  7. Zero valent iron simultaneously enhances methane production and sulfate reduction in anaerobic granular sludge reactors.

    PubMed

    Liu, Yiwen; Zhang, Yaobin; Ni, Bing-Jie

    2015-05-15

    Zero valent iron (ZVI) packed anaerobic granular sludge reactors have been developed for improved anaerobic wastewater treatment. In this work, a mathematical model is developed to describe the enhanced methane production and sulfate reduction in anaerobic granular sludge reactors with the addition of ZVI. The model is successfully calibrated and validated using long-term experimental data sets from two independent ZVI-enhanced anaerobic granular sludge reactors with different operational conditions. The model satisfactorily describes the chemical oxygen demand (COD) removal, sulfate reduction and methane production data from both systems. Results show ZVI directly promotes propionate degradation and methanogenesis to enhance methane production. Simultaneously, ZVI alleviates the inhibition of un-dissociated H2S on acetogens, methanogens and sulfate reducing bacteria (SRB) through buffering pH (Fe(0) + 2H(+) = Fe(2+) + H2) and iron sulfide precipitation, which improve the sulfate reduction capacity, especially under deterioration conditions. In addition, the enhancement of ZVI on methane production and sulfate reduction occurs mainly at relatively low COD/ [Formula: see text] ratio (e.g., 2-4.5) rather than high COD/ [Formula: see text] ratio (e.g., 16.7) compared to the reactor without ZVI addition. The model proposed in this work is expected to provide support for further development of a more efficient ZVI-based anaerobic granular system.

  8. Anaerobic O-demethylation of phenylmethylethers

    SciTech Connect

    Frazer, A.C.; Young, L.Y.

    1990-01-01

    Anaerobic O-demethylation (AOD) of phenylmethylethers is a process of both basic and applied significance. The aryl-O-methyl ethers are abundant in natural products, particularly as components of lignin. They are present as methoxylated lignin monomers in anaerobic environments and can be completely degraded there by mixed microbial populations. AOD is an essential early step in this process, and it is also a key reaction in the utilization of the O-methyl substituent as a C-one substrate by acetogens. An understanding of the AOD reaction mechanism might suggest new ways in which chemicals could be derived from lignocellulosic materials. The biochemical mechanism for the anaerobic cleavage of the aryl-O-methyl ether bond is an intriguing, but relatively unexplored process. In contrast to aerobic O-demethylating enzymes, AOD appears to involve methyl group transfer. Thus, novel biochemical information on an important biotransformation reaction will be gained from the research proposed. Recently, we have shown that AOD activity is inducible and have developed an assay for detecting AOD activity in cell-free extracts of Acetobacterium woodii. AOD activity is stimulated in vitro by the addition of ATP (1mM) and pyruvate (30 mM), the K{sub M} for vanillate being 0.4 mM. In collaboration with protein purification experts, we proposed to purify the AOD enzyme and characterize the protein(s) and the enzymatic reaction involved. 8 figs., 5 tabs.

  9. Effect of hydraulic retention time on pretreatment of blended municipal sludge.

    PubMed

    Koyunluoglu-Aynur, S; Riffat, R; Murthy, S

    2011-01-01

    The objective of the present work was to evaluate the effect of hydraulic retention time (HRT) on hydrolysis and acidogenesis for the pretreatment processes: acid phase digestion (APD) and autothermal thermophilic aerobic digestion (ATAD) using blended municipal sludge. The effect of the different pretreatment steps on mesophilic anaerobic digestion (MAD) was evaluated in terms of methane yield, keeping the operating conditions of the MAD the same for all systems. Best operating conditions for both APD and ATAD were observed for 2.5 d HRT with high total volatile fatty acids (tVFA), and the highest methane yield observed for MAD. No significant difference was observed between the two processes in terms of overall volatile solids (VS) reduction with same total HRT. The autothermal process produced heat of 14,300 J/g VS removed from hydrolytic and acetogenic reactions without compromising overall methane yields when the HRT was 2.5 d or lower and the total O2 used was 0.10 m3 O2/g VS added or lower. However, the process needs the input of oxygen and engineering analysis should balance these differences when considering the relative merits of the two pretreatment processes. This is the first study of its kind directly comparing these two viable pretreatment processes with the same sludge.

  10. Analysis of the Microbial Community in an Acidic Hollow-Fiber Membrane Biofilm Reactor (Hf-MBfR) Used for the Biological Conversion of Carbon Dioxide to Methane.

    PubMed

    Shin, Hyun Chul; Ju, Dong-Hun; Jeon, Byoung Seung; Choi, Okkyoung; Kim, Hyun Wook; Um, Youngsoon; Lee, Dong-Hoon; Sang, Byoung-In

    2015-01-01

    Hydrogenotrophic methanogens can use gaseous substrates, such as H2 and CO2, in CH4 production. H2 gas is used to reduce CO2. We have successfully operated a hollow-fiber membrane biofilm reactor (Hf-MBfR) for stable and continuous CH4 production from CO2 and H2. CO2 and H2 were diffused into the culture medium through the membrane without bubble formation in the Hf-MBfR, which was operated at pH 4.5-5.5 over 70 days. Focusing on the presence of hydrogenotrophic methanogens, we analyzed the structure of the microbial community in the reactor. Denaturing gradient gel electrophoresis (DGGE) was conducted with bacterial and archaeal 16S rDNA primers. Real-time qPCR was used to track changes in the community composition of methanogens over the course of operation. Finally, the microbial community and its diversity at the time of maximum CH4 production were analyzed by pyrosequencing methods. Genus Methanobacterium, related to hydrogenotrophic methanogens, dominated the microbial community, but acetate consumption by bacteria, such as unclassified Clostridium sp., restricted the development of acetoclastic methanogens in the acidic CH4 production process. The results show that acidic operation of a CH4 production reactor without any pH adjustment inhibited acetogenic growth and enriched the hydrogenotrophic methanogens, decreasing the growth of acetoclastic methanogens.

  11. A novel mode of lactate metabolism in strictly anaerobic bacteria.

    PubMed

    Weghoff, Marie Charlotte; Bertsch, Johannes; Müller, Volker

    2015-03-01

    Lactate is a common substrate for major groups of strictly anaerobic bacteria, but the biochemistry and bioenergetics of lactate oxidation is obscure. The high redox potential of the pyruvate/lactate pair of E0 ' = -190 mV excludes direct NAD(+) reduction (E0 ' = -320 mV). To identify the hitherto unknown electron acceptor, we have purified the lactate dehydrogenase (LDH) from the strictly anaerobic, acetogenic bacterium Acetobacterium woodii. The LDH forms a stable complex with an electron-transferring flavoprotein (Etf) that exhibited NAD(+) reduction only when reduced ferredoxin (Fd(2-) ) was present. Biochemical analyses revealed that the LDH/Etf complex of A. woodii uses flavin-based electron confurcation to drive endergonic lactate oxidation with NAD(+) as oxidant at the expense of simultaneous exergonic electron flow from reduced ferredoxin (E0 ' ≈ -500 mV) to NAD(+) according to: lactate + Fd(2-)  + 2 NAD(+)  → pyruvate + Fd + 2 NADH. The reduced Fd(2-) is regenerated from NADH by a sequence of events that involves conversion of chemical (ATP) to electrochemical ( Δ μ ˜ Na + ) and finally redox energy (Fd(2-) from NADH) via reversed electron transport catalysed by the Rnf complex. Inspection of genomes revealed that this metabolic scenario for lactate oxidation may also apply to many other anaerobes. © 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.

  12. Performance of mesophilic anaerobic granules for removal of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) from aqueous solution.

    PubMed

    An, Chun-jiang; He, Yan-ling; Huang, Guo-he; Liu, Yong-hong

    2010-07-15

    The performance of mesophilic anaerobic granules to degrade octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) was investigated under various conditions. The results of batch experiments showed that anaerobic granules were capable of removing HMX from aqueous solution with high efficiency. Both biotic and abiotic mechanisms contributed to the removal of HMX by anaerobic granules under mesophilic conditions. Adsorption appeared to play a significant role in the abiotic process. Furthermore, HMX could be biodegraded by anaerobic granules as the sole substrate. After 16 days of incubation, 99.04% and 96.42% of total HMX could be removed by 1g VSS/L acclimated and unacclimated granules, respectively. Vancomycin, an inhibitor of acetogenic bacteria, caused a significant inhibition of HMX biotransformation, while 2-bromoethanesulfonic acid, an inhibitor of methanogenic bacteria, only resulted in a slight decrease of metabolic activity. The presence of the glucose, as a suitable electron donor and carbon source, was found to enhance the degradation of HMX by anaerobic granules. Our study showed that sulfate had little adverse effects on biotransformation of HMX by anaerobic granules. However, nitrate had significant inhibitory effect on the extent of HMX removal especially in the initial period. This study offered good prospects of using high-rate anaerobic technology in the treatment of munition wastewater. 2010 Elsevier B.V. All rights reserved.

  13. Anaerobic growth of Methanosarcina acetivorans C2A on carbon monoxide: An unusual way of life for a methanogenic archaeon

    PubMed Central

    Rother, Michael; Metcalf, William W.

    2004-01-01

    All methanogenic Archaea examined to date rely on methanogenesis as their sole means of energy conservation. Among these are ones that use carbon monoxide as a growth substrate, producing methane via a pathway that involves hydrogen as an intermediate. To further examine the role of hydrogen in this process, we tested the ability of Methanosarcina acetivorans C2A, a metabolically versatile methanogen devoid of significant hydrogen metabolism, to use CO as a growth substrate. M. acetivorans grew on CO to high cell densities (≈1 × 108 per ml) with a doubling time of ≈24 h. Surprisingly, acetate and formate, rather than methane, were the major metabolic end products as shown by 13C NMR studies and enzymatic analysis of culture supernatants. Methane formation surpassed acetate/formate formation only when the cultures entered stationary growth phase, strongly suggesting that M. acetivorans conserves energy by means of this acetogenic and formigenic process. Resting cell experiments showed that methane production decreased linearly with increasing CO partial pressures, consistent with inhibition of methanogenesis by CO. Transposon-induced M. acetivorans mutants with lesions in the operon encoding phosphotransacetylase and acetate kinase failed to use either acetate or CO as growth substrates, indicating that these enzymes are required for both aceticlastic methanogenesis and carboxidotrophic acetogenesis. These findings greatly extend our concept of energy conservation and metabolic versatility in the methanogenic Archaea. PMID:15550538

  14. Ethanol and Volatile Fatty Acid Production from Lignocellulose by Clostridium cellulolyticum

    PubMed Central

    Williams, K.; Zheng, Y.; McGarvey, J.; Fan, Z.; Zhang, R.

    2013-01-01

    Clostridium cellulolyticum is capable of producing glycosyl hydrolase enzymes as well as fermentation products including ethanol and acetate. In this study, the potential of using C. cellulolyticum for ethanol and volatile fatty acid production from straw and grape pomace was examined. For rice straw, the effects of alkaline pretreatment and substrate sterilization prior to fermentation on products yields were also investigated. Effects of alkaline pretreatment and necessity for subsequent washing were tested for two types of grape pomace. For rice straw, the highest ethanol yield was 0.16 g/gVS from the straw pretreated with 10% sodium hydroxide loading at 121°C for 1 hour. Sterilization of the straw prior to fermentation was found to be not significant for ethanol production. Sterilization appeared to decrease native acetogen populations in the rice straw, resulting in lower acetic acid yields. The highest ethanol yield from grape pomace was of 0.09 g/gVS from the pretreated pomace. Pomace type (red or white) and washing were found to be not significant. Ethanol yields by C. cellulolyticum were lower than those from yeast in a simultaneous saccharification and fermentation system, but overall conversion of cellulose and hemicellulose was high, between 68 and 79%. PMID:25969767

  15. Biological conversion of methane to liquid fuels: status and opportunities.

    PubMed

    Ge, Xumeng; Yang, Liangcheng; Sheets, Johnathon P; Yu, Zhongtang; Li, Yebo

    2014-12-01

    Methane is the main component of natural gas and biogas. As an abundant energy source, methane is crucial not only to meet current energy needs but also to achieve a sustainable energy future. Conversion of methane to liquid fuels provides energy-dense products and therefore reduces costs for storage, transportation, and distribution. Compared to thermochemical processes, biological conversion has advantages such as high conversion efficiency and using environmentally friendly processes. This paper is a comprehensive review of studies on three promising groups of microorganisms (methanotrophs, ammonia-oxidizing bacteria, and acetogens) that hold potential in converting methane to liquid fuels; their habitats, biochemical conversion mechanisms, performance in liquid fuels production, and genetic modification to enhance the conversion are also discussed. To date, methane-to-methanol conversion efficiencies (moles of methanol produced per mole methane consumed) of up to 80% have been reported. A number of issues that impede scale-up of this technology, such as mass transfer limitations of methane, inhibitory effects of H2S in biogas, usage of expensive chemicals as electron donors, and lack of native strains capable of converting methane to liquid fuels other than methanol, are discussed. Future perspectives and strategies in addressing these challenges are also discussed.

  16. Formate-derived H2 , a driver of hydrogenotrophic processes in the root-zone of a methane-emitting fen.

    PubMed

    Hunger, Sindy; Schmidt, Oliver; Gößner, Anita S; Drake, Harold L

    2016-09-01

    Wetlands are important sources of globally emitted methane. Plants mediate much of that emission by releasing root-derived organic carbon, including formate, a direct precursor of methane. Thus, the objective of this study was to resolve formate-driven processes potentially linked to methanogenesis in the fen root-zone. Although, formate was anticipated to directly trigger methanogenesis, the rapid anaerobic consumption of formate by Carex roots unexpectedly yielded H2 and CO2 via enzymes such as formate-H2 -lyase (FHL), and likewise appeared to enhance the utilization of organic carbon. Collectively, 57 [FeFe]- and [NiFe]-hydrogenase-containing family level phylotypes potentially linked to FHL activity were detected. Under anoxic conditions, root-derived fermentative Citrobacter and Hafnia isolates produced H2 from formate via FHL. Formate-derived H2 fueled methanogenesis and acetogenesis, and methanogenic (Methanoregula, Methanobacterium, Methanocella) and acetogenic (Acetonema, Clostridum, Sporomusa) genera potentially linked to these hydrogenotrophic activities were identified. The findings (i) provide novel insights on highly diverse root-associated FHL-containing taxa that can augment secondary hydrogenotrophic processes via the production of formate-derived H2 , (ii) demonstrate that formate can have a 'priming' effect on the utilization of organic carbon, and (iii) raise questions regarding the fate of formate-derived H2 when it diffuses away from the root-zone.

  17. Tetrachloroethene-dehalogenating bacteria.

    PubMed

    Damborský, J

    1999-01-01

    Tetrachloroethene is a frequent groundwater contaminant often persisting in the subsurface environments. It is recalcitrant under aerobic conditions because it is in a highly oxidized state and is not readily susceptible to oxidation. Nevertheless, at least 15 organisms from different metabolic groups, viz. halorespirators (9), acetogens (2), methanogens (3) and facultative anaerobes (2), that are able to metabolize tetrachloroethene have been isolated as axenic cultures to-date. Some of these organisms couple dehalo-genation to energy conservation and utilize tetrachloroethene as the only source of energy while others dehalogenate tetrachloroethene fortuitously. Halorespiring organisms (halorespirators) utilize halogenated organic compounds as electron acceptors in an anaerobic respiratory process. Different organisms exhibit differences in the final products of tetrachloroethene dehalogenation, some strains convert tetrachloroethene to trichloroethene only, while others also carry out consecutive dehalogenation to dichloroethenes and vinyl chloride. Thus far, only a single organism, 'Dehalococcoides ethenogenes' strain 195, has been isolated which dechlorinates tetrachloroethene all the way down to ethylene. The majority of tetrachloroethene-dehalogenating organisms have been isolated only in the past few years and several of them, i.e., Dehalobacter restrictus, Desulfitobacterium dehalogenans, 'Dehalococcoides ethenogenes', 'Dehalospirillum multivorans', Desulfuromonas chloroethenica, and Desulfomonile tiedjei, are representatives of new taxonomic groups. This contribution summarizes the available information regarding the axenic cultures of the tetrachloroethene-dehalogenating bacteria. The present knowledge about the isolation of these organisms, their physiological characteristics, morphology, taxonomy and their ability to dechlorinate tetrachloroethene is presented to facilitate a comprehensive comparison.

  18. Alamethicin Suppresses Methanogenesis and Promotes Acetogenesis in Bioelectrochemical Systems

    PubMed Central

    Zhu, Xiuping; Siegert, Michael; Yates, Matthew D.

    2015-01-01

    Microbial electrosynthesis (MES) systems with mixed cultures often generate a variety of gaseous and soluble chemicals. Methane is the primary end product in mixed-culture MES because it is the thermodynamically most favorable reduction product of CO2. Here, we show that the peptaibol alamethicin selectively suppressed the growth of methanogens in mixed-culture MES systems, resulting in a shift of the solution and cathode communities to an acetate-producing system dominated by Sporomusa, a known acetogenic genus in MES systems. Archaea in the methane-producing control were dominated by Methanobrevibacter species, but no Archaea were detected in the alamethicin-treated reactors. No methane was detected in the mixed-culture reactors treated with alamethicin over 10 cycles (∼3 days each). Instead, acetate was produced at an average rate of 115 nmol ml−1 day−1, similar to the rate reported previously for pure cultures of Sporomusa ovata on biocathodes. Mixed-culture control reactors without alamethicin generated methane at nearly 100% coulombic recovery, and no acetate was detected. These results show that alamethicin is effective for the suppression of methanogen growth in MES systems and that its use enables the production of industrially relevant organic compounds by the inhibition of methanogenesis. PMID:25819972

  19. Reactor performance of a 750 m(3) anaerobic digestion plant: varied substrate input conditions impacting methanogenic community.

    PubMed

    Wagner, Andreas Otto; Malin, Cornelia; Lins, Philipp; Gstraunthaler, Gudrun; Illmer, Paul

    2014-10-01

    A 750 m(3) anaerobic digester was studied over a half year period including a shift from good reactor performance to a reduced one. Various abiotic parameters like volatile fatty acids (VFA) (formic-, acetic-, propionic-, (iso-)butyric-, (iso-)valeric-, lactic acid), total C, total N, NH4 -N, and total proteins, as well as the organic matter content and dry mass were determined. In addition several process parameters such as temperature, pH, retention time and input of substrate and the concentrations of CH4, H2, CO2 and H2S within the reactor were monitored continuously. The present study aimed at the investigation of the abundance of acetogens and total cell numbers and the microbial methanogenic community as derived from PCR-dHPLC analysis in order to put it into context with the determined abiotic parameters. An influence of substrate quantity on the efficiency of the anaerobic digestion process was found as well as a shift from a hydrogenotrophic in times of good reactor performance towards an acetoclastic dominated methanogenic community in times of reduced reactor performance. After the change in substrate conditions it took the methano-archaeal community about 5-6 weeks to be affected but then changes occurred quickly.

  20. Composting potential of different inoculum sources in the modified SEBAC system treatment of municipal solid wastes.

    PubMed

    Forster-Carneiro, T; Pérez, M; Romero, L I

    2007-12-01

    The aim of the work described here was to analyse the biomethanization process for three types of organic fraction of municipal solid waste (OFMSW) using two different inoculum sources in a sequential leach-bed anaerobic reactor under thermophilic (55 degrees C) and dry conditions (20%TS). The OFMSWs studied were: separately collected food waste (SC_OFMSW) from restaurants, synthetic waste (ST_OFMSW) and mechanically selected municipal waste (MS_OFMSW). The inoculum studied were digested mesophilic sludge (SLUDGE) and digested SC_OFMSW. The results show that SLUDGE gave the best performance and this was therefore selected for the rest of the experiments. Three assays were performed to analyse the biomethanization processes of three types of organic waste. The results suggest that all three wastes give rise to an acclimation stage with acidogenic/acetogenic activity between days 5 and 20-30 and a stabilization phase associated with methanogenic activity. In conclusion, a mixture of OFMSW (regardless of its origin) and SWINE arranged in layers in the reactor, as well as SLUDGE used with an inoculum source, enhanced the fast start up phase of a modified sequential leach-bed system under dry thermophilic conditions.

  1. Sulfide-driven microbial electrosynthesis.

    PubMed

    Gong, Yanming; Ebrahim, Ali; Feist, Adam M; Embree, Mallory; Zhang, Tian; Lovley, Derek; Zengler, Karsten

    2013-01-02

    Microbial electrosynthesis, the conversion of carbon dioxide to organic molecules using electricity, has recently been demonstrated for acetogenic microorganisms, such as Sporomusa ovata. The energy for reduction of carbon dioxide originates from the hydrolysis of water on the anode, requiring a sufficiently low potential. Here we evaluate the use of sulfide as an electron source for microbial electrosynthesis. Abiotically oxidation of sulfide on the anode yields two electrons. The oxidation product, elemental sulfur, can be further oxidized to sulfate by Desulfobulbus propionicus, generating six additional electrons in the process. The eight electrons generated from the combined abiotic and biotic steps were used to reduce carbon dioxide to acetate on a graphite cathode by Sporomusa ovata at a rate of 24.8 mmol/day · m(2). Using a strain of Desulfuromonas as biocatalyst on the anode resulted in an acetate production rate of 49.9 mmol/day · m(2), with a Coulombic efficiency of over 90%. These results demonstrate that sulfide can serve effectively as an alternative electron donor for microbial electrosynthesis.

  2. Performance of different Sporomusa species for the microbial electrosynthesis of acetate from carbon dioxide.

    PubMed

    Aryal, Nabin; Tremblay, Pier-Luc; Lizak, Dawid M; Zhang, Tian

    2017-06-01

    Sporomusa ovata DSM-2662 produces high rate of acetate during microbial electrosynthesis (MES) by reducing CO2 with electrons coming from a cathode. Here, we investigated other Sporomusa for MES with cathode potential set at -690mVvsSHE to establish if this capacity is conserved among this genus and to identify more performant strains. S. ovata DSM-2663 produced acetate 1.8-fold faster than S. ovata DSM-2662. On the contrary, S. ovata DSM-3300 was 2.7-fold slower whereas Sporomusa aerivorans had no MES activity. These results indicate that MES performance varies among Sporomusa. During MES, electron transfer from cathode to microbes often occurs via H2. To establish if efficient coupling between H2 oxidation and CO2 reduction may explain why specific acetogens are more productive MES catalysts, the metabolisms of the investigated Sporomusa were characterized under H2:CO2. Results suggest that other phenotypic traits besides the capacity to oxidize H2 efficiently are involved. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Isolation of a cytochrome-deficient mutant strain of Sporomusa sphaeroides not capable of oxidizing methyl groups.

    PubMed Central

    Kamlage, B; Blaut, M

    1993-01-01

    The homoacetogenic anaerobic bacterium Sporomusa sphaeroides was mutagenized with UV light. Taking advantage of the ampicillin enrichment technique and a newly developed test for the detection of heme in bacterial colonies, the cytochrome-deficient mutant strain S. sphaeroides BK824 was isolated. In contrast to the wild type, this mutant strain failed to grow on betaine, betaine plus methanol, H2 plus CO2, and methanol plus CO2. Growth on betaine plus formate, betaine plus H2, betaine plus pyruvate, methanol plus H2 and CO2, and acetoin was not impaired. All enzymes of the Wood pathway as well as hydrogenase and carbon monoxide dehydrogenase were detectable at comparable activities in both the wild type and the cytochrome-deficient mutant. Labeling experiments with [14C]methanol demonstrated the inability of S. sphaeroides BK824 to oxidize methyl groups. The role of cytochromes in electron transport steps associated with the Wood pathway enzymes and their possible role in energy conservation during autotrophic growth in acetogens are discussed. PMID:8491723

  4. Growth inhibition of Sporomusa ovata by incorporation of benzimidazole bases into cobamides.

    PubMed

    Mok, Kenny C; Taga, Michiko E

    2013-05-01

    Phenolyl cobamides are unique members of a class of cobalt-containing cofactors that includes vitamin B12 (cobalamin). Cobamide cofactors facilitate diverse reactions in prokaryotes and eukaryotes. Phenolyl cobamides are structurally and chemically distinct from the more commonly used benzimidazolyl cobamides such as cobalamin, as the lower axial ligand is a phenolic group rather than a benzimidazole. The functional significance of this difference is not well understood. Here we show that in the bacterium Sporomusa ovata, the only organism known to synthesize phenolyl cobamides, several cobamide-dependent acetogenic metabolisms have a requirement or preference for phenolyl cobamides. The addition of benzimidazoles to S. ovata cultures results in a decrease in growth rate when grown on methanol, 3,4-dimethoxybenzoate, H2 plus CO2, or betaine. Suppression of native p-cresolyl cobamide synthesis and production of benzimidazolyl cobamides occur upon the addition of benzimidazoles, indicating that benzimidazolyl cobamides are not functionally equivalent to the phenolyl cobamide cofactors produced by S. ovata. We further show that S. ovata is capable of incorporating other phenolic compounds into cobamides that function in methanol metabolism. These results demonstrate that S. ovata can incorporate a wide range of compounds as cobamide lower ligands, despite its preference for phenolyl cobamides in the metabolism of certain energy substrates. To our knowledge, S. ovata is unique among cobamide-dependent organisms in its preferential utilization of phenolyl cobamides.

  5. Sulfide-Driven Microbial Electrosynthesis

    SciTech Connect

    Gong, YM; Ebrahim, A; Feist, AM; Embree, M; Zhang, T; Lovley, D; Zengler, K

    2013-01-01

    Microbial electrosynthesis, the conversion of carbon dioxide to organic molecules using electricity, has recently been demonstrated for acetogenic microorganisms, such as Sporomusa ovata. The energy for reduction of carbon dioxide originates from the hydrolysis of water on the anode, requiring a sufficiently low potential. Here we evaluate the use of sulfide as an electron source for microbial electrosynthesis. Abiotically oxidation of sulfide on the anode yields two electrons. The oxidation product, elemental sulfur, can be further oxidized to sulfate by Desulfobulbus propionicus, generating six additional electrons in the process. The eight electrons generated from the combined abiotic and biotic steps were used to reduce carbon dioxide to acetate on a graphite cathode by Sporomusa ovata at a rate of 24.8 mmol/day.m(2). Using a strain of Desulfuromonas as biocatalyst on the anode resulted in an acetate production rate of 49.9 mmol/day.m(2), with a Coulombic efficiency of over 90%. These results demonstrate that sulfide can serve effectively as an alternative electron donor for microbial electrosynthesis.

  6. Alamethicin suppresses methanogenesis and promotes acetogenesis in bioelectrochemical systems.

    PubMed

    Zhu, Xiuping; Siegert, Michael; Yates, Matthew D; Logan, Bruce E

    2015-06-01

    Microbial electrosynthesis (MES) systems with mixed cultures often generate a variety of gaseous and soluble chemicals. Methane is the primary end product in mixed-culture MES because it is the thermodynamically most favorable reduction product of CO2. Here, we show that the peptaibol alamethicin selectively suppressed the growth of methanogens in mixed-culture MES systems, resulting in a shift of the solution and cathode communities to an acetate-producing system dominated by Sporomusa, a known acetogenic genus in MES systems. Archaea in the methane-producing control were dominated by Methanobrevibacter species, but no Archaea were detected in the alamethicin-treated reactors. No methane was detected in the mixed-culture reactors treated with alamethicin over 10 cycles (∼ 3 days each). Instead, acetate was produced at an average rate of 115 nmol ml(-1) day(-1), similar to the rate reported previously for pure cultures of Sporomusa ovata on biocathodes. Mixed-culture control reactors without alamethicin generated methane at nearly 100% coulombic recovery, and no acetate was detected. These results show that alamethicin is effective for the suppression of methanogen growth in MES systems and that its use enables the production of industrially relevant organic compounds by the inhibition of methanogenesis. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  7. Enrichment of amino acid-oxidizing, acetate-reducing bacteria.

    PubMed

    Ato, Makoto; Ishii, Masaharu; Igarashi, Yasuo

    2014-08-01

    In anaerobic condition, amino acids are oxidatively deaminated, and decarboxylated, resulting in the production of volatile fatty acids. In this process, excess electrons are produced and their consumption is necessary for the accomplishment of amino acid degradation. In this study, we anaerobically constructed leucine-degrading enrichment cultures from three different environmental samples (compost, excess sludge, and rice field soil) in order to investigate the diversity of electron-consuming reaction coupled to amino acid oxidation. Constructed enrichment cultures oxidized leucine to isovalerate and their activities were strongly dependent on acetate. Analysis of volatile fatty acids (VFAs) profiles and community structure analysis during batch culture of each enrichment indicated that Clostridium cluster I coupled leucine oxidation to acetate reduction in the enrichment from the compost and the rice field soil. In these cases, acetate was reduced to butyrate. On the other hand, Clostridium cluster XIVb coupled leucine oxidation to acetate reduction in the enrichment from the excess sludge. In this case, acetate was reduced to propionate. To our surprise, the enrichment from rice field soil oxidized leucine even in the absence of acetate and produced butyrate. The enrichment would couple leucine oxidation to reductive butyrate synthesis from CO2. The coupling reaction would be achieved based on trophic link between hydrogenotrophic acetogenic bacteria and acetate-reducing bacteria by sequential reduction of CO2 and acetate. Our study suggests anaerobic degradation of amino acids is achieved yet-to-be described reactions.

  8. Microbial methane formation in deep aquifers of a coal-bearing sedimentary basin, Germany.

    PubMed

    Gründger, Friederike; Jiménez, Núria; Thielemann, Thomas; Straaten, Nontje; Lüders, Tillmann; Richnow, Hans-Hermann; Krüger, Martin

    2015-01-01

    Coal-bearing sediments are major reservoirs of organic matter potentially available for methanogenic subsurface microbial communities. In this study the specific microbial community inside lignite-bearing sedimentary basin in Germany and its contribution to methanogenic hydrocarbon degradation processes was investigated. The stable isotope signature of methane measured in groundwater and coal-rich sediment samples indicated methanogenic activity. Analysis of 16S rRNA gene sequences showed the presence of methanogenic Archaea, predominantly belonging to the orders Methanosarcinales and Methanomicrobiales, capable of acetoclastic or hydrogenotrophic methanogenesis. Furthermore, we identified fermenting, sulfate-, nitrate-, and metal-reducing, or acetogenic Bacteria clustering within the phyla Proteobacteria, complemented by members of the classes Actinobacteria, and Clostridia. The indigenous microbial communities found in the groundwater as well as in the coal-rich sediments are able to degrade coal-derived organic components and to produce methane as the final product. Lignite-bearing sediments may be an important nutrient and energy source influencing larger compartments via groundwater transport.

  9. Archaea and Bacteria Acclimate to High Total Ammonia in a Methanogenic Reactor Treating Swine Waste

    PubMed Central

    Parameswaran, Prathap; Rittmann, Bruce E.

    2016-01-01

    Inhibition by ammonium at concentrations above 1000 mgN/L is known to harm the methanogenesis phase of anaerobic digestion. We anaerobically digested swine waste and achieved steady state COD-removal efficiency of around 52% with no fatty-acid or H2 accumulation. As the anaerobic microbial community adapted to the gradual increase of total ammonia-N (NH3-N) from 890 ± 295 to 2040 ± 30 mg/L, the Bacterial and Archaeal communities became less diverse. Phylotypes most closely related to hydrogenotrophic Methanoculleus (36.4%) and Methanobrevibacter (11.6%), along with acetoclastic Methanosaeta (29.3%), became the most abundant Archaeal sequences during acclimation. This was accompanied by a sharp increase in the relative abundances of phylotypes most closely related to acetogens and fatty-acid producers (Clostridium, Coprococcus, and Sphaerochaeta) and syntrophic fatty-acid Bacteria (Syntrophomonas, Clostridium, Clostridiaceae species, and Cloacamonaceae species) that have metabolic capabilities for butyrate and propionate fermentation, as well as for reverse acetogenesis. Our results provide evidence countering a prevailing theory that acetoclastic methanogens are selectively inhibited when the total ammonia-N concentration is greater than ~1000 mgN/L. Instead, acetoclastic and hydrogenotrophic methanogens coexisted in the presence of total ammonia-N of ~2000 mgN/L by establishing syntrophic relationships with fatty-acid fermenters, as well as homoacetogens able to carry out forward and reverse acetogenesis. PMID:27725793

  10. Anaerobes: a piece in the puzzle for alternative biofuels.

    PubMed

    Lawson, Paul A; Allen, Toby D; Caldwell, Matthew E; Tanner, Ralph S

    2011-08-01

    Although much newsprint is devoted to the subject of reducing the United States and other major developed countries dependence on their respective foreign energy sources; the most challenging issues for society is to provide long-term, sustainable energy sources to accommodate the global population as a whole. The projected population of planet Earth for the year 2050 is estimated to be in excess of 9 billion. With hydrocarbon-based energy becoming limiting it is unlikely that one type of energy will alone replace our dependence on this source. So-called "green" technologies that include solar, wind and wave powers are now being explored to reduce on traditional hydrocarbon-based fuel sources. The diverse and functional properties of microbes, and in particular anaerobes, are now being utilized in the production of biofuels and may provide one piece of the jigsaw for future energy requirements. Here we present some results of a screening program to identify and characterize a number of carbon monoxide oxidizing, ethanol-producing acetogenic anaerobes phylogenetically located within the Clostridiales.

  11. Autotrophic acetyl coenzyme A biosynthesis in Methanococcus maripaludis.

    PubMed Central

    Shieh, J; Whitman, W B

    1988-01-01

    To detect autotrophic CO2 assimilation in cell extracts of Methanococcus maripaludis, lactate dehydrogenase and NADH were added to convert pyruvate formed from autotrophically synthesized acetyl coenzyme A to lactate. The lactate produced was determined spectrophotometrically. When CO2 fixation was pulled in the direction of lactate synthesis, CO2 reduction to methane was inhibited. Bromoethanesulfonate (BES), a potent inhibitor of methanogenesis, enhanced lactate synthesis, and methyl coenzyme M inhibited it in the absence of BES. Lactate synthesis was dependent on CO2 and H2, but H2 + CO2-independent synthesis was also observed. In cell extracts, the rate of lactate synthesis was about 1.2 nmol min-1 mg of protein-1. When BES was added, the rate of lactate synthesis increased to 2.3 nmol min-1 mg of protein-1. Because acetyl coenzyme A did not stimulate lactate synthesis, pyruvate synthase may have been the limiting activity in these assays. Radiolabel from 14CO2 was incorporated into lactate. The percentages of radiolabel in the C-1, C-2, and C-3 positions of lactate were 73, 33, and 11%, respectively. Both carbon monoxide and formaldehyde stimulated lactate synthesis. 14CH2O was specifically incorporated into the C-3 of lactate, and 14CO was incorporated into the C-1 and C-2 positions. Low concentrations of cyanide also inhibited autotrophic growth, CO dehydrogenase activity, and autotrophic lactate synthesis. These observations are in agreement with the acetogenic pathway of autotrophic CO2 assimilation. PMID:3133359

  12. Growth of Yersinia pseudotuberculosis in human plasma: impacts on virulence and metabolic gene expression

    PubMed Central

    Rosso, Marie-Laure; Chauvaux, Sylvie; Dessein, Rodrigue; Laurans, Caroline; Frangeul, Lionel; Lacroix, Céline; Schiavo, Angèle; Dillies, Marie-Agnès; Foulon, Jeannine; Coppée, Jean-Yves; Médigue, Claudine; Carniel, Elisabeth; Simonet, Michel; Marceau, Michaël

    2008-01-01

    Background In man, infection by the Gram-negative enteropathogen Yersinia pseudotuberculosis is usually limited to the terminal ileum. However, in immunocompromised patients, the microorganism may disseminate from the digestive tract and thus cause a systemic infection with septicemia. Results To gain insight into the metabolic pathways and virulence factors expressed by the bacterium at the blood stage of pseudotuberculosis, we compared the overall gene transcription patterns (the transcriptome) of bacterial cells cultured in either human plasma or Luria-Bertani medium. The most marked plasma-triggered metabolic consequence in Y. pseudotuberculosis was the switch to high glucose consumption, which is reminiscent of the acetogenic pathway (known as "glucose overflow") in Escherichia coli. However, upregulation of the glyoxylate shunt enzymes suggests that (in contrast to E. coli) acetate may be further metabolized in Y. pseudotuberculosis. Our data also indicate that the bloodstream environment can regulate major virulence genes (positively or negatively); the yadA adhesin gene and most of the transcriptional units of the pYV-encoded type III secretion apparatus were found to be upregulated, whereas transcription of the pH6 antigen locus was strongly repressed. Conclusion Our results suggest that plasma growth of Y. pseudotuberculosis is responsible for major transcriptional regulatory events and prompts key metabolic reorientations within the bacterium, which may in turn have an impact on virulence. PMID:19055764

  13. A novel process simulation model (PSM) for anaerobic digestion using Aspen Plus.

    PubMed

    Rajendran, Karthik; Kankanala, Harshavardhan R; Lundin, Magnus; Taherzadeh, Mohammad J

    2014-09-01

    A novel process simulation model (PSM) was developed for biogas production in anaerobic digesters using Aspen Plus®. The PSM is a library model of anaerobic digestion, which predicts the biogas production from any substrate at any given process condition. A total of 46 reactions were used in the model, which include inhibitions, rate-kinetics, pH, ammonia, volume, loading rate, and retention time. The hydrolysis reactions were based on the extent of the reaction, while the acidogenic, acetogenic, and methanogenic reactions were based on the kinetics. The PSM was validated against a variety of lab and industrial data on anaerobic digestion. The P-value after statistical analysis was found to be 0.701, which showed that there was no significant difference between discrete validations and processing conditions. The sensitivity analysis for a ±10% change in composition of substrate and extent of reaction results in 5.285% higher value than the experimental value. The model is available at http://hdl.handle.net/2320/12358 (Rajendran et al., 2013b). Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. (Summer investigations into the isolation, cultivation and metabolism of anaerobes involved in biodegradation): Progress report, year 4, summer 1988

    SciTech Connect

    Not Available

    1988-01-01

    In the laboratory our students were trained in modern techniques for the isolation and study of a wide variety of microbes from marine and brackish environments. Special emphasis was placed on anaerobes and archaebacteria. Microbial groups that were studied included the propionic bacteria, clostridia, methanogens, acetogens, hydrogen oxidizing anaerobes and aerobes, sulfate-reducing bacteria and sulfur-reducing bacteria, anoxic photosynthetic bacteria, cyanobacteria, spirochetes, symbiotic and non-symbiotic nitrogen fixing bacteria, luminescent bacteria, iron bacteria, magnetic bacteria, and sulfur oxidizing bacteria. The permanent staff led discussions and presented lectures on the metabolism, physiology and biochemistry of the groups listed above. Material was also presented on motility and chemotaxis of bacteria, and particular emphasis was given to molecular approaches to studying evolution of bacteria. We also had five successful Microbiology Mini-symposia (see attached schedule). These one-day symposia involved lecture/seminar presentations by investigators involved in state-of-the-art working particularly exciting areas within the scope of our course.

  15. Anaerobic waste-activated sludge digestion - A bioconversion mechanism and kinetic model

    SciTech Connect

    Shimizu, Tatsuo; Kudo, Kenzo; Nasu, Yoshikazu )

    1993-05-01

    The anaerobic bioconversion of raw and mechanically lysed waste-activated sludge was kinetically investigated. The hydrolysis of the biopolymers, such as protein, which leaked out from the biological sludge with ultrasonic lysis, was a first-order reaction in anaerobic digestion and the rate constant was much higher than the decay rate constant of the raw waste activated sludge. An anaerobic digestion model that is capable of evaluating the effect of the mechanical sludge lysis on digestive performance was developed. The present model includes four major biological processes - the release of intracellular matter with sludge lysis; hydrolysis of biopolymers to volatile acids; the degradation of various volatile acids to acetate; and the conversion of acetate and hydrogen to methane. Each process was assumed to follow first-order kinetics. The model approximately simulated the overall process performance of the anaerobic digestion of waste-activated sludge. The model suggested that when the lysed waste-activated sludge was fed, the overall digestive performance remarkably increased in the two-phase system consisting of an acid forming process and a methanogenic process, which ensured the symbiotic growth of acetogenic and methanogenic bacteria.

  16. Molecular analysis of deep subsurface Cretaceous rock indicates abundant Fe(III)- and S°-reducing bacteria in a sulfate-rich environment

    SciTech Connect

    Kovacik, William P.; Takai, Ken; Mormile, Melanie R.; McKinley, James P.; Brockman, Fred J.; Fredrickson, Jim K.; Holben, William E.

    2006-01-01

    A multi-level sampler (MLS) was emplaced in a borehole straddling anaerobic, sulfate-rich Cretaceous-era shale and sandstone rock formations {approx}200 m below ground surface at Cerro Negro, New Mexico. Sterile quartzite sand contained in chambers in the sampler allowed in situ colonization and recovery of nucleic acids for molecular analyses. DGGE and 16S rRNA gene cloning results indicated a homogeneously distributed bacterial community across the shale/sandstone interface. ?-Proteobacteria sequences were common at all depths, and were dominated by members of the Geobacteraceae family (Pelobacter, Desulfuromonas, and Geobacter). Other members of this group are capable of dissimilatory Fe(III) and/or S0 reduction, but not sulfate reduction. RNA hybridization data also suggested that Fe(III)/S0 reducing bacteria were predominant. These findings are striking considering the lack of significant concentrations of these electron acceptors in this environment. The next most abundant bacterial group indicated was the sulfate reducers, including Desulfobacterium, Desulfocapsa and Desulfobulbus. Sequences related to fermenters, denitrifiers and acetogens were also recovered. The presence of a phylogenetically and functionally diverse microbial community in this deep subsurface environment likely reflects the complex nature of the primary energy and carbon sources, kerogen associated with the shale.

  17. Cultivation of high-rate sulfate reducing sludge by pH-based electron donor dosage.

    PubMed

    Paulo, Paula L; Kleerebezem, Robbert; Lettinga, Gatze; Lens, Piet N L

    2005-07-21

    A novel self-regulating bioreactor concept for sulfate reduction is proposed aiming for high biomass concentrations and treatment capacities. The system consists of a cell suspension of sulfate reducing bacteria in a continuous stirred tank reactor (30 degrees C) fed with a mixture of both electron donor and electron acceptor (formic acid and sulfuric acid, respectively), nutrients and phosphate buffer via a pH controller. The pH rise due to sulfate reduction is balanced with dosage of the sulfate reducing substrates as acids. The reactor concept was shown to be capable of full sulfate reduction without competition for the electron donor by methanogens and acetogens. Activity assays revealed that hardly any methanogenic activity on formate was left in the suspension by the end of the continuous run (130 days). In addition, the sulfidogenic activity with formate and H2/CO2 had increased, respectively, 3.9 and 11.6 times at the end of the experimental run. The evolution of the particle size distribution of the cell suspension over time indicated that newly grown cells have the tendency to attach together in flocs or to the existing agglomerates.

  18. Microbial consortium and its spatial distribution in a compartmentalized anaerobic reactor.

    PubMed

    Xing, Ya-juan; Ji, Jun-yuan; Zheng, Ping; Zhang, Ji-qiang; Ghulam, Abbas

    2014-02-01

    The compartmentalized anaerobic reactor (CAR) is a patent novel high-rate reactor, which consists of three compartments. The reactor has a great potential for application due to its many advantages. In this work, the microbial consortium, spatial distribution, and their relationship with performance of CAR were investigated by means of polymerase chain reaction, denaturing gradient gel electrophoresis, and fluorescence in situ hybridization. The results showed that the predominant archaea were Methanobacterium, Methanosaeta, and Methanospirillum, and the predominant bacteria were Firmicutes, Deltaproteobacteria, Spirochaetes, Actinobacteria, and Gammaproteobacteria in the microbial consortium. The methanogenic archaea (MA), the hydrogen-producing acetogenic bacteria (HAB), and the hydrolytic fermentative bacteria (HFB) were found to be predominant in the upper, middle, and bottom compartments, respectively. The results revealed that the granular sludge took on a stratified microbial structure. The HFB, HAB, and MA were located in the outer shell, middle layer, and core, respectively. The microbial populations from the bottom compartment were relatively homogeneous in the granular sludge, and from the middle and upper compartments, they were relatively heterogeneous in the granular sludge. The microbial consortia and their spatial distribution were in accordance with the organic loading rate and chemical components in the three compartments.

  19. Conversion of acetic acid to methane: thermophilic bacteria and their symbiotic interactions

    SciTech Connect

    Lee, M.J.

    1987-01-01

    A thermophilic (60/sup 0/C), anaerobic coculture conspired of an acetate-oxidizing eubacterial rod (AOR) and a H/sub 2/-using methanogen, Methanobacterium sp. strain THF, was studied. The AOR was isolated from the coculture by dilution into medium, with ethylene glycol. It grew on ethylene glycol, 1,2 propanediol, formate, pyruvate, glycinebetaine, and H/sub 2/-CO/sub 2/, usually forming acetate as the main product. Evidence was obtained consistent with the hypothesis that the AOR oxidized acetate via a series of reactions resembling the reversal of the acetyl CoA acetogenesis pathway. Two key enzymes were present in high activity; carbon monoxide dehydrogenase (CODH) and formate dehydrogenase (FDH), although formyltetrahydrofolate synthetase was not detectable. Tetrahydrofolate, an important C/sub 1/ carrier in acetogens, was not detectable indirectly via enzyme assays, inhibitor studies, or fluorescence spectra, suggesting an alternate carries in the AOR. Anaerobic activity stains for CODH in native polyacrylamide gels showed a novel major band in the coculture not detected in the component organisms, suggesting regulation of this enzyme and the reversal of the pathway. Enzyme assays also indicated regulate of CODH and FDH in Methanobacterium. THF.

  20. Bioremediation of chlorinated ethenes in fractured bedrock and associated changes in dechlorinating and nondechlorinating microbial populations.

    PubMed

    Pérez-de-Mora, Alfredo; Zila, Anna; McMaster, Michaye L; Edwards, Elizabeth A

    2014-05-20

    The use of enhanced in situ anaerobic bioremediation (EISB) and bioaugmentation in fractured bedrock is limited compared to its use in granular media. We evaluated EISB for the treatment of trichloroethene (TCE)-impacted groundwater in fractured carbonate rock at a site in Southern Ontario, Canada, with cool average groundwater temperature (∼ 13 °C). Borehole-connectivity, contaminant concentrations, and groundwater properties were investigated. Changes in dechlorinating and nondechlorinating populations (fermenters, acetogens, methanogens, and sulfate reducers) were assessed via quantitative PCR (qPCR). During biostimulation with ethanol, concentrations of TCE daughter products cis-dichloroethene (cDCE) and vinyl chloride (VC) decreased in association with an enrichment of vcrA (VC reductive dehalogenase)-carrying Dehalococcoides, whereas ethene production was only moderate. Following bioaugmentation with the mixed dechlorinating culture KB-1, greater concentrations of chloride-a product of dechlorination-was observed in most wells; in addition, ethene production increased significantly in monitoring well locations that had strong hydraulic connectivity to the groundwater recirculation system, while Dehalococcoides and vcrA concentrations did not appreciably vary. Interestingly, increases of 3-4 orders of magnitude of an ethanol-fermenting Bacteroidetes population also present in KB-1 were correlated to improved conversion to ethene, an observation which suggests there could be a causal relationship-for example, better syntrophy and/or synergy among bacterial populations.

  1. Physiologically anaerobic microorganisms of the deep subsurface. Final performance report, June 1, 1990--August 31, 1993

    SciTech Connect

    Stevens, S.E. Jr.; Chung, K.T.

    1993-10-01

    Anaerobic bacteria were isolated from deep subsurface sediment samples taken at study sites in Idaho (INEL) and Washington (HR) by culturing on dilute and concentrated medium. Morphologically distinct colonies were purified, and their responses to 21 selected physiological tests were determined. Although the number of isolates was small (18 INEL, 27 HR) some general patterns could be determined. Most strains could utilize all the carbon sources, however the glycerol and melizitose utilization was positive for 50% or less of the HR isolates. Catalase activity (27.78% at INEL, 74.07% at HR) and tryptophan metabolism (11.12% at INEL, 40.74% at HR) were significantly different between the two study sites. MPN and viable counts indicate that sediments near the water table yield the greatest numbers of anaerobes. Deeper sediments also appear to be more selective with the greatest number of viable counts on low-nutrient mediums. Likewise, only strictly obligate anaerobes were found in the deepest sediment samples. Selective media indicated the presence of methanogens, acetogens, and sulfate reducers at only the HR site.

  2. Microbial extracellular electron transfer and its relevance to iron corrosion.

    PubMed

    Kato, Souichiro

    2016-03-01

    Extracellular electron transfer (EET) is a microbial metabolism that enables efficient electron transfer between microbial cells and extracellular solid materials. Microorganisms harbouring EET abilities have received considerable attention for their various biotechnological applications, including bioleaching and bioelectrochemical systems. On the other hand, recent research revealed that microbial EET potentially induces corrosion of iron structures. It has been well known that corrosion of iron occurring under anoxic conditions is mostly caused by microbial activities, which is termed as microbiologically influenced corrosion (MIC). Among diverse MIC mechanisms, microbial EET activity that enhances corrosion via direct uptake of electrons from metallic iron, specifically termed as electrical MIC (EMIC), has been regarded as one of the major causative factors. The EMIC-inducing microorganisms initially identified were certain sulfate-reducing bacteria and methanogenic archaea isolated from marine environments. Subsequently, abilities to induce EMIC were also demonstrated in diverse anaerobic microorganisms in freshwater environments and oil fields, including acetogenic bacteria and nitrate-reducing bacteria. Abilities of EET and EMIC are now regarded as microbial traits more widespread among diverse microbial clades than was thought previously. In this review, basic understandings of microbial EET and recent progresses in the EMIC research are introduced.

  3. Iron corrosion activity of anaerobic hydrogen-consuming microorganisms isolated from oil facilities.

    PubMed

    Mori, Koji; Tsurumaru, Hirohito; Harayama, Shigeaki

    2010-10-01

    The purpose of the present study was to test the hypothesis that anaerobic hydrogen-consuming microorganisms generally promote iron corrosion. We isolated 26 hydrogen-consuming microorganisms (acetogens, sulfate-reducing bacteria, and methanogens) from oil facilities in Japan using hydrogen as an electron donor. The iron corrosion activities of these microorganisms were examined using iron (Fe0) granules as the sole electron donor. Almost all the isolates consumed hydrogen that was chemically generated from iron granules but did not induce significant iron corrosion. The amount of corroded iron in the cultures of these organisms was less than 2-fold that in an abiotic chemical corrosion reaction. These results indicated that hydrogen consumption did not strongly stimulate iron corrosion. On the other hand, one isolate, namely, Methanococcus maripaludis Mic1c10, considerably corroded iron: this phenomenon was not accompanied by hydrogen consumption, methane formation, or cell growth. This finding also provided strong evidence that M. maripaludis Mic1c10 produced some material that caused iron to corrode.

  4. Analyses of n-alkanes degrading community dynamics of a high-temperature methanogenic consortium enriched from production water of a petroleum reservoir by a combination of molecular techniques.

    PubMed

    Zhou, Lei; Li, Kai-Ping; Mbadinga, Serge Maurice; Yang, Shi-Zhong; Gu, Ji-Dong; Mu, Bo-Zhong

    2012-08-01

    Despite the knowledge on anaerobic degradation of hydrocarbons and signature metabolites in the oil reservoirs, little is known about the functioning microbes and the related biochemical pathways involved, especially about the methanogenic communities. In the present study, a methanogenic consortium enriched from high-temperature oil reservoir production water and incubated at 55 °C with a mixture of long chain n-alkanes (C(15)-C(20)) as the sole carbon and energy sources was characterized. Biodegradation of n-alkanes was observed as methane production in the alkanes-amended methanogenic enrichment reached 141.47 μmol above the controls after 749 days of incubation, corresponding to 17 % of the theoretical total. GC-MS analysis confirmed the presence of putative downstream metabolites probably from the anaerobic biodegradation of n-alkanes and indicating an incomplete conversion of the n-alkanes to methane. Enrichment cultures taken at different incubation times were subjected to microbial community analysis. Both 16S rRNA gene clone libraries and DGGE profiles showed that alkanes-degrading community was dynamic during incubation. The dominant bacterial species in the enrichment cultures were affiliated with Firmicutes members clustering with thermophilic syntrophic bacteria of the genera Moorella sp. and Gelria sp. Other represented within the bacterial community were members of the Leptospiraceae, Thermodesulfobiaceae, Thermotogaceae, Chloroflexi, Bacteroidetes and Candidate Division OP1. The archaeal community was predominantly represented by members of the phyla Crenarchaeota and Euryarchaeota. Corresponding sequences within the Euryarchaeota were associated with methanogens clustering with orders Methanomicrobiales, Methanosarcinales and Methanobacteriales. On the other hand, PCR amplification for detection of functional genes encoding the alkylsuccinate synthase α-subunit (assA) was positive in the enrichment cultures. Moreover, the appearance of a new ass

  5. Thermophile bacteria in permafrost: model for astrobiology

    NASA Astrophysics Data System (ADS)

    Gilichinsky, D.; Rivkina, E.; Shcherbakova, V.; Laurinavichius, K.; Kholodov, A.; Abramov, A.

    2003-04-01

    According the NASA point of view, one way to have liquid water on Mars at shallow depths would be through subglacial volcanism. Such volcano-ice interactions could be going on beneath the polar caps of Mars today, or even within the adjacent permafrost around the margins of the ice caps. This is why one of the Earth's models, close to extraterrestrial environment, represented by active volcanoes in permafrost areas and the main question is - does such econishes as volcanoes and associated environment contain recently microbial communities? The first step of this study was carried out on volcano Stromboli (Italy), using the marine water samples extracted from the borehole near the island marine coast, surrounding the volcano. According the temperatures (45^oC), this thermal water has the hydraulic connection with volcano. Microscopy analyses of studied water shown the presence of different morphological types of microorganisms: small mobile roads, coccoid and sarcina-like organisms and long fixed roads, as well as rest forms (spores and cysts). To separate this community on marine and volcano microorganisms, the common mineral media with added CO_2, acetate or glucose-peptone as a source of carbon were used for culturing, and Fe3+, S^o, SO_42- were added as a electron acceptors. We attempt to isolate thermophilic anaerobic microorganisms of different metabolic groups - methanogens, acetogens, iron-, sulfur- and sulfate-reducers, and to test each group of microorganisms on the presence of halophilic forms. After 24 hours of incubation at temperatures varied 55 to 85^o, the grow relatively the control media was observed at CO_2+H_2 and glucose-peptone media. Microscopy study of preparations showed small coccus of irregular shape that was unable to reduce S^o or SO_42-. During the subsequent re-seeding were obtained the enrichment cultures of themophilic bacteria, genetically closed to genera Thermococcus: heterotrophic, growing up to 95^oC with the growth optimum at

  6. A Genetic System for Clostridium ljungdahlii: a Chassis for Autotrophic Production of Biocommodities and a Model Homoacetogen

    PubMed Central

    Ueki, Toshiyuki; Nevin, Kelly P.; Lovley, Derek R.

    2013-01-01

    Methods for genetic manipulation of Clostridium ljungdahlii are of interest because of the potential for production of fuels and other biocommodities from carbon dioxide via microbial electrosynthesis or more traditional modes of autotrophy with hydrogen or carbon monoxide as the electron donor. Furthermore, acetogenesis plays an important role in the global carbon cycle. Gene deletion strategies required for physiological studies of C. ljungdahlii have not previously been demonstrated. An electroporation procedure for introducing plasmids was optimized, and four different replicative origins for plasmid propagation in C. ljungdahlii were identified. Chromosomal gene deletion via double-crossover homologous recombination with a suicide vector was demonstrated initially with deletion of the gene for FliA, a putative sigma factor involved in flagellar biogenesis and motility in C. ljungdahlii. Deletion of fliA yielded a strain that lacked flagella and was not motile. To evaluate the potential utility of gene deletions for functional genomic studies and to redirect carbon and electron flow, the genes for the putative bifunctional aldehyde/alcohol dehydrogenases, adhE1 and adhE2, were deleted individually or together. Deletion of adhE1, but not adhE2, diminished ethanol production with a corresponding carbon recovery in acetate. The double deletion mutant had a phenotype similar to that of the adhE1-deficient strain. Expression of adhE1 in trans partially restored the capacity for ethanol production. These results demonstrate the feasibility of genetic investigations of acetogen physiology and the potential for genetic manipulation of C. ljungdahlii to optimize autotrophic biocommodity production. PMID:23204413

  7. Syngas fermentation to biofuel: evaluation of carbon monoxide mass transfer coefficient (kLa) in different reactor configurations.

    PubMed

    Munasinghe, Pradeep Chaminda; Khanal, Samir Kumar

    2010-01-01

    Lignocellulosic biomass such as agri-residues, agri-processing by-products, and energy crops do not compete with food and feed, and is considered to be the ideal renewable feedstocks for biofuel production. Gasification of biomass produces synthesis gas (syngas), a mixture primarily consisting of CO and H(2). The produced syngas can be converted to ethanol by anaerobic microbial catalysts especially acetogenic bacteria such as various clostridia species.One of the major drawbacks associated with syngas fermentation is the mass transfer limitation of these sparingly soluble gases in the aqueous phase. One way of addressing this issue is the improvement in reactor design to achieve a higher volumetric mass transfer coefficient (k(L)a). In this study, different reactor configurations such as a column diffuser, a 20-μm bulb diffuser, gas sparger, gas sparger with mechanical mixing, air-lift reactor combined with a 20-μm bulb diffuser, air-lift reactor combined with a single gas entry point, and a submerged composite hollow fiber membrane (CHFM) module were employed to examine the k(L) a values. The k(L) a values reported in this study ranged from 0.4 to 91.08 h(-1). The highest k(L) a of 91.08 h(-1) was obtained in the air-lift reactor combined with a 20-μm bulb diffuser, whereas the reactor with the CHFM showed the lowest k(L) a of 0.4 h(-1). By considering both the k(L) a value and the statistical significance of each configuration, the air-lift reactor combined with a 20-μm bulb diffuser was found to be the ideal reactor configuration for carbon monoxide mass transfer in an aqueous phase. Copyright © 2010 American Institute of Chemical Engineers (AIChE).

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

    PubMed Central

    Ohtomo, Yoko; Ijiri, Akira; Ikegawa, Yojiro; Tsutsumi, Masazumi; Imachi, Hiroyuki; Uramoto, Go-Ichiro; Hoshino, Tatsuhiko; Morono, Yuki; Sakai, Sanae; Saito, Yumi; Tanikawa, Wataru; Hirose, Takehiro; Inagaki, Fumio

    2013-01-01

    Geological CO2 sequestration in unmineable subsurface oil/gas fields and coal formations has been proposed as a means of reducing anthropogenic greenhouse gasses in the atmosphere. However, the feasibility of injecting CO2 into subsurface depends upon a variety of geological and economic conditions, and the ecological consequences are largely unpredictable. In this study, we developed a new flow-through-type reactor system to examine potential geophysical, geochemical and microbiological impacts associated with CO2 injection by simulating in-situ pressure (0–100 MPa) and temperature (0–70°C) conditions. Using the reactor system, anaerobic artificial fluid and CO2 (flow rate: 0.002 and 0.00001 ml/min, respectively) were continuously supplemented into a column comprised of bituminous coal and sand under a pore pressure of 40 MPa (confined pressure: 41 MPa) at 40°C for 56 days. 16S rRNA gene analysis of the bacterial components showed distinct spatial separation of the predominant taxa in the coal and sand over the course of the experiment. Cultivation experiments using sub-sampled fluids revealed that some microbes survived, or were metabolically active, under CO2-rich conditions. However, no methanogens were activated during the experiment, even though hydrogenotrophic and methylotrophic methanogens were obtained from conventional batch-type cultivation at 20°C. During the reactor experiment, the acetate and methanol concentration in the fluids increased while the δ13Cacetate, H2 and CO2 concentrations decreased, indicating the occurrence of homo-acetogenesis. 16S rRNA genes of homo-acetogenic spore-forming bacteria related to the genus Sporomusa were consistently detected from the sandstone after the reactor experiment. Our results suggest that the injection of CO2 into a natural coal-sand formation preferentially stimulates homo-acetogenesis rather than methanogenesis, and that this process is accompanied by biogenic CO2 conversion to acetate. PMID

  9. Application of gas diffusion biocathode in microbial electrosynthesis from carbon dioxide.

    PubMed

    Bajracharya, Suman; Vanbroekhoven, Karolien; Buisman, Cees J N; Pant, Deepak; Strik, David P B T B

    2016-11-01

    Microbial catalysis of carbon dioxide (CO2) reduction to multi-carbon compounds at the cathode is a highly attractive application of microbial electrosynthesis (MES). The microbes reduce CO2 by either taking the electrons or reducing the equivalents produced at the cathode. While using gaseous CO2 as the carbon source, the biological reduction process depends on the dissolution and mass transfer of CO2 in the electrolyte. In order to deal with this issue, a gas diffusion electrode (GDE) was investigated by feeding CO2 through the GDE into the MES reactor for its reduction at the biocathode. A combination of the catalyst layer (porous activated carbon and Teflon binder) and the hydrophobic gas diffusion layer (GDL) creates a three-phase interface at the electrode. So, CO2 and reducing equivalents will be available to the biocatalyst on the cathode surface. An enriched inoculum consisting of acetogenic bacteria, prepared from an anaerobic sludge, was used as a biocatalyst. The cathode potential was maintained at -1.1 V vs Ag/AgCl to facilitate direct and/or hydrogen-mediated CO2 reduction. Bioelectrochemical CO2 reduction mainly produced acetate but also extended the products to ethanol and butyrate. Average acetate production rates of 32 and 61 mg/L/day, respectively, with 20 and 80 % CO2 gas mixture feed were achieved with 10 cm(2) of GDE. The maximum acetate production rate remained 238 mg/L/day for 20 % CO2 gas mixture. In conclusion, a gas diffusion biocathode supported bioelectrochemical CO2 reduction with enhanced mass transfer rate at continuous supply of gaseous CO2. Graphical abstract ᅟ.

  10. Carbon flow from volcanic CO2 into soil microbial communities of a wetland mofette

    USGS Publications Warehouse

    Beulig, Felix; Heuer, Verena B.; Akob, Denise M.; Viehweger, Bernhard; Elvert, Marcus; Herrmann, Martina; Hinrichs, Kai-Uwe; Küsel, Kirsten

    2015-01-01

    Effects of extremely high carbon dioxide (CO2) concentrations on soil microbial communities and associated processes are largely unknown. We studied a wetland area affected by spots of subcrustal CO2 degassing (mofettes) with focus on anaerobic autotrophic methanogenesis and acetogenesis because the pore gas phase was largely hypoxic. Compared with a reference soil, the mofette was more acidic (ΔpH ~0.8), strongly enriched in organic carbon (up to 10 times), and exhibited lower prokaryotic diversity. It was dominated by methanogens and subdivision 1Acidobacteria, which likely thrived under stable hypoxia and acidic pH. Anoxic incubations revealed enhanced formation of acetate and methane (CH4) from hydrogen (H2) and CO2 consistent with elevated CH4 and acetate levels in the mofette soil. 13CO2 mofette soil incubations showed high label incorporations with ~512 ng13C g (dry weight (dw)) soil−1 d−1 into the bulk soil and up to 10.7 ng 13C g (dw) soil−1 d−1 into almost all analyzed bacterial lipids. Incorporation of CO2-derived carbon into archaeal lipids was much lower and restricted to the first 10 cm of the soil. DNA-SIP analysis revealed that acidophilic methanogens affiliated withMethanoregulaceae and hitherto unknown acetogens appeared to be involved in the chemolithoautotrophic utilization of 13CO2. Subdivision 1 Acidobacteriaceae assimilated 13CO2 likely via anaplerotic reactions because Acidobacteriaceae are not known to harbor enzymatic pathways for autotrophic CO2 assimilation. We conclude that CO2-induced geochemical changes promoted anaerobic and acidophilic organisms and altered carbon turnover in affected soils.

  11. A genetic system for Clostridium ljungdahlii: a chassis for autotrophic production of biocommodities and a model homoacetogen.

    PubMed

    Leang, Ching; Ueki, Toshiyuki; Nevin, Kelly P; Lovley, Derek R

    2013-02-01

    Methods for genetic manipulation of Clostridium ljungdahlii are of interest because of the potential for production of fuels and other biocommodities from carbon dioxide via microbial electrosynthesis or more traditional modes of autotrophy with hydrogen or carbon monoxide as the electron donor. Furthermore, acetogenesis plays an important role in the global carbon cycle. Gene deletion strategies required for physiological studies of C. ljungdahlii have not previously been demonstrated. An electroporation procedure for introducing plasmids was optimized, and four different replicative origins for plasmid propagation in C. ljungdahlii were identified. Chromosomal gene deletion via double-crossover homologous recombination with a suicide vector was demonstrated initially with deletion of the gene for FliA, a putative sigma factor involved in flagellar biogenesis and motility in C. ljungdahlii. Deletion of fliA yielded a strain that lacked flagella and was not motile. To evaluate the potential utility of gene deletions for functional genomic studies and to redirect carbon and electron flow, the genes for the putative bifunctional aldehyde/alcohol dehydrogenases, adhE1 and adhE2, were deleted individually or together. Deletion of adhE1, but not adhE2, diminished ethanol production with a corresponding carbon recovery in acetate. The double deletion mutant had a phenotype similar to that of the adhE1-deficient strain. Expression of adhE1 in trans partially restored the capacity for ethanol production. These results demonstrate the feasibility of genetic investigations of acetogen physiology and the potential for genetic manipulation of C. ljungdahlii to optimize autotrophic biocommodity production.

  12. A Genetic System for Clostridium ljungdahlii: a Chassis for Autotrophic Production of Biocommodities and a Model Homoacetogen

    SciTech Connect

    Leang, C; Ueki, T; Nevin, KP; Lovley, DR

    2013-02-04

    Methods for genetic manipulation of Clostridium ljungdahlii are of interest because of the potential for production of fuels and other biocommodities from carbon dioxide via microbial electrosynthesis or more traditional modes of autotrophy with hydrogen or carbon monoxide as the electron donor. Furthermore, acetogenesis plays an important role in the global carbon cycle. Gene deletion strategies required for physiological studies of C. ljungdahlii have not previously been demonstrated. An electroporation procedure for introducing plasmids was optimized, and four different replicative origins for plasmid propagation in C. ljungdahlii were identified. Chromosomal gene deletion via double-crossover homologous recombination with a suicide vector was demonstrated initially with deletion of the gene for FliA, a putative sigma factor involved in flagellar biogenesis and motility in C. ljungdahlii. Deletion of fliA yielded a strain that lacked flagella and was not motile. To evaluate the potential utility of gene deletions for functional genomic studies and to redirect carbon and electron flow, the genes for the putative bifunctional aldehyde/alcohol dehydrogenases, adhE1 and adhE2, were deleted individually or together. Deletion of adhE1, but not adhE2, diminished ethanol production with a corresponding carbon recovery in acetate. The double deletion mutant had a phenotype similar to that of the adhE1-deficient strain. Expression of adhE1 in trans partially restored the capacity for ethanol production. These results demonstrate the feasibility of genetic investigations of acetogen physiology and the potential for genetic manipulation of C. ljungdahlii to optimize autotrophic biocommodity production.

  13. Microbial electron transport and energy conservation – the foundation for optimizing bioelectrochemical systems

    PubMed Central

    Kracke, Frauke; Vassilev, Igor; Krömer, Jens O.

    2015-01-01

    Microbial electrochemical techniques describe a variety of emerging technologies that use electrode–bacteria interactions for biotechnology applications including the production of electricity, waste and wastewater treatment, bioremediation and the production of valuable products. Central in each application is the ability of the microbial catalyst to interact with external electron acceptors and/or donors and its metabolic properties that enable the combination of electron transport and carbon metabolism. And here also lies the key challenge. A wide range of microbes has been discovered to be able to exchange electrons with solid surfaces or mediators but only a few have been studied in depth. Especially electron transfer mechanisms from cathodes towards the microbial organism are poorly understood but are essential for many applications such as microbial electrosynthesis. We analyze the different electron transport chains that nature offers for organisms such as metal respiring bacteria and acetogens, but also standard biotechnological organisms currently used in bio-production. Special focus lies on the essential connection of redox and energy metabolism, which is often ignored when studying bioelectrochemical systems. The possibility of extracellular electron exchange at different points in each organism is discussed regarding required redox potentials and effect on cellular redox and energy levels. Key compounds such as electron carriers (e.g., cytochromes, ferredoxin, quinones, flavins) are identified and analyzed regarding their possible role in electrode–microbe interactions. This work summarizes our current knowledge on electron transport processes and uses a theoretical approach to predict the impact of different modes of transfer on the energy metabolism. As such it adds an important piece of fundamental understanding of microbial electron transport possibilities to the research community and will help to optimize and advance bioelectrochemical

  14. Geochemical and hydrological constraints on the deep subsurface terrestrial ecosystems

    NASA Astrophysics Data System (ADS)

    Silver, B.; Onstott, T.; Hinton, S.; King, H.; Sherwood Lollar, B.; Lippmann-Pipke, J.

    2008-12-01

    acid utilizing metabolic reactions consistent with phylogenetic data showing few acetogens, no hydrocarbon oxidizers and a significant abundance of acetoclastic methanogens.

  15. A fast linear predictive adaptive model of packed bed coupled with UASB reactor treating onion waste to produce biofuel.

    PubMed

    Milquez-Sanabria, Harvey; Blanco-Cocom, Luis; Alzate-Gaviria, Liliana

    2016-10-03

    Agro-industrial wastes are an energy source for different industries. However, its application has not reached small industries. Previous and current research activities performed on the acidogenic phase of two-phase anaerobic digestion processes deal particularly with process optimization of the acid-phase reactors operating with a wide variety of substrates, both soluble and complex in nature. Mathematical models for anaerobic digestion have been developed to understand and improve the efficient operation of the process. At present, lineal models with the advantages of requiring less data, predicting future behavior and updating when a new set of data becomes available have been developed. The aim of this research was to contribute to the reduction of organic solid waste, generate biogas and develop a simple but accurate mathematical model to predict the behavior of the UASB reactor. The system was maintained separate for 14 days during which hydrolytic and acetogenic bacteria broke down onion waste, produced and accumulated volatile fatty acids. On this day, two reactors were coupled and the system continued for 16 days more. The biogas and methane yields and volatile solid reduction were 0.6 ± 0.05 m(3) (kg VSremoved)(-1), 0.43 ± 0.06 m(3) (kg VSremoved)(-1) and 83.5 ± 9.8 %, respectively. The model application showed a good prediction of all process parameters defined; maximum error between experimental and predicted value was 1.84 % for alkalinity profile. A linear predictive adaptive model for anaerobic digestion of onion waste in a two-stage process was determined under batch-fed condition. Organic load rate (OLR) was maintained constant for the entire operation, modifying effluent hydrolysis reactor feed to UASB reactor. This condition avoids intoxication of UASB reactor and also limits external buffer addition.

  16. Biofilm Formation by Clostridium ljungdahlii Is Induced by Sodium Chloride Stress: Experimental Evaluation and Transcriptome Analysis

    PubMed Central

    Rabaey, Korneel; Lovley, Derek R.; Vargas, Madeline

    2017-01-01

    The acetogen Clostridium ljungdahlii is capable of syngas fermentation and microbial electrosynthesis. Biofilm formation could benefit both these applications, but was not yet reported for C. ljungdahlii. Biofilm formation does not occur under standard growth conditions, but attachment or aggregation could be induced by different stresses. The strongest biofilm formation was observed with the addition of sodium chloride. After 3 days of incubation, the biomass volume attached to a plastic surface was 20 times higher with than without the addition of 200 mM NaCl to the medium. The addition of NaCl also resulted in biofilm formation on glass, graphite and glassy carbon, the latter two being often used electrode materials for microbial electrosynthesis. Biofilms were composed of extracellular proteins, polysaccharides, as well as DNA, while pilus-like appendages were observed with, but not without, the addition of NaCl. A transcriptome analysis comparing planktonic (no NaCl) and biofilm (NaCl addition) cells showed that C. ljungdahlii coped with the salt stress by the upregulation of the general stress response, Na+ export and osmoprotectant accumulation. A potential role for poly-N-acetylglucosamines and D-alanine in biofilm formation was found. Flagellar motility was downregulated, while putative type IV pili biosynthesis genes were not expressed. Moreover, the gene expression analysis suggested the involvement of the transcriptional regulators LexA, Spo0A and CcpA in stress response and biofilm formation. This study showed that NaCl addition might be a valuable strategy to induce biofilm formation by C. ljungdahlii, which can improve the efficacy of syngas fermentation and microbial electrosynthesis applications. PMID:28118386

  17. Trophic Status Controls Mercury Methylation Pathways in Northern Peats

    NASA Astrophysics Data System (ADS)

    Hines, M. E.; Zhang, L.; Barkay, T.; Krabbenhoft, D. P.; Schaefer, J.; Hu, H.; Sidelinger, W.; Liu, X.; Wang, Y.

    2015-12-01

    Methyl mercury (MeHg) can be produced by a variety of microbes including syntrophs, methanogens, acetogens, and fermenters, besides sulfate (SO42-, SRB) and iron- reducing bacteria. Many freshwater wetlands are deficient in electron acceptors that support the traditional respiratory pathways of methylation, yet they accumulate high levels of MeHg. To investigate methylation in these wetlands and to connect these pathways with vegetation and microbial communities, incubation experiments were conducted using peats from 26 sites in Alaska. The sites were clustered using multiple factor analysis based on pH, temp, CH4 and volatile fatty acids production rates, and surface vegetation composition. Three clusters were generated and corresponded to three trophic levels that were manifested by three pH levels (3.5, 4.5, and 5). Hg methylation activity in laboratory incubations was determined using the short-lived radioisotope 197Hg. In the low pH, Sphagnum-dominated cluster, methylation rates were less than 1% day-1 and likely conducted by primary fermenters. Conversely, the high pH trophic cluster dominated by Carex aquatilis and active syntrophy exhibited Hg methylation rates as high as 12% day-1. In intermediate sites, rich in Sphagnum magellanicum with less Carex, a gradient in syntrophy and Hg methylation paths was observed. Amendments with process-stimulators and inhibitors revealed no evidence of SO42- reduction, but suggested that SRB, metabolizing either syntrophically with methanogens and/or by fermentation, likely methylated Hg. While on going metatranscriptomics studies are required to verify the role of syntrophs, fermenters, and methanogens as methylators, these results revealed that Hg methylation pathways change greatly along trophic gradients with a dominance of respiratory pathways in mineral-rich sites, syntrophy dominance in intermediate sites, and fermentation dominance in nutrient-poor sites.

  18. Mercury Methylation in Alaskan Peatlands Spanning a Large Range of Trophic Structure

    NASA Astrophysics Data System (ADS)

    Krabbenhoft, D. P.; Zhang, L.; Hines, M. E.; Barkay, T.; Schaefer, J.; Aiken, G.

    2015-12-01

    The process of mercury (Hg) methylation has long been recognized as a key area of research in order to understand spatial and temporal variability of toxic methylmercury (MeHg) on the landscape. Numerous factors affect MeHg production, the most important generally falling into those that affect inorganic Hg(II) bioavailability (e.g., Hg(II) concentration and ligand composition), and those that affect microbial community composition and activity. The principal goal of this project is to decipher the details of MeHg production in Alaskan peatlands exhibiting a range of trophic status, including those lacking in electron acceptors that support the traditional respiratory pathway of MeHg production (e.g., sulfate reduction). MeHg production is carried out by a diverse group of microorganisms that possess the gene cluster (hgcAB), including the well-studied sulfate and iron- reducing bacteria (SRB and FeRB). However, less well known bacteria also possess the hgcAB genes, including: syntrophs, methanogens, acetogens, and fermenters. Methylation and demethylation activities were determined by injecting trace levels of the stable isotope (198Hg and Me204Hg) into intact peat cores. In addition, the short-lived radioisotope 197Hg was used in laboratory incubations. Laboratory studies also included assays for changes in diagnostic gas concentrations (CH4, CO2, H2) and LMW organic acids (formate, acetate, propionate, butyrate) to infer specific microbial processes, and the use of genomics to confirm microbial assemblages and the presence/absence of hgcAB genes. Overall, we observed Hg methylation rates were greatest at minerotrophic sites with active syntrophy and methanogenesis. Methylation and demethylation rates corresponded significantly across sites. There was no evidence of SO4- reduction in these samples, and addition of SO4- did not stimulate methylation suggesting that methylation was conducted by SRB that were metabolizing syntrophically and/or by fermentation.

  19. The transcriptional response of microbial communities in thawing Alaskan permafrost soils

    PubMed Central

    Coolen, Marco J. L.; Orsi, William D.

    2015-01-01

    Thawing of permafrost soils is expected to stimulate microbial decomposition and respiration of sequestered carbon. This could, in turn, increase atmospheric concentrations of greenhouse gasses, such as carbon dioxide and methane, and create a positive feedback to climate warming. Recent metagenomic studies suggest that permafrost has a large metabolic potential for carbon processing, including pathways for fermentation and methanogenesis. Here, we performed a pilot study using ultrahigh throughput Illumina HiSeq sequencing of reverse transcribed messenger RNA to obtain a detailed overview of active metabolic pathways and responsible organisms in up to 70 cm deep permafrost soils at a moist acidic tundra location in Arctic Alaska. The transcriptional response of the permafrost microbial community was compared before and after 11 days of thaw. In general, the transcriptional profile under frozen conditions suggests a dominance of stress responses, survival strategies, and maintenance processes, whereas upon thaw a rapid enzymatic response to decomposing soil organic matter (SOM) was observed. Bacteroidetes, Firmicutes, ascomycete fungi, and methanogens were responsible for largest transcriptional response upon thaw. Transcripts indicative of heterotrophic methanogenic pathways utilizing acetate, methanol, and methylamine were found predominantly in the permafrost table after thaw. Furthermore, transcripts involved in acetogenesis were expressed exclusively after thaw suggesting that acetogenic bacteria are a potential source of acetate for acetoclastic methanogenesis in freshly thawed permafrost. Metatranscriptomics is shown here to be a useful approach for inferring the activity of permafrost microbes that has potential to improve our understanding of permafrost SOM bioavailability and biogeochemical mechanisms contributing to greenhouse gas emissions as a result of permafrost thaw. PMID:25852660

  20. Physicochemical Conditions and Microbial Activities in the Highly Alkaline Gut of the Humus-Feeding Larva of Pachnoda ephippiata (Coleoptera: Scarabaeidae)

    PubMed Central

    Lemke, Thorsten; Stingl, Ulrich; Egert, Markus; Friedrich, Michael W.; Brune, Andreas

    2003-01-01

    The soil macrofauna plays an important role in the carbon and nitrogen cycle of terrestrial ecosystems. In order to gain more insight into the role of the intestinal microbiota in transformation and mineralization of organic matter during gut passage, we characterized the physicochemical conditions, microbial activities, and community structure in the gut of our model organism, the humus-feeding larva of the cetoniid beetle Pachnoda ephippiata. Microsensor measurements revealed an extreme alkalinity in the midgut, with highest values (pH > 10) between the second and third crown of midgut ceca. Both midgut and hindgut were largely anoxic, but despite the high pH, the redox potential of the midgut content was surprisingly high even in the largest instar. However, reducing conditions prevailed in the hindgut paunch of all instars (Eh ∼ −100 mV). Both gut compartments possessed a pronounced gut microbiota, with highest numbers in the hindgut, and microbial fermentation products were present in high concentrations. The stimulation of hindgut methanogenesis by exogenous electron donors, such as H2, formate, and methanol, together with considerable concentrations of formate in midgut and hemolymph, suggests that midgut fermentations are coupled to methanogenesis in the hindgut by an intercompartmental transfer of reducing equivalents via the hemolymph. The results of a cultivation-based enumeration of the major metabolic groups in midgut and hindgut, which yielded high titers of lactogenic, propionigenic, and acetogenic bacteria, are in good agreement not only with the accumulation of microbial fermentation products in the respective compartments but also with the results of a cultivation-independent characterization of the bacterial communities reported in the companion paper (M. Egert, B. Wagner, T. Lemke, A. Brune, and M. W. Friedrich, Appl. Environ. Microbiol. 69:6659-6668, 2003). PMID:14602625

  1. Acetone production with metabolically engineered strains of Acetobacterium woodii.

    PubMed

    Hoffmeister, Sabrina; Gerdom, Marzena; Bengelsdorf, Frank R; Linder, Sonja; Flüchter, Sebastian; Öztürk, Hatice; Blümke, Wilfried; May, Antje; Fischer, Ralf-Jörg; Bahl, Hubert; Dürre, Peter

    2016-07-01

    Expected depletion of oil and fossil resources urges the development of new alternative routes for the production of bulk chemicals and fuels beyond petroleum resources. In this study, the clostridial acetone pathway was used for the formation of acetone in the acetogenic bacterium Acetobacterium woodii. The acetone production operon (APO) containing the genes thlA (encoding thiolase A), ctfA/ctfB (encoding CoA transferase), and adc (encoding acetoacetate decarboxylase) from Clostridium acetobutylicum were cloned under the control of the thlA promoter into four vectors having different replicons for Gram-positives (pIP404, pBP1, pCB102, and pCD6). Stable replication was observed for all constructs. A. woodii [pJIR_actthlA] achieved the maximal acetone concentration under autotrophic conditions (15.2±3.4mM). Promoter sequences of the genes ackA from A. woodii and pta-ack from C. ljungdahlii were determined by primer extension (PEX) and cloned upstream of the APO. The highest acetone production in recombinant A. woodii cells was achieved using the promoters PthlA and Ppta-ack. Batch fermentations using A. woodii [pMTL84151_actthlA] in a bioreactor revealed that acetate concentration had an effect on the acetone production, due to the high Km value of the CoA transferase. In order to establish consistent acetate concentration within the bioreactor and to increase biomass, a continuous fermentation process for A. woodii was developed. Thus, acetone productivity of the strain A. woodii [pMTL84151_actthlA] was increased from 1.2mgL(-1)h(-1) in bottle fermentation to 26.4mgL(-1)h(-1) in continuous gas fermentation.

  2. Response of Deep Subsurface Microbial Community to Different Carbon Sources and Electron Acceptors during ∼2 months Incubation in Microcosms.

    PubMed

    Purkamo, Lotta; Bomberg, Malin; Nyyssönen, Mari; Ahonen, Lasse; Kukkonen, Ilmo; Itävaara, Merja

    2017-01-01

    Acetate plays a key role as electron donor and acceptor and serves as carbon source in oligotrophic deep subsurface environments. It can be produced from inorganic carbon by acetogenic microbes or through breakdown of more complex organic matter. Acetate is an important molecule for sulfate reducers that are substantially present in several deep bedrock environments. Aceticlastic methanogens use acetate as an electron donor and/or a carbon source. The goal of this study was to shed light on carbon cycling and competition in microbial communities in fracture fluids of Finnish crystalline bedrock groundwater system. Fracture fluid was anaerobically collected from a fracture zone at 967 m depth of the Outokumpu Deep Drill Hole and amended with acetate, acetate + sulfate, sulfate only or left unamended as a control and incubated up to 68 days. The headspace atmosphere of microcosms consisted of 80% hydrogen and 20% CO2. We studied the changes in the microbial communities with community fingerprinting technique as well as high-throughput 16S rRNA gene amplicon sequencing. The amended microcosms hosted more diverse bacterial communities compared to the intrinsic fracture zone community and the control treatment without amendments. The majority of the bacterial populations enriched with acetate belonged to clostridial hydrogenotrophic thiosulfate reducers and Alphaproteobacteria affiliating with groups earlier found from subsurface and groundwater environments. We detected a slight increase in the number of sulfate reducers after the 68 days of incubation. The microbial community changed significantly during the experiment, but increase in specifically acetate-cycling microbial groups was not observed.

  3. Invited review: anaerobic fermentation of dairy food wastewater.

    PubMed

    Hassan, A N; Nelson, B K

    2012-11-01

    Dairy food wastewater disposal represents a major environmental problem. This review discusses microorganisms associated with anaerobic digestion of dairy food wastewater, biochemistry of the process, factors affecting anaerobic digestion, and efforts to develop defined cultures. Anaerobic digestion of dairy food wastewater offers many advantages over other treatments in that a high level of waste stabilization is achieved with much lower levels of sludge. In addition, the process produces readily usable methane with low nutrient requirements and no oxygen. Anaerobic digestion is a series of complex reactions that broadly involve 2 groups of anaerobic or facultative anaerobic microorganisms: acidogens and methanogens. The first group of microorganisms breaks down organic compounds into CO(2) and volatile fatty acids. Some of these organisms are acetogenic, which convert long-chain fatty acids to acetate, CO(2), and hydrogen. Methanogens convert the acidogens' products to methane. The imbalance among the different microbial groups can lead not only to less methane production, but also to process failure. This is due to accumulation of intermediate compounds, such as volatile fatty acids, that inhibit methanogens. The criteria used for evaluation of the anaerobic digestion include levels of hydrogen and volatile fatty acids, methane:carbon ratio, and the gas production rate. A steady state is achieved in an anaerobic digester when the pH, chemical oxygen demand of the effluent, the suspended solids of the effluent, and the daily gas production remain constant. Factors affecting efficiency and stability of the process are types of microorganisms, feed C:N ratio, hydraulic retention time, reactor design, temperature, pH control, hydrogen pressure, and additives such as manure and surfactants. As anaerobic digesters become increasingly used in dairy plants, more research should be directed toward selecting the best cultures that maximize methane production from dairy

  4. Exogenous addition of H2 for an in situ biogas upgrading through biological reduction of carbon dioxide into methane.

    PubMed

    Mulat, Daniel Girma; Mosbæk, Freya; Ward, Alastair James; Polag, Daniela; Greule, Markus; Keppler, Frank; Nielsen, Jeppe Lund; Feilberg, Anders

    2017-06-13

    Biological reduction of CO2 into CH4 by exogenous addition of H2 is a promising technology for upgrading biogas into higher CH4 content. The aim of this work was to study the feasibility of exogenous H2 addition for an in situ biogas upgrading through biological conversion of the biogas CO2 into CH4. Moreover, this study employed systematic study with isotope analysis for providing comprehensive evidence on the underlying pathways of CH4 production and upstream processes. Batch reactors were inoculated with digestate originating from a full-scale biogas plant and fed once with maize leaf substrate. Periodic addition of H2 into the headspace resulted in a completely consumption of CO2 and a concomitant increase in CH4 content up to 89%. The microbial community and isotope analysis shows an enrichment of hydrogenotrophic Methanobacterium and the key role of hydrogenotrophic methanogenesis for biogas upgrading to higher CH4 content. Excess H2 was also supplied to evaluate its effect on overall process performance. The results show that excess H2 addition resulted in accumulation of H2, depletion of CO2 and inhibition of the degradation of acetate and other volatile fatty acids (VFA). A systematic isotope analysis revealed that excess H2 supply led to an increase in dissolved H2 to the level that thermodynamically inhibit the degradation of VFA and stimulate homo-acetogens for production of acetate from CO2 and H2. The inhibition was a temporary effect and acetate degradation resumed when the excess H2 was removed as well as in the presence of stoichiometric amount of H2 and CO2. This inhibition mechanism underlines the importance of carefully regulating the H2 addition rate and gas retention time to the CO2 production rate, H2-uptake rate and growth of hydrogenotrophic methanogens in order to achieve higher CH4 content without the accumulation of acetate and other VFA. Copyright © 2017 Elsevier Ltd. All rights reserved.

  5. Response of Deep Subsurface Microbial Community to Different Carbon Sources and Electron Acceptors during ∼2 months Incubation in Microcosms

    PubMed Central

    Purkamo, Lotta; Bomberg, Malin; Nyyssönen, Mari; Ahonen, Lasse; Kukkonen, Ilmo; Itävaara, Merja

    2017-01-01

    Acetate plays a key role as electron donor and acceptor and serves as carbon source in oligotrophic deep subsurface environments. It can be produced from inorganic carbon by acetogenic microbes or through breakdown of more complex organic matter. Acetate is an important molecule for sulfate reducers that are substantially present in several deep bedrock environments. Aceticlastic methanogens use acetate as an electron donor and/or a carbon source. The goal of this study was to shed light on carbon cycling and competition in microbial communities in fracture fluids of Finnish crystalline bedrock groundwater system. Fracture fluid was anaerobically collected from a fracture zone at 967 m depth of the Outokumpu Deep Drill Hole and amended with acetate, acetate + sulfate, sulfate only or left unamended as a control and incubated up to 68 days. The headspace atmosphere of microcosms consisted of 80% hydrogen and 20% CO2. We studied the changes in the microbial communities with community fingerprinting technique as well as high-throughput 16S rRNA gene amplicon sequencing. The amended microcosms hosted more diverse bacterial communities compared to the intrinsic fracture zone community and the control treatment without amendments. The majority of the bacterial populations enriched with acetate belonged to clostridial hydrogenotrophic thiosulfate reducers and Alphaproteobacteria affiliating with groups earlier found from subsurface and groundwater environments. We detected a slight increase in the number of sulfate reducers after the 68 days of incubation. The microbial community changed significantly during the experiment, but increase in specifically acetate-cycling microbial groups was not observed. PMID:28265265

  6. Investigation of availability of a high throughput screening method for predicting butanol solvent -producing ability of Clostridium beijerinckii.

    PubMed

    Su, HaiFeng; Zhu, Jun; Liu, Gang; Tan, Furong

    2016-07-22

    indicator of approach to select mutant solventogenic clostridia with improved fermentation of starch to increase butanol concentrations. The applicability can easily be broadened to a wide range of interesting microbes such as cellulolytic or acetogenic microorganisms, which produce biofuels from feedstock rich in starch.

  7. Efficient butanol-ethanol (B-E) production from carbon monoxide fermentation by Clostridium carboxidivorans.

    PubMed

    Fernández-Naveira, Ánxela; Abubackar, Haris Nalakath; Veiga, María C; Kennes, Christian

    2016-04-01

    The fermentation of waste gases rich in carbon monoxide using acetogens is an efficient way to obtain valuable biofuels like ethanol and butanol. Different experiments were carried out with the bacterial species Clostridium carboxidivorans as biocatalyst. In batch assays with no pH regulation, after complete substrate exhaustion, acetic acid, butyric acid, and ethanol were detected while only negligible butanol production was observed. On the other side, in bioreactors, with continuous carbon monoxide supply and pH regulation, both C2 and C4 fatty acids were initially formed as well as ethanol and butanol at concentrations never reported before for this type of anaerobic bioconversion of gaseous C1 compounds, showing that the operating conditions significantly affect the metabolic fermentation profile and butanol accumulation. Maximum ethanol and butanol concentrations in the bioreactors were obtained at pH 5.75, reaching values of 5.55 and 2.66 g/L, respectively. The alcohols were produced both from CO fermentation as well as from the bioconversion of previously accumulated acetic and butyric acids, resulting in low residual concentrations of such acids at the end of the bioreactor experiments. CO consumption was often around 50% and reached up to more than 80%. Maximum specific rates of ethanol and butanol production were reached at pH 4.75, with values of 0.16 g/h*g of biomass and 0.07 g/h*g of biomass, respectively, demonstrating that a low pH was more favorable to solventogenesis in this process, although it negatively affects biomass growth which does also play a role in the final alcohol titer.

  8. Methylmercury decomposition in sediments and bacterial cultures: Involvement of methanogens and sulfate reducers in oxidative demethylation

    USGS Publications Warehouse

    Oremland, R.S.; Culbertson, C.W.; Winfrey, M.R.

    1991-01-01

    Demethylation of monomethylmercury in freshwater and estuarine sediments and in bacterial cultures was investigated with 14CH3HgI. Under anaerobiosis, results with inhibitors indicated partial involvement of both sulfate reducers and methanogens, the former dominating estuarine sediments, while both were active in freshwaters. Aerobes were the most significant demethylators in estuarine sediments, but were unimportant in freshwater sediments. Products of anaerobic demethylation were mainly 14CO2 as well as lesser amounts of 14CH4. Acetogenic activity resulted in fixation of some 14CO2 produced from 14CH3HgI into acetate. Aerobic demethylation in estuarine sediments produced only 14CH4, while aerobic demethylation in freshwater sediments produced small amounts of both 14CH4 and 14CO2. Two species of Desulfovibrio produced only traces of 14CH4 from 14CH3HgI, while a culture of a methylotrophic methanogen formed traces of 14CO2 and 14CH4 when grown on trimethylamine in the presence of the 14CH3HgI. These results indicate that both aerobes and anaerobes demethylate mercury in sediments, but that either group may dominate in a particular sediment type. Aerobic demethylation in the estuarine sediments appeared to proceed by the previously characterized organomercurial-lyase pathway, because methane was the sole product. However, aerobic demethylation in freshwater sediments as well as anaerobic demethylation in all sediments studied produced primarily carbon dioxide. This indicates the presence of an oxidative pathway, possibly one in which methylmercury serves as an analog of one-carbon substrates.

  9. Carbon flow from volcanic CO2 into soil microbial communities of a wetland mofette.

    PubMed

    Beulig, Felix; Heuer, Verena B; Akob, Denise M; Viehweger, Bernhard; Elvert, Marcus; Herrmann, Martina; Hinrichs, Kai-Uwe; Küsel, Kirsten

    2015-03-01

    Effects of extremely high carbon dioxide (CO2) concentrations on soil microbial communities and associated processes are largely unknown. We studied a wetland area affected by spots of subcrustal CO2 degassing (mofettes) with focus on anaerobic autotrophic methanogenesis and acetogenesis because the pore gas phase was largely hypoxic. Compared with a reference soil, the mofette was more acidic (ΔpH ∼0.8), strongly enriched in organic carbon (up to 10 times), and exhibited lower prokaryotic diversity. It was dominated by methanogens and subdivision 1 Acidobacteria, which likely thrived under stable hypoxia and acidic pH. Anoxic incubations revealed enhanced formation of acetate and methane (CH4) from hydrogen (H2) and CO2 consistent with elevated CH4 and acetate levels in the mofette soil. (13)CO2 mofette soil incubations showed high label incorporations with ∼512 ng (13)C g (dry weight (dw)) soil(-1) d(-1) into the bulk soil and up to 10.7 ng (13)C g (dw) soil(-1) d(-1) into almost all analyzed bacterial lipids. Incorporation of CO2-derived carbon into archaeal lipids was much lower and restricted to the first 10 cm of the soil. DNA-SIP analysis revealed that acidophilic methanogens affiliated with Methanoregulaceae and hitherto unknown acetogens appeared to be involved in the chemolithoautotrophic utilization of (13)CO2. Subdivision 1 Acidobacteriaceae assimilated (13)CO2 likely via anaplerotic reactions because Acidobacteriaceae are not known to harbor enzymatic pathways for autotrophic CO2 assimilation. We conclude that CO2-induced geochemical changes promoted anaerobic and acidophilic organisms and altered carbon turnover in affected soils.

  10. Targeting Methanopterin Biosynthesis To Inhibit Methanogenesis

    PubMed Central

    Dumitru, Razvan; Palencia, Hector; Schroeder, Scott D.; DeMontigny, Bree A.; Takacs, James M.; Rasche, Madeline E.; Miner, Jess L.; Ragsdale, Stephen W.

    2003-01-01

    This paper describes the design, synthesis, and successful employment of inhibitors of 4-(β-d-ribofuranosyl)aminobenzene-5′-phosphate (RFA-P) synthase, which catalyzes the first committed step in the biosynthesis of methanopterin, to specifically halt the growth of methane-producing microbes. RFA-P synthase catalyzes the first step in the synthesis of tetrahydromethanopterin, a key cofactor required for methane formation and for one-carbon transformations in methanogens. A number of inhibitors, which are N-substituted derivatives of p-aminobenzoic acid (pABA), have been synthesized and their inhibition constants with RFA-P synthase have been determined. Based on comparisons of the inhibition constants among various inhibitors, we propose that the pABA binding site in RFA-P synthase has a relatively large hydrophobic pocket near the amino group. These enzyme-targeted inhibitors arrest the methanogenesis and growth of pure cultures of methanogens. Supplying pABA to the culture relieves the inhibition, indicating a competitive interaction between pABA and the inhibitor at the cellular target, which is most likely RFAP synthase. The inhibitors do not adversely affect the growth of pure cultures of the bacteria (acetogens) that play a beneficial role in the rumen. Inhibitors added to dense ruminal fluid cultures (artificial rumena) halt methanogenesis; however, they do not inhibit volatile fatty acid (VFA) production and, in some cases, VFA levels are slightly elevated in the methanogenesis-inhibited cultures. We suggest that inhibiting methanopterin biosynthesis could be considered in strategies to decrease anthropogenic methane emissions, which could have an environmental benefit since methane is a potent greenhouse gas. PMID:14660371

  11. Optimisation of sewage sludge anaerobic digestion through co-digestion with OFMSW: Effect of collection system and particle size

    SciTech Connect

    Silvestre, Gracia; Bonmatí, August; Fernández, Belén

    2015-09-15

    Highlights: • Methane production rate increased between 56% and 208% during OFMSW–SS codigestion. • The OFMSW particle size reduction from 20 to 8 mm did not affect the methane yield. • OFMSW–SS codigestion promoted β-oxidation and acetoclastic methanogenic activity. • The evolution of specific activity was a feasible tool to control the process. - Abstract: The effect of organic fraction of municipal solid waste (OFMSW) loading rate and particulate size on the sewage sludge (SS) mesophilic anaerobic co-digestion was assessed in continuous stirred tank reactor at hydraulic retention time of 20 days. The SS–OFMSW mixture composed by 54% of the volatile solids fed (inlet-VS), at OLR of 3.1 kg{sub COD} m{sup −3} d{sup −1} (1.9 kg{sub VS} m{sup −3} d{sup −1}), showed the highest increment on the volumetric methane production and yield of +200% and +59% respectively, under stable conditions. The effect of particulate size was assessed with the same mixture and same operational conditions but reducing the OFMSW particulate size from 20 mm to 8 mm with the aim to improve the hydrolysis step, but the results showed any influence in the OFMSW particulate size range analysed. In addition, specific biomass activity was assessed at the end of each co-digestion period. Results showed that OFMSW promoted β-oxidation syntrophic acetogens and the acetoclastic methanogens activity; although the last increase of the OFMSW percentage (from 47% to 54% inlet-VS) affected negatively the specific substrate activity, but not inhibitory effect was observed. Therefore, the results obtained in the continuous experiment could be related with some inhibitory or toxic effect and not due to hydrolysis limitation. The specific biomass activity test was demonstrated to be an interesting tool to evaluate and control the co-digestion process, especially when conventional parameters did not explain the behaviour of the biological system.

  12. Integration of ozonation and an anaerobic sequencing batch reactor (AnSBR) for the treatment of cherry stillage.

    PubMed

    Alvarez, Pedro M; Beltrán, Fernando J; Rodríguez, Eva M

    2005-01-01

    Cherry stillage is a high strength organic wastewater arising from the manufacture of alcoholic products by distillation of fermented cherries. It is made up of biorefractory polyphenols in addition to readily biodegradable organic matter. An anaerobic sequencing batch reactor (AnSBR) was used to treat cherry stillage at influent COD ranging from 5 to 50 g/L. Different cycle times were selected to test biomass organic loading rates (OLR(B)), from 0.3 to 1.2 g COD/g VSS.d. COD and TOC efficiency removals higher than 80% were achieved at influent COD up to 28.5 g/L but minimum OLR(B) tested. However, as a result of the temporary inhibition of acetogens and methanogens, volatile fatty acids (VFA) noticeably accumulated and methane production came to a transient standstill when operating at influent COD higher than 10 g/L. At these conditions, the AnSBR showed signs of instability and could not operate efficiently at OLR(B) higher than 0.3 g COD/g VSS.d. A feasible explanation for this inhibition is the presence of toxic polyphenols in cherry stillage. Thus, an ozonation step prior to the AnSBR was observed to be useful, since more than 75% of polyphenols could be removed by ozone. The integrated process was shown to be a suitable treatment technology as the following advantages compared to the single AnSBR treatment were observed: greater polyphenols and color removals, higher COD and TOC removal rates thus enabling the process to effectively operate at higher OLR, higher degree of biomethanation, and good stability with less risk of acidification.

  13. Characterization of the Core Rumen Microbiome in Cattle during Transition from Forage to Concentrate as Well as during and after an Acidotic Challenge

    PubMed Central

    Petri, Renee M.; Schwaiger, Tyler; Penner, Greg B.; Beauchemin, Karen A.; Forster, Robert J.; McKinnon, John J.; McAllister, Tim A.

    2013-01-01

    This study investigated the effect of diet and host on the rumen bacterial microbiome and the impact of an acidotic challenge on its composition. Using parallel pyrosequencing of the V3 hypervariable region of 16S rRNA gene, solid and liquid associated bacterial communities of 8 heifers were profiled. Heifers were exclusively fed forage, before being transitioned to a concentrate diet, subjected to an acidotic challenge and allowed to recover. Samples of rumen digesta were collected when heifers were fed forage, mixed forage, high grain, during challenge (4 h and 12 h) and recovery. A total of 560,994 high-quality bacterial sequences were obtained from the solid and liquid digesta. Using cluster analysis, prominent bacterial populations differed (P≤0.10) in solid and liquid fractions between forage and grain diets. Differences among hosts and diets were not revealed by DGGE, but real time qPCR showed that several bacteria taxon were impacted by changes in diet, with the exception of Streptococcus bovis. Analysis of the core rumen microbiome identified 32 OTU's representing 10 distinct bacterial taxa including Bacteroidetes (32.8%), Firmicutes (43.2%) and Proteobacteria (14.3%). Diversity of OTUs was highest with forage with 38 unique OTUs identified as compared to only 11 with the high grain diet. Comparison of the microbial profiles of clincial vs. subclinical acidotic heifers found a increases in the relative abundances of Acetitomaculum, Lactobacillus, Prevotella, and Streptococcus. Increases in Streptococcus and Lactobacillus likely reflect the tolerance of these species to low pH and their ability to proliferate on surplus fermentable carbohydrate. The acetogen, Acetitomaculum may thereforeplay a role in the conversion of lactate to acetate in acidotic animals. Further profiling of the bacterial populations associated with subclinical and clinical acidosis could establish a microbial fingerprint for these disorders and provide insight into whether there are

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

    PubMed Central

    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

  15. Optimisation of sewage sludge anaerobic digestion through co-digestion with OFMSW: Effect of collection system and particle size.

    PubMed

    Silvestre, Gracia; Bonmatí, August; Fernández, Belén

    2015-09-01

    The effect of organic fraction of municipal solid waste (OFMSW) loading rate and particulate size on the sewage sludge (SS) mesophilic anaerobic co-digestion was assessed in continuous stirred tank reactor at hydraulic retention time of 20days. The SS-OFMSW mixture composed by 54% of the volatile solids fed (inlet-VS), at OLR of 3.1kgCODm(-3)d(-1) (1.9kgVSm(-3)d(-1)), showed the highest increment on the volumetric methane production and yield of +200% and +59% respectively, under stable conditions. The effect of particulate size was assessed with the same mixture and same operational conditions but reducing the OFMSW particulate size from 20mm to 8mm with the aim to improve the hydrolysis step, but the results showed any influence in the OFMSW particulate size range analysed. In addition, specific biomass activity was assessed at the end of each co-digestion period. Results showed that OFMSW promoted β-oxidation syntrophic acetogens and the acetoclastic methanogens activity; although the last increase of the OFMSW percentage (from 47% to 54% inlet-VS) affected negatively the specific substrate activity, but not inhibitory effect was observed. Therefore, the results obtained in the continuous experiment could be related with some inhibitory or toxic effect and not due to hydrolysis limitation. The specific biomass activity test was demonstrated to be an interesting tool to evaluate and control the co-digestion process, especially when conventional parameters did not explain the behaviour of the biological system.

  16. Mechanistic enzymology of CO dehydrogenase from Clostridium thermoaceticum. Progress report, March 25, 1993--March 24, 1994

    SciTech Connect

    Ragsdale, S.W.

    1994-04-01

    Anaerobic acetogenic bacteria can convert carbon dioxide and carbon monoxide to cell carbon by a pathway which is called the acetyl-CoA pathway. With this pathway they convert monosaccharides and the methoxy group of lignin derived aromatics into acetic acid. The acetic acid is then used by a number of organisms, including methanogens, as a carbon and energy source. Therefore, the acetyl-CoA pathway links the biodegradation of complex macromolecules like cellulose and lignin to the utilization of simple two carbon units. The final steps in acetyl-CoA biosynthesis by anaerobic bacteria are performed by carbon monoxide dehydrogenase (CODH), a nickel/iron-sulfur protein. We have previously demonstrate that the conversion of CH{sub 3}-H{sub 4} folate, CO and CoA to acetyl-CoA involves enzyme-bound intermediates that the one- and two-carbon precursors of acetyl-CoA are organometallic complexes, and that the site for assembly of acetyl-CoA is a novel Ni-Fe-S cluster which binds CO as a terminal carbonyl, i.e., M-C {equivalent_to} O. In the past year we have shown that the activities for CO oxidation and acetyl-CoA synthesis sites occur at separate sites, that it is a Fe, not a Ni, site in the Ni-Fe-S cluster which binds CO for acetyl-CoA synthesis, and that carbon disulfide (CS2) reacts with CODH at the Ni-Fe-S site to generate an isolated Ni(I) species. We also determined that CS{sub 2} is competitive with CO at the acetyl-CoA synthesis site and does not bind to the CO oxidation/CO{sub 2} reduction site.

  17. Uropathogenic Escherichia coli CFT073 Is Adapted to Acetatogenic Growth but Does Not Require Acetate during Murine Urinary Tract Infection▿ §

    PubMed Central

    Anfora, Andrew T.; Halladin, David K.; Haugen, Brian J.; Welch, Rodney A.

    2008-01-01

    In vivo accumulation of d-serine by Escherichia coli CFT073 leads to elevated expression of PAP fimbriae and hemolysin by an unknown mechanism. Loss of d-serine catabolism by CFT073 leads to a competitive advantage during murine urinary tract infection (UTI), but loss of both d- and l-serine catabolism results in attenuation. Serine is the first amino acid to be consumed in closed tryptone broth cultures and precedes the production of acetyl phosphate, a high-energy molecule involved in intracellular signaling, and the eventual secretion of acetate. We propose that the colonization defect associated with the loss of serine catabolism is due to perturbations of acetate metabolism. CFT073 grows more rapidly on acetogenic substrates than does E. coli K-12 isolate MG1655. As shown by transcription microarray results, d-serine is catabolized into acetate via the phosphotransacetylase (pta) and acetate kinase (ackA) genes while downregulating expression of acetyl coenzyme A synthase (acs). CFT073 acs, which is unable to reclaim secreted acetate, colonized mouse bladders and kidneys in the murine model of UTI indistinguishably from the wild type. Both pta and ackA are involved in the maintenance of intracellular acetyl phosphate. CFT073 pta and ackA mutants were screened to investigate the role of acetyl phosphate in UTI pathogenesis. Both single mutants are at a competitive disadvantage relative to the wild type in the kidneys but normally colonize the bladder. CFT073 ackA pta was attenuated in both the bladder and the kidneys. Thus, we demonstrate that CFT073 is adapted to acetate metabolism as a result of requiring a proper cycling of the acetyl phosphate pathway for colonization of the upper urinary tract. PMID:18838520

  18. Accelerated rates of in situ microbial activity after permafrost collapse estimated from a porewater isotope model

    NASA Astrophysics Data System (ADS)

    Waldrop, M. P.; Neumann, R. B.; Jones, M.; Manies, K.; Mcfarland, J. W.; Blazewicz, S.; Turetsky, M. R.

    2016-12-01

    Permafrost thaw is expected to become widespread in interior Alaska over the coming century, resulting in increased CO2 and CH4 fluxes from soils and a positive feedback to global warming. However much of our understanding of the microbial response to thaw is predicated on simple laboratory incubations that preclude the multitude of interactions occurring in soils under field situations. Here, we utilize a time series of 13CO2 and 13CH4 measured in porewater collected from thermokarst bogs of different ages to estimate in-situ reaction rates of microbial respiration, methanogenesis from acetate, methanogenesis from CO2, homoacetogenesis, and methane oxidation from porewater concentrations and 13CO2 and 13CH4. We utilized this modeling technique to test the hypothesis that microbial activities are stimulated soon after permafrost thaw and this effect declines over time. Our field site is a chronosequence of thermokarst bogs at the Alaska Peatland Experiment (APEX) in interior AK where we have observed significant losses of peatland carbon since permafrost collapse over the last half century. Concentrations of dissolved CO2 and CH4 in porewater increased with depth, and were higher in the youngest bog compared to the older bogs. With increasing depth 13CH4 became more depleted while 13CO2 became more enriched. Preliminary modeling results, based upon these porewater gas concentrations and isotope values, indicate that microbial activities are higher in the youngest bogs compared to the older bogs, supporting the hypothesis that accelerated rates of microbial activities in young thermokarst features are responsible for high rates of C losses from these systems. Additionally, model results will be compared to variation in the abundance of methanogens, methane oxidizers, and acetogens as well as process rates measured in lab incubations, providing insights into the mechanisms responsible for these losses.

  19. Pan-genome of the dominant human gut-associated archaeon, Methanobrevibacter smithii, studied in twins.

    PubMed

    Hansen, Elizabeth E; Lozupone, Catherine A; Rey, Federico E; Wu, Meng; Guruge, Janaki L; Narra, Aneesha; Goodfellow, Jonathan; Zaneveld, Jesse R; McDonald, Daniel T; Goodrich, Julia A; Heath, Andrew C; Knight, Rob; Gordon, Jeffrey I

    2011-03-15

    The human gut microbiota harbors three main groups of H(2)-consuming microbes: methanogens including the dominant archaeon, Methanobrevibacter smithii, a polyphyletic group of acetogens, and sulfate-reducing bacteria. Defining their roles in the gut is important for understanding how hydrogen metabolism affects the efficiency of fermentation of dietary components. We quantified methanogens in fecal samples from 40 healthy adult female monozygotic (MZ) and 28 dizygotic (DZ) twin pairs, analyzed bacterial 16S rRNA datasets generated from their fecal samples to identify taxa that co-occur with methanogens, sequenced the genomes of 20 M. smithii strains isolated from families of MZ and DZ twins, and performed RNA-Seq of a subset of strains to identify their responses to varied formate concentrations. The concordance rate for methanogen carriage was significantly higher for MZ versus DZ twin pairs. Co-occurrence analysis revealed 22 bacterial species-level taxa positively correlated with methanogens: all but two were members of the Clostridiales, with several being, or related to, known hydrogen-producing and -consuming bacteria. The M. smithii pan-genome contains 987 genes conserved in all strains, and 1,860 variably represented genes. Strains from MZ and DZ twin pairs had a similar degree of shared genes and SNPs, and were significantly more similar than strains isolated from mothers or members of other families. The 101 adhesin-like proteins (ALPs) in the pan-genome (45 ± 6 per strain) exhibit strain-specific differences in expression and responsiveness to formate. We hypothesize that M. smithii strains use their different repertoires of ALPs to create diversity in their metabolic niches, by allowing them to establish syntrophic relationships with bacterial partners with differing metabolic capabilities and patterns of co-occurrence.

  20. Diversity of planktonic and attached bacterial communities in a phenol-contaminated sandstone aquifer.

    PubMed

    Rizoulis, Athanasios; Elliott, David R; Rolfe, Stephen A; Thornton, Steven F; Banwart, Steven A; Pickup, Roger W; Scholes, Julie D

    2013-07-01

    Polluted aquifers contain indigenous microbial communities with the potential for in situ bioremediation. However, the effect of hydrogeochemical gradients on in situ microbial communities (especially at the plume fringe, where natural attenuation is higher) is still not clear. In this study, we used culture-independent techniques to investigate the diversity of in situ planktonic and attached bacterial communities in a phenol-contaminated sandstone aquifer. Within the upper and lower plume fringes, denaturing gradient gel electrophoresis profiles indicated that planktonic community structure was influenced by the steep hydrogeochemical gradient of the plume rather than the spatial location in the aquifer. Under the same hydrogeochemical conditions (in the lower plume fringe, 30 m below ground level), 16S rRNA gene cloning and sequencing showed that planktonic and attached bacterial communities differed markedly and that the attached community was more diverse. The 16S rRNA gene phylogeny also suggested that a phylogenetically diverse bacterial community operated at this depth (30 mbgl), with biodegradation of phenolic compounds by nitrate-reducing Azoarcus and Acidovorax strains potentially being an important process. The presence of acetogenic and sulphate-reducing bacteria only in the planktonic clone library indicates that some natural attenuation processes may occur preferentially in one of the two growth phases (attached or planktonic). Therefore, this study has provided a better understanding of the microbial ecology of this phenol-contaminated aquifer, and it highlights the need for investigating both planktonic and attached microbial communities when assessing the potential for natural attenuation in contaminated aquifers.

  1. Control of Interspecies Electron Flow during Anaerobic Digestion: Significance of Formate Transfer versus Hydrogen Transfer during Syntrophic Methanogenesis in Flocs

    PubMed Central

    Thiele, Jurgen H.; Zeikus, J. Gregory

    1988-01-01

    Microbial formate production and consumption during syntrophic conversion of ethanol or lactate to methane was examined in purified flocs and digestor contents obtained from a whey-processing digestor. Formate production by digestor contents or purified digestor flocs was dependent on CO2 and either ethanol or lactate but not H2 gas as an electron donor. During syntrophic methanogenesis, flocs were the primary site for formate production via ethanol-dependent CO2 reduction, with a formate production rate and methanogenic turnover constant of 660 μM/h and 0.044/min, respectively. Floc preparations accumulated fourfold-higher levels of formate (40 μM) than digestor contents, and the free flora was the primary site for formate cleavage to CO2 and H2 (90 μM formate per h). Inhibition of methanogenesis by CHCl3 resulted in formate accumulation and suppression of syntrophic ethanol oxidation. H2 gas was an insignificant intermediary metabolite of syntrophic ethanol conversion by flocs, and its exogenous addition neither stimulated methanogenesis nor inhibited the initial rate of ethanol oxidation. These results demonstrated that >90% of the syntrophic ethanol conversion to methane by mixed cultures containing primarily Desulfovibrio vulgaris and Methanobacterium formicicum was mediated via interspecies formate transfer and that <10% was mediated via interspecies H2 transfer. The results are discussed in relation to biochemical thermodynamics. A model is presented which describes the dynamics of a bicarbonate-formate electron shuttle mechanism for control of carbon and electron flow during syntrophic methanogenesis and provides a novel mechanism for energy conservation by syntrophic acetogens. PMID:16347526

  2. Studies on the Mechanism of Electron Bifurcation Catalyzed by Electron Transferring Flavoprotein (Etf) and Butyryl-CoA Dehydrogenase (Bcd) of Acidaminococcus fermentans*

    PubMed Central

    Chowdhury, Nilanjan Pal; Mowafy, Amr M.; Demmer, Julius K.; Upadhyay, Vikrant; Koelzer, Sebastian; Jayamani, Elamparithi; Kahnt, Joerg; Hornung, Marco; Demmer, Ulrike; Ermler, Ulrich; Buckel, Wolfgang

    2014-01-01

    Electron bifurcation is a fundamental strategy of energy coupling originally discovered in the Q-cycle of many organisms. Recently a flavin-based electron bifurcation has been detected in anaerobes, first in clostridia and later in acetogens and methanogens. It enables anaerobic bacteria and archaea to reduce the low-potential [4Fe-4S] clusters of ferredoxin, which increases the efficiency of the substrate level and electron transport phosphorylations. Here we characterize the bifurcating electron transferring flavoprotein (EtfAf) and butyryl-CoA dehydrogenase (BcdAf) of Acidaminococcus fermentans, which couple the exergonic reduction of crotonyl-CoA to butyryl-CoA to the endergonic reduction of ferredoxin both with NADH. EtfAf contains one FAD (α-FAD) in subunit α and a second FAD (β-FAD) in subunit β. The distance between the two isoalloxazine rings is 18 Å. The EtfAf-NAD+ complex structure revealed β-FAD as acceptor of the hydride of NADH. The formed β-FADH− is considered as the bifurcating electron donor. As a result of a domain movement, α-FAD is able to approach β-FADH− by about 4 Å and to take up one electron yielding a stable anionic semiquinone, α-FAD⨪, which donates this electron further to Dh-FAD of BcdAf after a second domain movement. The remaining non-stabilized neutral semiquinone, β-FADH•, immediately reduces ferredoxin. Repetition of this process affords a second reduced ferredoxin and Dh-FADH− that converts crotonyl-CoA to butyryl-CoA. PMID:24379410

  3. Biofilm Formation by Clostridium ljungdahlii Is Induced by Sodium Chloride Stress: Experimental Evaluation and Transcriptome Analysis.

    PubMed

    Philips, Jo; Rabaey, Korneel; Lovley, Derek R; Vargas, Madeline

    2017-01-01

    The acetogen Clostridium ljungdahlii is capable of syngas fermentation and microbial electrosynthesis. Biofilm formation could benefit both these applications, but was not yet reported for C. ljungdahlii. Biofilm formation does not occur under standard growth conditions, but attachment or aggregation could be induced by different stresses. The strongest biofilm formation was observed with the addition of sodium chloride. After 3 days of incubation, the biomass volume attached to a plastic surface was 20 times higher with than without the addition of 200 mM NaCl to the medium. The addition of NaCl also resulted in biofilm formation on glass, graphite and glassy carbon, the latter two being often used electrode materials for microbial electrosynthesis. Biofilms were composed of extracellular proteins, polysaccharides, as well as DNA, while pilus-like appendages were observed with, but not without, the addition of NaCl. A transcriptome analysis comparing planktonic (no NaCl) and biofilm (NaCl addition) cells showed that C. ljungdahlii coped with the salt stress by the upregulation of the general stress response, Na+ export and osmoprotectant accumulation. A potential role for poly-N-acetylglucosamines and D-alanine in biofilm formation was found. Flagellar motility was downregulated, while putative type IV pili biosynthesis genes were not expressed. Moreover, the gene expression analysis suggested the involvement of the transcriptional regulators LexA, Spo0A and CcpA in stress response and biofilm formation. This study showed that NaCl addition might be a valuable strategy to induce biofilm formation by C. ljungdahlii, which can improve the efficacy of syngas fermentation and microbial electrosynthesis applications.

  4. The transcriptional response of microbial communities in thawing Alaskan permafrost soils.

    PubMed

    Coolen, Marco J L; Orsi, William D

    2015-01-01

    Thawing of permafrost soils is expected to stimulate microbial decomposition and respiration of sequestered carbon. This could, in turn, increase atmospheric concentrations of greenhouse gasses, such as carbon dioxide and methane, and create a positive feedback to climate warming. Recent metagenomic studies suggest that permafrost has a large metabolic potential for carbon processing, including pathways for fermentation and methanogenesis. Here, we performed a pilot study using ultrahigh throughput Illumina HiSeq sequencing of reverse transcribed messenger RNA to obtain a detailed overview of active metabolic pathways and responsible organisms in up to 70 cm deep permafrost soils at a moist acidic tundra location in Arctic Alaska. The transcriptional response of the permafrost microbial community was compared before and after 11 days of thaw. In general, the transcriptional profile under frozen conditions suggests a dominance of stress responses, survival strategies, and maintenance processes, whereas upon thaw a rapid enzymatic response to decomposing soil organic matter (SOM) was observed. Bacteroidetes, Firmicutes, ascomycete fungi, and methanogens were responsible for largest transcriptional response upon thaw. Transcripts indicative of heterotrophic methanogenic pathways utilizing acetate, methanol, and methylamine were found predominantly in the permafrost table after thaw. Furthermore, transcripts involved in acetogenesis were expressed exclusively after thaw suggesting that acetogenic bacteria are a potential source of acetate for acetoclastic methanogenesis in freshly thawed permafrost. Metatranscriptomics is shown here to be a useful approach for inferring the activity of permafrost microbes that has potential to improve our understanding of permafrost SOM bioavailability and biogeochemical mechanisms contributing to greenhouse gas emissions as a result of permafrost thaw.

  5. The Ether-Cleaving Methyltransferase System of the Strict Anaerobe Acetobacterium dehalogenans: Analysis and Expression of the Encoding Genes▿

    PubMed Central

    Schilhabel, Anke; Studenik, Sandra; Vödisch, Martin; Kreher, Sandra; Schlott, Bernhard; Pierik, Antonio Y.; Diekert, Gabriele

    2009-01-01

    Anaerobic O-demethylases are inducible multicomponent enzymes which mediate the cleavage of the ether bond of phenyl methyl ethers and the transfer of the methyl group to tetrahydrofolate. The genes of all components (methyltransferases I and II, CP, and activating enzyme [AE]) of the vanillate- and veratrol-O-demethylases of Acetobacterium dehalogenans were sequenced and analyzed. In A. dehalogenans, the genes for methyltransferase I, CP, and methyltransferase II of both O-demethylases are clustered. The single-copy gene for AE is not included in the O-demethylase gene clusters. It was found that AE grouped with COG3894 proteins, the function of which was unknown so far. Genes encoding COG3894 proteins with 20 to 41% amino acid sequence identity with AE are present in numerous genomes of anaerobic microorganisms. Inspection of the domain structure and genetic context of these orthologs predicts that these are also reductive activases for corrinoid enzymes (RACEs), such as carbon monoxide dehydrogenase/acetyl coenzyme A synthases or anaerobic methyltransferases. The genes encoding the O-demethylase components were heterologously expressed with a C-terminal Strep-tag in Escherichia coli, and the recombinant proteins methyltransferase I, CP, and AE were characterized. Gel shift experiments showed that the AE comigrated with the CP. The formation of other protein complexes with the O-demethylase components was not observed under the conditions used. The results point to a strong interaction of the AE with the CP. This is the first report on the functional heterologous expression of acetogenic phenyl methyl ether-cleaving O-demethylases. PMID:19011025

  6. Characterization of an O-Demethylase of Desulfitobacterium hafniense DCB-2

    PubMed Central

    Studenik, Sandra; Vogel, Michaela

    2012-01-01

    Besides acetogenic bacteria, only Desulfitobacterium has been described to utilize and cleave phenyl methyl ethers under anoxic conditions; however, no ether-cleaving O-demethylases from the latter organisms have been identified and investigated so far. In this study, genes of an operon encoding O-demethylase components of Desulfitobacterium hafniense strain DCB-2 were cloned and heterologously expressed in Escherichia coli. Methyltransferases I and II were characterized. Methyltransferase I mediated the ether cleavage and the transfer of the methyl group to the superreduced corrinoid of a corrinoid protein. Desulfitobacterium methyltransferase I had 66% identity (80% similarity) to that of the vanillate-demethylating methyltransferase I (OdmB) of Acetobacterium dehalogenans. The substrate spectrum was also similar to that of the latter enzyme; however, Desulfitobacterium methyltransferase I showed a higher level of activity for guaiacol and used methyl chloride as a substrate. Methyltransferase II catalyzed the transfer of the methyl group from the methylated corrinoid protein to tetrahydrofolate. It also showed a high identity (∼70%) to methyltransferases II of A. dehalogenans. The corrinoid protein was produced in E. coli as cofactor-free apoprotein that could be reconstituted with hydroxocobalamin or methylcobalamin to function in the methyltransferase I and II assays. Six COG3894 proteins, which were assumed to function as activating enzymes mediating the reduction of the corrinoid protein after an inadvertent oxidation of the corrinoid cofactor, were studied with respect to their abilities to reduce the recombinant reconstituted corrinoid protein. Of these six proteins, only one was found to catalyze the reduction of the corrinoid protein. PMID:22522902

  7. Volatile Fatty Acid production by the hindgut microbiota of xylophagous termites.

    PubMed

    Odelson, D A; Breznak, J A

    1983-05-01

    Acetate dominated the extracellular pool of volatile fatty acids (VFAs) in the hindgut fluid of Reticulitermes flavipes, Zootermopsis angusticollis, and Incisitermes schwarzi, where it occurred at concentrations of 57.9 to 80.6 mM and accounted for 94 to 98 mol% of all VFAs. Small amounts of C(3) to C(5) VFAs were also observed. Acetate was also the major VFA in hindgut homogenates of Schedorhinotermes lamanianus, Prorhinotermes simplex, Coptotermes formosanus, and Nasutitermes corniger. Estimates of in situ acetogenesis by the hindgut microbiota of R. flavipes (20.2 to 43.3 nmol . termite . h) revealed that this activity could support 77 to 100% of the respiratory requirements of the termite (51.6 to 63.6 nmol of O(2) . termite . h). This conclusion was buttressed by the demonstration of acetate in R. flavipes hemolymph (at 9.0 to 11.6 mM), but not in feces, and by the ability of termite tissues to readily oxidize acetate to CO(2). About 85% of the acetate produced by the hindgut microbiota was derived from cellulose C; the remainder was derived from hemicellulose C. Selective removal of major groups of microbes from the hindgut of R. flavipes indicated that protozoa were primarily responsible for acetogenesis but that bacteria also functioned in this capacity. H(2) and CH(4) were evolved by R. flavipes (usually about 0.4 nmol . termite . h), but these compounds represented a minor fate of electrons derived from wood dissimilation within R. flavipes. A working model is proposed for symbiotic wood polysaccharide degradation in R. flavipes, and the possible roles of individual gut microbes, including CO(2)-reducing acetogenic bacteria, are discussed.

  8. The Origin of Life in Alkaline Hydrothermal Vents.

    PubMed

    Sojo, Victor; Herschy, Barry; Whicher, Alexandra; Camprubí, Eloi; Lane, Nick

    2016-02-01

    Over the last 70 years, prebiotic chemists have been very successful in synthesizing the molecules of life, from amino acids to nucleotides. Yet there is strikingly little resemblance between much of this chemistry and the metabolic pathways of cells, in terms of substrates, catalysts, and synthetic pathways. In contrast, alkaline hydrothermal vents offer conditions similar to those harnessed by modern autotrophs, but there has been limited experimental evidence that such conditions could drive prebiotic chemistry. In the Hadean, in the absence of oxygen, alkaline vents are proposed to have acted as electrochemical flow reactors, in which alkaline fluids saturated in H2 mixed with relatively acidic ocean waters rich in CO2, through a labyrinth of interconnected micropores with thin inorganic walls containing catalytic Fe(Ni)S minerals. The difference in pH across these thin barriers produced natural proton gradients with equivalent magnitude and polarity to the proton-motive force required for carbon fixation in extant bacteria and archaea. How such gradients could have powered carbon reduction or energy flux before the advent of organic protocells with genes and proteins is unknown. Work over the last decade suggests several possible hypotheses that are currently being tested in laboratory experiments, field observations, and phylogenetic reconstructions of ancestral metabolism. We analyze the perplexing differences in carbon and energy metabolism in methanogenic archaea and acetogenic bacteria to propose a possible ancestral mechanism of CO2 reduction in alkaline hydrothermal vents. Based on this mechanism, we show that the evolution of active ion pumping could have driven the deep divergence of bacteria and archaea.

  9. Methylmercury decomposition in sediments and bacterial cultures: Involvement of methanogens and sulfate reducers in oxidative demethylation

    SciTech Connect

    Oremland, R.S.; Culbertson, C.W. ); Winfrey, M.R. )

    1991-01-01

    The biogeochemical cycling of mercury has received considerable attention because of the toxicity of methylmercury, its bioaccumulation in biota, and its biomagnification in aquatic food chains. The formation of methylmercury is mediated primarily by microorganisms. Demethylation of monomethylmercury in freshwater and estuarine sediments and in bacterial cultures was investigated with {sup 14}CH{sub 3}HgI. Under anaerobiosis, results with inhibitors indicated partial involvement of both sulfate reducers and methanogens, the former dominated estuarine sediments, while both were active in freshwaters. Aerobes were the most significant demethylators in estuarine sediments, but were unimportant in freshwater sediments. Products of anaerobic demthylation were mainly {sup 14}CO{sub 2} as well as lesser amounts of {sup 14}CH{sub 4}. Acetogenic activity resulted in fixation of some {sup 14}CO{sub 2} produced from {sup 14}CH{sub 3}HgI into acetate. Aerobic demethylation in estuarine sediments produced only {sup 14}CH{sub 4}, while aerobic demethylation in freshwater sediments produced small amounts of both {sup 14}CH{sub 4} and {sup 14}CO{sub 2}. Two species of Desulfovibrio produced only traces of {sup 14}CH{sub 4} from {sup 14}CH{sub 3}HgI, while a culture of a methylotrophic methanogen formed traces of {sup 14}CO{sub 2} and {sup 14}CH{sub 4} when grown on trimethylamine in the presence of the {sup 14}CH{sub 3}HgI. These results indicate that both aerobes and anaerobes demethylate mercury in sediments, but that either group may dominate in a particular sediment type. Aerobic demethylation in the estuarine sediments appeared to proceed by the previously characterized organomercurial-lyase pathway, because methane was the sole product. This indicates the presence of an oxidative pathway, possibly one in which methylmercury serves as an analog of one-carbon substrates.

  10. The core populations and co-occurrence patterns of prokaryotic communities in household biogas digesters.

    PubMed

    Rui, Junpeng; Li, Jiabao; Zhang, Shiheng; Yan, Xuefeng; Wang, Yuanpeng; Li, Xiangzhen

    2015-01-01

    Household biogas digesters are widely used to harvest energy in rural areas of developing countries. Understanding core prokaryotic communities, their co-occurrence patterns, and their relationships to environmental factors is important to manage these small-scale anaerobic digestion systems effectively. In this study, 43 household biogas digesters were collected across eight provinces in China. Prokaryotic communities were investigated using 454 pyrosequencing of 16S rRNA genes. Fourteen core genera and ten core OTUs were identified in household biogas digesters. They were mainly affiliated with the phylum Firmicutes, Synergistetes, Actinobacteria, Chloroflexi, and Spirochaetes. Core prokaryotic genera were mainly composed of Clostridium, Clostridium XI, Syntrophomonas, Cloacibacillus, Sedimentibacter, and Turicibacter. Prokaryotic communities in the 43 samples were clearly divided into two clusters. Cluster I was dominated by Clostridium, while Cluster II was dominated by members of Spirochaetes, Bacteroidales, Clostridia, and abundant syntrophs and methanogens. NH4 (+)-N and COD contributed significantly to the assembly of the prokaryotic community in Cluster I, while NH4 (+)-N, pH, and phosphate contributed significantly to Cluster II. Correlation-based network analysis showed that the prokaryotic communities in the biogas digesters were dominated by some functional modules. Cluster I was dominated by acetotrophic methanogenic modules and the Clostridium-driven primary fermentation module, while the network of Cluster II was dominated by hydrogenotrophic and acetogenic methanogenesis modules and multi-group-driven (Spirochaetes, Bacteroidales, and Clostridia) primary fermentation modules. The network of Cluster II was more complex and functionally redundant. Prokaryotic communities identified in the household biogas digesters varied significantly and were affected by environmental factors, such as NH4 (+)-N, pH, and COD. However, core prokaryotic communities

  11. Control of interspecies electron flow during anaerobic digestion: role of floc formation in syntrophic methanogenesis

    SciTech Connect

    Thiele, J.H.; Chartrain, M.; Zeikus, J.G.

    1988-01-01

    The flora of an anaerobic whey-processing chemostat was separated by anaerobic sedimentation techniques into a free-living bacterial fraction and a bacterial floc fraction. The floc fraction constituted a major part (i.e., 57% total protein) of the total microbial population in the digestor, and it accounted for 87% of the total CO/sub 2/-dependent methanogenic activity and 76% of the total ethanol-consuming acetogenic activity. Lactose was degraded by both cellular fractions, but in the free flora fraction it was associated with higher intermediary levels of H/sub 2/, ethanol, butyrate, and propionate production. Electron microscopic analysis of flocs showed bacterial diversity and juxtapositioning of tentative Desulfovibrio and Methanobacterium species without significant microcolony formation. Ethanol, an intermediary product of lactose-hydrolyzing bacteria, was converted to acetate and methane within the flocs by interspecies electron transfer. Ethanol-dependent methane formation was compartmentalized and closely coupled kinetically within the flocs but without significant formation of H/sub 2/ gas. Physical disruption of flocs into fragments of 10- to 20-..mu..m diameter initially increased the H/sub 2/ partial pressure but did not change the carbon transformation kinetic patterns of ethanol metabolism or demonstrate a significant role for H/sub 2/ in CO/sub 2/ reduction to methane. The data demonstrate that floc formation in a whey-processing anaerobic digestor functions in juxtapositioning cells for interspecies electron transfer during syntrophic ethanol conversion into acetate and methane but by a mechanism which was independent of the available dissolved H/sub 2/ gas pool in the ecosystem.

  12. The completely annotated genome and comparative genomics of the Peptoniphilaceae bacterium str. ING2-D1G, a novel acidogenic bacterium isolated from a mesophilic biogas reactor.

    PubMed

    Tomazetto, Geizecler; Hahnke, Sarah; Langer, Thomas; Wibberg, Daniel; Blom, Jochen; Maus, Irena; Pühler, Alfred; Klocke, Michael; Schlüter, Andreas

    2017-09-10

    The strictly anaerobic Peptoniphilaceae bacterium str. ING2-D1G (=DSM 28672=LMG 28300) was isolated from a mesophilic laboratory-scale completely stirred tank biogas reactor (CSTR) continuously co-digesting maize silage, pig and cattle manure. Based on 16S rRNA gene sequence comparison, the closest described relative to this strain is Peptoniphilus obesi ph1 showing 91.2% gene sequence identity. The most closely related species with a validly published name is Peptoniphilus indolicus DSM 20464(T) whose 16S rRNA gene sequence is 90.6% similar to the one of strain ING2-D1G. The genome of the novel strain was completely sequenced and manually annotated to reconstruct its metabolic potential regarding anaerobic digestion of biomass. The strain harbors a circular chromosome with a size of 1.6 Mb that contains 1466 coding sequences, 53 tRNA genes and 4 ribosomal RNA (rrn) operons. The genome carries a 28,261bp prophage insertion comprising 47 phage-related coding sequences. Reconstruction of fermentation pathways revealed that strain ING2-D1G encodes all enzymes for hydrogen, lactate and acetate production, corroborating that it is involved in the acido- and acetogenic phase of the biogas process. Comparative genome analyses of Peptoniphilaceae bacterium str. ING2-D1G and its closest relative Peptoniphilus obesi ph1 uncovered rearrangements, deletions and insertions within the chromosomes of both strains substantiating a divergent evolution. In addition to genomic analyses, a physiological and phenotypic characterization of the novel isolate was performed. Grown in Brain Heart Infusion Broth with added yeast extract, cells were spherical to ovoid, catalase- and oxidase-negative and stained Gram-positive. Optimal growth occurred between 35 and 37°C and at a pH value of 7.6. Fermentation products were acetate, butanoate and carbon dioxide. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Characterization of microbial activities and U reduction in a shallow aquifer contaminated by uranium mill tailings.

    PubMed

    Elias, D A; Krumholz, L R; Wong, D; Long, P E; Suflita, J M

    2003-07-01

    A characterization of the Shiprock, NM, uranium mill tailing site focused on the geochemical and microbiological factors governing in-situ uranium-redox reactions. Groundwater and aqueous extracts of sediment samples contained a wide concentration range of sulfate, nitrate, and U(VI) with median values of 21.2 mM, 16.1 micro M, and 2.7 micro M, respectively. Iron(III) was not detected in groundwater, but a median value of 0.3 mM in sediment extracts was measured. Bacterial diversity down gradient from the disposal pile reflected the predominant geochemistry with relatively high numbers of sulfate- and nitrate-reducing microorganisms, and smaller numbers of acetogenic, methanogenic, nitrate-dependent Fe(II)-oxidizing, Fe(III)-reducing, and sulfide-oxidizing bacteria. In aquifer slurry incubations, nitrate reduction was always preferred and had a negative impact on sulfate-, Fe(III)-, and U-reduction rates. We also found that sulfate-reduction rates decreased sharply in the presence of clay, while Fe(III)-reduction increased with no clear impact on U reduction. In the absence of clay, iron and sulfate reduction correlated with concentrations of Fe(III) and sulfate, respectively. Rates of U(VI) loss did not correlate with the concentration of any electron acceptor. With the exception of Fe(III), electron donor amendment was largely unsuccessful in stimulating electron acceptor loss over a 2-week incubation period, suggesting that endogenous forms of organic matter were sufficient to support microbial activity. Our findings suggest that efforts to accelerate biological U reduction should initially focus on stimulating nitrate removal.

  14. Metagenomic Insights into Anaerobic Metabolism along an Arctic Peat Soil Profile

    PubMed Central

    Lipson, David A.; Haggerty, John Matthew; Srinivas, Archana; Raab, Theodore K.; Sathe, Shashank; Dinsdale, Elizabeth A.

    2013-01-01

    A metagenomic analysis was performed on a soil profile from a wet tundra site in northern Alaska. The goal was to link existing biogeochemical knowledge of the system with the organisms and genes responsible for the relevant metabolic pathways. We specifically investigated how the importance of iron (Fe) oxides and humic substances (HS) as terminal electron acceptors in this ecosystem is expressed genetically, and how respiratory and fermentative processes varied with soil depth into the active layer and into the upper permafrost. Overall, the metagenomes reflected a microbial community enriched in a diverse range of anaerobic pathways, with a preponderance of known Fe reducing species at all depths in the profile. The abundance of sequences associated with anaerobic metabolic processes generally increased with depth, while aerobic cytochrome c oxidases decreased. Methanogenesis genes and methanogen genomes followed the pattern of CH4 fluxes : they increased steeply with depth into the active layer, but declined somewhat over the transition zone between the lower active layer and the upper permafrost. The latter was relatively enriched in fermentative and anaerobic respiratory pathways. A survey of decaheme cytochromes (MtrA, MtrC and their homologs) revealed that this is a promising approach to identifying potential reducers of Fe(III) or HS, and indicated a possible role for Acidobacteria as Fe reducers in these soils. Methanogens appear to coexist in the same layers, though in lower abundance, with Fe reducing bacteria and other potential competitors, including acetogens. These observations provide a rich set of hypotheses for further targeted study. PMID:23741360

  15. Dynamics of a microbial community exposed to several concentrations of 2-chlorophenol in an anaerobic sequencing batch reactor.

    PubMed

    Beristain-Montiel, Lizeth; Martínez-Hernández, Sergio; de María Cuervo-López, Flor; Ramírez-Vives, Florina

    2015-01-01

    The aim of this study was to contribute to the knowledge on the dynamic of the microbial community involved in anaerobic degradation of different concentrations of 2-chlorophenol (2CP, from 28 to 196 mg 2CP-C/L) and a mixture of 2CP and phenol (from 28 to 196 mg phenol-C/L) and its relationship with the respiratory process in two anaerobic sequencing batch reactors (ASBR). The dynamic of the microbial community was evaluated by denaturant gradient gel electrophoresis (DGGE) and ecological indices (S and J indices). The respiratory process was evaluated by means of substrate consumption efficiency, biogas yield, and specific consumption rates as response variables. The high consumption efficiency (90%) and the constant biogas yields obtained at concentrations up to 140 mg C/L may be related with the evenness of microbial populations (J index=0.97±0.2) present in both reactors. Pseudomonas genus was present in all concentrations tested, suggesting a possible relationship with the dehalogenation observed in both reactors. The decrease in specific consumption rate and biogas yield as well as the accumulation of phenol and volatile fatty acids observed in both reactors at 196 mg 2CP-C/L might be associated with the disappearance of the bands related to Caulobacter and Bacillus. At these conditions, the disappearance of fermentative or acetogenic bacteria resulted in reduction of substrates required to carry out methanogenesis, which eventually might cause the declination in methanogenic populations present in the reactors.

  16. On the possibility of chemosynthetic ecosystems in subsurface habitats on Mars.

    PubMed

    Boston, P J; Ivanov, M V; McKay, C P

    1992-01-01

    We have reexamined the question of extant microbial life on Mars in light of the most recent information about the planet and recently discovered nonphotosynthetic microbial ecosystems on Earth--deep sea hydrothermal vent communities and deep subsurface aquifer communities. On Mars, protected subsurface niches associated with hydrothermal activity could have continued to support life even after surface conditions became inhospitable. Geochemical evidence from the SNC meteorites and geomorphological evidence for recent volcanism suggest that such habitats could persist to the present time. There are also extensive geological features that attest to the ubiquitous nature of volcano-ground ice interactions on Mars. We suggest a possible deep subsurface microbial ecology similar to those discovered to depths of several kilometers below the surface of the Earth. We focus on anaerobic systems utilizing CO2 as the primary source of carbon. Liquid water could be provided by the heat of geothermal or volcanic activity melting permafrost or other subsurface water sources. Gases from volcanic activity deep in the planet could provide reducing power (as CH4, H2, or H2S) percolating up from below and enabling the development of a microbial community based upon chemolithoautotrophy. We suggest a methanogen (or acetogen) and sulfur-based microbial ecology as one possible basis for microbial primary production. Our hypothetical ecosystem is neither supported, nor excluded, by current observations of Mars. Tests for such a subsurface system involve locating active geothermal areas associated with ground ice or detecting trace quantities of reduced atmospheric gases that would leak from such a system.

  17. Ecophysiological adaptations of anaerobic bacteria to low pH: analysis of anaerobic digestion in acidic bog sediments. [Lactobacillus; Clostridium; Sarcina ventriculi

    SciTech Connect

    Goodwin, S.; Zeikus, G.J.

    1987-01-01

    The dynamics of anaerobic digestion were examined in the low-pH sediments of Crystal Bog in Wisconsin. The sediments (pH 4.9) contained 71% organic matter and the following concentrations of dissolved gases (micromoles per liter):CO/sub 2/, 1140; CH/sub 4/, 490; and H/sub 2/, 0.01. The rate of methane production was 6.2 ..mu..mol/liter of sediment per h, which is slower than eutrophic, neutral sediments. Microbial metabolic processes displayed the following pH optima: hydrolysis reactions, between 4.2 and 5.6; aceticlastic methanogenesis, 5.2; and hydrogen-consuming reactions, 5.6. The turnover rate constants for key intermediary metabolites were (h/sup -1/): glucose, 1.10; lactate, 0.277; acetate, 0.118; and ethanol, 0.089. The populations of anaerobes were low, with hydrolytic groups (10/sup 6//ml) several orders of magnitude higher than methanogens (10/sup 2//ml). The addition of carbon electron donors to the sediment resulted in the accumulation of hydrogen, whereas the addition of hydrogen resulted in the accumulation of fatty acids and the inhibition of hydrogen-producing acetogenic reactions. Strains of Lactobacillus, Clostridium, and Sarcina ventriculi were isolated from the bog, and their physiological attributes were characterized in relation to hydrolytic process functions in the sediments. The present studies provide evidence that the pH present in the bog sediments alter anaerobic digestion processes s, that total biocatalytic activity is lower bu the general carbon and electron flow pathways are similar to those of neutral anoxic sediments.

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

    PubMed

    Ohtomo, Yoko; Ijiri, Akira; Ikegawa, Yojiro; Tsutsumi, Masazumi; Imachi, Hiroyuki; Uramoto, Go-Ichiro; Hoshino, Tatsuhiko; Morono, Yuki; Sakai, Sanae; Saito, Yumi; Tanikawa, Wataru; Hirose, Takehiro; Inagaki, Fumio

    2013-01-01

    Geological CO2 sequestration in unmineable subsurface oil/gas fields and coal formations has been proposed as a means of reducing anthropogenic greenhouse gasses in the atmosphere. However, the feasibility of injecting CO2 into subsurface depends upon a variety of geological and economic conditions, and the ecological consequences are largely unpredictable. In this study, we developed a new flow-through-type reactor system to examine potential geophysical, geochemical and microbiological impacts associated with CO2 injection by simulating in-situ pressure (0-100 MPa) and temperature (0-70°C) conditions. Using the reactor system, anaerobic artificial fluid and CO2 (flow rate: 0.002 and 0.00001 ml/min, respectively) were continuously supplemented into a column comprised of bituminous coal and sand under a pore pressure of 40 MPa (confined pressure: 41 MPa) at 40°C for 56 days. 16S rRNA gene analysis of the bacterial components showed distinct spatial separation of the predominant taxa in the coal and sand over the course of the experiment. Cultivation experiments using sub-sampled fluids revealed that some microbes survived, or were metabolically active, under CO2-rich conditions. However, no methanogens were activated during the experiment, even though hydrogenotrophic and methylotrophic methanogens were obtained from conventional batch-type cultivation at 20°C. During the reactor experiment, the acetate and methanol concentration in the fluids increased while the δ(13)Cacetate, H2 and CO2 concentrations decreased, indicating the occurrence of homo-acetogenesis. 16S rRNA genes of homo-acetogenic spore-forming bacteria related to the genus Sporomusa were consistently detected from the sandstone after the reactor experiment. Our results suggest that the injection of CO2 into a natural coal-sand formation preferentially stimulates homo-acetogenesis rather than methanogenesis, and that this process is accompanied by biogenic CO2 conversion to acetate.

  19. Denitrification by Pseudomonas stutzeri coupled with CO2 reduction by Sporomusa ovata with hydrogen as an electron donor assisted by solid-phase humin.

    PubMed

    Xiao, Zhixing; Awata, Takanori; Zhang, Dongdong; Katayama, Arata

    2016-09-01

    A co-culture system comprising an acetogenic bacterium, Sporomusa ovata DSMZ2662, and a denitrifying bacterium, Pseudomonas stutzeri JCM20778, enabled denitrification using H2 as the sole external electron donor and CO2 as the sole external carbon source. Acetate produced by S. ovata supported the heterotrophic denitrification of P. stutzeri. A nitrogen balance study showed the reduction of nitrate to nitrogen gas without the accumulation of nitrite and nitrous oxide in the co-culture system. S. ovata did not show nitrate reduction to ammonium in the co-culture system. Significant proportions of the consumed H2 were utilized for denitrification: 79.9 ± 4.6% in the co-culture system containing solid-phase humin and 62.9±11.1% in the humin-free co-culture system. The higher utilization efficiency of hydrogen in the humin-containing system was attributed to the higher denitrification activity of P. stutzeri under the acetate deficient conditions. The nitrogen removal rate of the humin-containing co-culture system reached 0.19 kg NO3(-)-N·m(-3)·d(-1). Stable denitrification activity for 61 days of successive sub-culturing suggested the robustness of this co-culture system. This study provides a novel strategy for the in situ enhancement of microbial denitrification. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  20. Carbon flow from volcanic CO2 into soil microbial communities of a wetland mofette

    PubMed Central

    Beulig, Felix; Heuer, Verena B; Akob, Denise M; Viehweger, Bernhard; Elvert, Marcus; Herrmann, Martina; Hinrichs, Kai-Uwe; Küsel, Kirsten

    2015-01-01

    Effects of extremely high carbon dioxide (CO2) concentrations on soil microbial communities and associated processes are largely unknown. We studied a wetland area affected by spots of subcrustal CO2 degassing (mofettes) with focus on anaerobic autotrophic methanogenesis and acetogenesis because the pore gas phase was largely hypoxic. Compared with a reference soil, the mofette was more acidic (ΔpH ∼0.8), strongly enriched in organic carbon (up to 10 times), and exhibited lower prokaryotic diversity. It was dominated by methanogens and subdivision 1 Acidobacteria, which likely thrived under stable hypoxia and acidic pH. Anoxic incubations revealed enhanced formation of acetate and methane (CH4) from hydrogen (H2) and CO2 consistent with elevated CH4 and acetate levels in the mofette soil. 13CO2 mofette soil incubations showed high label incorporations with ∼512 ng 13C g (dry weight (dw)) soil−1 d−1 into the bulk soil and up to 10.7 ng 13C g (dw) soil−1 d−1 into almost all analyzed bacterial lipids. Incorporation of CO2-derived carbon into archaeal lipids was much lower and restricted to the first 10 cm of the soil. DNA-SIP analysis revealed that acidophilic methanogens affiliated with Methanoregulaceae and hitherto unknown acetogens appeared to be involved in the chemolithoautotrophic utilization of 13CO2. Subdivision 1 Acidobacteriaceae assimilated 13CO2 likely via anaplerotic reactions because Acidobacteriaceae are not known to harbor enzymatic pathways for autotrophic CO2 assimilation. We conclude that CO2-induced geochemical changes promoted anaerobic and acidophilic organisms and altered carbon turnover in affected soils. PMID:25216086

  1. Towards a paleo-salinity proxy: Decreasing D/H fractionation in algal and bacterial lipids with increasing salinity in Christmas Island saline ponds

    NASA Astrophysics Data System (ADS)

    Sachse, D.; Sachs, J. P.

    2007-12-01

    We investigated the effect of a wide range of salinities (13 -149 PSU) on the D/H ratio of lipids in microbial mat sediments from hypersaline ponds on Christmas Island. The hydrogen isotope ratios (expressed as δD values) of total lipid extracts, and the individual hydrocarbons heptadecane, heptadecene, octadecane, octadecene, diploptene and phytene from algae and bacteria, became increasingly enriched in deuterium as salinity increased, spanning a range of 100‰ while lake water δD values spanned a range of just 12‰. D/H fractionation between lipids and source water thus decreased as salinity increased. Isotope fractionation factors (αlipid-water) were strongly correlated with salinity and increased in all compound classes studied. The apparent isotope fractionation (ɛlipid-water) decreased by 0.8 to 1.1‰ per PSU increase in salinity. Differences in the hydrogen isotopic composition of lipids derived from three biosynthetic pathways (acetogenic, MVA and DOXP/MEP) remained similar irrespective of the salinity, suggesting that the mechanism responsible for the observed αlipid-water - salinity relationship originates prior to the last common biosynthetic branching point, the Calvin Cycle. These findings imply that caution must be exercised when attempting to reconstruct source water δD values using lipid δD values from aquatic environments that may have experienced salinity variations of ~3 PSU or more (based on a 1‰ per PSU response of D/H fractionation to salinity changes, and a lipid δD measurement precision of 3‰). On the other hand our results can be used to establish a paleo-salinity proxy based on algal and bacterial lipid δD values if salinity variations exceeded ~3 PSU and/or if additional constraints on source water δD values can be made.

  2. Inverse relationship between D/H fractionation in cyanobacterial lipids and salinity in Christmas Island saline ponds

    NASA Astrophysics Data System (ADS)

    Sachse, Dirk; Sachs, Julian P.

    2008-02-01

    Sediments from 28 saline and hypersaline (salinity 13.6-149.2) ponds on Christmas Island (Kiritimati), in the Central tropical Pacific Ocean, were investigated for the effect of salinity on the D/H ratios of lipid biomarkers. Hydrogen isotope ratios (expressed as δD values) of total lipid extracts, and individual hydrocarbons heptadecane, heptadecene, octadecane, octadecene, diploptene, and phytene from cyanobacteria, became increasingly enriched in deuterium as salinity increased, spanning a range of 100‰, while lake water δD values spanned a range of just 12‰. Net D/H fractionation between lipids and source water thus decreased as salinity increased. Isotope fractionation factors ( αlipid-water) were strongly correlated with salinity, and increased in all compound classes studied by up to 0.0967 over a salinity range of 136. Differences in the hydrogen isotopic composition of lipids derived from three biosynthetic pathways (acetogenic, mevalonate, and non-mevalonate) remained similar irrespective of the salinity. This suggests that the mechanism responsible for the observed αlipid-water-salinity relationship originates prior to the last common biosynthetic branching point, the Calvin Cycle. We propose that a decrease in the exchange of intra- and extra-cellular (ambient) water resulting from down-regulation or closure of water channels (aquaporins) within cyanobacterial cell membranes, and subsequent isotopic enrichment of the intracellular water, likely resulting from metabolic reactions. These findings imply that caution must be exercised when attempting to reconstruct source water δD values using lipid δD values from environments that may have experienced salinity variations. On the other, hand our results can be used to establish a paleo-salinity proxy based on lipid δD, if additional constraints on source water δD values can be made.

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

    PubMed Central

    Fermoso, Fernando G.; Collins, Gavin; Bartacek, Jan; O’Flaherty, Vincent

    2008-01-01

    The effect of nickel deprivation from the influent of a mesophilic (30°C) methanol fed upflow anaerobic sludge bed (UASB) reactor was investigated by coupling the reactor performance to the evolution of the Methanosarcina population of the bioreactor sludge. The reactor was operated at pH 7.0 and an organic loading rate (OLR) of 5–15 g COD l−1 day−1 for 191 days. A clear limitation of the specific methanogenic activity (SMA) on methanol due to the absence of nickel was observed after 129 days of bioreactor operation: the SMA of the sludge in medium with the complete trace metal solution except nickel amounted to 1.164 (±0.167) g CH4-COD g VSS−1 day−1 compared to 2.027 (±0.111) g CH4-COD g VSS−1 day−1 in a medium with the complete (including nickel) trace metal solution. The methanol removal efficiency during these 129 days was 99%, no volatile fatty acid (VFA) accumulation was observed and the size of the Methanosarcina population increased compared to the seed sludge. Continuation of the UASB reactor operation with the nickel limited sludge lead to incomplete methanol removal, and thus methanol accumulation in the reactor effluent from day 142 onwards. This methanol accumulation subsequently induced an increase of the acetogenic activity in the UASB reactor on day 160. On day 165, 77% of the methanol fed to the system was converted to acetate and the Methanosarcina population size had substantially decreased. Inclusion of 0.5 μM Ni (dosed as NiCl2) to the influent from day 165 onwards lead to the recovery of the methanol removal efficiency to 99% without VFA accumulation within 2 days of bioreactor operation. PMID:18247139

  4. Microbial electron transport and energy conservation - the foundation for optimizing bioelectrochemical systems.

    PubMed

    Kracke, Frauke; Vassilev, Igor; Krömer, Jens O

    2015-01-01

    Microbial electrochemical techniques describe a variety of emerging technologies that use electrode-bacteria interactions for biotechnology applications including the production of electricity, waste and wastewater treatment, bioremediation and the production of valuable products. Central in each application is the ability of the microbial catalyst to interact with external electron acceptors and/or donors and its metabolic properties that enable the combination of electron transport and carbon metabolism. And here also lies the key challenge. A wide range of microbes has been discovered to be able to exchange electrons with solid surfaces or mediators but only a few have been studied in depth. Especially electron transfer mechanisms from cathodes towards the microbial organism are poorly understood but are essential for many applications such as microbial electrosynthesis. We analyze the different electron transport chains that nature offers for organisms such as metal respiring bacteria and acetogens, but also standard biotechnological organisms currently used in bio-production. Special focus lies on the essential connection of redox and energy metabolism, which is often ignored when studying bioelectrochemical systems. The possibility of extracellular electron exchange at different points in each organism is discussed regarding required redox potentials and effect on cellular redox and energy levels. Key compounds such as electron carriers (e.g., cytochromes, ferredoxin, quinones, flavins) are identified and analyzed regarding their possible role in electrode-microbe interactions. This work summarizes our current knowledge on electron transport processes and uses a theoretical approach to predict the impact of different modes of transfer on the energy metabolism. As such it adds an important piece of fundamental understanding of microbial electron transport possibilities to the research community and will help to optimize and advance bioelectrochemical

  5. Physiology and Sporulation in Clostridium.

    PubMed

    Dürre, Peter

    2014-08-01

    Clostridia are Gram-positive, anaerobic, endospore-forming bacteria, incapable of dissimilatory sulfate reduction. Comprising approximately 180 species, the genus Clostridium is one of the largest bacterial genera. Physiology is mostly devoted to acid production. Numerous pathways are known, such as the homoacetate fermentation by acetogens, the propionate fermentation by Clostridium propionicum, and the butyrate/butanol fermentation by C. acetobutylicum, a well-known solvent producer. Clostridia degrade sugars, alcohols, amino acids, purines, pyrimidines, and polymers such as starch and cellulose. Energy conservation can be performed by substrate-level phosphorylation as well as by the generation of ion gradients. Endospore formation resembles the mechanism elucidated in Bacillus. Morphology, contents, and properties of spores are very similar to bacilli endospores. Sporulating clostridia usually form swollen mother cells and accumulate the storage substance granulose. However, clostridial sporulation differs by not employing the so-called phosphorelay. Initiation starts by direct phosphorylation of the master regulator Spo0A. The cascade of sporulation-specific sigma factors is again identical to what is known from Bacillus. The onset of sporulation is coupled in some species to either solvent (acetone, butanol) or toxin (e.g., C. perfringens enterotoxin) formation. The germination of spores is often induced by various amino acids, often in combination with phosphate and sodium ions. In medical applications, C. butyricum spores are used as a C. difficile prophylaxis and as treatment against diarrhea. Recombinant spores are currently under investigation and testing as antitumor agents, because they germinate only in hypoxic tissues (i.e., tumor tissue), allowing precise targeting and direct killing of tumor cells.

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

    SciTech Connect

    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.

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

    PubMed

    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. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Characterization of anaerobic chloroethene-dehalogenating activity in several subsurface sediments

    SciTech Connect

    Skeen, R.S.; Gao, J.; Hooker, B.S.; Quesenberry, R.D.

    1996-11-01

    Anaerobic microcosms of subsurface soils from four locations were used to investigate the separate effects of several electron donors on tetrachloroethylene (PCE) dechlorination activity. The substrates tested were methanol, formate, lactate, acetate, and sucrose. Various levels of sulfate-reducing, acetogenic, fermentative, and methanogenic activity were observed in all sediments. PCE dechlorination was detected in all microcosms, but the amount of dehalogenation varied by several orders of magnitude. Trichloroethylene was the primary dehalogenation product; however, small amounts of cis-1,2-dichloroethylene, 1,1-dichloroethylene, and vinyl chloride were also detected in several microcosms. Lactate-amended microcosms showed large amounts of dehalogenation. in three of the four sediments. One of the two sediments which showed positive activity with lactate also had large amounts of delialogenation with methanol. Sucrose, formate, and acetate also stimulated large amounts of delialogenation in one sediment that showed activity with lactate. These results suggest that lactate may be an appropriate substrate for screening sediments for PCE or TCE delialogenation activity, but that the microbial response is not sufficient for complete in situ bioremediation. A detailed study of the Victoria activity revealed that delialogenation rates were more similar to the Cornell culture than to rates measured for methanogens, or a methanol-enriched sediment culture. This may suggest that these sediments contain a highly efficient delialogenation activity similar to the Cornell culture. This assertion is supported further by the fact that an average of 3% of added reducing equivalents could be diverted to dehalogenation in tests which were conducted using PCE-saturated hexadecane as a constant source of PCE during incubation. Further evidence is needed to confirm this premise. The application of these results to in situ bioremediation of highly contaminated areas are discussed.

  9. Volatile Fatty Acid Production by the Hindgut Microbiota of Xylophagous Termites †

    PubMed Central

    Odelson, David A.; Breznak, John A.

    1983-01-01

    Acetate dominated the extracellular pool of volatile fatty acids (VFAs) in the hindgut fluid of Reticulitermes flavipes, Zootermopsis angusticollis, and Incisitermes schwarzi, where it occurred at concentrations of 57.9 to 80.6 mM and accounted for 94 to 98 mol% of all VFAs. Small amounts of C3 to C5 VFAs were also observed. Acetate was also the major VFA in hindgut homogenates of Schedorhinotermes lamanianus, Prorhinotermes simplex, Coptotermes formosanus, and Nasutitermes corniger. Estimates of in situ acetogenesis by the hindgut microbiota of R. flavipes (20.2 to 43.3 nmol · termite−1 · h−1) revealed that this activity could support 77 to 100% of the respiratory requirements of the termite (51.6 to 63.6 nmol of O2 · termite−1 · h−1). This conclusion was buttressed by the demonstration of acetate in R. flavipes hemolymph (at 9.0 to 11.6 mM), but not in feces, and by the ability of termite tissues to readily oxidize acetate to CO2. About 85% of the acetate produced by the hindgut microbiota was derived from cellulose C; the remainder was derived from hemicellulose C. Selective removal of major groups of microbes from the hindgut of R. flavipes indicated that protozoa were primarily responsible for acetogenesis but that bacteria also functioned in this capacity. H2 and CH4 were evolved by R. flavipes (usually about 0.4 nmol · termite−1 · h−1), but these compounds represented a minor fate of electrons derived from wood dissimilation within R. flavipes. A working model is proposed for symbiotic wood polysaccharide degradation in R. flavipes, and the possible roles of individual gut microbes, including CO2-reducing acetogenic bacteria, are discussed. PMID:16346296

  10. Autotrophic fixation of geogenic CO2 by microorganisms contributes to soil organic matter formation and alters isotope signatures in a wetland mofette

    NASA Astrophysics Data System (ADS)

    Nowak, M. E.; Beulig, F.; von Fischer, J.; Muhr, J.; Küsel, K.; Trumbore, S. E.

    2015-09-01

    quantification of cbbL/cbbM marker genes encoding for RubisCO by quantitative polymerase chain reaction (qPCR) and by acetogenic and methanogenic microorganisms, shown present in the moffettes by previous studies. Combined Δ14C and δ13C isotope mass balances indicated that microbially derived carbon accounted for 8 to 27 % of bulk SOM in this soil layer. The findings imply that autotrophic organisms can recycle significant amounts of carbon in wetland soils and might contribute to observed reservoir effects influencing radiocarbon signatures in peat deposits.

  11. Lipid D/H Ratios from Multiple Sources and Deposits Indicate Drier Little Ice Age at Washington Island (4°43`N, 160°25`W), Central Pacific

    NASA Astrophysics Data System (ADS)

    Muegler, I.; Sachse, D.; Sachs, J. P.

    2010-12-01

    To compare the sensitivity of biomarker D/H ratios from two distinct climate archives, a lake and a peat bog on the Tropical Pacific Island of Terrania, compound-specific hydrogen isotope ratios (expressed as δD values) were determined on lipid biomarkers from various biological sources deposited in the two climate archives. At present, Terrania or Washington Island (4°43`N, 160°25`W) permanently lies in the intertropical convergence zone (ITCZ) and receives an annual precipitation of 2,903 mm. The interior of this lens shaped island contains a freshwater lake and peat bogs. Previous studies on the lake sediments found evidence for a substantially drier climate at times during the Little Ice Age (AD 1400-1850) based on the lithologic transition from modern freshwater sediments to a sequence of pure cyanobacterial mat in concert with δD values from total lipid extracts (Sachs et al., 2009). Here we report on δD values from lipids of various sources: dinoflagellate algae (dinosterol and a saturated C30 sterol (4α -methyl-24-ethyl-5α -cholestan-3β-ol), microbial sources (diploptene and nC21 alkane) and higher plants (fern-7-ene, β-sitosterol and stigmastanol). Mean δD values from all lipids, measured in both archives, are significantly enriched in deuterium by between 22 and 86‰ during previously inferred drier climate conditions and simultaneously record the transition towards a freshwater lake at around AD1550. Measured δD values of all lipids cover a wide range from -281‰ to -105‰ during freshwater deposition and from -185‰ to -50‰ when climate was drier. In agreement with the observed isotopic difference between lipids produced via the acetogenic and the mevalonic acid biosynthetic pathway δD values for algae and higher plant sterols are depleted in deuterium relative to the nC21 alkane by 150‰ on average. The consistent δD values from dinosterol and 4α -methyl-24-ethyl-5α -cholestan-3β-ol from the lake and peat deposits signify the

  12. Converting Carbon Dioxide to Butyrate with an Engineered Strain of Clostridium ljungdahlii

    SciTech Connect

    Ueki, T; Nevin, KP; Woodard, TL; Lovley, DR

    2014-08-26

    Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70% of the carbon and electron flow was diverted to the production of butyrate with H-2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. IMPORTANCE The development of a microbial chassis for efficient conversion of carbon dioxide directly to desired organic products would greatly advance the environmentally sustainable production of biofuels and other commodities. Clostridium ljungdahlii is an effective catalyst for microbial electrosynthesis, a technology in which electricity generated with renewable technologies, such as solar or wind, powers the conversion of carbon dioxide and water to organic products. Other electron donors

  13. Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii.

    PubMed

    Ueki, Toshiyuki; Nevin, Kelly P; Woodard, Trevor L; Lovley, Derek R

    2014-10-21

    Microbial conversion of carbon dioxide to organic commodities via syngas metabolism or microbial electrosynthesis is an attractive option for production of renewable biocommodities. The recent development of an initial genetic toolbox for the acetogen Clostridium ljungdahlii has suggested that C. ljungdahlii may be an effective chassis for such conversions. This possibility was evaluated by engineering a strain to produce butyrate, a valuable commodity that is not a natural product of C. ljungdahlii metabolism. Heterologous genes required for butyrate production from acetyl-coenzyme A (CoA) were identified and introduced initially on plasmids and in subsequent strain designs integrated into the C. ljungdahlii chromosome. Iterative strain designs involved increasing translation of a key enzyme by modifying a ribosome binding site, inactivating the gene encoding the first step in the conversion of acetyl-CoA to acetate, disrupting the gene which encodes the primary bifunctional aldehyde/alcohol dehydrogenase for ethanol production, and interrupting the gene for a CoA transferase that potentially represented an alternative route for the production of acetate. These modifications yielded a strain in which ca. 50 or 70% of the carbon and electron flow was diverted to the production of butyrate with H2 or CO as the electron donor, respectively. These results demonstrate the possibility of producing high-value commodities from carbon dioxide with C. ljungdahlii as the catalyst. Importance: The development of a microbial chassis for efficient conversion of carbon dioxide directly to desired organic products would greatly advance the environmentally sustainable production of biofuels and other commodities. Clostridium ljungdahlii is an effective catalyst for microbial electrosynthesis, a technology in which electricity generated with renewable technologies, such as solar or wind, powers the conversion of carbon dioxide and water to organic products. Other electron donors

  14. Hydrocarbon bio-jet fuel from bioconversion of poplar biomass: life cycle assessment.

    PubMed

    Budsberg, Erik; Crawford, Jordan T; Morgan, Hannah; Chin, Wei Shan; Bura, Renata; Gustafson, Rick

    2016-01-01

    Bio-jet fuels compatible with current aviation infrastructure are needed as an alternative to petroleum-based jet fuel to lower greenhouse gas emissions and reduce dependence on fossil fuels. Cradle to grave life cycle analysis is used to investigate the global warming potential and fossil fuel use of converting poplar biomass to drop-in bio-jet fuel via a novel bioconversion platform. Unique to the biorefinery designs in this research is an acetogen fermentation step. Following dilute acid pretreatment and enzymatic hydrolysis, poplar biomass is fermented to acetic acid and then distilled, hydroprocessed, and oligomerized to jet fuel. Natural gas steam reforming and lignin gasification are proposed to meet hydrogen demands at the biorefineries. Separate well to wake simulations are performed using the hydrogen production processes to obtain life cycle data. Both biorefinery designs are assessed using natural gas and hog fuel to meet excess heat demands. Global warming potential of the natural gas steam reforming and lignin gasification bio-jet fuel scenarios range from CO2 equivalences of 60 to 66 and 32 to 73 g MJ(-1), respectively. Fossil fuel usage of the natural gas steam reforming and lignin gasification bio-jet fuel scenarios range from 0.78 to 0.84 and 0.71 to 1.0 MJ MJ(-1), respectively. Lower values for each impact category result from using hog fuel to meet excess heat/steam demands. Higher values result from using natural gas to meet the excess heat demands. Bio-jet fuels produced from the bioconversion of poplar biomass reduce the global warming potential and fossil fuel use compared with petroleum-based jet fuel. Production of hydrogen is identified as a major source of greenhouse gas emissions and fossil fuel use in both the natural gas steam reforming and lignin gasification bio-jet simulations. Using hog fuel instead of natural gas to meet heat demands can help lower the global warming potential and fossil fuel use at the biorefineries.

  15. Would acetate (or its derivatives) be the most reliable guide to life on terraqueous globes?

    NASA Astrophysics Data System (ADS)

    Russell, Michael; Martin-Torres, Javier; Yung, Yuk; Kanik, Isik

    2010-05-01

    At bottom life hydrogenates carbon dioxide. But so does serpentinization-to methane-hence the problem of diagnosing its source (Mumma et al. 2009). However, this abiotic process does not appear to produce acetate or acetic acid (CH3COOH) in measurable quantities-only the acetogenic and sulfate-reducing bacteria do that. On the early Earth it seems that the homoacetogens were the first to resolve the tension between CO2 and H2 via the autotrophic acetyl coenzyme-A pathway. The acetyl co-A pathway employs two separate redox controlled tributaries-one Ni-Fe-directed, merely reduces CO2 to CO, while the other, initially molybdenum-directed, reduces CO2 through to a methyl group. The CO and the -CH3 are then assembled on the nickel-bearing acetyl coenzyme-A synthase. Such a complex dual delivery system from contrasting redox conditions could not be prefigured by serpentinization but required a chemiosmotic drive, as did the origin of life itself (Nitschke and Russell 2009). Homoacetogens can compete successfully against the methanoarchaea for H2 and CO2 in the cold, as can the sulfate-reducing acetate-generating bacteria (Krumholz et al. 1999). Thus we argue that acetate or acetic acid effluent (depending on pH) from putative microbes on wet rocky planets would be a more reliable indicator of life. What are the difficulties? The most critical is that in ground-waters and oceans with pH >5 acetate remains in solution and would therefore not be detectable remotely. Even were the waters acidic enough to release volatile acetic acid, it would be prone to photo- and chemical oxidation. However, apart from CO2 and CH4, the products are formic (HCOOH), glycolic (HOCH2.COOH) and tartaric (HOOC.HCOH.HOCH.COOH) acids (Ogata et al. 1981). Remote sensing in the ultraviolet to near-infrared might be used for detection of all these acids, especially when their concentrations are enhanced in plumes. In situ techniques would be required for acetate detection. Krumholz, L.R. et al

  16. Salt-tolerant and high-pH-resistant hydrogenase from the haloalkaliphilic, sulfate-reducing bacterium Desulfonatronum thiodismutans

    NASA Astrophysics Data System (ADS)

    Detkova, Ekaterina N.; Pikuta, Elena V.; Hoover, Richard B.

    2004-11-01

    Hydrogenase is the key enzyme of energetic metabolism in cells, catalyzing the converse reaction of hydrogen oxidation and responsible for the consumption and excretion of hydrogen in bacteria. Hydrogenases are proteins, most of which contain either nickel and iron or iron alone in their active center. Hydrogenases have been found in many microorganisms, such as methanogenic, acetogenic, nitrogen-fixing, sulfate-reducing, photosynthetic bacteria, and algae that use the hydrogen as an energy source or as an electron sink. Hydrogenases are the subject of wide physiological, biochemical, physico-chemical and genetic studies due to their abilities to produce molecular hydrogen as an alternative source of energy. Despite the large quantity of work dealing with the intracellular and extracellular enzymes of halophilic bacteria, the data about the response of hydrogenases to salts are practically absent. The study of hydrogenase in cell-free extracts of the extremely halophilic eubacterium Acetohalobium arabaticum showed a dramatic increase in the activity of the enzyme at high concentrations of NaCl and KCl (near saturated solutions). Here we present data about hydrogenase in a free-cell extract from the new halo-alkaliphilic sulfate-reducing bacterium Desulfonatronum thiodismutans, which grows on a highly mineralized carbonate-bicarbonate medium in the salinity range from 1 to 7 % NaCl and at pH 8.0-10.0. The studied enzyme was active in concentration range from 0.0 to 4.3 M NaCl with the optimum at 1.0 M NaCl. At 1.0 M NaCl the enzyme expressed 20 % additional activity, with NaCl concentration changing from 2.1 M to 3.4 M, and then the activity decreased and reached a constant level. Although sodium bicarbonate decreases the hydrogenase activity, the enzyme still showed activity at 60 % of the maximum level at concentration in a near saturated solution (1.2 M NaHCO3). The maximum enzyme activity was observed at pH 9.5 with limits of 7.5 and 11.5, which is practically

  17. Variations in microbial carbon sources and cycling in the deep continental subsurface

    NASA Astrophysics Data System (ADS)

    Simkus, Danielle N.; Slater, Greg F.; Lollar, Barbara Sherwood; Wilkie, Kenna; Kieft, Thomas L.; Magnabosco, Cara; Lau, Maggie C. Y.; Pullin, Michael J.; Hendrickson, Sarah B.; Wommack, K. Eric; Sakowski, Eric G.; van Heerden, Esta; Kuloyo, Olukayode; Linage, Borja; Borgonie, Gaetan; Onstott, Tullis C.

    2016-01-01

    Deep continental subsurface fracture water systems, ranging from 1.1 to 3.3 km below land surface (kmbls), were investigated to characterize the indigenous microorganisms and elucidate microbial carbon sources and their cycling. Analysis of phospholipid fatty acid (PLFA) abundances and direct cell counts detected varying biomass that was not correlated with depth. Compound-specific carbon isotope analyses (δ13C and Δ14C) of the phospholipid fatty acids (PLFAs) and carbon substrates combined with genomic analyses did identify, however, distinct carbon sources and cycles between the two depth ranges studied. In the shallower boreholes at circa 1 kmbls, isotopic evidence indicated microbial incorporation of biogenic CH4 by the in situ microbial community. At the shallowest site, 1.05 kmbls in Driefontein mine, this process clearly dominated the isotopic signal. At slightly deeper depths, 1.34 kmbls in Beatrix mine, the isotopic data indicated the incorporation of both biogenic CH4 and dissolved inorganic carbon (DIC) derived from CH4 oxidation. In both of these cases, molecular genetic analysis indicated that methanogenic and methanotrophic organisms together comprised a small component (<5%) of the microbial community. Thus, it appears that a relatively minor component of the prokaryotic community is supporting a much larger overall bacterial community in these samples. In the samples collected from >3 kmbls in Tau Tona mine (TT107, TT109 Bh2), the CH4 had an isotopic signature suggesting a predominantly abiogenic origin with minor inputs from microbial methanogenesis. In these samples, the isotopic enrichments (δ13C and Δ14C) of the PLFAs relative to CH4 were consistent with little incorporation of CH4 into the biomass. The most 13C-enriched PLFAs were observed in TT107 where the dominant CO2-fixation pathway was the acetyl-CoA pathway by non-acetogenic bacteria. The differences in the δ13C of the PLFAs and the DIC and DOC for TT109 Bh2 were ∼-24‰ and 0

  18. The stable carbon isotope biogeochemistry of acetate and other dissolved carbon species in deep subseafloor sediments at the northern Cascadia Margin

    USGS Publications Warehouse

    Heuer, Verena B.; Pohlman, John W.; Torres, Marta E.; Elvert, Marcus; Hinrichs, Kai-Uwe

    2009-01-01

    Ocean drilling has revealed the existence of vast microbial populations in the deep subseafloor, but to date little is known about their metabolic activities. To better understand the biogeochemical processes in the deep biosphere, we investigate the stable carbon isotope chemistry of acetate and other carbon-bearing metabolites in sediment pore-waters. Acetate is a key metabolite in the cycling of carbon in anoxic sediments. Its stable carbon isotopic composition provides information on the metabolic processes dominating acetate turnover in situ. This study reports our findings for a methane-rich site at the northern Cascadia Margin (NE Pacific) where Expedition 311 of the Integrated Ocean Drilling Program (IODP) sampled the upper 190 m of sediment. At Site U1329, δ13C values of acetate span a wide range from −46.0‰ to −11.0‰ vs. VPDB and change systematically with sediment depth. In contrast, δ13C values of both the bulk dissolved organic carbon (DOC) (−21.6 ± 1.3‰ vs. VPDB) and the low-molecular-weight compound lactate (−20.9 ± 1.8‰ vs. VPDB) show little variability. These species are interpreted to represent the carbon isotopic composition of fermentation products. Relative to DOC, acetate is up to 23.1‰ depleted and up to 9.1‰ enriched in 13C. Broadly, 13C-depletions of acetate relative to DOC indicate flux of carbon from acetogenesis into the acetate pool while 13C-enrichments of pore-water acetate relative to DOC suggest consumption of acetate by acetoclastic methanogenesis. Isotopic relationships between acetate and lactate or DOC provide new information on the carbon flow and the presence and activity of specific functional microbial communities in distinct biogeochemical horizons of the sediment. In particular, they suggest that acetogenic CO2-reduction can coexist with methanogenic CO2-reduction, a notion contrary to the hypothesis that hydrogen levels are controlled by the thermodynamically most favorable electron

  19. Linked metatranscriptomic and geochemical data indicate microbial succession in naturally reduced aquifer sediments dominated by H2-oxidizing Comamonadaceae

    NASA Astrophysics Data System (ADS)

    Jewell, T. N. M.; Karaoz, U.; Bill, M.; Chakraborty, R.; Brodie, E.; Williams, K. H.; Beller, H. R.

    2015-12-01

    In this study, we sought to better understand what natural organic matter fuels heterotrophic microbial communities in the anoxic subsurface at the Rifle (CO) site and what genes may be diagnostic of that activity. We conducted a 20-day microcosm experiment with naturally reduced zone (NRZ) sediments and collected replicate samples every 5 days for omics (metagenome and metatranscriptome) and biogeochemical measurements (e.g., continuous CO2 production, H2, CH4, acetate, DOC, Fe(II), sulfate, NH4+, spectroscopic analyses of sediment OM). No electron donors were added other than the NRZ sediment, which is enriched in organic matter relative to typical Rifle aquifer material. The microcosms were constructed and incubated under anaerobic conditions in serum bottles with a N2headspace. Biogeochemical measurements indicate that the decomposition of native organic matter occurred in different phases, including depletion of DOC and release of CO2 during the first week of incubation, followed by a pulse of acetogenesis and methanogenesis after 2 weeks (with acetogenesis dominating carbon flux after 2 weeks). While H2 remained below detection levels throughout the study, a peak of [NiFe] uptake hydrogenase, acetyl-CoA synthetase, urease, and nitrate reductase transcripts belonging to the Comamonadaceae family occurred at day 15. Some members of Comamonadaceae are facultative H2-oxidizing chemolithoautotrophs and fix carbon via the acetogenic Wood-Ljungdahl pathway. Comamonadaceae plateaued at 73% of the metagenome at this time and represented 69% of the metatranscriptome, succeeding the S-oxidizing Sulfurimonas genus. Sulfurimonas species were the dominant group at day 0, accounting for 43% of the metagenome and 25% of the metatranscript