Metabolic interactions in methanogenic and sulfate-reducing bioreactors.

Science.gov (United States)

Stams, A J M; Plugge, C M; de Bok, F A M; van Houten, B H G W; Lens, P; Dijkman, H; Weijma, J

2005-01-01

In environments where the amount of electron acceptors 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 consortia of acetogenic bacteria and methanogenic archaea. Hydrogen consumption by methanogens is essential for acetogenic bacteria to convert organic acids to acetate and hydrogen. Several syntrophic cocultures growing on propionate and butyrate have been described. These syntrophic fatty acid-degrading consortia are affected by the presence of sulfate. When sulfate is present sulfate-reducing bacteria compete with methanogenic archaea for hydrogen and acetate, and with acetogenic bacteria for propionate and butyrate. Sulfate-reducing bacteria easily outcompete methanogens for hydrogen, but the presence of acetate as carbon source may influence the outcome of the competition. By contrast, acetoclastic methanogens can compete reasonably well with acetate-degrading sulfate reducers. Sulfate-reducing bacteria grow much faster on propionate and butyrate than syntrophic consortia.

Sulfate reduction with methanol by a thermophilic consortium obtained from a methanogenic reactor

Energy Technology Data Exchange (ETDEWEB)

Davidova, I.A. [Wageningen Agricultural Univ. (Netherlands). Dept. of Microbiology; Stams, A.J.M. [Wageningen Agricultural Univ. (Netherlands). Dept. of Microbiology

1996-12-31

An enrichment culture obtained from anaerobic granular sludge of a bench-scale anarobic reactor degraded methanol at 65 C via sulfate reduction and acetogenesis. Sulfate reduction was the dominant process (S{sup 2-}/acetate=2.5). No methane formation was observed. Approximately 30% of the methanol was converted by acetogenic bacteria to acetate, while the remainder was degraded by these bacteria to H{sub 2} and CO{sub 2} in syntrophy with hydrogen-consuming sulfate-reducing bacteria. Pure cultures of sulfate-reducing and acetogenic bacteria were isolated and characterized. (orig.)

A deeply branching thermophilic bacterium with an ancient acetyl-CoA pathway dominates a subsurface ecosystem.

Directory of Open Access Journals (Sweden)

Hideto Takami

Full Text Available 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 CO(2 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 H(2-dependent acetogens and methanogenes living in hydrothermal environments.

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

Science.gov (United States)

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

Metabolic engineering of Clostridium autoethanogenum for selective alcohol production.

Science.gov (United States)

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

2017-03-01

Gas fermentation using acetogenic bacteria such as Clostridium autoethanogenum offers an attractive route for production of fuel ethanol from industrial waste gases. Acetate reduction to acetaldehyde and further to ethanol via an aldehyde: ferredoxin oxidoreductase (AOR) and alcohol dehydrogenase has been postulated alongside the classic pathway of ethanol formation via a bi-functional aldehyde/alcohol dehydrogenase (AdhE). Here we demonstrate that AOR is critical to ethanol formation in acetogens and inactivation of AdhE led to consistently enhanced autotrophic ethanol production (up to 180%). Using ClosTron and allelic exchange mutagenesis, which was demonstrated for the first time in an acetogen, we generated single mutants as well as double mutants for both aor and adhE isoforms to confirm the role of each gene. The aor1+2 double knockout strain lost the ability to convert exogenous acetate, propionate and butyrate into the corresponding alcohols, further highlighting the role of these enzymes in catalyzing the thermodynamically unfavourable reduction of carboxylic acids into alcohols. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Next-Gen³: Sequencing, Modeling, and Advanced Biofuels - Final Technical Report

Energy Technology Data Exchange (ETDEWEB)

Zengler, Karsten [Univ. of California, San Diego, CA (United States). Dept. of Pediatrics; Palsson, Bernhard [Univ. of California, San Diego, CA (United States). Dept. of Bioengineering; Lewis, Nathan [Univ. of California, San Diego, CA (United States). Dept. of Pediatrics

2017-12-27

Successful, scalable implementation of biofuels is dependent on the efficient and near complete utilization of diverse biomass sources. One approach is to utilize the large recalcitrant biomass fraction (or any organic waste stream) through the thermochemical conversion of organic compounds to syngas, a mixture of carbon monoxide (CO), carbon dioxide (CO₂), and hydrogen (H₂), which can subsequently be metabolized by acetogenic microorganisms to produce next-gen biofuels. The goal of this proposal was to advance the development of the acetogen Clostridium ljungdahlii as a chassis organism for next-gen biofuel production from cheap, renewable sources and to detail the interconnectivity of metabolism, energy conservation, and regulation of acetogens using next-gen sequencing and next-gen modeling. To achieve this goal we determined optimization of carbon and energy utilization through differential translational efficiency in C. ljungdahlii. Furthermore, we reconstructed a next-generation model of all major cellular processes, such as macromolecular synthesis and transcriptional regulation and deployed this model to predicting proteome allocation, overflow metabolism, and metal requirements in this model acetogen. In addition we explored the evolutionary significance of tRNA operon structure using the next-gen model and determined the optimal operon structure for bioproduction. Our study substantially enhanced the knowledgebaase for chemolithoautotrophs and their potential for advanced biofuel production. It provides next-generation modeling capability, offer innovative tools for genome-scale engineering, and provide novel methods to utilize next-generation models for the design of tunable systems that produce commodity chemicals from inexpensive sources.

Optimisation of the two-phase dry-thermophilic anaerobic digestion process of sulphate-containing municipal solid waste: population dynamics.

Science.gov (United States)

Zahedi, S; Sales, D; Romero, L I; Solera, R

2013-11-01

Microbial population dynamics and anaerobic digestion (AD) process to eight different hydraulic retention times (HRTs) (from 25d to 3.5d) in two-phase dry-thermophilic AD from sulphate-containing solid waste were investigated. Maximum values of gas production (1.9 ± 0.2 l H2/l/d; 5.4 ± 0.3 l CH4/l/d and 82 ± 9 ml H2S/l/d) and microbial activities were obtained at 4.5d HRT; where basically comprised hydrolysis step in the first phase (HRT=1.5d) and acidogenic step finished in the second phase as well as acetogenic-methanogenic steps (HRT=3d). In the first phase, hydrolytic-acidogenic bacteria (HABs) was the main group (44-77%) and Archaea, acetogens and sulphate-reducing bacteria (SRBs) contents were not significant; in the second phase (except to 2d HRT), microbial population was able to adapt to change in substrate and HRTs to ensure the proper functioning of the system and both acetogens and Archaea were dominated over SRBs. Decreasing HRT resulted in an increase in microbial activities. Copyright © 2013 Elsevier Ltd. All rights reserved.

Acetogenesis in the energy-starved deep biosphere – a paradox?

Directory of Open Access Journals (Sweden)

Mark Alexander Lever

2012-01-01

Full Text Available 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.

Metagenomic analysis of the rumen microbial community following inhibition of methane formation by a halogenated methane analogue

Directory of Open Access Journals (Sweden)

Stuart E Denman

2015-10-01

Full Text Available 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 shifts in the rumen. Microbial ecology methods identified many 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 hydrogen-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.

Abundance and Diversity of Hydrogenotrophic Microorganisms in the Infant Gut before the Weaning Period Assessed by Denaturing Gradient Gel Electrophoresis and Quantitative PCR

Directory of Open Access Journals (Sweden)

Valeria Sagheddu

2017-06-01

Full Text Available Delivery mode (natural vs. cesarean and feeding type (breast vs. formula feeding are relevant factors for neonatal gut colonization. Biomolecular methods have shown that the ecological structure of infant microbiota is more complex than previously proposed, suggesting a relevant presence of unculturable bacteria. It has also been postulated that among unculturable bacteria, hydrogenotrophic populations might play a key role in infant health. Sulfate-reducing bacteria (SRB, acetogens, and methanogenic archaea use hydrogenotrophic pathways within the human colon. However, to date, few studies have reported detection of hydrogenotrophic microorganisms in newborns, possibly because of limitations on available group-specific, culture-independent quantification procedures. In the present work, we analyzed 16 fecal samples of healthy babies aged 1–6 months by means of quantitative PCR (qPCR targeting the 16S rRNA or metabolic functional genes and by denaturing gradient gel electrophoresis (DGGE. qPCR data showed quantifiable levels of methanogens, SRB, and acetogens in all samples, indicating that the relative abundances of these microbial groups were not affected by delivery mode (natural vs. caesarian. DGGE revealed a high prevalence of the Blautia genus within the acetogenic bacteria despite strong interindividual variability. Our preliminary results suggest that hydrogenotrophic microorganisms, which have been a neglected group to date, should be included in future ecological and metabolic studies evaluating the infant intestinal microbiota.

Bacterial study of the anaerobic bioreactor for distillery effluent

International Nuclear Information System (INIS)

Shah, F. A.; Pathan, M. I.

2006-01-01

This study relates with anaerobic bioreactors of Habib Sugar Mills, Nawabshah. Bacterial growth was studied through microscope along with its effect on the production of methane gas (Biogas) at all HRTs (Hydraulic Retention Times) between 15 and 28 days. The bacterium has the efficiency to convert 12% glucose within 24 hours to final product and cell mass. The acetogenic organisms also show their maximum growth on glucose in BGP-1 and BPG-2 at both the corks, where as Methanogenic organisms have shown their zero shown their zero growth on glucose. The efforts have been taken to determine the methanogenic, acetogenic and syntrophomonas sp. data of anaerobic bioreactors of BGP (Biogas Plant) I and II, when these samples were cultured on acetate, methanol, formate, butyrate, propionate and glucose. (author)

Isolation and characterization of a new hydrogen-utilizing bacterium from the rumen.

Science.gov (United States)

Rieu-Lesme, F; Fonty, G; Doré, J

1995-01-01

A new H2/CO2-utilizing acetogenic bacterium was isolated from the rumen of a mature deer. This is the first report of a spore-forming Gram-negative bacterial species from the rumen. The organism was a strictly anaerobic, motile rod and was able to grow autotrophically on hydrogen and carbon dioxide. Acetate was the major product detected. Glucose, fructose and lactate were also fermented heterotrophically. The optimum pH for growth was 7.0-7.5, and the optimum temperature was 37-42 degrees C. Yeast extract was required for growth and rumen fluid was highly stimulatory. The DNA base ratio was 52.9 +/- 0.5 mol% G+C. On the basis of these characteristics and fermentation products, the isolate was considered to be different from acetogenic bacteria described previously.

Anaerobic Co-digestion of Fresh Maize Leaves with Elephant Grass ...

African Journals Online (AJOL)

MICHAEL HORSFALL

antagonistic effect of co-digestion of the substrates on biogas production in order to establish the best blend. Six different ... obligate hydrogen-producing acetogens. Finally in .... Impact of food industrial waste on anaerobic co- digestion of ...

Role of formate and hydrogen in the syntrophic degradation of propionate and butyrate

NARCIS (Netherlands)

Xiuzhu Dong,

1994-01-01

Under methanogenic conditions, complex organic matter is mineralized by fermentative, acetogenic and methanogenic bacteria. Propionate and butyrate are two important intermediates; they account for 35% and 8% of the total methane formation, respectively. Propionate and butyrate are

The deep-subsurface sulfate reducer Desulfotomaculum kuznetsovii employs two methanol-degrading pathways

NARCIS (Netherlands)

Sousa, Diana Z.; Visser, Michael; Gelder, Van Antonie H.; Boeren, Sjef; Pieterse, Mervin M.; Pinkse, Martijn W.H.; Verhaert, Peter D.E.M.; Vogt, Carsten; Franke, Steffi; Kümmel, Steffen; Stams, Alfons J.M.

2018-01-01

Methanol is generally metabolized through a pathway initiated by a cobalamine-containing methanol methyltransferase by anaerobic methylotrophs (such as methanogens and acetogens), or through oxidation to formaldehyde using a methanol dehydrogenase by aerobes. Methanol is an important substrate in

Methane fermentation process as anaerobic digestion of biomass ...

African Journals Online (AJOL)

Anaerobic decomposition of organic compounds is conducted in close cooperation of specialized bacteria of different types, including mostly hydrolyzing, digestive, acetogenic, homoacetogenic, sulfate-reducing (VI) and methanogenic bacteria. A great interest in the anaerobic digestion process results mainly from its ...

The anaerobic treatment of sulfate containing wastewater

NARCIS (Netherlands)

Visser, A.

1995-01-01

In the anaerobic treatment of sulfate containing wastewater sulfate reducing bacteria (SRB) will compete with methanogenic- (MB) and acetogenic bacteria (AB) for the available substrates such as hydrogen, acetate, propionate and butyrate. The outcome of this competition will

Biogas, as a renewable energy source, produced during the anaerobic digestion of organic waste

CSIR Research Space (South Africa)

Greben, H

2009-10-01

Full Text Available of the process is dependent on the balance between symbiotic growth of the different groups of bacteria, such as the acid forming bacteria, obligate hydrogen producing acetogens and methanogens (Angelidaki, et al., 2009). This symbiotic relationship can...

The fate of methanol in anaerobic bioreactors

NARCIS (Netherlands)

Florencio, L.

1994-01-01

Methanol is an important component of certain industrial wastewaters. In anaerobic environments, methanol can be utilized by methanogens and acetogens. In wastewater treatment plants, the conversion of methanol into methane is preferred because this conversion is responsible for chemical

Quantifying electron fluxes in methanogenic microbial communities

NARCIS (Netherlands)

Junicke, H.

2015-01-01

Anaerobic digestion is a widely applied process in which close interactions between different microbial groups result in the formation of renewable energy in the form of biogas. Nevertheless, the regulatory mechanisms of the electron transfer between acetogenic bacteria and methanogenic archaea in

Moorella Strains for the Production of Biochemicals from Syngas

DEFF Research Database (Denmark)

Redl, Stephanie; Jensen, Torbjørn Ølshøj; Nielsen, Alex Toftgaard

. In the syngasfermentation process even the recalcitrant lignin portion can be fully converted into higher valuecompounds.However, cost-effectiveness of this process requires better understanding of the metabolism andmodification of the acetogenic strains. The present project lays the basis for production of higher...

Metabolic interactions in methanogenic and sulfate-reducing bioreactors

NARCIS (Netherlands)

Stams, A.J.M.; Plugge, C.M.; Bok, de F.A.M.; Houten, van B.H.G.W.; Lens, P.N.L.; Dijkman, H.; Weijma, J.

2005-01-01

In environments where the amount of electron acceptors 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 consortia of acetogenic bacteria and methanogenic archaea.

Interspecies electron transfer in methanogenic propionate degrading consortia

NARCIS (Netherlands)

Bok, de F.A.M.; Plugge, C.M.; Stams, A.J.M.

2004-01-01

Propionate is a key intermediate in the conversion of complex organic matter under methanogenic conditions. Oxidation of this compound requires obligate syntrophic consortia of acetogenic proton- and bicarbonate reducing bacteria and methanogenic archaea. Although H-2 acts as an electron-carrier in

HIGH-RATE ANAEROBIC TREATMENT OF ALCOHOLIC WASTEWATERS

Directory of Open Access Journals (Sweden)

Florencio L.

1997-01-01

Full Text Available Modern high-rate anaerobic wastewater treatment processes are rapidly becoming popular for industrial wastewater treatment. However, until recently stable process conditions could not be guaranteed for alcoholic wastewaters containing higher concentrations of methanol. Although methanol can be directly converted into methane by methanogens, under specific conditions it can also be converted into acetate and butyrate by acetogens. The accumulation of volatile fatty acids can lead to reactor instability in a weakly buffered reactor. Since this process was insufficiently understood, the application of high-rate anaerobic reactors was highly questionable. This research investigated the environmental factors that are of importance in the predominance of methylotrophic methanogens over acetogens in a natural mixed culture during anaerobic wastewater treatment in upflow anaerobic sludge bed reactors. Technological and microbiological aspects were investigated. Additionally, the route by which methanol is converted into methane is also presented

A mathematical model for the interactive behavior of sulfate-reducing bacteria and methanogens during anaerobic digestion.

Science.gov (United States)

Ahammad, S Ziauddin; Gomes, James; Sreekrishnan, T R

2011-09-01

Anaerobic degradation of waste involves different classes of microorganisms, and there are different types of interactions among them for substrates, terminal electron acceptors, and so on. A mathematical model is developed based on the mass balance of different substrates, products, and microbes present in the system to study the interaction between methanogens and sulfate-reducing bacteria (SRB). The performance of major microbial consortia present in the system, such as propionate-utilizing acetogens, butyrate-utilizing acetogens, acetoclastic methanogens, hydrogen-utilizing methanogens, and SRB were considered and analyzed in the model. Different substrates consumed and products formed during the process also were considered in the model. The experimental observations and model predictions showed very good prediction capabilities of the model. Model prediction was validated statistically. It was observed that the model-predicted values matched the experimental data very closely, with an average error of 3.9%.

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

Energy Technology Data Exchange (ETDEWEB)

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.

Physiologically anaerobic microorganisms of the deep subsurface

Energy Technology Data Exchange (ETDEWEB)

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.

Microbial kinetic for In-Storage-Psychrophilic Anaerobic Digestion (ISPAD).

Science.gov (United States)

Madani-Hosseini, Mahsa; Mulligan, Catherine N; Barrington, Suzelle

2014-12-15

In-Storage-Psychrophilic-Anaerobic-Digestion (ISPAD) is a wastewater storage tank converted into an anaerobic digestion (AD) system by means of an airtight floating geo-membrane. For process optimization, ISPAD requires modelling with well-established microbial kinetics coefficients. The present objectives were to: obtain kinetics coefficients for the modelling of ISPAD; compare the prediction of the conventional and decomposition fitting approach, an innovative fitting technique used in other fields of science, and; obtain equations to predict the maximum growth rate (μmax) of microbial communities as a function of temperature. The method consisted in conducting specific Substrate Activity Tests (SAT) using ISPAD inoculum to monitor the rate of degradation of specific substrates at 8, 18 and 35 °C. Microbial kinetics coefficients were obtained by fitting the Monod equations to SAT. The statistical procedure of Least Square Error analysis was used to minimize the Sum of Squared Errors (SSE) between the measured ISPAD experimental data and the Monod equation values. Comparing both fitting methods, the decomposition approach gave higher correlation coefficient (R) for most kinetics values, as compared to the conventional approach. Tested to predict μmax with temperature, the Square Root equation better predicted temperature dependency of both acidogens and propionate degrading acetogens, while the Arrhenius equation better predicted that of methanogens and butyrate degrading acetogens. Increasing temperature from 18 to 35 °C did not affect butyrate degrading acetogens, likely because of their dominance, as demonstrated by microbial population estimation. The estimated ISPAD kinetics coefficients suggest a robust psychrophilic and mesophilic coexisting microbial community demonstrating acclimation to ambient temperature. Copyright © 2014 Elsevier Ltd. All rights reserved.

[Long-term storage of obligate anaerobic microorganisms in glycerol].

Science.gov (United States)

Briukhanov, A I; Netrusov, A I

2006-01-01

We evaluated the possibility of storing the cultures of obligate anaerobic microorganisms (clostridia. acetogenic and sulfate-reducing bacteria, and methanogenic archaea) in 25% glycerol at -70 degrees C for a long time (up to 3 years). This method of storage is adequate to preserve cell viability in most obligate anaerobes.

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

DEFF Research Database (Denmark)

Lever, Mark

2012-01-01

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...... to be taken into account to understand microbial survival in the energy-depleted deep biosphere....

Microbial reduction of Fe(III) and turnover of acetate in Hawaiian soils.

Science.gov (United States)

Küsel, Kirsten; Wagner, Christine; Trinkwalter, Tanja; Gössner, Anita S; Bäumler, Rupert; Drake, Harold L

2002-04-01

Soils contain anoxic microzones, and acetate is an intermediate during the turnover of soil organic carbon. Due to negligible methanogenic activities in well-drained soils, acetate accumulates under experimentally imposed short-term anoxic conditions. In contrast to forest, agricultural, and prairie soils, grassland soils from Hawaii rapidly consumed rather than formed acetate when incubated under anoxic conditions. Thus, alternative electron acceptors that might be linked to the anaerobic oxidation of soil organic carbon in Hawaiian soils were assessed. Under anoxic conditions, high amounts of Fe(II) were formed by Hawaiian soils as soon as soils were depleted of nitrate. Rates of Fe(II) formation for different soils ranged from 0.01 to 0.31 micromol (g dry weight soil)(-1) h(-1), but were not positively correlated to increasing amounts of poorly crystallized iron oxides. In general, sulfate-reducing and methanogenic activities were negligible. Supplemental acetate was rapidly oxidized to CO2 via the sequential reduction of nitrate and Fe(III) in grassland soil (obtained near Kaena State Park). Supplemental H2 stimulated the formation of Fe(II), but H2-utilizing acetogens appeared to also be involved in the consumption of H2. Approximately 270 micromol Fe(III) (g dry weight soil)(-1) was available for Fe(III)-reducing bacteria, and acetate became a stable end product when Fe(III) was depleted in long-term incubations. Most-probable-number estimates of H2- and acetate-utilizing Fe(III) reducers and of H2-utilizing acetogens were similar. These results indicate that (i) the microbial reduction of Fe(III) is an important electron-accepting process for the anaerobic oxidation of organic matter in Fe(III)-rich Hawaiian soils of volcanic origin, and (ii) acetate, formed by the combined activity of fermentative and acetogenic bacteria, is an important trophic link in anoxic microsites of these soils.

Biomethanation and Its Potential

DEFF Research Database (Denmark)

Angelidaki, Irini; Karakashev, Dimitar Borisov; Batstone, Damien J.

2011-01-01

matter via cascades of biochemical conversions to methane and carbon dioxide. Syntrophic relationships between hydrogen producers (acetogens) and hydrogen scavengers (homoacetogens, hydrogenotrophic methanogens, etc.) are critical to the process. Determination of practical and theoretical methane...... residues and wastes. It will help to reduce the use of fossil fuels and thus reduce CO2 emission....

Improved biogas production from whole stillage by co-digestion with cattle manure.

Science.gov (United States)

Westerholm, Maria; Hansson, Mikael; Schnürer, Anna

2012-06-01

Whole stillage, as sole substrate or co-digested with cattle manure, was evaluated as substrate for biogas production in five mesophilic laboratory-scale biogas reactors, operating semi-continuously for 640 days. The process performance was monitored by chemical parameters and by quantitative analysis of the methanogenic and acetogenic population. With whole stillage as sole substrate the process showed clear signs of instability after 120 days of operation. However, co-digestion with manure clearly improved biogas productivity and process stability and indicated increased methane yield compared with theoretical values. The methane yield at an organic loading rate (OLR) at 2.8 g VS/(L×day) and a hydraulic retention time (HRT) of 45 days with a substrate mixture 85% whole stillage and 15% manure (based on volatile solids [VS]) was 0.31 N L CH(4)/gVS. Surprisingly, the abundance of the methanogenic and acetogenic populations remained relatively stable throughout the whole operation and was not influenced by process performance. Copyright © 2012 Elsevier Ltd. All rights reserved.

Characterization of two subsurface H2-utilizing bacteria, Desulfomicrobium hypogeium sp. nov. and Acetobacterium psammolithicum sp. nov., and their ecological roles.

Science.gov (United States)

Krumholz, L R; Harris, S H; Tay, S T; Suflita, J M

1999-06-01

We examined the relative roles of acetogenic and sulfate-reducing bacteria in H2 consumption in a previously characterized subsurface sandstone ecosystem. Enrichment cultures originally inoculated with ground sandstone material obtained from a Cretaceous formation in central New Mexico were grown with hydrogen in a mineral medium supplemented with 0.02% yeast extract. Sulfate reduction and acetogenesis occurred in these cultures, and the two most abundant organisms carrying out the reactions were isolated. Based on 16S rRNA analysis data and on substrate utilization patterns, these organisms were named Desulfomicrobium hypogeium sp. nov. and Acetobacterium psammolithicum sp. nov. The steady-state H2 concentrations measured in sandstone-sediment slurries (threshold concentration, 5 nM), in pure cultures of sulfate reducers (threshold concentration, 2 nM), and in pure cultures of acetogens (threshold concentrations 195 to 414 nM) suggest that sulfate reduction is the dominant terminal electron-accepting process in the ecosystem examined. In an experiment in which direct competition for H2 between D. hypogeium and A. psammolithicum was examined, sulfate reduction was the dominant process.

Reaction engineering analysis of the autotrophic energy metabolism of Clostridium aceticum.

Science.gov (United States)

Mayer, Alexander; Weuster-Botz, Dirk

2017-12-01

Acetogenesis with CO2:H2 or CO via the reductive acetyl-CoA pathway does not provide any net ATP formation in homoacetogenic bacteria. Autotrophic energy conservation is coupled to the generation of chemiosmotic H+ or Na+ gradients across the cytoplasm membrane using either a ferredoxin:NAD+ oxidoreductase (Rnf), a ferredoxin:H+ oxidoreductase (Ech) or substrate-level phosphorylation via cytochromes. The first isolated acetogenic bacterium Clostridium aceticum shows both cytochromes and Rnf complex, putting it into an outstanding position. Autotrophic batch processes with continuous gas supply were performed in fully controlled stirred-tank bioreactors to elucidate energy metabolism of C. aceticum. Varying the initial Na+ concentration in the medium showed sodium-dependent growth of C. aceticum with a growth optimum between 60 and 90 mM Na+. The addition of the Na+-selective ionophore ETH2120 or the protonophore CCCP or the H+/cation-antiporter monensin revealed that an H+ gradient is used as primary energy conservation mechanism, which strengthens the exceptional position of C. aceticum as acetogenic bacterium showing an H+-dependent energy conservation mechanism as well as Na+-dependent growth. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Optimisation of single-phase dry-thermophilic anaerobic digestion under high organic loading rates of industrial municipal solid waste: population dynamics.

Science.gov (United States)

Zahedi, S; Sales, D; Romero, L I; Solera, R

2013-10-01

Different high feed organic loading rates (OLRs) (from 5.7 g to 46.0 g TVS/l/d) or hydraulic retention times (HRTs) (from 15 d to 2 d) in single-phase dry-thermophilic anaerobic digestion (AD) of organic fraction municipal solid waste (OFMSW) were investigated. The specific gas production (SGP) values (0.25-0.53 m(3)/kg TVS) and the percentages of Eubacteria, Archaea, H2-utilising methanogens (HUMs) and acetate-utilising methanogens (AUMs) were stable within the ranges 80.2-91.1%, 12.4-18.5%, 4.4-9.8% and 5.5-10.9%, respectively. A HUM/AUM ratio greater than 0.7 seems to be necessary to maintain very low partial pressures of H2 required for dry AD process. Increasing OLR resulted in an increase in all the populations, except for propionate-utilising acetogens (PUAs). Optimal conditions were obtained at 3d HRT (OLR=30.7 g TVS/l/d), which is lower than the doubling time of acetogens and methanogens. The methane production (MP) was clearly higher than those reported in AD of OFMSW. Copyright © 2013 Elsevier Ltd. All rights reserved.

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

Energy Technology Data Exchange (ETDEWEB)

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.

Regional distribution of microbes in groundwater from Haestholmen, Kivetty, Olkiluoto and Romuvaara, Finland

Energy Technology Data Exchange (ETDEWEB)

Haveman, S.A.; Nilsson, E.L.; Pedersen, K. [Goeteborg University (Sweden)

2000-06-01

Groundwater was sampled with the PAVE groundwater sampling system from eight boreholes at Haestholmen, Kivetty, Olkiluoto and Romuvaara, Finland, in 1998 and 1999, for investigation of microbial populations. The groundwater samples had a wide range of salinity and chemistry and contained 104-105 cells per ml, which is typical for subsurface groundwater. In preparing culture media, two approaches were used and compared. Natural, groundwater-based media were prepared from groundwater from the same section of each borehole tested, and synthetic media were prepared based on groundwater chemistry data. No significant difference was observed between the two types of media for brackish and saline groundwater. The groundwater to a depth of 750 m contained mainly sulphate-reducing bacteria (SRB), ironreducing bacteria (IRB) and heterotrophic acetogenic (HA) bacteria. Autotrophic acetogenic (AA) bacteria and methanogenic archaea were found in some samples. Iron-reducing and HA bacteria predominated in brackish groundwater from Haestholmen, with SRB present in smaller numbers. A different microbial population was found in deep saline groundwater from Haestholmen and Olkiluoto that consists of a large proportion of a saline or brine end member. No SRB or AA bacteria were cultured; instead, the microbial population consisted of HA bacteria and either IRB or methanogens. In Olkiluoto, SRB predominated in the brackish and saline groundwater at depths to about 500 m, while methanogens were found in deeper saline groundwater. Stable isotope data (C-13) indicated that the methanogens are part of an autotrophic population consuming dissolved inorganic carbon (DIC) and hydrogen and producing methane and organic carbon. This deep ecosystem may be independent of surface life processes. A high-level radioactive waste (HLW) repository at 500 m depth in the Fennoscandian Shield will be inhabited by SRB, IRB and acetogens. Methanogens may also be present. These anaerobic micro

The fate of methanol in anaerobic bioreactors

OpenAIRE

Florencio, L.

1994-01-01

Methanol is an important component of certain industrial wastewaters. In anaerobic environments, methanol can be utilized by methanogens and acetogens. In wastewater treatment plants, the conversion of methanol into methane is preferred because this conversion is responsible for chemical oxygen demand (COD) removal, whereas with the formation of volatile fatty acids (VFA) little COD removal is achieved. Moreover, the accumulation of VFA can lead to reactor instability due to pH drops...

Characterization of 16S rRNA genes from oil field microbial communities indicates the presence of a variety of sulfate-reducing, fermentative, and sulfide-oxidizing bacteria.

OpenAIRE

Voordouw, G; Armstrong, S M; Reimer, M F; Fouts, B; Telang, A J; Shen, Y; Gevertz, D

1996-01-01

Oil field bacteria were characterized by cloning and sequencing of PCR-amplified 16S rRNA genes. A variety of gram-negative, sulfate-reducing bacteria was detected (16 members of the family Desulfovibrionaceae and 8 members of the family Desulfobacteriaceae). In contrast, a much more limited number of anaerobic, fermentative, or acetogenic bacteria was found (one Clostridium sp., one Eubacterium sp., and one Synergistes sp.). Potential sulfide oxidizers and/or microaerophiles (Thiomicrospira,...

Methane-yielding microbial communities processing lactate-rich substrates: a piece of the anaerobic digestion puzzle.

Science.gov (United States)

Detman, Anna; Mielecki, Damian; Pleśniak, Łukasz; Bucha, Michał; Janiga, Marek; Matyasik, Irena; Chojnacka, Aleksandra; Jędrysek, Mariusz-Orion; Błaszczyk, Mieczysław K; Sikora, Anna

2018-01-01

Anaerobic digestion, whose final products are methane and carbon dioxide, ensures energy flow and circulation of matter in ecosystems. This naturally occurring process is used for the production of renewable energy from biomass. Lactate, a common product of acidic fermentation, is a key intermediate in anaerobic digestion of biomass in the environment and biogas plants. Effective utilization of lactate has been observed in many experimental approaches used to study anaerobic digestion. Interestingly, anaerobic lactate oxidation and lactate oxidizers as a physiological group in methane-yielding microbial communities have not received enough attention in the context of the acetogenic step of anaerobic digestion. This study focuses on metabolic transformation of lactate during the acetogenic and methanogenic steps of anaerobic digestion in methane-yielding bioreactors. Methane-yielding microbial communities instead of pure cultures of acetate producers were used to process artificial lactate-rich media to methane and carbon dioxide in up-flow anaerobic sludge blanket reactors. The media imitated the mixture of acidic products found in anaerobic environments/digesters where lactate fermentation dominates in acidogenesis. Effective utilization of lactate and biogas production was observed. 16S rRNA profiling was used to examine the selected methane-yielding communities. Among Archaea present in the bioreactors, the order Methanosarcinales predominated. The acetoclastic pathway of methane formation was further confirmed by analysis of the stable carbon isotope composition of methane and carbon dioxide. The domain Bacteria was represented by Bacteroidetes , Firmicutes , Proteobacteria , Synergistetes , Actinobacteria , Spirochaetes , Tenericutes , Caldithrix , Verrucomicrobia , Thermotogae , Chloroflexi , Nitrospirae, and Cyanobacteria. Available genome sequences of species and/or genera identified in the microbial communities were searched for genes encoding the lactate

An Adaptive Laboratory Evolution Method to Accelerate Autotrophic Metabolism

DEFF Research Database (Denmark)

Zhang, Tian; Tremblay, Pier-Luc

2018-01-01

Adaptive laboratory evolution (ALE) is an approach enabling the development of novel characteristics in microbial strains via the application of a constant selection pressure. This method is also an efficient tool to acquire insights on molecular mechanisms responsible for specific phenotypes. ALE...... autotrophically and reducing CO2 into acetate more efficiently. Strains developed via this ALE method were also used to gain knowledge on the autotrophic metabolism of S. ovata as well as other acetogenic bacteria....

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

Science.gov (United States)

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.

Microbial ecophysiology of whey biomethanation

International Nuclear Information System (INIS)

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

Methanogenesis at low temperatures by microflora of tundra wetland soil.

Science.gov (United States)

Kotsyurbenko, O R; Nozhevnikova, A N; Soloviova, T I; Zavarzin, G A

1996-01-01

Active methanogenesis from organic matter contained in soil samples from tundra wetland occurred even at 6 degrees C. Methane was the only end product in balanced microbial community with H2/CO2 as a substrate, besides acetate was produced as an intermediate at temperatures below 10 degrees C. The activity of different microbial groups of methanogenic community in the temperature range of 6-28 degrees C was investigated using 5% of tundra soil as inoculum. Anaerobic microflora of tundra wetland fermented different organic compounds with formation of hydrogen, volatile fatty acids (VFA) and alcohols. Methane was produced at the second step. Homoacetogenic and methanogenic bacteria competed for such substrates as hydrogen, formate, carbon monoxide and methanol. Acetogens out competed methanogens in an excess of substrate and low density of microbial population. Kinetic analysis of the results confirmed the prevalence of hydrogen acetogenesis on methanogenesis. Pure culture of acetogenic bacteria was isolated at 6 degrees C. Dilution of tundra soil and supply with the excess of substrate disbalanced the methanoigenic microbial community. It resulted in accumulation of acetate and other VFA. In balanced microbial community obviously autotrophic methanogens keep hydrogen concentration below a threshold for syntrophic degradation of VFA. Accumulation of acetate- and H2/CO2-utilising methanogens should be very important in methanogenic microbial community operating at low temperatures.

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

Energy Technology Data Exchange (ETDEWEB)

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.

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

Directory of Open Access Journals (Sweden)

Lisa-Maree Gulino

Full Text Available 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.

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

Science.gov (United States)

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.

A bacterial population analysis of granular sludge from an anaerobic digester treating a maize-processing waste

Energy Technology Data Exchange (ETDEWEB)

Howgrave-Graham, A.R.; Wallis, F.M. (Natal Univ., Pietermaritzburg (ZA). Dept. of Microbiology and Plant Pathology); Steyn, P.L. (Pretoria Univ. (South Africa))

1991-01-01

Microbial population studies were conducted on a dense granular sludge, with excellent settling, thickening and nutrient removal properties, from a South African clarigester treating effluent from a factory producing glucose and other carbohydrates from maize. The bacterial population comprised a heterogeneous group including acetogens, enterobacteria, sulphate-reducers, spirochaetes, heterofermentative lactobacilli and methanogens. The presence of these bacteria and lack of propionic acid and butyric acid bacteria suggests that the microbial activity of this anaerobic digester involved acetate and lactate metabolism rather than propionate or butyrate catabolism as a source of precursors for methane production. (author).

Anaerobic bacterial quantitation of Yucca Mountain, Nevada DOE site samples

International Nuclear Information System (INIS)

Clarkson, W.W.; Krumholz, L.R.; Suflita, J.M.

1996-01-01

Anaerobic bacteria were studied from samples of excavated rock material as one phase of the overall Yucca Mountain site characterization effort. An indication of the abundance of important groups of anaerobic bacteria would enable inferences to be made regarding the natural history of the site and allow for more complete risk evaluation of the site as a nuclear repository. Six bacterial groups were investigated including anaerobic heterotrophs, acetogens, methanogens, sulfate-, nitrate-, and iron-reducing bacteria. The purpose of this portion of the study was to detect and quantify the aforementioned bacterial groups

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

Science.gov (United States)

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.

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

Energy Technology Data Exchange (ETDEWEB)

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

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.

Genome-guided analysis of physiological capacities of Tepidanaerobacter acetatoxydans provides insights into environmental adaptations and syntrophic acetate oxidation.

Directory of Open Access Journals (Sweden)

Bettina Müller

Full Text Available 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.

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

Science.gov (United States)

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. Copyright © 2015 Elsevier Ltd. All rights reserved.

Carbon dioxide conversion to fuels and chemicals using a hybrid green process

International Nuclear Information System (INIS)

Ramachandriya, Karthikeyan D.; Kundiyana, Dimple K.; Wilkins, Mark R.; Terrill, Jennine B.; Atiyeh, Hasan K.; Huhnke, Raymond L.

2013-01-01

Highlights: • A unique CO 2 conversion technology using microorganisms was demonstrated. • Corn steep liquor medium enhanced production of n-butanol and n-hexanol. • Cotton seed extract (CSE) medium promoted ethanol formation. • CSE medium without morpholinoethanesulfonic acid buffer reduced the cost by 99%. - Abstract: A unique hybrid technology that uses renewable hydrogen (H 2 ) and carbon dioxide (CO 2 ) sequestered from large point sources, to produce fuels and chemicals has been proposed and tested. The primary objective of this research was to determine the feasibility of using two acetogenic bacteria to metabolize H 2 and CO 2 for the production of ethanol. Three experiments were conducted in small scale reactors to select a bacterium, feed gas composition and nutrient medium source to produce ethanol. The results indicated that Clostridium carboxidivorans produced 33% more ethanol and 66% less acetic acid compared to Clostridium ragsdalei, making C. carboxidivorans the better candidate for ethanol production. Furthermore, the removal of morpholinoethanesulfonic acid (MES) buffer from cotton seed extract (CSE) medium offered a low-cost medium for fermentations. Additionally, we observed that corn steep liquor (CSL) in the medium diversified the product range with both bacteria. Maximum concentrations of ethanol, n-butanol, n-hexanol, acetic acid, butyric acid, and hexanoic acid from different fermentation treatments were 2.78 g L −1 , 0.70 g L −1 , 0.52 g L −1 , 4.06 g L −1 , 0.13 g L −1 and 0.42 g L −1 , respectively. This study highlights the important role that acetogenic microbes can offer for CO 2 conversion into valuable fuels and chemicals

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

Science.gov (United States)

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.

Metabolic Flexibility of Sulfate Reducing Bacteria

Directory of Open Access Journals (Sweden)

Caroline M. Plugge

2011-05-01

Full Text Available Dissimilatory sulfate-reducing prokaryotes (SRB are a very diverse group of anaerobic bacteria that are omnipresent in nature and play an imperative role in the global cycling of carbon and sulfur. In anoxic marine sediments sulfate reduction accounts for up to 50% of the entire organic mineralization in coastal and shelf ecosystems where sulfate diffuses several meters deep into the sediment. As a consequence, SRB would be expected in the sulfate-containing upper sediment layers, whereas methanogenic Archaea would be expected to succeed in the deeper sulfate-depleted layers of the sediment. Where sediments are high in organic matter, sulfate is depleted at shallow sediment depths, and biogenic methane production will occur. In the absence of sulfate, many SRB ferment organic acids and alcohols, producing hydrogen, acetate, and carbon dioxide, and may even rely on hydrogen- and acetate-scavenging methanogens to convert organic compounds to methane. SRB can establish two different life styles, and these can be termed as sulfidogenic and acetogenic, hydrogenogenic metabolism. The advantage of having different metabolic capabilities is that it raises the chance of survival in environments when electron acceptors become depleted. In marine sediments, SRB and methanogens do not compete but rather complement each other in the degradation of organic matter.Also in freshwater ecosystems with sulfate concentrations of only 10-200 μM, sulfate is consumed efficiently within the top several cm of the sediments. Here, many of the δ-Proteobacteria present have the genetic machinery to perform dissimilatory sulfate reduction, yet they have an acetogenic, hydrogenogenic way of life.In this review we evaluate the physiology and metabolic mode of SRB in relation with their environment.

Draft genome of an Aerophobetes bacterium reveals a facultative lifestyle in deep-sea anaerobic sediments

KAUST Repository

Wang, Yong

2016-07-01

Aerophobetes (or CD12) is a recently defined bacterial phylum, of which the metabolic processes and ecological importance remain unclear. In the present study, we obtained the draft genome of an Aerophobetes bacterium TCS1 from saline sediment near the Thuwal cold seep in the Red Sea using a genome binning method. Analysis of 16S rRNA genes of TCS1 and close relatives revealed wide distribution of Aerophobetes in deep-sea sediments. Phylogenetic relationships showed affinity between Aerophobetes TCS1 and some thermophilic bacterial phyla. The genome of TCS1 (at least 1.27 Mbp) contains a full set of genes encoding core metabolic pathways, including glycolysis and pyruvate fermentation to produce acetyl-CoA and acetate. The identification of cross-membrane sugar transporter genes further indicates its potential ability to consume carbohydrates preserved in the sediment under the microbial mat. Aerophobetes bacterium TCS1 therefore probably carried out saccharolytic and fermentative metabolism. The genes responsible for autotrophic synthesis of acetyl-CoA via the Wood–Ljungdahl pathway were also found in the genome. Phylogenetic study of the essential genes for the Wood–Ljungdahl pathway implied relative independence of Aerophobetes bacterium from the known acetogens and methanogens. Compared with genomes of acetogenic bacteria, Aerophobetes bacterium TCS1 genome lacks the genes involved in nitrogen metabolism, sulfur metabolism, signal transduction and cell motility. The metabolic activities of TCS1 might depend on geochemical conditions such as supplies of CO2, hydrogen and sugars, and therefore the TCS1 might be a facultative bacterium in anaerobic saline sediments near cold seeps. © 2016, Science China Press and Springer-Verlag Berlin Heidelberg.

Prerequisites for Amplicon Pyrosequencing of Microbial Methanol Utilizers in the Environment

Directory of Open Access Journals (Sweden)

Steffen eKolb

2013-09-01

Full Text Available The commercial availability of next generation sequencing (NGS technologies facilitated the assessment of functional groups of microorganisms in the environment with high coverage, resolution, and reproducibility. Soil methylotrophs were among the first microorganisms in the environment that were assessed with molecular tools, and nowadays, as well with NGS technologies. Studies in the past years re-attracted notice to the pivotal role of methylotrophs in global conversions of methanol, which mainly originates from plants, and is involved in oxidative reactions and ozone formation in the atmosphere. Aerobic methanol utilizers belong to Bacteria, yeasts, Ascomycota, and molds. Numerous bacterial methylotrophs are facultatively aerobic, and also contribute to anaerobic methanol oxidation in the environment, whereas strict anaerobic methanol utilizers belong to methanogens and acetogens. The diversity of enzymes catalyzing the initial oxidation of methanol is considerable, and comprises at least five different enzyme types in aerobes, and one in strict anaerobes. Only the gene of the large subunit of PQQ-dependent methanol dehydrogenase (mxaF has been analyzed by environmental pyrosequencing. To enable a comprehensive assessment of methanol utilizers in the environment, new primers targeting genes of the PQQ MDH in Methylibium (mdh2, of the NAD-dependent MDH (mdh, of the methanol oxidoreductase of Actinobacteria (mdo, of the fungal FAD-dependent alcohol oxidase (mod1, mod2, and homologues, and of the gene of the large subunit of the methanol:corrinoid methyltransferases (mtaC in methanogens and acetogens need to be developed. Combined stable isotope probing of nucleic acids or proteins with amplicon-based NGS are straightforward approaches to reveal insights into functions of certain methylotrophic taxa in the global methanol cycle.

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

Directory of Open Access Journals (Sweden)

Sam D. Molenaar

2017-01-01

Full Text Available 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.

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

Science.gov (United States)

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

2017-01-19

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.

Attenuation of landfill leachate by UK Triassic sandstone aquifer materials. 1. Fate of inorganic pollutants in laboratory columns

Science.gov (United States)

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

2000-05-01

The attenuation of inorganic contaminants in acetogenic and methanogenic landfill leachate by calcareous and carbonate-deficient, oxide-rich Triassic sandstone aquifer materials from the English Midlands was examined in laboratory columns. Aqueous equilibrium speciation modelling, simple transport modelling and chemical mass balance approaches are used to evaluate the key processes and aquifer geochemical properties controlling contaminant fate. The results indicate that leachate-rock interactions are dominated by ion-exchange processes, acid-base and redox reactions and sorption/precipitation of metal species. Leachate NH 4 is attenuated by cation exchange with the aquifer sediments; however, NH 4 migration could be described with a simple model using retardation factors. Organic acids in the acetogenic leachate buffered the system pH at low levels during flushing of the calcareous aquifer material. In contrast, equilibrium with Al oxyhydroxide phases initially buffered pH (˜4.5) during flushing of the carbonate-deficient sandstone with methanogenic leachate. This led to the mobilisation of sorbed and oxide-bound heavy metals from the aquifer sediment which migrated as a concentrated pulse at the leachate front. Abiotic reductive dissolution of Mn oxyhydroxides on each aquifer material by leachate Fe 2+ maintains high concentrations of dissolved Mn and buffers the leachate inorganic redox system. This feature is analogous to the Mn-reducing zones found in leachate plumes and in the experiments provides a sink for the leachate Fe load and other heavy metals. The availability of reactive solid phase Mn oxyhydroxides limits the duration of redox buffering and Fe attenuation by these aquifer sediments. Aquifer pH and redox buffering capacity exert a fundamental influence on leachate inorganic contaminant fate in these systems. The implications for the assessment of aquifer vulnerability at landfills are discussed and simple measurements of aquifer properties which

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

Energy Technology Data Exchange (ETDEWEB)

None

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.

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

KAUST Repository

Balk, Melike

2010-08-03

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.

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

KAUST Repository

Balk, Melike; 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 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.

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

Science.gov (United States)

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.

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

OpenAIRE

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 yi...

Exploiting the potential of gas fermentation

DEFF Research Database (Denmark)

Redl, Stephanie Maria Anna; Diender, Martijn; Jensen, Torbjørn Ølshøj

2017-01-01

The use of gas fermentation for production of chemicals and fuels with lower environmental impact is a technology that is gaining increasing attention. Over 38 Gt of CO2 is annually being emitted from industrial processes, thereby contributing significantly to the concentration of greenhouse gases...... in the atmosphere. Together with the gasification of biomass and different waste streams, these gases have the potential for being utilized for production of chemicals through fermentation processes. Acetogens are among the most studied organisms capable of utilizing waste gases. Although engineering...... focus on the advantages of alternative fermentation scenarios, including thermophilic production strains, multi-stage fermentations, mixed cultures, as well as mixotrophy. Such processes have the potential to significantly broaden the product portfolio, increase the product concentrations and yields...

Inhibition of the anaerobic digestion process by linear alkylbenzene sulfonates

DEFF Research Database (Denmark)

Gavala, Hariklia N.; Ahring, Birgitte Kiær

2002-01-01

Linear Alkylbenzene Sulfonates (LAS) are the most widely used synthetic anionic surfactants. They are anthropogenic, toxic compounds and are found in the primary sludge generated in municipal wastewater treatment plants. Primary sludge is usually stabilized anaerobically and therefore it is impor......Linear Alkylbenzene Sulfonates (LAS) are the most widely used synthetic anionic surfactants. They are anthropogenic, toxic compounds and are found in the primary sludge generated in municipal wastewater treatment plants. Primary sludge is usually stabilized anaerobically and therefore...... it is important to investigate the effect of these xenobiotic compounds on an anaerobic environment. The inhibitory effect of Linear Alkylbenzene Sulfonates (LAS) on the acetogenic and methanogenic step of the anaerobic digestion process was studied. LAS inhibit both acetogenesis from propionate...

Enhanced sulfate reduction with acidogenic sulfate-reducing bacteria

International Nuclear Information System (INIS)

Wang Aijie; Ren Nanqi; Wang Xu; Lee Duujong

2008-01-01

Sulfate reduction in a continuous flow, acidogenic reactor using molasses wastewater as the carbon source was studied at varying chemical oxygen demand/sulfate (COD/SO 4 2- ) ratios. At a critical COD/SO 4 2- ratio of 2.7, neither COD nor sulfate were in excess for extra production of ethanol or acetate in the reactor. An acetic-type microbial metabolism was established with sulfate-reducing bacteria (SRB) significantly consuming hydrogen and volatile fatty acids produced by acidogenic bacteria and hydrogen producing acetogens in degrading COD, thereby yielding sulfate removal rate >94.6%. A low critical COD/SO 4 2- ratio of 1.6 was also observed with the enriched ASRB population in reactor which overcomes the barrier to the treatment capability of sulfate-laden wastewater treatment with limited COD supply

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

Science.gov (United States)

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.

My Lifelong Passion for Biochemistry and Anaerobic Microorganisms.

Science.gov (United States)

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.

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

Energy Technology Data Exchange (ETDEWEB)

NONE

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.

Properties of thermophilic microorganisms

International Nuclear Information System (INIS)

Ljungdahl, L.G.

1984-01-01

Microorganisms are called thermophilic or extreme thermophilic (caldo-active) if they grow and reproduce over 47 0 C and 70 0 C, respectively. A survey of growth characteristics of thermophiles is presented and it includes those which also live at extreme pH. The prevalent but not completely emcompassing theory of the ability of thermophiles to grow at high temperatures is that they have macromolecules and cell organelles with high thermostability. Work on some proteins and cell organelles from thermophiles is reviewed. The thermostabilities of these components are compared with those of the living cells, and factors which may govern optimum as well as minimum growth temperatures of microorganisms are discussed. Examples are from the literature but also include enzymes involved in tetrahydrofolate metabolism and other proteins of acetogenic therhmophilic bacteria which are presently studied in the author's laboratory

Advances in metabolic engineering in the microbial production of fuels and chemicals from C1 gas.

Science.gov (United States)

Humphreys, Christopher M; Minton, Nigel P

2018-04-01

The future sustainable production of chemicals and fuels from non-petrochemical sources, while at the same time reducing greenhouse gas (GHG) emissions, represent two of society's greatest challenges. Microbial chassis able to grow on waste carbon monoxide (CO) and carbon dioxide (CO 2 ) can provide solutions to both. Ranging from the anaerobic acetogens, through the aerobic chemoautotrophs to the photoautotrophic cyanobacteria, they are able to convert C1 gases into a range of chemicals and fuels which may be enhanced and extended through appropriate metabolic engineering. The necessary improvements will be facilitated by the increasingly sophisticated gene tools that are beginning to emerge as part of the Synthetic Biology revolution. These tools, in combination with more accurate metabolic and genome scale models, will enable C1 chassis to deliver their full potential. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Antibacterial activity of red algae (Gracilaria verrucosa) extract against Escherichia coli and Salmonella typhimurium

Science.gov (United States)

Dayuti, S.

2018-04-01

Red alga was widely used in several fields, including food, feed, phamacy and industrial point of view. The chemical analysis showed that red alga contained terpenoid, acetogenic, and aromatic compounds, which have a wide range of biological activities, such as anti-micobial, anti-inflammatory and anti-viral. The objectives of this research was to evaluate the effect of extraction solvent and time on antibacterial activity of red alga (Gracilaria verrucosa), and to explore the bioactive compound contained within Gracilaria verrucosa. The method in this study used descriptive reseach. These findings revealed that the highest inhibition activity among all extracts was obtained with the ratio of methanol:aquades (75:25) and extraction time around 72 hours against Escherichia coli and Salmonella typhimurium. The bioactive compounds of Gracilaria verrucosa tested by phytochemical analysisi consisted of flavonoid, alkaloid, and saponin. Those secondary metabolites may be approximated as antibactial substances.

The deep-subsurface sulfate reducer Desulfotomaculum kuznetsovii employs two methanol-degrading pathways.

Science.gov (United States)

Sousa, Diana Z; Visser, Michael; van Gelder, Antonie H; Boeren, Sjef; Pieterse, Mervin M; Pinkse, Martijn W H; Verhaert, Peter D E M; Vogt, Carsten; Franke, Steffi; Kümmel, Steffen; Stams, Alfons J M

2018-01-16

Methanol is generally metabolized through a pathway initiated by a cobalamine-containing methanol methyltransferase by anaerobic methylotrophs (such as methanogens and acetogens), or through oxidation to formaldehyde using a methanol dehydrogenase by aerobes. Methanol is an important substrate in deep-subsurface environments, where thermophilic sulfate-reducing bacteria of the genus Desulfotomaculum have key roles. Here, we study the methanol metabolism of Desulfotomaculum kuznetsovii strain 17 T , isolated from a 3000-m deep geothermal water reservoir. We use proteomics to analyze cells grown with methanol and sulfate in the presence and absence of cobalt and vitamin B12. The results indicate the presence of two methanol-degrading pathways in D. kuznetsovii, a cobalt-dependent methanol methyltransferase and a cobalt-independent methanol dehydrogenase, which is further confirmed by stable isotope fractionation. This is the first report of a microorganism utilizing two distinct methanol conversion pathways. We hypothesize that this gives D. kuznetsovii a competitive advantage in its natural environment.

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

Science.gov (United States)

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. Copyright © 2016 Elsevier Ltd. All rights reserved.

Methanogenesis of carbohydrates and their fermentation products by syntrophic methane producing bacteria isolated from freshwater sediments

Energy Technology Data Exchange (ETDEWEB)

Tabassum, R; Ibrahim Rajoka, M [National Inst. for Biotechnology and Genetic Engineering, Faisalabad (Pakistan)

2000-05-01

Anaerobic conversion of substrates namely cellulose, cellobiose, glucose, volatile fatty acids, and methanol with a co-culture of fermentative, acidogenic, acetogenic, and methanogenic organisms isolated from freshwater sediments was performed. Maximum reduction of volatile solids (VS) was from cellulose, cellobiose and glucose followed by methanol and other compounds with a product yield coefficient (Y{sub p/s}) of 0.59 m{sup 3}/kg VS consumed with a volumetric productivity (Q{sub p}) of 15.7 mmol/l/d after 12 d fermentation of cellulose. Maximum methane content in the gas mixture was 86.1% with an average of 82.5 {+-} 3.6%. Batch culture methane production characteristics were analyzed and compared. The maximum values of Y{sub p/s}, from cellobiose, glucose, methanol, formate, acetate, propionate, and butyrate were 4.0, 2.2, 0.71. 0.22, 0.90. 1.6 and 1.43 mmol/M substrate used and are higher than those values reported in the literature. (Author)

In-situ biogas upgrading process: Modeling and simulations aspects.

Science.gov (United States)

Lovato, Giovanna; Alvarado-Morales, Merlin; Kovalovszki, Adam; Peprah, Maria; Kougias, Panagiotis G; Rodrigues, José Alberto Domingues; Angelidaki, Irini

2017-12-01

Biogas upgrading processes by in-situ hydrogen (H 2 ) injection are still challenging and could benefit from a mathematical model to predict system performance. Therefore, a previous model on anaerobic digestion was updated and expanded to include the effect of H 2 injection into the liquid phase of a fermenter with the aim of modeling and simulating these processes. This was done by including hydrogenotrophic methanogen kinetics for H 2 consumption and inhibition effect on the acetogenic steps. Special attention was paid to gas to liquid transfer of H 2 . The final model was successfully validated considering a set of Case Studies. Biogas composition and H 2 utilization were correctly predicted, with overall deviation below 10% compared to experimental measurements. Parameter sensitivity analysis revealed that the model is highly sensitive to the H 2 injection rate and mass transfer coefficient. The model developed is an effective tool for predicting process performance in scenarios with biogas upgrading. Copyright © 2017 Elsevier Ltd. All rights reserved.

Labelled precursors for biosynthetic studies on naphthylisoquinoline alkaloids

International Nuclear Information System (INIS)

Bringmann, Gerhard; Pokorny, Frank; Wenzel, Matthias; Wurm, Kathi; Schneider, Christoph

1997-01-01

The isotope labelled monocyclic ketones 5 and 8, postulated precursors to the presumably acetogenic naphthylisoquinoline alkaloids, have been synthesized for biogenetic experiments to Ancistrocladaceae and Dioncophyllaceae plants. Key step of the preparation of 1-(2'-[carbonyl- 14 C] acetyl-3',5'-dibenzyloxyphenyl-2-propanone ([ 14 C]-13 is the C-acetylation of the arylpropanone 10 with the mixed pivalic acetic anhydride ([ 14 C]-11). The resulting pyrylium salt [ 14 C]-12, which is stable and can be stored, is cleaved directly before the feeding experiment to give the diketone [ 14 C]-13 and deprotected to give the free phenolic target molecule [ 14 C]-5. This synthetic route is applicable also to the preparation of 1-(2'-[ 13 C 2 ]acetyl-3'hydroxyphenyl)-2-propanone ([ 13 C 2 ]-5) for biosynthetic experiments with NMR analysis. For the preparation of the oxygen-poorer 13 C-labelled diketone 1-(2'-[methyl- 13 C] acetyl-3'-hydr oxyphenyl)-2-propanone [ 13 C]-8, an 'indanone-route' has been elaborated. (Author)

Long-term high-solids anaerobic digestion of food waste: Effects of ammonia on process performance and microbial community.

Science.gov (United States)

Peng, Xuya; Zhang, ShangYi; Li, Lei; Zhao, Xiaofei; Ma, Yao; Shi, Dezhi

2018-04-22

A long-term high solids anaerobic digestion of food waste was conducted to identify microbial mechanisms of ammonia inhibition during digestion and to clarify correlations between ammonia accumulation, microbial community dynamics (diversity, composition, and interactions), and process stability. Results show that the effects of ammonia on process performance and microbial community were indirectly caused by volatile fatty acid accumulation. Excess free ammonia blocked acetate metabolism, leading to process instability. Accumulated acetate caused feedback inhibition at the acetogenesis stage, which resulted in considerable accumulation of propionate, valerate, and other long-chain fatty acids. This high concentration of volatile fatty acids reduced the abundance of syntrophic acetogenic bacteria and allowed hydrolytic fermentative bacteria to dominate. The normally interactive and orderly metabolic network was broken, which further exacerbated the process instability. These results improve the understanding of microbial mechanisms which contribute to process instability and provide guidance for the microbial management of anaerobic digesters. Copyright © 2018 Elsevier Ltd. All rights reserved.

Microbial ecology of anaerobic digesters: the key players of anaerobiosis.

Science.gov (United States)

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

2014-01-01

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

Microbial Ecology of Anaerobic Digesters: The Key Players of Anaerobiosis

Science.gov (United States)

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

2014-01-01

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

Commercial Biomass Syngas Fermentation

Directory of Open Access Journals (Sweden)

James Daniell

2012-12-01

Full Text Available The use of gas fermentation for the production of low carbon biofuels such as ethanol or butanol from lignocellulosic biomass is an area currently undergoing intensive research and development, with the first commercial units expected to commence operation in the near future. In this process, biomass is first converted into carbon monoxide (CO and hydrogen (H2-rich synthesis gas (syngas via gasification, and subsequently fermented to hydrocarbons by acetogenic bacteria. Several studies have been performed over the last few years to optimise both biomass gasification and syngas fermentation with significant progress being reported in both areas. While challenges associated with the scale-up and operation of this novel process remain, this strategy offers numerous advantages compared with established fermentation and purely thermochemical approaches to biofuel production in terms of feedstock flexibility and production cost. In recent times, metabolic engineering and synthetic biology techniques have been applied to gas fermenting organisms, paving the way for gases to be used as the feedstock for the commercial production of increasingly energy dense fuels and more valuable chemicals.

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

Science.gov (United States)

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.

Physiologically anaerobic microorganisms of the deep subsurface

International Nuclear Information System (INIS)

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

Characterization of 16S rRNA genes from oil field microbial communities indicates the presence of a variety of sulfate-reducing, fermentative, and sulfide-oxidizing bacteria.

Science.gov (United States)

Voordouw, G; Armstrong, S M; Reimer, M F; Fouts, B; Telang, A J; Shen, Y; Gevertz, D

1996-05-01

Oil field bacteria were characterized by cloning and sequencing of PCR-amplified 16S rRNA genes. A variety of gram-negative, sulfate-reducing bacteria was detected (16 members of the family Desulfovibrionaceae and 8 members of the family Desulfobacteriaceae). In contrast, a much more limited number of anaerobic, fermentative, or acetogenic bacteria was found (one Clostridium sp., one Eubacterium sp., and one Synergistes sp.). Potential sulfide oxidizers and/or microaerophiles (Thiomicrospira, Arcobacter, Campylobacter, and Oceanospirillum spp.) were also detected. The first two were prominently amplified from uncultured production water DNA and represented 28 and 47% of all clones, respectively. Growth on media containing sulfide as the electron donor and nitrate as the electron acceptor and designed for the isolation of Thiomicrospira spp. gave only significant enrichment of the Campylobacter sp., which was shown to be present in different western Canadian oil fields. This newly discovered sulfide oxidizer may provide a vital link in the oil field sulfur cycle by reoxidizing sulfide formed by microbial sulfate or sulfur reduction.

Sulfate reduction at low pH to remediate acid mine drainage

International Nuclear Information System (INIS)

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

2014-01-01

Highlights: • Acid mine drainage (AMD) is an important environmental concern. • Remediation through biological sulfate reduction and metal recovery can be applied for AMD. • Microbial community composition has a major impact on the performance of bioreactors to treat AMD. • Acidophilic SRB are strongly influenced by proton, sulfide and organic acids concentration. - Abstract: 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

Anaerobic C1 metabolism of the O-methyl-14C-labeled substituent of vanillate

International Nuclear Information System (INIS)

Frazer, A.C.; Young, L.Y.

1986-01-01

The O-methyl substituents of aromatic compounds constitute a C 1 growth substrate for a number of taxonomically diverse anaerobic acetogens. In this study, strain TH-001, an O-demethylating obligate anaerobe, was chosen to represent this physiological group, and the carbon flow when cells were grown on O-methyl substituents as a C 1 substrate was determined by 14 C radiotracer techniques. O-[methyl- 14 C]vanillate (4-hydroxy-3-methoxy-benzoate) was used as the labeled C 1 substrate. The data showed that for every O-methyl carbon converted to [ 14 C]acetate, two were oxidized to 14 CO 2 . Quantitation of the carbon recovered in the two products, acetate and CO 2 , indicated that acetate was formed in part by the fixation of unlabeled CO 2 . The specific activity of 14 C in acetate was 70% of that in the O-methyl substrate, suggesting that only one carbon of acetate was derived from the O-methyl group. Thus, it is postulated that the carboxyl carbon of the product acetate is derived from CO 2 and the methyl carbon is derived from the O-methyl substituent of vanillate

Exocellular electron transfer in anaerobic microbial communities.

Science.gov (United States)

Stams, Alfons J M; de Bok, Frank A M; Plugge, Caroline M; van Eekert, Miriam H A; Dolfing, Jan; Schraa, Gosse

2006-03-01

Exocellular electron transfer plays an important role in anaerobic microbial communities that degrade organic matter. Interspecies hydrogen transfer between microorganisms is the driving force for complete biodegradation in methanogenic environments. Many organic compounds are degraded by obligatory syntrophic consortia of proton-reducing acetogenic bacteria and hydrogen-consuming methanogenic archaea. Anaerobic microorganisms that use insoluble electron acceptors for growth, such as iron- and manganese-oxide as well as inert graphite electrodes in microbial fuel cells, also transfer electrons exocellularly. Soluble compounds, like humic substances, quinones, phenazines and riboflavin, can function as exocellular electron mediators enhancing this type of anaerobic respiration. However, direct electron transfer by cell-cell contact is important as well. This review addresses the mechanisms of exocellular electron transfer in anaerobic microbial communities. There are fundamental differences but also similarities between electron transfer to another microorganism or to an insoluble electron acceptor. The physical separation of the electron donor and electron acceptor metabolism allows energy conservation in compounds as methane and hydrogen or as electricity. Furthermore, this separation is essential in the donation or acceptance of electrons in some environmental technological processes, e.g. soil remediation, wastewater purification and corrosion.

Microbial Anaerobic Digestion (Bio-Digesters as an Approach to the Decontamination of Animal Wastes in Pollution Control and the Generation of Renewable Energy

Directory of Open Access Journals (Sweden)

Golden Makaka

2013-09-01

Full Text Available 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.

Improving the yield from fermentative hydrogen production.

Science.gov (United States)

Kraemer, Jeremy T; Bagley, David M

2007-05-01

Efforts to increase H(2) yields from fermentative H(2) production include heat treatment of the inoculum, dissolved gas removal, and varying the organic loading rate. Although heat treatment kills methanogens and selects for spore-forming bacteria, the available evidence indicates H(2) yields are not maximized compared to bromoethanesulfonate, iodopropane, or perchloric acid pre-treatments and spore-forming acetogens are not killed. Operational controls (low pH, short solids retention time) can replace heat treatment. Gas sparging increases H(2) yields compared to un-sparged reactors, but no relationship exists between the sparging rate and H(2) yield. Lower sparging rates may improve the H(2) yield with less energy input and product dilution. The reasons why sparging improves H(2) yields are unknown, but recent measurements of dissolved H(2) concentrations during sparging suggest the assumption of decreased inhibition of the H(2)-producing enzymes is unlikely. Significant disagreement exists over the effect of organic loading rate (OLR); some studies show relatively higher OLRs improve H(2) yield while others show the opposite. Discovering the reasons for higher H(2) yields during dissolved gas removal and changes in OLR will help improve H(2) yields.

Interaction of organic contaminants with natural clay type geosorbents: potential use as geologic barrier in urban landfill.

Science.gov (United States)

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.

Strategies for Reducing the Start-up Operation of Microbial Electrochemical Treatments of Urban Wastewater

Directory of Open Access Journals (Sweden)

Zulema Borjas

2015-12-01

Full Text Available Microbial electrochemical technologies (METs constitute the core of a number of emerging technologies with a high potential for treating urban wastewater due to a fascinating reaction mechanism—the electron transfer between bacteria and electrodes to transform metabolism into electrical current. In the current work, we focus on the model electroactive microorganism Geobacter sulfurreducens to explore both the design of new start-up procedures and electrochemical operations. Our chemostat-grown plug and play cells, were able to reduce the start-up period by 20-fold while enhancing chemical oxygen demand (COD removal by more than 6-fold during this period. Moreover, a filter-press based bioreactor was successfully tested for both acetate-supplemented synthetic wastewater and real urban wastewater. This proof-of-concept pre-pilot treatment included a microbial electrolysis cell (MEC followed in time by a microbial fuel cell (MFC to finally generate electrical current of ca. 20 A·m−2 with a power of 10 W·m−2 while removing 42 g COD day−1·m−2. The effective removal of acetate suggests a potential use of this modular technology for treating acetogenic wastewater where Geobacter sulfurreducens outcompetes other organisms.

The Acimet{reg_sign} Process: An innovative approach to biogasification of municipal sludge

Energy Technology Data Exchange (ETDEWEB)

Ghosh, S. [Univ. of Utah, Salt Lake City, UT (United States); Buoy, K. [DuPage County Dept. of Public Works, Wheaton, IL (United States)

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.

Microbial Anaerobic Digestion (Bio-Digesters) as an Approach to the Decontamination of Animal Wastes in Pollution Control and the Generation of Renewable Energy

Science.gov (United States)

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

Microbial anaerobic digestion (bio-digesters) as an approach to the decontamination of animal wastes in pollution control and the generation of renewable energy.

Science.gov (United States)

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.

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

Science.gov (United States)

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.

Inhibitors degradation and microbial response during continuous anaerobic conversion of hydrothermal liquefaction wastewater.

Science.gov (United States)

Si, Buchun; Li, Jiaming; Zhu, Zhangbing; Shen, Mengmeng; Lu, Jianwen; Duan, Na; Zhang, Yuanhui; Liao, Qiang; Huang, Yun; Liu, Zhidan

2018-07-15

One critical challenge of hydrothermal liquefaction (HTL) is its complex aqueous product, which has a high concentration of organic pollutants (up to 100gCOD/L) and diverse fermentation inhibitors, such as furfural, phenolics and N-heterocyclic compounds. Here we report continuous anaerobic digestion of HTL wastewater via an up-flow anaerobic sludge bed reactor (UASB) and packed bed reactor (PBR). Specifically, we investigated the transformation of fermentation inhibitors and microbial response. GC-MS identified the complete degradation of furfural and 5-hydroxymethylfurfural (5-HMF), and partial degradation (54.0-74.6%) of organic nitrogen and phenolic compounds, including 3-hydroxypyridine, phenol and 4-ethyl-phenol. Illumina MiSeq sequencing revealed that the bacteria families related to detoxification increased in response to the HTL aqueous phase. In addition, the increase of acetate-oxidizing bacteria in UASB and acetogens in PBR showed a strengthened acetogenesis. As for the archaeal communities, an increase in hydrogenotrophic methanogens was observed. Based on GC-MS/HPLC and microbial analysis, we speculate that dominant fermentation inhibitors were transformed into intermediates (Acetyl-CoA and acetate), further contributing to biomethane formation. Copyright © 2018 Elsevier B.V. All rights reserved.

Sulfate reduction at low pH to remediate acid mine drainage

Energy Technology Data Exchange (ETDEWEB)

Sánchez-Andrea, Irene, E-mail: irene.sanchezandrea@wur.nl [Departamento de Biología Molecular, Universidad Autónoma de Madrid, 28049 Madrid (Spain); Laboratory of Microbiology, Wageningen University, Dreijenplein 10, 6703 HB Wageningen (Netherlands); Sanz, Jose Luis [Departamento de Biología Molecular, Universidad Autónoma de Madrid, 28049 Madrid (Spain); Bijmans, Martijn F.M. [Wetsus, Centre of Sustainable Water Technology, P.O. Box 1113, 8900 CC Leeuwarden (Netherlands); Stams, Alfons J.M. [Laboratory of Microbiology, Wageningen University, Dreijenplein 10, 6703 HB Wageningen (Netherlands); IBB – Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, University of Minho, 4710-057 Braga (Portugal)

2014-03-01

Highlights: • Acid mine drainage (AMD) is an important environmental concern. • Remediation through biological sulfate reduction and metal recovery can be applied for AMD. • Microbial community composition has a major impact on the performance of bioreactors to treat AMD. • Acidophilic SRB are strongly influenced by proton, sulfide and organic acids concentration. - Abstract: 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.

Conversion of Corn Stover Hydrolysates to Acids: Comparison Between Clostridium carboxidivorans P7 and Microbial Communities Developed from Lake Sediment and an Anaerobic Digester

Energy Technology Data Exchange (ETDEWEB)

Chen, Xiaowen [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Tucker, Melvin P [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Xia, Chunjie [Southern Illinois University; Kumar, Aditi [Carbondale Community High School; Liang, Yanna [Southern Illinois University

2017-01-18

Anaerobic fermentation is an environmentally sustainable technology for converting a variety of feedstocks to biofuels and bioproducts. Considering the complex nature of lignocellulosic hydrolysates, we aimed to investigate product formation from corn stover hydrolysates by using microbial communities under anaerobic conditions. A community developed from lake sediment was able to produce lactic acid from only glucose in the raw or overlimed hydrolysates. Another community from an anaerobic digester, however, was capable of using all hexose and pentose sugars in the raw and undetoxified hydrolysates and released lactic acid at 26.76 g/L. A pure acetogen, Clostridium carboxidivorans P7, was able to grow on the raw and overlimed hydrolysates, too. But the consumption of sugars was minimal and the total released acid concentrations were less than 2 g/L. Next generation sequencing of the enriched community derived from the anaerobic digester revealed the presence of Lactobacillus strains. The predominant species were Lactobacillus parafarraginis (72.6%) and L. buchneri (13.4%). Product titer from using this enriched community can be further enhanced by cultivating at fed-batch or continuous fermentation modes. Results from this study widened the door for producing valuable products from lignocellulosic feedstocks through using mixed cultures.

The effect of mixing on fermentation of primary solids, glycerol, and biodiesel waste.

Science.gov (United States)

Ghasemi, Marzieh; Randall, Andrew A

2018-03-01

In this study, the effect of mixing on volatile fatty acid (VFA) production and composition was investigated through running five identical bench-scale reactors that were filled with primary solid and dosed with either pure glycerol or biodiesel waste. Experimental results revealed that there was an inverse correlation between the mixing intensity and the VFA production. The total VFA production in the un-mixed reactor was 9,787 ± 3,601 mg COD/L, whereas in the reactor mixed at 100 rpm this dropped to 3,927 ± 1,175 mg COD/L, while both types of reactor were dosed with pure glycerol at the beginning of each cycle to reach the initial concentration of 1,000 mg/L (1,217 mg COD/L). Propionic acid was the dominant VFA in all the reactors except the reactor mixed at 30 rpm. It is hypothesized that low mixing facilitated hydrogen transfer between obligate hydrogen producing acetogens (OHPA) and hydrogen consuming acidogens in these non-methanogenic reactors. Also, in a narrower range of mixing (0 or 7 rpm), the total VFA production in biodiesel waste-fed reactors was considerably higher than that of pure glycerol-fed reactors.

Biological acetate production from carbon dioxide by Acetobacterium woodii and Clostridium ljungdahlii: The effect of cell immobilization.

Science.gov (United States)

Cheng, Hai-Hsuan; Syu, Jyun-Cyuan; Tien, Shih-Yuan; Whang, Liang-Ming

2018-08-01

This study investigated the acetate production from gas mixture of hydrogen (H 2 ) and carbon dioxide (CO 2 ) in the ratio of 7:3 using two acetogens: Acetobacterium woodii and Clostridium ljungdahlii. Batch result shows A. woodii performed two-phase degradation with the presence of glucose that lactate was produced from glucose and was reutilized for the production of butyrate and few acetate, while only acetate was detected when providing gas mixture. C. ljungdahlii produced butyrate and ethanol along with acetate when glucose was introduced, while only ethanol and acetate were found by feeding gas mixture. The acetate-to-ethanol (A/E) ratio can be enhanced by cell immobilization, while GAC immobilization produced only acetate and the production rate reached 0.072 mmol/d under fed-batch operation. Acetate production rate increased from 18 to 28 mmol/L/d with GAC immobilization when gas flowrate increased from 100 to 300 mL/min in anaerobic fluidized membrane bioreactor (AFMBR), and a highest A/E ratio of 30 implies the possible application of acetate recovery from H 2 and CO 2 . Copyright © 2018 Elsevier Ltd. All rights reserved.

Results report. Sampling and analyses of gases and microorganisms in the water from MINICAN in 2007, 2008 and 2010

Energy Technology Data Exchange (ETDEWEB)

Lydmark, Sara; Hallbeck, Lotta (Microbial Analytics Sweden AB (Sweden))

2011-04-15

The MINICAN project is located at the depth of 450 m in the Aespoe Hard Rock Laboratory and was initiated to study how corrosion of the cast iron insert inside a perforated copper canister would evolve with time. Miniature canisters with different perforations and with and without bentonite buffer in steel cages were installed and monitored. Samples for microbiological and gas composition together with samples for groundwater chemistry have been analysed at three occasions in 2007, 2008 and 2010. The results show how the microbial populations outside the canisters have evolved from a mixture of microorganism able to grow on organic material, like heterotrophic organisms, and acetogens that grow on hydrogen gas and carbon dioxide in 2007, to populations with a large proportion of sulphate-reducing bacteria in 2010. The highest number of sulphate-reducing bacteria was found in MINICAN experiment A02C, canister with one hole at the top of the copper canister, in 2010 with 2.4 x 104 mL-1 followed by 8 x 103 mL-1 in A03 (hole in the bottom of the canister) and 7 x 103 mL-1 in A06 (two holes at the top and no bentonite). The numbers of culturable heterotrophic bacteria were between 200 and 530 mL-1 in the experiments with bentonite in 2007 but below detection in all experiments in 2010. The same trend was shown for acetogenes. Measurable amounts of hydrogen gas were found in all experiments at all sampling occasions. There was no general trend for the amounts of hydrogen but there was an increase in three of the experiments and in the groundwater outside MINICAN. It was found that the water chemistry differed between A06 and A02-A04 experiment by higher sulphate and chloride concentrations in A06 compared to the others. By plotting the concentrations of chloride, sulphate against time, a decrease in sulphate concentration was found in all canister experiments. The chloride concentrations were stable during the same period. On the other hand, an increase in sulphate

Forsmark site investigation. Microorganisms in groundwater from boreholes KFM10A, KFM11A and KFM08D - numbers, viability, and metabolic diversity. Results from five sections 298.0-305.1 m and 478.0-487.5 m in KFM10A, 447.5-454.6 m in KFM11A, and 669.7-676.8 m and 828.4-835.5 m in KFM08D

Energy Technology Data Exchange (ETDEWEB)

Pedersen, Karsten (Microbial Analytics Sweden AB, Goeteborg (SE))

2007-09-15

Microorganisms and their characteristic features were investigated while geochemically characterizing the groundwater, as part of the site investigation programme at Forsmark. The investigation consists of determining the total numbers of microorganisms, the concentration of adenosine-tri-phosphate (ATP), and the number of culturable heterotrophic aerobic bacteria (CHAB); also included is a method for determining the numbers of organisms belonging to different physiological groups, the most probable number (MPN) method. This investigation covered eight different groups, namely, nitrate-, iron-, manganese-, and sulphate-reducing bacteria, auto- and heterotrophic acetogens, and auto- and heterotrophic methanogens. The reproducibility of the MPN method was tested using groundwater from a depth of 450 m at the Aspo Hard Rock Laboratory and was found to be excellent. Samples were taken from boreholes KFM10A at 298.305 m and 478.487 m, KFM11A at 447.454 m, and KFM08D at 669.676 m and 828.835 m; the sampling dates were 2006-11-28, 2006-10-31, 2007-03-13, 2007-06-19, and 2007-05-02, respectively. The total number of cells (TNC) found in KFM10A groundwater was the highest so far found in a total of 19 analysed sections in the Forsmark area. In contrast, KFM08D-828 m and KFM11A-447 m had among the lowest numbers of cells found thus far. A large amount of ATP per cell indicates large, active cells. The average of all previous ATP/TNC ratios (n approx = 100) in deep groundwater was determined to be 0.43. The analysed groundwater samples from KFM10A-478 m and KFM08D-669 m had ATP/TNC ratios exceeding the overall average of 0.43 for deep groundwater. This suggests that the microorganisms in these groundwaters possessed viability and activity levels above the average for deep groundwater microorganisms. The ratios between the CHAB and NRB numbers found here suggest that there was no surface water contamination. The percentages of TNC culturable using the MPN method were in the 1

Results report. Sampling and analyses of gases and microorganisms in the water from MINICAN in 2007, 2008 and 2010

International Nuclear Information System (INIS)

Lydmark, Sara; Hallbeck, Lotta

2011-04-01

The MINICAN project is located at the depth of 450 m in the Aespoe Hard Rock Laboratory and was initiated to study how corrosion of the cast iron insert inside a perforated copper canister would evolve with time. Miniature canisters with different perforations and with and without bentonite buffer in steel cages were installed and monitored. Samples for microbiological and gas composition together with samples for groundwater chemistry have been analysed at three occasions in 2007, 2008 and 2010. The results show how the microbial populations outside the canisters have evolved from a mixture of microorganism able to grow on organic material, like heterotrophic organisms, and acetogens that grow on hydrogen gas and carbon dioxide in 2007, to populations with a large proportion of sulphate-reducing bacteria in 2010. The highest number of sulphate-reducing bacteria was found in MINICAN experiment A02C, canister with one hole at the top of the copper canister, in 2010 with 2.4 x 10 4 mL -1 followed by 8 x 10 3 mL -1 in A03 (hole in the bottom of the canister) and 7 x 10 3 mL -1 in A06 (two holes at the top and no bentonite). The numbers of culturable heterotrophic bacteria were between 200 and 530 mL-1 in the experiments with bentonite in 2007 but below detection in all experiments in 2010. The same trend was shown for acetogenes. Measurable amounts of hydrogen gas were found in all experiments at all sampling occasions. There was no general trend for the amounts of hydrogen but there was an increase in three of the experiments and in the groundwater outside MINICAN. It was found that the water chemistry differed between A06 and A02-A04 experiment by higher sulphate and chloride concentrations in A06 compared to the others. By plotting the concentrations of chloride, sulphate against time, a decrease in sulphate concentration was found in all canister experiments. The chloride concentrations were stable during the same period. On the other hand, an increase in sulphate

Forsmark site investigation. Microorganisms in groundwater from boreholes KFM10A, KFM11A and KFM08D - numbers, viability, and metabolic diversity. Results from five sections 298.0-305.1 m and 478.0-487.5 m in KFM10A, 447.5-454.6 m in KFM11A, and 669.7-676.8 m and 828.4-835.5 m in KFM08D

International Nuclear Information System (INIS)

Pedersen, Karsten

2007-09-01

Microorganisms and their characteristic features were investigated while geochemically characterizing the groundwater, as part of the site investigation programme at Forsmark. The investigation consists of determining the total numbers of microorganisms, the concentration of adenosine-tri-phosphate (ATP), and the number of culturable heterotrophic aerobic bacteria (CHAB); also included is a method for determining the numbers of organisms belonging to different physiological groups, the most probable number (MPN) method. This investigation covered eight different groups, namely, nitrate-, iron-, manganese-, and sulphate-reducing bacteria, auto- and heterotrophic acetogens, and auto- and heterotrophic methanogens. The reproducibility of the MPN method was tested using groundwater from a depth of 450 m at the Aspo Hard Rock Laboratory and was found to be excellent. Samples were taken from boreholes KFM10A at 298.305 m and 478.487 m, KFM11A at 447.454 m, and KFM08D at 669.676 m and 828.835 m; the sampling dates were 2006-11-28, 2006-10-31, 2007-03-13, 2007-06-19, and 2007-05-02, respectively. The total number of cells (TNC) found in KFM10A groundwater was the highest so far found in a total of 19 analysed sections in the Forsmark area. In contrast, KFM08D-828 m and KFM11A-447 m had among the lowest numbers of cells found thus far. A large amount of ATP per cell indicates large, active cells. The average of all previous ATP/TNC ratios (n ≅ 100) in deep groundwater was determined to be 0.43. The analysed groundwater samples from KFM10A-478 m and KFM08D-669 m had ATP/TNC ratios exceeding the overall average of 0.43 for deep groundwater. This suggests that the microorganisms in these groundwaters possessed viability and activity levels above the average for deep groundwater microorganisms. The ratios between the CHAB and NRB numbers found here suggest that there was no surface water contamination. The percentages of TNC culturable using the MPN method were in the 1

Control of amphibious weed ipomoea (Ipomoea carnea) by utilizing it for the extraction of volatile fatty acids as energy precursors.

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Rafiq Kumar, M; Tauseef, S M; Abbasi, Tasneem; Abbasi, S A

2015-01-01

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 10(th) or 11(th) 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.

Control of amphibious weed ipomoea (Ipomoea carnea by utilizing it for the extraction of volatile fatty acids as energy precursors

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M. Rafiq Kumar

2015-01-01

Full Text Available 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.

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

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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.

Electro-Fermentation in Aid of Bioenergy and Biopolymers

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Prasun Kumar

2018-02-01

Full Text Available The soaring levels of industrialization and rapid progress towards urbanization across the world have elevated the demand for energy besides generating a massive amount of waste. The latter is responsible for poisoning the ecosystem in an exponential manner, owing to the hazardous and toxic chemicals released by them. In the past few decades, there has been a paradigm shift from “waste to wealth”, keeping the value of high organic content available in the wastes of biological origin. The most practiced processes are that of anaerobic digestion, leading to the production of methane. However; such bioconversion has limited net energy yields. Industrial fermentation targeting value-added bioproducts such as—H2, butanediols; polyhydroxyalkanoates, citric acid, vitamins, enzymes, etc. from biowastes/lignocellulosic substrates have been planned to flourish in a multi-step process or as a “Biorefinery”. Electro-fermentation (EF is one such technology that has attracted much interest due to its ability to boost the microbial metabolism through extracellular electron transfer during fermentation. It has been studied on various acetogens and methanogens, where the enhancement in the biogas yield reached up to 2-fold. EF holds the potential to be used with complex organic materials, leading to the biosynthesis of value-added products at an industrial scale.

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

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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.

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

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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.

Alamethicin Suppresses Methanogenesis and Promotes Acetogenesis in Bioelectrochemical Systems

KAUST Repository

Zhu, Xiuping; Siegert, Michael; Yates, Matthew D.; Logan, Bruce E.

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.

Functional structure of the bromeliad tank microbiome is strongly shaped by local geochemical conditions.

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Louca, Stilianos; Jacques, Saulo M S; Pires, Aliny P F; Leal, Juliana S; González, Angélica L; Doebeli, Michael; Farjalla, Vinicius F

2017-08-01

Phytotelmata in tank-forming Bromeliaceae plants are regarded as potential miniature models for aquatic ecology, but detailed investigations of their microbial communities are rare. Hence, the biogeochemistry in bromeliad tanks remains poorly understood. Here we investigate the structure of bacterial and archaeal communities inhabiting the detritus within the tanks of two bromeliad species, Aechmea nudicaulis and Neoregelia cruenta, from a Brazilian sand dune forest. We used metagenomic sequencing for functional community profiling and 16S sequencing for taxonomic profiling. We estimated the correlation between functional groups and various environmental variables, and compared communities between bromeliad species. In all bromeliads, microbial communities spanned a metabolic network adapted to oxygen-limited conditions, including all denitrification steps, ammonification, sulfate respiration, methanogenesis, reductive acetogenesis and anoxygenic phototrophy. Overall, CO2 reducers dominated in abundance over sulfate reducers, and anoxygenic phototrophs largely outnumbered oxygenic photoautotrophs. Functional community structure correlated strongly with environmental variables, between and within a single bromeliad species. Methanogens and reductive acetogens correlated with detrital volume and canopy coverage, and exhibited higher relative abundances in N. cruenta. A comparison of bromeliads to freshwater lake sediments and soil from around the world, revealed stark differences in terms of taxonomic as well as functional microbial community structure. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

Alamethicin Suppresses Methanogenesis and Promotes Acetogenesis in Bioelectrochemical Systems

KAUST Repository

Zhu, Xiuping

2015-03-27

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.

Microbiological Diversity of the Anaerobic Sludge During Treatment of Venezuelan Oilfield Produced Waters

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Cajacuri María Patricia

2013-06-01

Full Text Available In the present investigation the microbial abundances in the granular sludge of two upflow anaerobic sludge blanket reactors (UASB were compared: the first one fed with production waters of light oil (31.1-39.0° API, from the zuliana region (Venezuela (APP and the second one with glucose. To this respect, the populations of glucose fermenting bacteria (BFG, acetogenic bacteria (BAC, metanogens (MET, sulfatereducing bacteria (BSR, nitrate-reducing bacteria (BNRand heterotrophic bacteria were monitored, using selective culture media. The microbial density was correlated with physicochemical parameters: pH, total alkalinity, COD, SO4 =, NO3-, as well as with the percentages of CH4, CO2 and N2in the biogas. The results exhibit significant differences between the microbial diversity of both reactors, with a proportion of BFG > BSR > MET > BAC > BNR for the glucose reactor and of MET > BNR > BAC > BSR > BFG for the APP. The abundance of bacteria in the glucose reactor was in the order of 108, whereas in the APP reactor was of 105, which ensues from the organic and mineral composition of effluents. The results presented in this study reach evidences on the population dynamics in sludge of UASB reactors, during the treatment of oilfield produced waters.

Suppression of the Escherichia coli dnaA46 mutation by changes in the activities of the pyruvate-acetate node links DNA replication regulation to central carbon metabolism.

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Tymecka-Mulik, Joanna; Boss, Lidia; Maciąg-Dorszyńska, Monika; Matias Rodrigues, João F; Gaffke, Lidia; Wosinski, Anna; Cech, Grzegorz M; Szalewska-Pałasz, Agnieszka; Węgrzyn, Grzegorz; Glinkowska, Monika

2017-01-01

To ensure faithful transmission of genetic material to progeny cells, DNA replication is tightly regulated, mainly at the initiation step. Escherichia coli cells regulate the frequency of initiation according to growth conditions. Results of the classical, as well as the latest studies, suggest that the DNA replication in E. coli starts at a predefined, constant cell volume per chromosome but the mechanisms coordinating DNA replication with cell growth are still not fully understood. Results of recent investigations have revealed a role of metabolic pathway proteins in the control of cell division and a direct link between metabolism and DNA replication has also been suggested both in Bacillus subtilis and E. coli cells. In this work we show that defects in the acetate overflow pathway suppress the temperature-sensitivity of a defective replication initiator-DnaA under acetogenic growth conditions. Transcriptomic and metabolic analyses imply that this suppression is correlated with pyruvate accumulation, resulting from alterations in the pyruvate dehydrogenase (PDH) activity. Consequently, deletion of genes encoding the pyruvate dehydrogenase subunits likewise resulted in suppression of the thermal-sensitive growth of the dnaA46 strain. We propose that the suppressor effect may be directly related to the PDH complex activity, providing a link between an enzyme of the central carbon metabolism and DNA replication.

Metagenome-Assembled Genome Sequences of Acetobacterium sp. Strain MES1 and Desulfovibrio sp. Strain MES5 from a Cathode-Associated Acetogenic Microbial Community.

Science.gov (United States)

Ross, Daniel E; Marshall, Christopher W; May, Harold D; Norman, R Sean

2017-09-07

Draft genome sequences of Acetobacterium sp. strain MES1 and Desulfovibrio sp. strain MES5 were obtained from the metagenome of a cathode-associated community enriched within a microbial electrosynthesis system (MES). The draft genome sequences provide insight into the functional potential of these microorganisms within an MES and a foundation for future comparative analyses. Copyright © 2017 Ross et al.

Comparison of expression of key sporulation, solventogenic and acetogenic genes in C. beijerinckii NRRL B-598 and its mutant strain overexpressing spo0A.

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Kolek, J; Diallo, M; Vasylkivska, M; Branska, B; Sedlar, K; López-Contreras, A M; Patakova, P

2017-11-01

The production of acetone, butanol and ethanol by fermentation of renewable biomass has potential to become a valuable industrial process. Mechanisms of solvent production and sporulation involve some common regulators in some ABE-producing clostridia, although details of the links between the pathways are not clear. In this study, we compare a wild-type (WT) Clostridium beijerinckii NRRL B-598 with its mutant strain OESpo0A, in which the gene encoding Spo0A, an important regulator of both sporulation and solventogenesis, is overexpressed in terms of solvent and acid production. We also compare morphologies during growth on two different media: TYA broth, where the WT culture sporulates, and RCM, where the WT culture does not. In addition, RT-qPCR-based analysis of expression profiles of spo0A, spoIIE, sigG, spoVD, ald and buk1 genes involved in sporulation or solvent production in these strains, were compared. The OESpo0A mutant did not produce spores and butanol titre was lower compared to the WT, but increased amounts of butyric acid and ethanol were produced. The gene spo0A had high levels of expression in the WT under non-sporulating culture conditions while other selected genes for sporulation factors were downregulated significantly. Similar observations were obtained for OESpo0A where spo0A overexpression and downregulation of other sporulation genes were demonstrated. Higher expression of spo0A led to higher expression of buk1 and ald, which could confirm the role of spo0A in activation of the solventogenic pathway, although solvent production was not affected significantly in the WT and was weakened in the OESpo0A mutant.

Comparison of expression of key sporulation, solventogenic and acetogenic genes in C. beijerinckii NRRL B-598 and its mutant strain overexpressing spo0A

NARCIS (Netherlands)

Kolek, J.; Diallo, M.; Vasylkivska, M.; Branska, B.; Sedlar, K.; López-Contreras, A.M.; Patakova, P.

2017-01-01

The production of acetone, butanol and ethanol by fermentation of renewable biomass has potential to become a valuable industrial process. Mechanisms of solvent production and sporulation involve some common regulators in some ABE-producing clostridia, although details of the links between the

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

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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

Hydrothermal focusing of chemical and chemiosmotic energy, supported by delivery of catalytic Fe, Ni, Mo/W, Co, S and Se, forced life to emerge.

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Nitschke, Wolfgang; Russell, Michael J

2009-11-01

Energised by the protonmotive force and with the intervention of inorganic catalysts, at base Life reacts hydrogen from a variety of sources with atmospheric carbon dioxide. It seems inescapable that life emerged to fulfil the same role (i.e., to hydrogenate CO(2)) on the early Earth, thus outcompeting the slow geochemical reduction to methane. Life would have done so where hydrothermal hydrogen interfaced a carbonic ocean through inorganic precipitate membranes. Thus we argue that the first carbon-fixing reaction was the molybdenum-dependent, proton-translocating formate hydrogenlyase system described by Andrews et al. (Microbiology 143:3633-3647, 1997), but driven in reverse. Alkaline on the inside and acidic and carbonic on the outside - a submarine chambered hydrothermal mound built above an alkaline hydrothermal spring of long duration - offered just the conditions for such a reverse reaction imposed by the ambient protonmotive force. Assisted by the same inorganic catalysts and potential energy stores that were to evolve into the active centres of enzymes supplied variously from ocean or hydrothermal system, the formate reaction enabled the rest of the acetyl coenzyme-A pathway to be followed exergonically, first to acetate, then separately to methane. Thus the two prokaryotic domains both emerged within the hydrothermal mound-the acetogens were the forerunners of the Bacteria and the methanogens were the forerunners of the Archaea.

Isolation and characterization of cultivable fermentative bacteria from the intestine of two edible snails, Helixpomatia and Cornu aspersum (Gastropoda: Pulmonata

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MAR YVONNE CHARRIER

2006-01-01

Full Text Available The intestinal microbiota of the edible snails Cornu aspersum fSyn: H. aspersa, and Helix pomatia were investigated by culture-based methods, 16S rRNA sequence analyses and phenotypic characterisations. The study was carried out on aestivating snails and two populations of H. pomatia were considered. The cultivable bacteria dominated in the distal part of the intestine, with up to 5.10(9 CFU g -1, but the Swedish H. pomatia appeared significantly less colonised, suggesting a higher sensitivity of its microbiota to climatic change. All the strains, but one, shared ≥ 97% sequence identity with reference strains. They were arranged into two taxa: the Gamma Proteobacteria with Buttiauxella, Citrobacter, Enterobacter, Kluyvera, Obesumbacterium, Raoultella and the Firmicutes with Enterococcus, Lactococcus, and Clostridium. According to the literature, these genera are mostly assigned to enteric environments or to phyllosphere, data in favour of culturing snails in contact with soil and plants. None of the strains were able to digest filter paper, Avicel cellulose or carboxymethyl cellulose (CMC. Acetogens and methanogenic archaea were not cultivated, so the fate of hydrogen remains questionable. This microbiota could play important roles in the digestive process (fermentation and the energy supply of the snail (L-lactate, acetate. The choice of cereals and plants by snail farmers should take into account the fermentative abilities of the intestinal microbiota

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

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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).

Activation of CO2-reducing methanogens in oil reservoir after addition of nutrient.

Science.gov (United States)

Yang, Guang-Chao; Zhou, Lei; Mbadinga, Serge Maurice; You, Jing; Yang, Hua-Zhen; Liu, Jin-Feng; Yang, Shi-Zhong; Gu, Ji-Dong; Mu, Bo-Zhong

2016-12-01

Nutrient addition as part of microbial enhanced oil recovery (MEOR) operations have important implications for more energy recovery from oil reservoirs, but very little is known about the in situ response of microorganisms after intervention. An analysis of two genes as biomarkers, mcrA encoding the key enzyme in methanogenesis and fthfs encoding the key enzyme in acetogenesis, was conducted during nutrient addition in oil reservoir. Clone library data showed that dominant mcrA sequences changed from acetoclastic (Methanosaetaceae) to CO 2 -reducing methanogens (Methanomicrobiales and Methanobacteriales), and the authentic acetogens affiliated to Firmicutes decreased after the intervention. Principal coordinates analysis (PCoA) and Jackknife environment clusters revealed evidence on the shift of the microbial community structure among the samples. Quantitative analysis of methanogens via qPCR showed that Methanobacteriales and Methanomicrobiales increased after nutrient addition, while acetoclastic methanogens (Methanosaetaceae) changed slightly. Nutrient treatment activated native CO 2 -reducing methanogens in oil reservoir. The high frequency of Methanobacteriales and Methanomicrobiales (CO 2 -reducers) after nutrient addition in this petroleum system suggested that CO 2 -reducing methanogenesis was involved in methane production. The nutrient addition could promote the methane production. The results will likely improve strategies of utilizing microorganisms in subsurface environments. Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

History of adaptation determines short-term shifts in performance and community structure of hydrogen-producing microbial communities degrading wheat straw.

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Valdez-Vazquez, Idania; Morales, Ana L; Escalante, Ana E

2017-11-01

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 H 2 -producing members, and Bacteroidetes with non-H 2 -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. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Archaea and Bacteria Acclimate to High Total Ammonia in a Methanogenic Reactor Treating Swine Waste

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Sofia Esquivel-Elizondo

2016-01-01

Full Text Available 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.

Ecophysiology of terminal carbon metabolizing bacteria in anoxic sedimentary environments

International Nuclear Information System (INIS)

Phelps, T.J.

1985-01-01

Chemical, radiotracer, and microbiological experiments were used to understand the transformation of simple carbon compounds by anaerobic bacteria in diverse aquatic sediments and laboratory cultures. The mildly acidic sediments of Knack Lake (pH 6.2), displayed low rates of organic decomposition, and methane formation occurred almost exclusively from acetate. Low pH inhibited methanogenesis and organic decomposition. Fall turnover in Lake Mendota sediments was associated with dramatic changes in environmental parameters including: elevated concentrations of sulfate and carbon metabolites, increased rates of sulfate reduction, decreased levels of methanogenesis, increased ratio (by viable counts) of sulfate reducing to methanogenic bacteria, and higher 14 CO 2 / 14 C 4 + 14 CO 2 gas ratios produced during the biodegradation of 14 C-carbon substrates (e.g., acetate and methanol). Hydrogen consumption by sulfate reducers in Lake Mendota sediments and in co-cultures of Desulfovibrio vulgaris and Methanosarcina barkeri led to an alteration in the carbon and electron flow pathway resulting in increased CO 2 , sulfide production, and decreased methanogenesis. These data agreed with the environmental observations in Lake Mendota that high sulfate concentrations resulted in higher ratios of CO 2 /CH 4 produced from the degradation of organic matter. A new glycine-metabolizing acetogenic species was isolated and characterized from Knaack Lake which further extended the known diversity of anaerobic bacteria in nature

Geomicrobial investigations of groundwaters from Olkiluoto, Haestholmen, Kivetty and Romuvaara, Finland

Energy Technology Data Exchange (ETDEWEB)

Haveman, S.A.; Pedersen, K. [Goeteborg Univ. (Sweden); Ruotsalainen, P. [Fintact Oy, Helsinki (Finland)

1998-08-01

Groundwater from four deep hard rock sites being considered for nuclear waste disposal in Finland (Olkiluoto, Haestholmen, Kivetty and Romuvaara) were investigated for microbial populations. Bacteria will be present in a waste disposal vault, so it is important to understand the microbiology of any potential site. Groundwater samples were collected from 200 to 950 m depth and included fresh, brackish and saline waters. Samples were collected with a pressurized groundwater sampler, PAVE, which is an excellent tool for microbiological sampling. Total cell numbers were typical for deep groundwater, 105 to 106 cells/ml. Growth media designed using groundwater chemistry data were used for enumeration of methanogens, acetogens, sulfate reducing bacteria (SRB) and iron reducing bacteria (IRB). Microbial populations varied between sites. Iron sulfide fracture minerals are common in the brackish high sulfate groundwaters of Olkiluoto, where SRB predominated. Haestholmen groundwater has high dissolved iron, iron hydroxide fracture minerals and IRB were the main microbial population. Kivetty and Romuvaara had mixed populations. It has been proposed that deep subsurface ecosystems are based on hydrogen and carbon dioxide which provide energy and carbon to support the food chain. Signs of such an ecosystem were seen in Olkiluoto. More study is needed to understand the basis for deep subsurface life. From a microbiological point of view, all sites investigated are equally suitable for nuclear waste disposal. (orig.) 66 refs.

Contributions of available substrates and activities of trophic microbial community to methanogenesis in vegetative and reproductive rice rhizospheric soil.

Science.gov (United States)

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.

Geomicrobial investigations of groundwaters from Olkiluoto, Haestholmen, Kivetty and Romuvaara, Finland

International Nuclear Information System (INIS)

Haveman, S.A.; Pedersen, K.; Ruotsalainen, P.

1998-08-01

Groundwater from four deep hard rock sites being considered for nuclear waste disposal in Finland (Olkiluoto, Haestholmen, Kivetty and Romuvaara) were investigated for microbial populations. Bacteria will be present in a waste disposal vault, so it is important to understand the microbiology of any potential site. Groundwater samples were collected from 200 to 950 m depth and included fresh, brackish and saline waters. Samples were collected with a pressurized groundwater sampler, PAVE, which is an excellent tool for microbiological sampling. Total cell numbers were typical for deep groundwater, 105 to 106 cells/ml. Growth media designed using groundwater chemistry data were used for enumeration of methanogens, acetogens, sulfate reducing bacteria (SRB) and iron reducing bacteria (IRB). Microbial populations varied between sites. Iron sulfide fracture minerals are common in the brackish high sulfate groundwaters of Olkiluoto, where SRB predominated. Haestholmen groundwater has high dissolved iron, iron hydroxide fracture minerals and IRB were the main microbial population. Kivetty and Romuvaara had mixed populations. It has been proposed that deep subsurface ecosystems are based on hydrogen and carbon dioxide which provide energy and carbon to support the food chain. Signs of such an ecosystem were seen in Olkiluoto. More study is needed to understand the basis for deep subsurface life. From a microbiological point of view, all sites investigated are equally suitable for nuclear waste disposal. (orig.)

Growth Inhibition of Sporomusa ovata by Incorporation of Benzimidazole Bases into Cobamides

Science.gov (United States)

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

EFICIENTIZAREA OBŢINERII SEDIMENTELOR FURAJERE B12 -VITAMINIZATE DIN APE REZIDUALE AGROINDUSTRIALE: 2. MODIFICĂRI ALE UTILAJULUI

Directory of Open Access Journals (Sweden)

Victor COVALIOV

2017-03-01

Full Text Available Prin modificări constructive ale dispozitivelor bioreactorului şi soluţii noi: separarea zonelor acetogenă şi meta­no­genă în interiorul bioreactorului, recircularea CO2 şi suplimentarea acestuia cu H2 exogen, adsorbţia vitaminei B12 din lichidul postfermentare cu diatomită s-a obţinut ridicarea conţinutului de vitamina B12 în sedimentele epurării fermentativ–metanogene (anaerobe a borhotului postalcoolic până la calitatea de con­centrat furajer B12-vitaminizat, concomitent cu intensificarea producerii biometanului, ca elemente ale ridicării eficienţei ecologo-economice a epurării anaerobe a borhotului.MORE EFFICIENT PRODUCITON OF VITAMINIZED FORAGE SLUDGE CONTAINING B12 FROM AGRO-INDUSTRIAL WASTES: 2. EQUIPMENT MODIFICAITON The increase in vitamin B12 contents in the sludge was obtained resulted from the methanogenic (anaerobic digestion of post-distillery vinasse, through the design modifications of bioreactor interiour, CO2 re-circulation and its interaction with additionally dosed exogenic hydrogen. Vitamin B12 was further adsorbed on diatomite surface from the post-digestion liquid, which made it possible to produce the cattle forage concentrate enriched with vitamin B12. At the same time, biomethane production was intensified, being the element of ecologically-economic efficiency of anaerobic treatment of vinasse (agro-industrial waste.

Performance of mesophilic anaerobic granules for removal of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) from aqueous solution

International Nuclear Information System (INIS)

An Chunjiang; He Yanling; Huang Guohe; Liu Yonghong

2010-01-01

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 1 g 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.

Microbial community analysis of ambient temperature anaerobic digesters

Energy Technology Data Exchange (ETDEWEB)

Ciotola, R. [Ohio State Univ., Columbus, OH (United States). Dept. of Food, Agriculture and Biological Engineering

2010-07-01

This paper reported on a study in which designs for Chinese and Indian fixed-dome anaerobic digesters were modified in an effort to produce smaller and more affordable digesters. While these types of systems are common in tropical regions of developing countries, they have not been used in colder climates because of the low biogas yield during the winter months. Although there is evidence that sufficient biogas production can be maintained in colder temperatures through design and operational changes, there is a lack of knowledge about the seasonal changes in the composition of the microbial communities in ambient temperature digesters. More knowledge is needed to design and operate systems for maximum biogas yield in temperate climates. The purpose of this study was to cultivate a microbial community that maximizes biogas production at psychrophilic temperatures. The study was conducted on a 300 gallon experimental anaerobic digester on the campus of Ohio State University. Culture-independent methods were used on weekly samples collected from the digester in order to examine microbial community response to changes in ambient temperature. Microbial community profiles were established using universal bacterial and archaeal primers that targeted the 16S rRNA gene. In addition to the methanogenic archaea, this analysis also targeted some of the other numerically and functionally important microbial taxa in anaerobic digesters, such as hydrolytic, fermentative, acetogenic and sulfate reducing bacteria. According to preliminary results, the composition of the microbial community shifts with changes in seasonal temperature.

Studies on the mechanism of electron bifurcation catalyzed by electron transferring flavoprotein (Etf) and butyryl-CoA dehydrogenase (Bcd) of Acidaminococcus fermentans.

Science.gov (United States)

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

2014-02-21

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.

Molecular analysis of the biomass of a fluidized bed reactor treating synthetic vinasse at anaerobic and micro-aerobic conditions.

Science.gov (United States)

Rodríguez, Elisa; Lopes, Alexandre; Fdz-Polanco, María; Stams, Alfons J M; García-Encina, Pedro A

2012-03-01

The microbial communities (Bacteria and Archaea) established in an anaerobic fluidized bed reactor used to treat synthetic vinasse (betaine, glucose, acetate, propionate, and butyrate) were characterized by denaturing gradient gel electrophoresis (DGGE) and phylogenetic analysis. This study was focused on the competitive and syntrophic interactions between the different microbial groups at varying influent substrate to sulfate ratios of 8, 4, and 2 and anaerobic or micro-aerobic conditions. Acetogens detected along the anaerobic phases at substrate to sulfate ratios of 8 and 4 seemed to be mainly involved in the fermentation of glucose and betaine, but they were substituted by other sugar or betaine degraders after oxygen application. Typical fatty acid degraders that grow in syntrophy with methanogens were not detected during the entire reactor run. Likely, sugar and betaine degraders outnumbered them in the DGGE analysis. The detected sulfate-reducing bacteria (SRB) belonged to the hydrogen-utilizing Desulfovibrio. The introduction of oxygen led to the formation of elemental sulfur (S(0)) and probably other sulfur compounds by sulfide-oxidizing bacteria (γ-Proteobacteria). It is likely that the sulfur intermediates produced from sulfide oxidation were used by SRB and other microorganisms as electron acceptors, as was supported by the detection of the sulfur respiring Wolinella succinogenes. Within the Archaea population, members of Methanomethylovorans and Methanosaeta were detected throughout the entire reactor operation. Hydrogenotrophic methanogens mainly belonging to the genus Methanobacterium were detected at the highest substrate to sulfate ratio but rapidly disappeared by increasing the sulfate concentration.

Aespoe Hard Rock Laboratory Canister Retrieval Test. Microorganisms in buffer from the Canister Retrieval Test - numbers and metabolic diversity

International Nuclear Information System (INIS)

Lydmark, Sara; Pedersen, Karsten

2011-03-01

'Canister Retrieval Test' (CRT) is an experiment that started at Aespoe Hard Rock Laboratory (HRL) 2000. CRT is a part of the investigations which evaluate a possible KBS-3 storage of nuclear waste. The primary aim was to see whether it is possible or not to retrieve a copper canister after storage under authentic KBS-3 conditions. However, CRT also provided a unique opportunity to investigate if bacteria survived in the bentonite buffer during storage. Therefore, in connection to the retrieval of the canister microbiological samples were extracted from the bentonite buffer and the bacterial composition was studied. In this report, microbiological analyses of a total of 66 samples at the C2, R10, R9 and R6 levels in the bentonite from CRT are presented and discussed. By culturing bacteria from the bentonite in specific media the following bacterial parameters were investigated: The total amount of culturable heterotrophic aerobic bacteria, sulphate-reducing bacteria, and bacteria that produce the organic compound acetate (acetogens). The biovolume in the bentonite was determined by detection of the ATP content. In addition, bacteria from the bentonite were cultured in different sulphate-reducing media. In these cultures, the presence of the biotic compounds sulphide and acetate was investigated, since these have potentially negative effect on the copper canister in a KBS-3 repository. The results were to some extent compared to density, water content, and temperature data provided by Clay Technology AB. The results showed that 10 0 -10 2 viable sulphate-reducing and acetogenic bacteria and 10 2 -10 4 heterotrophic aerobic bacteria g -1 bentonite were present after five years of storage in the rock. Bacteria with several morphologies could be found in the cultures with bentonite. The most bacteria were detected in the bentonite buffer close to the rock but in a few samples also in bentonite close to the copper canister. When the presence of bacteria in the bentonite

Aespoe Hard Rock Laboratory Canister Retrieval Test. Microorganisms in buffer from the Canister Retrieval Test - numbers and metabolic diversity

Energy Technology Data Exchange (ETDEWEB)

Lydmark, Sara; Pedersen, Karsten (Microbial Analytics Sweden AB (Sweden))

2011-03-15

'Canister Retrieval Test' (CRT) is an experiment that started at Aespoe Hard Rock Laboratory (HRL) 2000. CRT is a part of the investigations which evaluate a possible KBS-3 storage of nuclear waste. The primary aim was to see whether it is possible or not to retrieve a copper canister after storage under authentic KBS-3 conditions. However, CRT also provided a unique opportunity to investigate if bacteria survived in the bentonite buffer during storage. Therefore, in connection to the retrieval of the canister microbiological samples were extracted from the bentonite buffer and the bacterial composition was studied. In this report, microbiological analyses of a total of 66 samples at the C2, R10, R9 and R6 levels in the bentonite from CRT are presented and discussed. By culturing bacteria from the bentonite in specific media the following bacterial parameters were investigated: The total amount of culturable heterotrophic aerobic bacteria, sulphate-reducing bacteria, and bacteria that produce the organic compound acetate (acetogens). The biovolume in the bentonite was determined by detection of the ATP content. In addition, bacteria from the bentonite were cultured in different sulphate-reducing media. In these cultures, the presence of the biotic compounds sulphide and acetate was investigated, since these have potentially negative effect on the copper canister in a KBS-3 repository. The results were to some extent compared to density, water content, and temperature data provided by Clay Technology AB. The results showed that 100-102 viable sulphate-reducing and acetogenic bacteria and 102-104 heterotrophic aerobic bacteria g-1 bentonite were present after five years of storage in the rock. Bacteria with several morphologies could be found in the cultures with bentonite. The most bacteria were detected in the bentonite buffer close to the rock but in a few samples also in bentonite close to the copper canister. When the presence of bacteria in the

The transcriptional response of microbial communities in thawing Alaskan permafrost soils

Science.gov (United States)

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

A Genetic System for Clostridium ljungdahlii: a Chassis for Autotrophic Production of Biocommodities and a Model Homoacetogen

Energy Technology Data Exchange (ETDEWEB)

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.

Modified batch anaerobic digestion assay for testing efficiencies of trace metal additives to enhance methane production of energy crops.

Science.gov (United States)

Brulé, Mathieu; Bolduan, Rainer; Seidelt, Stephan; Schlagermann, Pascal; Bott, Armin

2013-01-01

Batch biochemical methane potential (BMP) assays to evaluate the methane yield of biogas substrates such as energy crops are usually carried out with undiluted inoculum. A BMP assay was performed on two energy crops (green cuttings and grass silage). Anaerobic digestion was performed both with and without supplementation of three commercial additives containing trace metals in liquid, solid or adsorbed form (on clay particles). In order to reveal positive effects of trace metal supplementation on the methane yield, besides undiluted inoculum, 3-fold and 10-fold dilutions of the inoculum were applied for substrate digestion. Diluted inoculum variants were supplemented with both mineral nutrients and pH-buffering substances to prevent a collapse of the digestion process. As expected, commercial additives had no effect on the digestion process performed with undiluted inoculum, while significant increases of methane production through trace element supplementation could be observed on the diluted variants. The effect of inoculum dilution may be twofold: (1) decrease in trace metal supplementation from the inoculum and (2) reduction in the initial number of bacterial cells. Bacteria require higher growth rates for substrate degradation and hence have higher trace element consumption. According to common knowledge of the biogas process, periods with volatile fatty acids accumulation and decreased pH may have occurred in the course ofanaerobic digestion. These effects may have led to inhibition, not only ofmethanogenes and acetogenes involved in the final phases of methane production, but also offibre-degrading bacterial strains involved in polymer hydrolysis. Further research is required to confirm this hypothesis.

The transcriptional response of microbial communities in thawing Alaskan permafrost soils

Directory of Open Access Journals (Sweden)

M J L Coolen

2015-03-01

Full Text Available Thawing of permafrost soils is expected to stimulate microbial decomposition and respiration of sequestered carbon. This could, in turn, increase atmospheric concentrations of greenhouse gases, 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 eleven 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.

Studies on the Mechanism of Electron Bifurcation Catalyzed by Electron Transferring Flavoprotein (Etf) and Butyryl-CoA Dehydrogenase (Bcd) of Acidaminococcus fermentans*

Science.gov (United States)

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

Numbers, biomass and cultivable diversity of microbial populations relate to depth and borehole-specific conditions in groundwater from depths of 4-450 m in Olkiluoto, Finland.

Science.gov (United States)

Pedersen, Karsten; Arlinger, Johanna; Eriksson, Sara; Hallbeck, Anna; Hallbeck, Lotta; Johansson, Jessica

2008-07-01

Microbiology, chemistry and dissolved gas in groundwater from Olkiluoto, Finland, were analysed over 3 years; samples came from 16 shallow observation tubes and boreholes from depths of 3.9-16.2 m and 14 deep boreholes from depths of 35-742 m. The average total number of cells (TNC) was 3.9 x 10(5) cells per ml in the shallow groundwater and 5.7 x 10(4) cells per ml in the deep groundwater. There was a significant correlation between the amount of biomass, analysed as ATP concentration, and TNC. ATP concentration also correlated with the stacked output of anaerobic most probable number cultivations of nitrate-, iron-, manganese- and sulphate-reducing bacteria, and acetogenic bacteria and methanogens. The numbers and biomass varied at most by approximately three orders of magnitude between boreholes, and TNC and ATP were positively related to the concentration of dissolved organic carbon. Two depth zones were found where the numbers, biomass and diversity of the microbial populations peaked. Shallow groundwater down to a depth of 16.2 m on average contained more biomass and cultivable microorganisms than did deep groundwater, except in a zone at a depth of approximately 300 m where the average biomass and number of cultivable microorganisms approached those of shallow groundwater. Starting at a depth of approximately 300 m, there were steep gradients of decreasing sulphate and increasing methane concentrations with depth; together with the peaks in biomass and sulphide concentration at this depth, these suggest that anaerobic methane oxidation may be a significant process at depth in Olkiluoto.

Characteristics and performance of anaerobic wastewater treatment (a review)

International Nuclear Information System (INIS)

Zeb, B.S.; Mahmood, Q.; Pervez, A.

2013-01-01

Summary: Pakistan's current population of 180 million is expected to grow to about 221 million by the year 2025. In developing countries such as Pakistan water pollution is a major threat to the livelihood of people. Pakistan is also currently experiencing profound demographic, economic changes and energy crisis that have major implications for water management. The contamination of aquatic and terrestrial ecosystems with heavy metals is a major environmental problem. Each pollution problem calls for specific optimal and cost effective solution so if one technology proves less or ineffective other takes its place. Every day the vast amounts of the municipal, industrial and agricultural wastes are released in to the environment and create serious problems. Anaerobic digestion is very attractive and cost-effective option and technology for the highly loaded waste water treatment and energy conversion. The anaerobic process is in many ways ideal for waste treatment. It has several significant advantages over other available methods. In this process organic matter is utilized as source of electron donor to reduce carbon dioxide to produce methane gas. It involves three bacterial groups namely: hydrolytic, acetogenic and methanogenic bacteria that work optimally at pH and temperature ranges of 6.8 to 7.5 and 30-35 degree C, respectively. The residence time in a digester varies with the amount and type of feed material, the configuration of the digestion system, and whether it be one-stage or two-stage. It is ideal for all kinds of wastewaters. Currently anaerobic technology is being operated at full scale in many industrialized nations. (author)

The potential of subterranean microbes in facilitating actinide migration at the Grimsel test site and waste isolation pilot plant

International Nuclear Information System (INIS)

Gillow, J.B.; Dunn, M.; Francis, A.J.; Lucero, D.A.; Papenguth, H.W.

2000-01-01

Microorganisms may affect the long-term stability and mobility of radionuclides disposed of in deep geological formations. Of particular concern is the association of radionuclides with subterranean microbial cells and their subsequent transport as biocolloids. We characterized the total microbial populations in two groundwater samples: one from the Culebra dolomite member of the Rustler formation at the waste isolation pilot plant (WIPP), NM, and the other from the granitic formation at the Grimsel test site (GTS), Switzerland. Culebra groundwater (ionic strength 2.8 M, pH 7) contained 1.51 ± 1.08 x 10 5 cells ml -1 , with a mean cell length of 0.75 ± 0.04 μm and width of 0.58 ± 0.02 μm. In contrast, low ionic-strength GTS groundwater (0.001 M, pH 10) contained 3.97 ± 0.37 x 10 3 cells ml -1 , with a mean cell length of 1.50 ± 0.14 μm and width of 0.37 ± 0.01 μm. Adding appropriate electron donors and acceptors to the groundwaters facilitated the growth of aerobic, denitrifying, fermentative, and acetogenic microorganisms. Uranium biosorption was studied in two isolates from these groundwaters, as well as several pure cultures from saline and non-saline environments. Halophilic and non-halophilic bacteria exhibited differences in the amount of U associated with the cells. Plutonium uptake by Acetobacterium sp. isolated from GTS varied from 30-145 pg of Pu mg -1 dry weight of cells. (orig.)

Autotrophic acetyl coenzyme A biosynthesis in Methanococcus maripaludis

International Nuclear Information System (INIS)

Shieh, J.; Whitman, W.B.

1988-01-01

To detect autotrophic CO 2 assimilation in cell extracts of Methanococcus maripaludis, lactate dehydrogenase and NADH were added to convert pyruvate formed from autotropically synthesized acetyl coenzyme A to lactate. The lactate produced was determined spectrophotometrically. When CO 2 fixation was pulled in the direction of lactate synthesis, CO 2 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 CO 2 and H 2 , but H 2 + CO 2 -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 2.1 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 14 CO 2 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. 14 CH 2 O was specifically incorporated into the C-3 of lactate, and 14 CO 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 CO 2 assimilation

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

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Jamie A FitzGerald

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

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

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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.

Metagenomic insights into anaerobic metabolism along an Arctic peat soil profile.

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David A Lipson

Full Text Available 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.

Early Microbial Evolution: The Age of Anaerobes.

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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.

Evaluating the microbial community and gene regulation involved in crystallization kinetics of ZnS formation in reduced environments

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Falk, Nicholas; Chaganti, Subba Rao; Weisener, Christopher G.

2018-01-01

In anoxic environments, sulfate-reducing bacteria (SRB) may precipitate sparingly-soluble, fine-grained sulfides as by-products of dissimilatory sulfate reduction. This bio-mechanism lends importance to acid rock drainage (ARD) remediation efforts for its ability to immobilize harmful metals from contaminant pathways, including Zn. However, SRB often coexist alongside multiple bacterial guilds in these environments, and may be sustained or hindered by the activities and metabolic by-products of their cohorts, driven by the commonly available substrates. Thus, the effectiveness of onset sulfate reduction and resultant metal-sulfide generation in ARD treatment can be enhanced by unravelling the complexities associated with these interactions. This research used material sourced from a passive bioreactor system located at the Stockton Coal Mine, New Zealand to investigate SRB activity and associated community function. RNA sequencing showed spore-forming Desulfitobacterium and Desulfotomaculum as the dominant SRB enriched from the reduced zone of the bioreactor. Metatranscriptomic analysis revealed acetogenic bacteria as syntrophic partners in substrate availability and Pseudomonas as metal-resistant community members. ZnS precipitates were observed by scanning electron microscopy (SEM) in short-term batch enrichments as well as long-term raw bioreactor material, with observed differences in mineral arrangement indicative of different nucleation scenarios. Syntrophy, metal response mechanisms, and the capacity for sporulation were observed as key microbial functions in mine waste reclamation settings. Here, Zn and S mass balance calculations coupled with RNA sequence data and microscopy illuminated favourable physicochemical and biological conditions for early metal sulfide precipitation in passive treatment systems for ARD and highlight the advantages of linking both lab and field-scale studies.

Ammonia threshold for inhibition of anaerobic digestion of thin stillage and the importance of organic loading rate.

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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. © 2015 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Integration of ozonation and an anaerobic sequencing batch reactor (AnSBR) for the treatment of cherry stillage.

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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.

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

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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 CO 2 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 (H 2 ase) 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 H 2 ase 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 H 2 ase activity photosynthesized acetic acid the fastest. These results suggest a two-pathway mechanism: a high quantum efficiency charge-transfer pathway to H 2 ase generating H 2 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.

Methylmercury decomposition in sediments and bacterial cultures: Involvement of methanogens and sulfate reducers in oxidative demethylation

International Nuclear Information System (INIS)

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 14 CH 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 14 CO 2 as well as lesser amounts of 14 CH 4 . Acetogenic activity resulted in fixation of some 14 CO 2 produced from 14 CH 3 HgI into acetate. Aerobic demethylation in estuarine sediments produced only 14 CH 4 , while aerobic demethylation in freshwater sediments produced small amounts of both 14 CH 4 and 14 CO 2 . Two species of Desulfovibrio produced only traces of 14 CH 4 from 14 CH 3 HgI, while a culture of a methylotrophic methanogen formed traces of 14 CO 2 and 14 CH 4 when grown on trimethylamine in the presence of the 14 CH 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

Carbon flow from volcanic CO2 into soil microbial communities of a wetland mofette.

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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.

Enteric methane mitigation technologies for ruminant livestock: a synthesis of current research and future directions.

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Patra, Amlan Kumar

2012-04-01

Enteric methane (CH(4)) emission in ruminants, which is produced via fermentation of feeds in the rumen and lower digestive tract by methanogenic archaea, represents a loss of 2% to 12% of gross energy of feeds and contributes to global greenhouse effects. Globally, about 80 million tonnes of CH(4) is produced annually from enteric fermentation mainly from ruminants. Therefore, CH(4) mitigation strategies in ruminants have focused to obtain economic as well as environmental benefits. Some mitigation options such as chemical inhibitors, defaunation, and ionophores inhibit methanogenesis directly or indirectly in the rumen, but they have not confirmed consistent effects for practical use. A variety of nutritional amendments such as increasing the amount of grains, inclusion of some leguminous forages containing condensed tannins and ionophore compounds in diets, supplementation of low-quality roughages with protein and readily fermentable carbohydrates, and addition of fats show promise for CH(4) mitigation. These nutritional amendments also increase the efficiency of feed utilization and, therefore, are most likely to be adopted by farmers. Several new potential technologies such as use of plant secondary metabolites, probiotics and propionate enhancers, stimulation of acetogens, immunization, CH(4) oxidation by methylotrophs, and genetic selection of low CH(4)-producing animals have emerged to decrease CH(4) production, but these require extensive research before they can be recommended to livestock producers. The use of bacteriocins, bacteriophages, and development of recombinant vaccines targeting archaeal-specific genes and cell surface proteins may be areas worthy of investigation for CH(4) mitigation as well. A combination of different CH(4) mitigation strategies should be adopted in farm levels to substantially decrease methane emission from ruminants. Evidently, comprehensive research is needed to explore proven and reliable CH(4) mitigation technologies

Microbial electron transport and energy conservation – the foundation for optimizing bioelectrochemical systems

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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

A fast linear predictive adaptive model of packed bed coupled with UASB reactor treating onion waste to produce biofuel.

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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 VS removed ) -1 , 0.43 ± 0.06 m 3 (kg VS removed ) -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.

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

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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

Carbon flow from volcanic CO2 into soil microbial communities of a wetland mofette

Science.gov (United States)

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.

Microbiological characterization and specific methanogenic activity of anaerobe sludges used in urban solid waste treatment

International Nuclear Information System (INIS)

Sandoval Lozano, Claudia Johanna; Vergara Mendoza, Marisol; Carreno de Arango, Mariela; Castillo Monroy, Edgar Fernando

2009-01-01

This study presents the microbiological characterization of the anaerobic sludge used in a two-stage anaerobic reactor for the treatment of organic fraction of urban solid waste (OFUSW). This treatment is one alternative for reducing solid waste in landfills at the same time producing a biogas (CH 4 and CO 2 ) and an effluent that can be used as biofertilizer. The system was inoculated with sludge from a wastewater treatment plant (WWTP) (Rio Frio Plant in Bucaramanga-Colombia) and a methanogenic anaerobic digester for the treatment of pig manure (Mesa de los Santos in Santander). Bacterial populations were evaluated by counting groups related to oxygen sensitivity, while metabolic groups were determined by most probable number (MPN) technique. Specific methanogenic activity (SMA) for acetate, formate, methanol and ethanol substrates was also determined. In the acidogenic reactor (R1), volatile fatty acids (VFA) reached values of 25,000 mg L -1 and a concentration of CO 2 of 90%. In this reactor, the fermentative population was predominant (10 5 -10 6 MPN mL -1 ). The acetogenic population was (10 5 MPN mL -1 ) and the sulphate-reducing population was (10 4 -10 5 MPN mL -1 ). In the methanogenic reactor (R2), levels of CH 4 (70%) were higher than CO 2 (25%), whereas the VFA values were lower than 4000 mg L -1 . Substrate competition between sulphate-reducing (10 4 -10 5 MPN mL -1 ) and methanogenic bacteria (10 5 MPN mL -1 ) was not detected. From the SMA results obtained, acetoclastic (2.39 g COD-CH 4 g -1 VSS -1 day -1 ) and hydrogenophilic (0.94 g COD-CH 4 g -1 VSS -1 day -1 ) transformations as possible metabolic pathways used by methanogenic bacteria is suggested from the SMA results obtained. Methanotrix sp., Methanosarcina sp., Methanoccocus sp. and Methanobacterium sp. were identified

Desulfofrigus sp. prevails in sulfate-reducing dilution cultures from sediments of the Benguela upwelling area.

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Kraft, Beate; Engelen, Bert; Goldhammer, Tobias; Lin, Yu-Shih; Cypionka, Heribert; Könneke, Martin

2013-04-01

Sediments of coastal upwelling areas are generally characterized by a high content of organic carbon that is mainly degraded via anaerobic microbial processes including sulfate reduction as a major terminal oxidation step. Despite the high importance of sulfate reduction in these sediments, the identity of sulfate-reducing bacteria (SRB) has remained almost unknown. Here, we applied a cultivation-based approach using selective enrichment conditions to study the diversity and distribution of active SRB in sediments along a transect perpendicular to the continental slope off the coast of Namibia (Meteor-cruise M76/1). To promote growth of the most abundant SRB, dilution series were prepared and amended with hydrogen, acetate, or a mixture of monomers representing typical substrates for SRB. Growth of SRB could be detected in the presence of all electron donors and from sediment down to 4 m depth. 16S rRNA gene-based DGGE analysis and sequencing revealed the predominance of SRB related to psychrophiles in particular to the genus Desulfofrigus, which made up 1 % of the total microbial community, accounting for an absolute abundance of up to 4.8 × 10(7)  cells mL(-1) . In general, the abundance of cultured SRB changed with depth and between the different sampling sites and correlated with the content of organic carbon as previously reported. Growth of chemolithotrophic SRB in relatively high dilution steps and the enrichment of methanogens as well as acetogens from deeper sediment point to a competition between hydrogen-utilizing microbial processes and their biogeochemical significance in deep sediment layers of the Benguela upwelling area. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Modelling an industrial anaerobic granular reactor using a multi-scale approach.

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Feldman, H; Flores-Alsina, X; Ramin, P; Kjellberg, K; Jeppsson, U; Batstone, D J; Gernaey, K V

2017-12-01

The objective of this paper is to show the results of an industrial project dealing with modelling of anaerobic digesters. A multi-scale mathematical approach is developed to describe reactor hydrodynamics, granule growth/distribution and microbial competition/inhibition for substrate/space within the biofilm. The main biochemical and physico-chemical processes in the model are based on the Anaerobic Digestion Model No 1 (ADM1) extended with the fate of phosphorus (P), sulfur (S) and ethanol (Et-OH). Wastewater dynamic conditions are reproduced and data frequency increased using the Benchmark Simulation Model No 2 (BSM2) influent generator. All models are tested using two plant data sets corresponding to different operational periods (#D1, #D2). Simulation results reveal that the proposed approach can satisfactorily describe the transformation of organics, nutrients and minerals, the production of methane, carbon dioxide and sulfide and the potential formation of precipitates within the bulk (average deviation between computer simulations and measurements for both #D1, #D2 is around 10%). Model predictions suggest a stratified structure within the granule which is the result of: 1) applied loading rates, 2) mass transfer limitations and 3) specific (bacterial) affinity for substrate. Hence, inerts (X I ) and methanogens (X ac ) are situated in the inner zone, and this fraction lowers as the radius increases favouring the presence of acidogens (X su ,X aa , X fa ) and acetogens (X c4 ,X pro ). Additional simulations show the effects on the overall process performance when operational (pH) and loading (S:COD) conditions are modified. Lastly, the effect of intra-granular precipitation on the overall organic/inorganic distribution is assessed at: 1) different times; and, 2) reactor heights. Finally, the possibilities and opportunities offered by the proposed approach for conducting engineering optimization projects are discussed. Copyright © 2017 Elsevier Ltd. All

Acetone production with metabolically engineered strains of Acetobacterium woodii.

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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. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Thermophilic bio-energy process study on hydrogen fermentation with vegetable kitchen waste

Energy Technology Data Exchange (ETDEWEB)

Lee, Ze-Kun; Li, Shiue-Lin; Kuo, Pei-Chen; Chen, I.-Chieh; Tien, Yu-Min; Huang, Yu-Jung; Chuang, Chung-Po; Wong, Son-Chi; Cheng, Sheng-Shung [Department of Environmental Engineering, National Cheng Kung University No. 1, University Road, Tainan 701, ROC (China)

2010-12-15

An intermittent-continuous stirred tank reactor (I-CSTR) was evaluated for thermophilic anaerobic hydrogen fermentation with vegetable kitchen waste (VKW). The seeding sludge was enriched from kitchen waste compost. Because of different seasonal dietary habits, the quality of vegetable kitchen waste was unstable, and all variations of composition were in the range from 20 to 40%. The I-CSTR process was conducted under different volumetric loading rates (VLR) with different VKW-diluted concentrations. The hydrogen production rate and yield in Run 2 (VLR as 28 g-COD L{sup -1} day{sup -1}) were 1.0 L-H{sub 2} L{sup -1} day{sup -1} and 1.7 mmol-H{sub 2} g-COD{sup -1}, which were higher than those in Run1 (VLR as 19 g-COD/L-day). The hydrolysis efficiency of organic solids (VSS) was about 45% in Run 1 better than the 32% in Run 2. The carbohydrate component of VKW was clearly degraded with the accumulation of butyrate, while the organic nitrogen component was converted to ammonia. The vegetable cellulose was degraded from 3.2 g L{sup -1} and 3.6-1.8 and 3.2 g L{sup -1} in Runs 1 and 2, respectively. In addition, the high concentration of lactate from the acidified VKW could be degraded completely both in Runs 1 and 2. According to the results of the time series profile in day 59, oil and grease were not degraded significantly. The removal of oil and grease was superficially caused by stacking on the wall, pipe, and propeller of the reactor, or by floating on the liquid surface. The 16S rDNA cloning library and sequence were applied for analyzing microbial communities. The dominant OTU was closely affiliated to Thermoanaerobacterium thermosaccharolyticum, which is considered as the predominant hydrogen-producing bacteria. The OTUs closely related to Moorella thermoacetica and Clostridiaceae bacterium FH052 were considered as acetogenic bacterium and hydrogen-producing bacteria in the I-CSTR system. (author)

Characteristics of adapted hydrogenotrophic community during biomethanation.

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Rachbauer, Lydia; Beyer, Reinhard; Bochmann, Günther; Fuchs, Werner

2017-10-01

The results presented in this study were carried out as concomitant experiments during the start-up and operation of a biomethanation unit to evaluate the effect of process parameters on carbon conversion, product formation (methane and acetate) and community composition. For that, two different samples were withdrawn from a trickle-bed reactor with immobilized enrichment culture of hydrogenotrophic methanogens adapted from sewage sludge. One sample was taken from the recirculation liquid during start-up phase while the other was withdrawn directly from the carrier material in the reactor. Elevated acid levels especially during start-up were shown to affect the overall carbon conversion. This effect was also seen during the acid tolerance testing reported here. Final acid concentrations of 1.6±0.3g/L resulted in a reduced conversion ratio of only 46%. Without acid addition complete conversion of CO 2 in the headspace was achieved. However, maximum methane production of 0.55±0.02mmol after 4days of incubation was monitored at moderate initial acetate concentration of 0.4g/L. In both analyzed inoculation materials Methanobacterium species were by far the most dominant Archaea with 21.8% in the recirculation liquid during start-up and 84.8% in the enrichment culture immobilized on the carrier material. The microbial composition of the two analyzed samples is in accordance with the results obtained for the carbon conversion and product formation. With approximately 50% of Bacteroidetes and Firmicutes present during reactor start-up the acetic acid production significantly contributed to the overall carbon conversion. In contrast, methane was produced almost exclusively in trials representing continuous operation where acetogenic bacteria accounted only up to 17.5%. In summary, the acid accumulation monitored during reactor start-up of a biomethanation unit is most likely to result from the microbial composition present. Nevertheless, complete adaptation to

Methane: Fuel or Exhaust at the Emergence of Life?

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Russell, Michael J; Nitschke, Wolfgang

2017-10-01

As many of the methanogens first encountered at hydrothermal vents were thermophilic to hyperthermophilic and comprised one of the lower roots of the evolutionary tree, it has been assumed that methanogenesis was one of the earliest, if not the earliest, pathway to life. It being well known that hydrothermal springs associated with serpentinization also bore abiotic methane, it had been further assumed that emergent biochemistry merely adopted and quickened this supposed serpentinization reaction. Yet, recent hydrothermal experiments simulating serpentinization have failed to generate methane so far, thus casting doubt on this assumption. The idea that the inverse view is worthy of debate, that is, that methanotrophy was the earlier, is stymied by the "fact" that methanotrophy itself has been termed "reverse methanogenesis," so allotting the methanogens the founding pedigree. Thus, attempting to suggest instead that methanogenesis might be termed reverse methanotrophy would require "unlearning"-a challenge to the subconscious! Here we re-examine the "impossibility" of methanotrophy predating methanogenesis as in what we have termed the "denitrifying methanotrophic acetogenic pathway." Advantages offered by such thinking are that methane would not only be a fuel but also a ready source of reduced carbon to combine with formate or carbon monoxide-available in hydrothermal fluids-to generate acetate, a target molecule of the first autotrophs. And the nitrate/nitrite required for the putative oxidation of methane with activated NO would also be a ready source of fixed nitrogen for amination reactions. Theoretical conditions for such a putative pathway would be met in a hydrothermal green rust-bearing exhalative pile and associated chimneys subject to proton and electron counter gradients. This hypothesis could be put to test in a high-pressure hydrothermal reaction chamber in which a cool carbonate/nitrate/nitrite-bearing early acidulous ocean simulant is juxtaposed

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

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Yoko eOhtomo

2013-12-01

Full Text Available 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

Exogenous addition of H2 for an in situ biogas upgrading through biological reduction of carbon dioxide into methane.

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Mulat, Daniel Girma; Mosbæk, Freya; Ward, Alastair James; Polag, Daniela; Greule, Markus; Keppler, Frank; Nielsen, Jeppe Lund; Feilberg, Anders

2017-10-01

Biological reduction of CO 2 into CH 4 by exogenous addition of H 2 is a promising technology for upgrading biogas into higher CH 4 content. The aim of this work was to study the feasibility of exogenous H 2 addition for an in situ biogas upgrading through biological conversion of the biogas CO 2 into CH 4. Moreover, this study employed systematic study with isotope analysis for providing comprehensive evidence on the underlying pathways of CH 4 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 H 2 into the headspace resulted in a completely consumption of CO 2 and a concomitant increase in CH 4 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 CH 4 content. Excess H 2 was also supplied to evaluate its effect on overall process performance. The results show that excess H 2 addition resulted in accumulation of H 2 , depletion of CO 2 and inhibition of the degradation of acetate and other volatile fatty acids (VFA). A systematic isotope analysis revealed that excess H 2 supply led to an increase in dissolved H 2 to the level that thermodynamically inhibit the degradation of VFA and stimulate homo-acetogens for production of acetate from CO 2 and H 2 . The inhibition was a temporary effect and acetate degradation resumed when the excess H 2 was removed as well as in the presence of stoichiometric amount of H 2 and CO 2 . This inhibition mechanism underlines the importance of carefully regulating the H 2 addition rate and gas retention time to the CO 2 production rate, H 2 -uptake rate and growth of hydrogenotrophic methanogens in order to achieve higher CH 4 content without the accumulation of acetate and other VFA. Copyright © 2017 Elsevier Ltd. All rights reserved.

Characterization of the core rumen microbiome in cattle during transition from forage to concentrate as well as during and after an acidotic challenge.

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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

Hydrocarbon bio-jet fuel from bioconversion of poplar biomass: life cycle assessment.

Science.gov (United States)

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.

Zinc deprivation of methanol fed anaerobic granular sludge bioreactors

Science.gov (United States)

Fermoso, Fernando G.; Collins, Gavin; Bartacek, Jan

2008-01-01

The effect of omitting zinc from the influent of mesophilic (30 °C) methanol fed upflow anaerobic sludge bed (UASB) reactors, and latter zinc supplementation to the influent to counteract the deprivation, was investigated by coupling the UASB reactor performance to the microbial ecology of the bioreactor sludge. Limitation of the specific methanogenic activity (SMA) on methanol due to the absence of zinc from the influent developed after 137 days of operation. At that day, the SMA in medium with a complete trace metal solution except Zn was 3.4 g CH4-COD g VSS−1 day−1, compared to 4.2 g CH4-COD g VSS−1 day−1 in a medium with a complete (including zinc) trace metal solution. The methanol removal capacity during these 137 days was 99% and no volatile fatty acids accumulated. Two UASB reactors, inoculated with the zinc-deprived sludge, were operated to study restoration of the zinc limitation by zinc supplementation to the bioreactor influent. In a first reactor, no changes to the operational conditions were made. This resulted in methanol accumulation in the reactor effluent after 12 days of operation, which subsequently induced acetogenic activity 5 days after the methanol accumulation started. Methanogenesis could not be recovered by the continuous addition of 0.5 μM ZnCl2 to the reactor for 13 days. In the second reactor, 0.5 μM ZnCl2 was added from its start-up. Although the reactor stayed 10 days longer methanogenically than the reactor operated without zinc, methanol accumulation was observed in this reactor (up to 1.1 g COD-MeOH L−1) as well. This study shows that zinc limitation can induce failure of methanol fed UASB reactors due to acidification, which cannot be restored by resuming the continuous supply of the deprived metal. PMID:18283507

Optimisation of sewage sludge anaerobic digestion through co-digestion with OFMSW: Effect of collection system and particle size

Energy Technology Data Exchange (ETDEWEB)

Silvestre, Gracia [IRTA, GIRO Joint Research Unit IRTA-UPC, Torre Marimon, Barcelona (Spain); Ainia, Departamento de Medio Ambiente, Bioenergía e Higiene Industrial, Paterna, Valencia (Spain); Bonmatí, August [IRTA, GIRO Joint Research Unit IRTA-UPC, Torre Marimon, Barcelona (Spain); Fernández, Belén, E-mail: belen.fernandez@irta.cat [IRTA, GIRO Joint Research Unit IRTA-UPC, Torre Marimon, Barcelona (Spain)

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.

Monitoring of microbial communities in anaerobic digestion sludge for biogas optimisation.

Science.gov (United States)

Lim, Jun Wei; Ge, Tianshu; Tong, Yen Wah

2018-01-01

This study characterised and compared the microbial communities of anaerobic digestion (AD) sludge using three different methods - (1) Clone library; (2) Pyrosequencing; and (3) Terminal restriction fragment length polymorphism (T-RFLP). Although high-throughput sequencing techniques are becoming increasingly popular and affordable, the reliance of such techniques for frequent monitoring of microbial communities may be a financial burden for some. Furthermore, the depth of microbial analysis revealed by high-throughput sequencing may not be required for monitoring purposes. This study aims to develop a rapid, reliable and economical approach for the monitoring of microbial communities in AD sludge. A combined approach where genetic information of sequences from clone library was used to assign phylogeny to T-RFs determined experimentally was developed in this study. In order to assess the effectiveness of the combined approach, microbial communities determined by the combined approach was compared to that characterised by pyrosequencing. Results showed that both pyrosequencing and clone library methods determined the dominant bacteria phyla to be Proteobacteria, Firmicutes, Bacteroidetes, and Thermotogae. Both methods also found that sludge A and B were predominantly dominated by acetogenic methanogens followed by hydrogenotrophic methanogens. The number of OTUs detected by T-RFLP was significantly lesser than that detected by the clone library. In this study, T-RFLP analysis identified majority of the dominant species of the archaeal consortia. However, many of the more highly diverse bacteria consortia were missed. Nevertheless, the combined approach developed in this study where clone sequences from the clone library were used to assign phylogeny to T-RFs determined experimentally managed to accurately predict the same dominant microbial groups for both sludge A and sludge B, as compared to the pyrosequencing results. Results showed that the combined approach of

Live Cells Decreased Methane Production in Intestinal Content of Pigs

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Y. L. Gong

2013-06-01

Full Text Available An in vitro gas production technique was used in this study to elucidate the effect of two strains of active live yeast on methane (CH4 production in the large intestinal content of pigs to provide an insight to whether active live yeast could suppress CH4 production in the hindgut of pigs. Treatments used in this study include blank (no substrate and no live yeast cells, control (no live yeast cells and yeast (YST supplementation groups (supplemented with live yeast cells, YST1 or YST2. The yeast cultures contained 1.8×1010 cells per g, which were added at the rates of 0.2 mg and 0.4 mg per ml of the fermented inoculum. Large intestinal contents were collected from 2 Duroc×Landrace×Yorkshire pigs, mixed with a phosphate buffer (1:2, and incubated anaerobically at 39°C for 24 h using 500 mg substrate (dry matter (DM basis. Total gas and CH4 production decreased (p<0.05 with supplementation of yeast. The methane production reduction potential (MRP was calculated by assuming net methane concentration for the control as 100%. The MRP of yeast 2 was more than 25%. Compared with the control group, in vitro DM digestibility (IVDMD and total volatile fatty acids (VFA concentration increased (p<0.05 in 0.4 mg/ml YST1 and 0.2 mg/ml YST2 supplementation groups. Proportion of propionate, butyrate and valerate increased (p<0.05, but that of acetate decreased (p<0.05, which led to a decreased (p<0.05 acetate: propionate (A: P ratio in the both YST2 treatments and the 0.4 mg/ml YST 1 supplementation groups. Hydrogen recovery decreased (p<0.05 with yeast supplementation. Quantity of methanogenic archaea per milliliter of inoculum decreased (p<0.05 with yeast supplementation after 24 h of incubation. Our results suggest that live yeast cells suppressed in vitro CH4 production when inoculated into the large intestinal contents of pigs and shifted the fermentation pattern to favor propionate production together with an increased population of acetogenic

Physiology and Genetics of Biogenic Methane-Production from Acetate

Energy Technology Data Exchange (ETDEWEB)

Sowers, Kevin R

2013-04-04

Biomass conversion catalyzed by methanogenic consortia is a widely available, renewable resource for both energy production and waste treatment. The efficiency of this process is directly dependent upon the interaction of three metabolically distinct groups of microorganisms; the fermentative and acetogenic Bacteria and the methanogenic Archaea. One of the rate limiting steps in the degradation of soluble organic matter is the dismutation of acetate, a predominant intermediate in the process, which accounts for 70 % or more of the methane produced by the methanogens. Acetate utilization is controlled by regulation of expression of carbon monoxide dehydrogensase (COdh), which catalyzes the dismutation of acetate. However, physiological and molecular factors that control differential substrate utilization have not been identified in these Archaea. Our laboratory has identified sequence elements near the promoter of the gene (cdh) encoding for COdh and we have confirmed that these sequences have a role in the in vivo expression of cdh. The current proposal focuses on identifying the regulatory components that interact with DNA and RNA elements, and identifying the mechanisms used to control cdh expression. We will determine whether expression is controlled at the level of transcription or if it is mediated by coordinate interaction of transcription initiation with other processes such as transcription elongation rate and differential mRNA stability. Utilizing recently sequenced methanosarcinal genomes and a DNA microarray currently under development genes that encode regulatory proteins and transcription factors will be identified and function confirmed by gene disruption and subsequent screening on different substrates. Functional interactions will be determined in vivo by assaying the effects of gene dosage and site-directed mutagenesis of the regulatory gene on the expression of a cdh::lacZ operon fusion. Results of this study will reveal whether this critical

Hydrocarbon bio-jet fuel from bioconversion of poplar biomass: techno-economic assessment.

Science.gov (United States)

Crawford, Jordan T; Shan, Chin Wei; Budsberg, Erik; Morgan, Hannah; Bura, Renata; Gustafson, Rick

2016-01-01

Infrastructure compatible hydrocarbon biofuel proposed to qualify as renewable transportation fuel under the U.S. Energy Independence and Security Act of 2007 and Renewable Fuel Standard (RFS2) is evaluated. The process uses a hybrid poplar feedstock, which undergoes dilute acid pretreatment and enzymatic hydrolysis. Sugars are fermented to acetic acid, which undergoes conversion to ethyl acetate, ethanol, ethylene, and finally a saturated hydrocarbon end product. An unfermentable lignin stream may be burned for steam and electricity production, or gasified to produce hydrogen. During biofuel production, hydrogen gas is required and may be obtained by various methods including lignin gasification. Both technical and economic aspects of the biorefinery are analyzed, with different hydrogen sources considered including steam reforming of natural gas and gasification of lignin. Cash operating costs for jet fuel production are estimated to range from 0.67 to 0.86 USD L -1 depending on facility capacity. Minimum fuel selling prices with a 15 % discount rate are estimated to range from 1.14 to 1.79 USD L -1 . Capacities of 76, 190, and 380 million liters of jet fuel per year are investigated. Capital investments range from 356 to 1026 million USD. A unique biorefinery is explored to produce a hydrocarbon biofuel with a high yield from bone dry wood of 330 L t -1 . This yield is achieved chiefly due to the use of acetogenic bacteria that do not produce carbon dioxide as a co-product during fermentation. Capital investment is significant in the biorefinery in part because hydrogen is required to produce a fully de-oxygenated fuel. Minimum selling price to achieve reasonable returns on investment is sensitive to capital financing options because of high capital costs. Various strategies, such as producing alternative, intermediate products, are investigated with the intent to reduce risk in building the proposed facility. It appears that producing and selling these

Biomethanation Of Syngas Using Anaerobic Sludge: Shift In The Catabolic Routes With The CO Partial Pressure Increase

Directory of Open Access Journals (Sweden)

Silvia Sancho-Navarro

2016-08-01

Full Text Available Syngas generated by thermal gasification of biomass or coal can be steam reformed and purified into methane, which could be used locally for energy needs, or re-injected in the natural gas grid. As an alternative to chemical catalysis, the main components of the syngas (CO, CO2, and H2 can be used as substrates by a wide range of microorganisms, to be converted into gas biofuels, including methane. This study evaluates the carboxydotrophic (CO-consuming methanogenic potential present in an anaerobic sludge from an upflow anaerobic sludge bed (UASB reactor treating waste water, and elucidates the CO conversion routes to methane at 35±3˚C. Kinetic activity tests under CO at partial pressures (pCO varying from 0.1 to 1.5 atm (0.09-1.31 mmol/L in the liquid phase showed a significant carboxydotrophic activity potential for growing conditions on CO alone. A maximum methanogenic activity of 1 mmol CH4 per g of volatile suspended solid and per day was achieved at 0.2 atm of CO (0.17 mmol/L, and then the rate decreased with the amount of CO supplied. The intermediary metabolites such as acetate, H2 and propionate started to accumulate at higher CO concentrations. Inhibition experiments with 2-bromoethanesulfonic acid (BES, fluoroacetate, and vancomycin showed that in a mixed culture CO was converted mainly to acetate by acetogenic bacteria, which was further transformed to methane by acetoclastic methanogens, while direct methanogenic CO conversion was negligible. Methanogenesis was totally blocked at high pCO in the bottles (≥ 1 atm. However it was possible to achieve higher methanogenic potential under a 100% CO atmosphere after acclimation of the sludge to CO. This adaptation to high CO concentrations led to a shift in the archaeal population, then dominated by hydrogen-utilizing methanogens, which were able to take over acetoclastic methanogens, while syntrophic acetate oxidizing (SAO bacteria oxidized acetate into CO2 and H2. The disaggregation

Biomethanation of Syngas Using Anaerobic Sludge: Shift in the Catabolic Routes with the CO Partial Pressure Increase.

Science.gov (United States)

Sancho Navarro, Silvia; Cimpoia, Ruxandra; Bruant, Guillaume; Guiot, Serge R

2016-01-01

Syngas generated by thermal gasification of biomass or coal can be steam reformed and purified into methane, which could be used locally for energy needs, or re-injected in the natural gas grid. As an alternative to chemical catalysis, the main components of the syngas (CO, CO2, and H2) can be used as substrates by a wide range of microorganisms, to be converted into gas biofuels, including methane. This study evaluates the carboxydotrophic (CO-consuming) methanogenic potential present in an anaerobic sludge from an upflow anaerobic sludge bed (UASB) reactor treating waste water, and elucidates the CO conversion routes to methane at 35 ± 3°C. Kinetic activity tests under CO at partial pressures (pCO) varying from 0.1 to 1.5 atm (0.09-1.31 mmol/L in the liquid phase) showed a significant carboxydotrophic activity potential for growing conditions on CO alone. A maximum methanogenic activity of 1 mmol CH4 per g of volatile suspended solid and per day was achieved at 0.2 atm of CO (0.17 mmol/L), and then the rate decreased with the amount of CO supplied. The intermediary metabolites such as acetate, H2, and propionate started to accumulate at higher CO concentrations. Inhibition experiments with 2-bromoethanesulfonic acid (BES), fluoroacetate, and vancomycin showed that in a mixed culture CO was converted mainly to acetate by acetogenic bacteria, which was further transformed to methane by acetoclastic methanogens, while direct methanogenic CO conversion was negligible. Methanogenesis was totally blocked at high pCO in the bottles (≥1 atm). However it was possible to achieve higher methanogenic potential under a 100% CO atmosphere after acclimation of the sludge to CO. This adaptation to high CO concentrations led to a shift in the archaeal population, then dominated by hydrogen-utilizing methanogens, which were able to take over acetoclastic methanogens, while syntrophic acetate oxidizing (SAO) bacteria oxidized acetate into CO2 and H2. The disaggregation of the

Comparative genomic analysis of single-molecule sequencing and hybrid approaches for finishing the Clostridium autoethanogenum JA1-1 strain DSM 10061 genome

Energy Technology Data Exchange (ETDEWEB)

Brown, Steven D [ORNL; Nagaraju, Shilpa [LanzaTech; Utturkar, Sagar M [ORNL; De Tissera, Sashini [LanzaTech; Segovia, Simón [LanzaTech; Mitchell, Wayne [LanzaTech; Land, Miriam L [ORNL; Dassanayake, Asela [LanzaTech; Köpke, Michael [LanzaTech

2014-01-01

Background Clostridium autoethanogenum strain JA1-1 (DSM 10061) is an acetogen capable of fermenting CO, CO2 and H2 (e.g. from syngas or waste gases) into biofuel ethanol and commodity chemicals such as 2,3-butanediol. A draft genome sequence consisting of 100 contigs has been published. Results A closed, high-quality genome sequence for C. autoethanogenum DSM10061 was generated using only the latest single-molecule DNA sequencing technology and without the need for manual finishing. It is assigned to the most complex genome classification based upon genome features such as repeats, prophage, nine copies of the rRNA gene operons. It has a low G + C content of 31.1%. Illumina, 454, Illumina/454 hybrid assemblies were generated and then compared to the draft and PacBio assemblies using summary statistics, CGAL, QUAST and REAPR bioinformatics tools and comparative genomic approaches. Assemblies based upon shorter read DNA technologies were confounded by the large number repeats and their size, which in the case of the rRNA gene operons were ~5 kb. CRISPR (Clustered Regularly Interspaced Short Paloindromic Repeats) systems among biotechnologically relevant Clostridia were classified and related to plasmid content and prophages. Potential associations between plasmid content and CRISPR systems may have implications for historical industrial scale Acetone-Butanol-Ethanol (ABE) fermentation failures and future large scale bacterial fermentations. While C. autoethanogenum contains an active CRISPR system, no such system is present in the closely related Clostridium ljungdahlii DSM 13528. A common prophage inserted into the Arg-tRNA shared between the strains suggests a common ancestor. However, C. ljungdahlii contains several additional putative prophages and it has more than double the amount of prophage DNA compared to C. autoethanogenum. Other differences include important metabolic genes for central metabolism (as an additional hydrogenase and the absence of a

Low fermentation pH is a trigger to alcohol production, but a killer to chain elongation

Directory of Open Access Journals (Sweden)

Ramon eGanigué

2016-05-01

Full Text Available 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 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 110 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

Heterologous Expression of the Clostridium carboxidivorans CO Dehydrogenase Alone or Together with the Acetyl Coenzyme A Synthase Enables both Reduction of CO2 and Oxidation of CO by Clostridium acetobutylicum.

Science.gov (United States)

Carlson, Ellinor D; Papoutsakis, Eleftherios T

2017-08-15

With recent advances in synthetic biology, CO 2 could be utilized as a carbon feedstock by native or engineered organisms, assuming the availability of electrons. Two key enzymes used in autotrophic CO 2 fixation are the CO dehydrogenase (CODH) and acetyl coenzyme A (acetyl-CoA) synthase (ACS), which form a bifunctional heterotetrameric complex. The CODH/ACS complex can reversibly catalyze CO 2 to CO, effectively enabling a biological water-gas shift reaction at ambient temperatures and pressures. The CODH/ACS complex is part of the Wood-Ljungdahl pathway (WLP) used by acetogens to fix CO 2 , and it has been well characterized in native hosts. So far, only a few recombinant CODH/ACS complexes have been expressed in heterologous hosts, none of which demonstrated in vivo CO 2 reduction. Here, functional expression of the Clostridium carboxidivorans CODH/ACS complex is demonstrated in the solventogen Clostridium acetobutylicum , which was engineered to express CODH alone or together with the ACS. Both strains exhibited CO 2 reduction and CO oxidation activities. The CODH reactions were interrogated using isotopic labeling, thus verifying that CO was a direct product of CO 2 reduction, and vice versa. CODH apparently uses a native C. acetobutylicum ferredoxin as an electron carrier for CO 2 reduction. Heterologous CODH activity depended on actively growing cells and required the addition of nickel, which is inserted into CODH without the need to express the native Ni insertase protein. Increasing CO concentrations in the gas phase inhibited CODH activity and altered the metabolite profile of the CODH-expressing cells. This work provides the foundation for engineering a complete and functional WLP in nonnative host organisms. IMPORTANCE Functional expression of CO dehydrogenase (CODH) from Clostridium carboxidivorans was demonstrated in C. acetobutylicum , which is natively incapable of CO 2 fixation. The expression of CODH, alone or together with the C. carboxidivorans

Microbial community analysis of shallow subsurface samples with PCR-DGGE

Energy Technology Data Exchange (ETDEWEB)

Itaevaara, M.; Suihko, M. -L.; Kapanen, A.; Piskonen, R.; Juvonen, R. [VTT Biotechnology, Espoo (Finland)

2005-11-15

This work is part of the site investigations for the disposal of spent nuclear fuel in Olkiluoto bedrock. The purpose of the research was to study the suitability of PCR-DGGE (polymerase chain reaction - denaturing gradient gel electrophoresis) method for monitoring of hydrogeomicrobiology of Olkiluoto repository site. PCR-DGGE method has been applied for monitoring microbial processes in several applications. The benefit of the method is that microorganisms are not cultivated but the presence of microbial communities can be monitored by direct DNA extractions from the environmental samples. Partial 16SrDNA gene sequence is specifically amplified by PCR (polymerase chain reaction) which detect bacteria as a group. The gene sequences are separated in DGGE, and the nucleotide bands are then cut out, extracted, sequenced and identified by the genelibraries by e.g. Blast program. PCR-DGGE method can be used to detect microorganisms which are present abundantly in the microbial communities because small quantities of genes cannot be separated reliably. However, generally the microorganisms involved in several environmental processes are naturally enriched and present as major population. This makes it possible to utilize PCRDGGE as a monitoring method. In this study, we studied the structure of microbial communities in ten ground water samples originating from Olkiluoto. Two universal bacterial primer sets were compared which amplified two different regions of the 16SrDNA gene. The longer sequence amplified resulted in fewer bands in DGGE, in addition there were problems with purification of the sequences after DGGE. The shorter sequence gave more bands in DGGE and more clear results without any amplification problems. Comparison of the sequences from the gene-libraries resulted in the detection of the same species by both primer sets, in addition some different species were detected. Several species were anaerobic bacteria, such as acetogenic and sulphate reducing

Microbial community analysis of shallow subsurface samples with PCR-DGGE

International Nuclear Information System (INIS)

Itaevaara, M.; Suihko, M.-L.; Kapanen, A.; Piskonen, R.; Juvonen, R.

2005-11-01

This work is part of the site investigations for the disposal of spent nuclear fuel in Olkiluoto bedrock. The purpose of the research was to study the suitability of PCR-DGGE (polymerase chain reaction - denaturing gradient gel electrophoresis) method for monitoring of hydrogeomicrobiology of Olkiluoto repository site. PCR-DGGE method has been applied for monitoring microbial processes in several applications. The benefit of the method is that microorganisms are not cultivated but the presence of microbial communities can be monitored by direct DNA extractions from the environmental samples. Partial 16SrDNA gene sequence is specifically amplified by PCR (polymerase chain reaction) which detect bacteria as a group. The gene sequences are separated in DGGE, and the nucleotide bands are then cut out, extracted, sequenced and identified by the genelibraries by e.g. Blast program. PCR-DGGE method can be used to detect microorganisms which are present abundantly in the microbial communities because small quantities of genes cannot be separated reliably. However, generally the microorganisms involved in several environmental processes are naturally enriched and present as major population. This makes it possible to utilize PCRDGGE as a monitoring method. In this study, we studied the structure of microbial communities in ten ground water samples originating from Olkiluoto. Two universal bacterial primer sets were compared which amplified two different regions of the 16SrDNA gene. The longer sequence amplified resulted in fewer bands in DGGE, in addition there were problems with purification of the sequences after DGGE. The shorter sequence gave more bands in DGGE and more clear results without any amplification problems. Comparison of the sequences from the gene-libraries resulted in the detection of the same species by both primer sets, in addition some different species were detected. Several species were anaerobic bacteria, such as acetogenic and sulphate reducing

The stable carbon isotope biogeochemistry of acetate and other dissolved carbon species in deep subseafloor sediments at the northern Cascadia Margin

Science.gov (United States)

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

Geodynamic and metabolic cycles in the Hadean

Directory of Open Access Journals (Sweden)

M. J. Russell

2005-01-01

Full Text Available High-degree melting of hot dry Hadean mantle at ocean ridges and plumes resulted in a crust about 30km thick, overlain in places by extensive and thick mafic volcanic plateaus. Continental crust, by contrast, was relatively thin and mostly submarine. At constructive and destructive plate boundaries, and above the many mantle plumes, acidic hydrothermal springs at ~400°C contributed Fe and other transition elements as well as P and H2 to the deep ocean made acidulous by dissolved CO2 and minor HCl derived from volcanoes. Away from ocean ridges, submarine hydrothermal fluids were cool (≤100°C, alkaline (pH ~10, highly reduced and also H2-rich. Reaction of solvents in this fluid with those in ocean water was catalyzed in a hydrothermal mound, a natural self-restoring flow reactor and fractionation column developed above the alkaline spring. The mound consisted of brucite, Mg-rich clays, ephemeral carbonates, Fe-Ni sulfide and green rust. Acetate and glycine were the main products, some of which were eluted to the ocean. The rest, along with other organic byproducts were retained and concentrated within Fe-Ni sulfide compartments. These compartments, comprising the natural hydrothermal reactor, consisted partly of greigite (Fe5NiS8. It was from reactions between organic modules confined within these inorganic compartments that the first prokaryotic organism evolved. These acetogenic precursors to the bacteria diversified and migrated down the mound and into the ocean floor to inaugurate the 'deep biosphere'. Once there they were protected from cataclysmic heating events caused by large meteoritic impacts. Geodynamic forces led to the eventual obduction of the deep biosphere into the photic zone where, initially protected by a thin veneer of sediment, the use of solar energy was mastered and photosynthesis emerged. The further evolution to oxygenic photosynthesis was effected as catalytic [Mn,Ca]-bearing molecules that otherwise would have been

Analysis of trickle-bed reactor for ethanol production from syngas using Clostridium ragsdalei

Science.gov (United States)

Devarapalli, Mamatha

The conversion of syngas components (CO, CO2 and H2) to liquid fuels such as ethanol involves complex biochemical reactions catalyzed by a group of acetogens such as Clostridium ljungdahlii, Clostridium carboxidivorans and Clostridium ragsdalei. The low ethanol productivity in this process is associated with the low solubility of gaseous substrates CO and H2 in the fermentation medium. In the present study, a 1-L trickle-bed reactor (TBR) was analyzed to understand its capabilities to improve the mass transfer of syngas in fermentation medium. Further, semi-continuous and continuous syngas fermentations were performed using C. ragsdalei to evaluate the ability of the TBR for ethanol production. In the mass transfer studies, using 6-mm glass beads, it was found that the overall mass transfer coefficient (kLa/V L) increased with the increase in gas flow rate from 5.5 to 130.5 sccm. Further, an increase in the liquid flow rate in the TBR decreased the kLa/VL due to the increase in liquid hold up volume (VL) in the packing. The highest kLa/VL values of 421 h-1 and 178 h-1 were achieved at a gas flow rate of 130.5 sccm for 6-mm and 3-mm glass beads, respectively. Semi-continuous fermentations were performed with repetitive medium replacement in counter-current and co-current modes. In semi-continuous fermentations with syngas consisting of 38% CO, 5% N2, 28.5% CO2 and 28.5% H2 (by volume), the increase in H2 conversion (from 18 to 55%) and uptake (from 0.7 to 2.2 mmol/h) were observed. This increase was attributed to more cell attachment in the packing that reduced CO inhibition to hydrogenase along the column length and increased the H2 uptake. The maximum ethanol produced during counter-current and co-current modes were 3.0 g/L and 5.7 g/L, respectively. In continuous syngas fermentation, the TBR was operated at dilution rates between 0.006 h-1and 0.012 h -1 and gas flow rates between 1.5 sccm and 18.9 sccm. The highest ethanol concentration of 13 g/L was achieved at

Microbiology of Olkiluoto and ONKALO groundwater results and interpretations, 2008-2009

Energy Technology Data Exchange (ETDEWEB)

Pedersen, K.; Arlinger, J.; Edlund, J.; Eriksson, L.; Lydmark, S.; Johansson, J.; Jaegevall, S.; Rabe, L. (Microbial Analytics Sweden AB, Moelnlycke (Sweden))

2010-08-15

Microbiology cultivation, DNA, and RNA data were assembled from 18 groundwater samples from Olkiluoto, from deep drillholes ranging in depth from 62 to 708 m, and from groundwater from eight ONKALO drillholes ranging in depth from 7.1 to 318 m. Biomass was determined by counting total numbers of microbial cells (TNC) and determining adenosine triphosphate (ATP) concentrations. The aerobic cultivation method used comprised aerobic plate counts. Anaerobic most probable number (MPN) methods were used to determine counts of nitrate-, iron-, manganese-, and sulphatereducing bacteria, acetogenic bacteria, and methanogens. Molecular methods for analysing the diversity and abundance of microorganisms have been continuously developed and applied to groundwater samples. These methods included the sampling of DNA and RNA, extraction of nucleic acids, cloning and sequencing of environmental nucleic acids, and real-time quantitative polymerase chain reaction (qPCR) for analysing amounts of DNA and RNA. The results of these analyses have been merged and interpreted, and the outcomes are reported here. The four methods for biomassrelated analysis correlated well. These methods focus on different characteristics of microbial cells: TNC analyses whole cells using a microscope, ATP analyses a cell component using a biochemical method, MPN is based on cultivation and qPCR analyses DNA (genes) and RNA (gene expression). The range of analytical focus encompassed by the methods ensures that the biomass-related information in this and previous reports from Olkiluoto and ONKALO is reliable and reflects a diverse range of the biomass-related characteristics of the analysed microorganisms. The distribution of the MPN data over depth from 2008 to 2009 followed the distribution found earlier. There were generally more cultivable microorganisms between depths of 200 and 400 m than in the shallower 50-200-m depth range. These new results agree with previous results, suggesting that

Advanced modelling, monitoring, and process control of bioconversion systems

Science.gov (United States)

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

Miniature Canister (MiniCan) Corrosion Experiment Progress Report 3 for 2008-2010

Energy Technology Data Exchange (ETDEWEB)

Smart, N.R.; Reddy, B.; Rance, A.P. (Serco (United Kingdom))

2011-08-15

obtained in 2007 and 2008, including gas composition and microbial activity. These data show an increase in the dissolved iron concentration inside the support cage, together with a decrease in the pH. Both these observations may be due to microbial activity, but it is also possible that abiotic processes could have contributed to changes in the local chemistry. Microbial analysis has shown the presence of sulphate reducing bacteria (SRBs) and autotrophic acetogens. The presence of low density bentonite appears to stimulate the growth of SRBs, but recent studies reported in the literature indicate that high density fully compacted bentonite inhibits SRB activity. A further set of water samples was taken for analysis at the end of 2010, but the full set of results from this sampling campaign is not currently available and will be included in a subsequent report

Microbiology of Olkiluoto and ONKALO groundwater results and interpretations, 2008-2009

International Nuclear Information System (INIS)

Pedersen, K.; Arlinger, J.; Edlund, J.; Eriksson, L.; Lydmark, S.; Johansson, J.; Jaegevall, S.; Rabe, L.

2010-08-01

Microbiology cultivation, DNA, and RNA data were assembled from 18 groundwater samples from Olkiluoto, from deep drillholes ranging in depth from 62 to 708 m, and from groundwater from eight ONKALO drillholes ranging in depth from 7.1 to 318 m. Biomass was determined by counting total numbers of microbial cells (TNC) and determining adenosine triphosphate (ATP) concentrations. The aerobic cultivation method used comprised aerobic plate counts. Anaerobic most probable number (MPN) methods were used to determine counts of nitrate-, iron-, manganese-, and sulphatereducing bacteria, acetogenic bacteria, and methanogens. Molecular methods for analysing the diversity and abundance of microorganisms have been continuously developed and applied to groundwater samples. These methods included the sampling of DNA and RNA, extraction of nucleic acids, cloning and sequencing of environmental nucleic acids, and real-time quantitative polymerase chain reaction (qPCR) for analysing amounts of DNA and RNA. The results of these analyses have been merged and interpreted, and the outcomes are reported here. The four methods for biomassrelated analysis correlated well. These methods focus on different characteristics of microbial cells: TNC analyses whole cells using a microscope, ATP analyses a cell component using a biochemical method, MPN is based on cultivation and qPCR analyses DNA (genes) and RNA (gene expression). The range of analytical focus encompassed by the methods ensures that the biomass-related information in this and previous reports from Olkiluoto and ONKALO is reliable and reflects a diverse range of the biomass-related characteristics of the analysed microorganisms. The distribution of the MPN data over depth from 2008 to 2009 followed the distribution found earlier. There were generally more cultivable microorganisms between depths of 200 and 400 m than in the shallower 50-200-m depth range. These new results agree with previous results, suggesting that

PA-1, a Versatile Anaerobe Obtained in Pure Culture, Catabolizes Benzenoids and Other Compounds in Syntrophy with Hydrogenotrophs, and P-2 plus Wolinella sp. Degrades Benzenoids

Science.gov (United States)

Barik, Sudhakar; Brulla, W. J.; Bryant, M. P.

1985-01-01

Methanogenic enrichments catabolizing 13 mM phenylacetate or 4 mM phenol were established at 37°C, using a 10% inoculum from a municipal anaerobic digester. By using agar roll tubes of the basal medium plus 0.1% yeast extract-25 mM fumarate, a hydrogenotrophic lawn of Wolinella succinogenes and phenol or phenylacetate, strains P-2 and PA-1, respectively, were isolated in coculture with W. succinogenes. With the lawn deleted, PA-1 was isolated in pure culture. Strain P-2 is apparently a new species of anaerobic, motile, gram-negative, spindle-shaped, small rod that as yet has been grown only in coculture with W. succinogenes. It used phenol, hydrocinnamate, benzoate, and phenylacetate as energy sources. Product recovery by the coculture, per mole of phenol and 4.4 mol of fumarate used, included 2.03, 0.12, 0.08, and 3.23 mol, respectively, of acetate, propionate, butyrate, and succinate. Carbon recovery was 75% and H recovery was 80%, although CO2 and a few other possible products were not determined. That P-2 is an obligate proton-reducing acetogen and possible pathways for its degradation of phenol are discussed. Strain PA-1 is apparently a new species of anaerobic, motile, relatively small, gram-negative rod. It utilized compounds such as phenylacetate, hydrocinnamate, benzoate, phenol, resorcinol, gallate, 4-aminophenol, 2-aminobenzoate, pyruvate, Casamino Acids, and aspartate as energy sources in coculture with W. succinogenes. Per mole of phenylacetate and 1.44 mol of fumarate used, 1.04, 0.53, and 0.78 mol of acetate, propionate, and succinate, respectively, were recovered from the coculture. Only about 50% of the carbon and H were recovered. In coculture with Methanospirillum hungatei, 0.96 mol of acetate and 0.25 mol of methane were recovered per mol of pyruvate used; 0.90 mol of acetate and 0.33 mol of methane, per mol of fumarate used; 0.93 mol of acetate and 0.54 mol of methane, per mol of aspartate used; and 1.71 mol of acetate and 0.57 mol of methane

Energy alternative for the use of organic waste (swine feces) in the state of Tabasco; Alternativa energetica para el aprovechamiento de los desechos organicos (excretas de ganado porcino) en el estado de Tabasco

Energy Technology Data Exchange (ETDEWEB)

Ramon Garcia, E.; Angles Falconi, S.I.; Vazquez Magana, L.R.; Tun Perez, A.C. [Instituto Tecnologico de Villahermosa, Villahermosa, Tabasco (Mexico)]. Email: bioleliza_ramgar@hotmail.com; sergio_falcorp@hotmail.com

2009-09-15

Biogas is obtained from decomposing organic material due to the action of four types of bacteria, in the absence of oxygen: hydrolytic, producers of moncarbonate compounds and acidogenics, producing organic fatty acids and other polycarbonate compounds; acetogenic, producers of hydrogen and CO{sub 2}; homoacetogenic, that can convert a considerable amount of multicarbonate or monocarbonate compounds into acetic acid and; metanogenic, producers of methane gas, the principal component of biogas. To design and build the pilot equipment proposed for this research, a 20-liter container was used with 10 kg of swine feces with identical volumes of inoculates. The aerobic fermentation process was immediate begun, followed by the anaerobic process; metabolic reactions were conducted. A total of 0.119 m3 of biogas was obtained from the anaerobic fermentation of 10 kg of swine feces, 60% (0.0714 m{sup 3}) of which was methane gas and the remaining 40%(0.0476 m{sup 3}) hydrogen sulfide (HS). During this investigation, it was concluded that obtaining this by-product (methane biogas), which can be used as a biofuel, provides an economically and ecologically sustainable alternative that reduces organic discharges (swine feces) into rivers and reduces the energy expenditures of swine farms. Because of the high generation of RSM, the creation of a biogas production plant in Tabasco is proposed. [Spanish] El biogas se obtiene al descomponerse la materia organica debido a la accion de cuatro tipos de bacterias, en ausencia de oxigeno: las hidroliticas, que producen compuestos monocarbonados, las acidogenicas producen acidos grasos organicos y otros compuestos policarbonados; las acetogenicas productoras de Hidrogeno y CO{sub 2}; las homoacetogenicas, que pueden convertir una cantidad considerable de compuestos multicarbonados o monocarbonados en acido acetico; y las metanogenicas, productoras del gas metano, principal componente del biogas. Para el diseno y construccion del equipo

Miniature Canister (MiniCan) Corrosion experiment progress report 4 for 2008-2011

International Nuclear Information System (INIS)

Smart, Nick; Reddy, Bharti; Rance, Andy

2012-06-01

2007, 2008 and 2010 including gas composition and microbial activity. These data show an increase in the dissolved iron concentration inside the support cage, together with a decrease in the pH. Both these observations appear to be mainly due to microbial activity, but it is also possible that abiotic processes could have contributed to changes in the local chemistry. Microbial analysis has shown the presence of sulphate-reducing bacteria (SRB) and autotrophic acetogens. The presence of low density bentonite appears to stimulate the growth of SRB, but recent studies reported in the literature indicate that high density fully compacted bentonite inhibits SRB activity. A further set of water samples was taken for analysis at the end of 2010 and these results are also included in this report. These data show a similar trend as before, with increases in the dissolved iron concentration inside the support cage and a decrease in the pH. Further water analysis on Experiment 3 was carried out before it was removed for analysis, but the results from this analysis are presented in a separate report

Impact on the deep biosphere of CO2 geological sequestration in (ultra)mafic rocks and retroactive consequences on its fate

Science.gov (United States)

Ménez, Bénédicte; Gérard, Emmanuelle; Rommevaux-Jestin, Céline; Dupraz, Sébastien; Guyot, François; Arnar Alfreősson, Helgi; Reynir Gíslason, Sigurőur; Sigurőardóttir, Hólmfríiur

2010-05-01

amplification of small subunit ribosomal RNA genes (SSU rDNAs). The stratigraphic levels targeted to store the injected CO2 as aqueous phase harbor numerous new species close to cultivable species belonging to the genus Thermus or Proteobacteria species known to be linked in particular with the hydrogen and iron cycles. After injection, the evolution of these microbial communities will be monitored using the Denaturing Gradient Gel Electrophoresis technique. Beyond the ecological impact of storing high levels of CO2 in deep environments, particularly important is the ability of intraterrestrial microbes to potentially interact with the injected fluids. For example, carbonation has been shown to be strongly influenced by microbiological activities that can locally modify pH and induce nucleation of solid carbonates. To improve the understanding of these processes and to better constrain the influence of deep biota on the evolving chemical and petrophysical properties of the reservoir, an experimental and numerical modeling is carried out in parallel, using model strains representative of the subsurface (including acetogens, sulphate and iron reducing bacteria), as single-species or consortia. A set of batch experiments in presence of crushed olivine or basalts was especially designed to evaluate how microbial activity could overcome the slow kinetics of mineral-fluid reactions and reduce the energy needed to hasten the carbonation process.

Miniature Canister (MiniCan) Corrosion experiment progress report 4 for 2008-2011

Energy Technology Data Exchange (ETDEWEB)

Smart, Nick; Reddy, Bharti; Rance, Andy [Serco, Hook (United Kingdom)

2012-06-15

obtained in 2007, 2008 and 2010 including gas composition and microbial activity. These data show an increase in the dissolved iron concentration inside the support cage, together with a decrease in the pH. Both these observations appear to be mainly due to microbial activity, but it is also possible that abiotic processes could have contributed to changes in the local chemistry. Microbial analysis has shown the presence of sulphate-reducing bacteria (SRB) and autotrophic acetogens. The presence of low density bentonite appears to stimulate the growth of SRB, but recent studies reported in the literature indicate that high density fully compacted bentonite inhibits SRB activity. A further set of water samples was taken for analysis at the end of 2010 and these results are also included in this report. These data show a similar trend as before, with increases in the dissolved iron concentration inside the support cage and a decrease in the pH. Further water analysis on Experiment 3 was carried out before it was removed for analysis, but the results from this analysis are presented in a separate report.

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

Science.gov (United States)

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

2013-12-01

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

Biogeochemical processes in a clay formation in situ experiment: Part A - Overview, experimental design and water data of an experiment in the Opalinus Clay at the Mont Terri Underground Research Laboratory, Switzerland

International Nuclear Information System (INIS)

Wersin, P.; Leupin, O.X.; Mettler, S.; Gaucher, E.C.; Maeder, U.; De Canniere, P.; Vinsot, A.; Gaebler, H.E.; Kunimaro, T.; Kiho, K.; Eichinger, L.

2011-01-01

Highlights: → The composition was affected by the complex interplay of diffusion, mineral and surface reactions. → The 13 C signals for carbon species showed significant variations which could only be partly explained. → The main cations remained remarkably constant during the experiment. → This underlines the strong buffering via cation exchange and carbonate dissolution/precipitation. - Abstract: An in situ test in the Opalinus Clay formation, termed porewater chemistry (PC) experiment, was carried out for a period of 5 years. It was based on the concept of diffusive equilibration whereby a traced water with a composition close to that expected in the formation was continuously circulated and monitored in a packed-off borehole. The main original focus was to obtain reliable data on the pH/pCO 2 conditions of the porewater, but because of unexpected microbiologically-induced redox reactions, the objective was extended to elucidate the biogeochemical processes occurring in the borehole and to understand their impact on pH/pCO 2 and porewater chemistry in the low permeability clay formation. The behaviour of the conservative tracers 2 H and Br - could be explained by diffusive dilution in the clay and moreover the results showed that diffusive equilibration between the borehole water and the formation occurred within about 3 year's time. However, the composition and pH/pCO 2 conditions differed considerably from those of the in situ porewater. Thus, pH was lower and pCO 2 was higher than indicated by complementary laboratory investigations. The noted differences are explained by microbiologically-induced redox reactions occurring in the borehole and in the interfacial wall area which were caused by an organic source released from the equipment material. The degradation of this source was accompanied by sulfate reduction and - to a lesser extent - by methane generation, which induced a high rate of acetogenic reactions corresponding to very high acetate

Biogeochemical processes in a clay formation in situ experiment: Part A - Overview, experimental design and water data of an experiment in the Opalinus Clay at the Mont Terri Underground Research Laboratory, Switzerland

Energy Technology Data Exchange (ETDEWEB)

Wersin, P., E-mail: paul.wersin@gruner.ch [NAGRA, Hardstrasse 73, 5430 Wettingen (Switzerland)] [Gruner Ltd., Gellertstrasse 55, 4020 Basel (Switzerland); Leupin, O.X. [NAGRA, Hardstrasse 73, 5430 Wettingen (Switzerland); Mettler, S. [NAGRA, Hardstrasse 73, 5430 Wettingen (Switzerland)] [Solexperts Ltd., Mettlenbachstrasse 25, 8617 Moenchaltorf (Switzerland); Gaucher, E.C. [BRGM, 3 avenue Claude Guillemin, B.P. 36009, 45060 Orleans Cedex 2 (France); Maeder, U. [University of Bern, Institute of Geological Sciences, Baltzerstrasse 3, CH-3012 Bern (Switzerland); De Canniere, P. [SCK.CEN, Waste and Disposal Project, Boeretang 200, 2400 Mol (Belgium); Vinsot, A. [ANDRA, Laboratoire de Recherche Souterrain de Meuse/Haute-Marne, RD960 BP9, 55290 Bure (France); Gaebler, H.E. [BGR, Stilleweg 2, 30655 Hannover (Germany); Kunimaro, T. [JAEA, Tokai-mura, Naka-gun, Ibaraki 319-1195 (Japan); Kiho, K. [CRIEPI, 1646 Abiko, Abiko-city Chiba 270-1194 (Japan); Eichinger, L. [Hydroisotop, 85301 Schweitenkirchen (Germany)

2011-06-15

Highlights: > The composition was affected by the complex interplay of diffusion, mineral and surface reactions. > The {sup 13}C signals for carbon species showed significant variations which could only be partly explained. > The main cations remained remarkably constant during the experiment. > This underlines the strong buffering via cation exchange and carbonate dissolution/precipitation. - Abstract: An in situ test in the Opalinus Clay formation, termed porewater chemistry (PC) experiment, was carried out for a period of 5 years. It was based on the concept of diffusive equilibration whereby a traced water with a composition close to that expected in the formation was continuously circulated and monitored in a packed-off borehole. The main original focus was to obtain reliable data on the pH/pCO{sub 2} conditions of the porewater, but because of unexpected microbiologically-induced redox reactions, the objective was extended to elucidate the biogeochemical processes occurring in the borehole and to understand their impact on pH/pCO{sub 2} and porewater chemistry in the low permeability clay formation. The behaviour of the conservative tracers {sup 2}H and Br{sup -} could be explained by diffusive dilution in the clay and moreover the results showed that diffusive equilibration between the borehole water and the formation occurred within about 3 year's time. However, the composition and pH/pCO{sub 2} conditions differed considerably from those of the in situ porewater. Thus, pH was lower and pCO{sub 2} was higher than indicated by complementary laboratory investigations. The noted differences are explained by microbiologically-induced redox reactions occurring in the borehole and in the interfacial wall area which were caused by an organic source released from the equipment material. The degradation of this source was accompanied by sulfate reduction and - to a lesser extent - by methane generation, which induced a high rate of acetogenic reactions

Aespoe Hard Rock Laboratory. Prototype Repository. Analyses of microorganisms, gases and water chemistry in buffer and backfill, 2009

Energy Technology Data Exchange (ETDEWEB)

Lydmark, Sara (Microbial Analytics Sweden AB (Sweden))

2010-09-15

The Prototype repository is an international project to build and study a full-scale model of the planned Swedish final repository for spent nuclear fuel. The Prototype repository differs from a real storage in that it is drained. For example, this makes the swelling pressure lower in the Prototype repository compared with a real storage. The project is being conducted at the Aespoe Hard Rock Laboratory (HRL) in crystalline rock at a depth of approximately 450 m. A monitoring programme is investigating the evolution of the water chemistry, gas, and microbial activity at the site, and one of the specific aims is to monitor the microbial consumption of oxygen in situ in the Prototype repository. This document describes the results of the analyses of microbes, gases, and chemistry inside and outside the Prototype in 2009. Hydrogen, helium, nitrogen, oxygen, carbon monoxide, carbon dioxide, methane, ethane, and ethene were analysed in the following sampling points in the Prototype repository: KBU10001, KBU10002, KBU10004, KBU10006, KBU10008, KFA01 and KFA04. Where the sampling points in the Prototype delivered pore water, the water was analysed for amount of ATP (i.e., the biovolume), cultivable heterotrophic aerobic bacteria (CHAB), sulphate-reducing bacteria (SRB), methane-oxidizing bacteria (MOB), autotrophic acetogens (AA) and in some cases iron-reducing bacteria (IRB). Cultivation methods were also compared with qPCR molecular techniques to evaluate these before next year's decommission of the Prototype repository. The collected pore water from the Prototype repository was subject to chemistry analysis (as many analyses were conducted as the amount of water allowed). In addition, groundwater from two borehole sections in the rock surrounding the Prototype was analysed regarding its gas composition, microbiology and redox. Chemistry data from a previous investigation of the groundwater outside the Prototype repository were compared with the pore water

Aespoe Hard Rock Laboratory. Prototype Repository. Analyses of microorganisms, gases and water chemistry in buffer and backfill, 2009

International Nuclear Information System (INIS)

Lydmark, Sara

2010-09-01

The Prototype repository is an international project to build and study a full-scale model of the planned Swedish final repository for spent nuclear fuel. The Prototype repository differs from a real storage in that it is drained. For example, this makes the swelling pressure lower in the Prototype repository compared with a real storage. The project is being conducted at the Aespoe Hard Rock Laboratory (HRL) in crystalline rock at a depth of approximately 450 m. A monitoring programme is investigating the evolution of the water chemistry, gas, and microbial activity at the site, and one of the specific aims is to monitor the microbial consumption of oxygen in situ in the Prototype repository. This document describes the results of the analyses of microbes, gases, and chemistry inside and outside the Prototype in 2009. Hydrogen, helium, nitrogen, oxygen, carbon monoxide, carbon dioxide, methane, ethane, and ethene were analysed in the following sampling points in the Prototype repository: KBU10001, KBU10002, KBU10004, KBU10006, KBU10008, KFA01 and KFA04. Where the sampling points in the Prototype delivered pore water, the water was analysed for amount of ATP (i.e., the biovolume), cultivable heterotrophic aerobic bacteria (CHAB), sulphate-reducing bacteria (SRB), methane-oxidizing bacteria (MOB), autotrophic acetogens (AA) and in some cases iron-reducing bacteria (IRB). Cultivation methods were also compared with qPCR molecular techniques to evaluate these before next year's decommission of the Prototype repository. The collected pore water from the Prototype repository was subject to chemistry analysis (as many analyses were conducted as the amount of water allowed). In addition, groundwater from two borehole sections in the rock surrounding the Prototype was analysed regarding its gas composition, microbiology and redox. Chemistry data from a previous investigation of the groundwater outside the Prototype repository were compared with the pore water chemistry