Interactions in Syntrophic Associations of Endospore-Forming, Butyrate-Degrading Bacteria and H2-Consuming Bacteria

Butyrate is an important intermediate in the anaerobic degradation of organic matter. In sulfate-depleted environments butyrate is oxidized to acetate and hydrogen by obligate proton reducers, in syntrophic association with hydrogen-consuming methanogens. This paper describes two enrichments of endospore-forming bacteria degrading butyrate in consortia with methanogens. The isolates are readily established in coculture with H₂-consuming, sulfate-reducing bacteria by pasteurizing the culture. The two original enrichments differed in that one grew to an optically dense culture while the second grew in clumps. Examination by scanning electron microscopy showed that clumping resulted from the production of large amounts of extracellular polymer. Several H₂-consuming methanogens were identified in the enrichments. Some of them grew closely associated to the butyrate degraders. This attachment to the hydrogen producer may permit some methanogens to compete for the growth substrate against other bacteria having higher substrate affinity.     

Isolation and partial characterization of bacteria in an anaerobic consortium that mineralizes 3-chlorobenzoic Acid

A methanogenic consortium able to use 3-chlorobenzoic acid as its sole energy and carbon source was enriched from anaerobic sewage sludge. Seven bacteria were isolated from the consortium in mono- or coculture. They included: one dechlorinating bacterium (strain DCB-1), one benzoate-oxidizing bacterium (strain BZ-2), two butyrate-oxidizing bacteria (strains SF-1 and NSF-2), two H(2)-consuming methanogens (Methanospirillum hungatei PM-1 and Methanobacterium sp. strain PM-2), and a sulfate-reducing bacterium (Desulfovibrio sp. strain PS-1). The dechlorinating bacterium (DCB-1) was a gram-negative, obligate anaerobe with a unique "collar" surrounding the cell. A medium containing rumen fluid supported minimal growth; pyruvate was the only substrate found to increase growth. The bacterium had a generation time of 4 to 5 days. 3-Chlorobenzoate was dechlorinated stoichiometrically to benzoate, which accumulated in the medium; the rate of dechlorination was ca. 0.1 pmol bacterium day. The benzoate-oxidizing bacterium (BZ-2) was a gram-negative, obligate anaerobe and could only be grown as a syntroph. Benzoate was the only substrate observed to support growth, and, when grown in coculture with M. hungatei, it was fermented to acetate and CH(4). One butyrate-oxidizing bacterium (NSF-2) was a gram-negative, non-sporeforming, obligate anaerobe; the other (SF-1) was a gram-positive, sporeforming, obligate anaerobe. Both could only be grown as syntrophs. The substrates observed to support growth of both bacteria were butyrate, 2-dl-methylbutyrate, valerate, and caproate; isobutyrate supported growth of only the sporeforming bacterium (SF-1). Fermentation products were acetate and CH(4) (from butyrate, isobutyrate, or caproate) or acetate, propionate, and CH(4) (from 2-dl-methylbutyrate or valerate) when grown in coculture with M. hungatei. A mutualism among at least the dechlorinating, benzoate-oxidizing, and methane-forming members was apparently required for utilization of the 3-chlorobenzoate substrate.     

Methanogenic bacteria from human dental plaque.

Samples of human dental plaque were examined for the presence of methanogenic bacteria. Of 54 samples from 36 patients, 20 yielded H2/CO2-using methanogenic enrichment cultures. All methanogen-positive samples were from patients with some degree of periodontal disease. The predominant populations in the enrichments had morphologies characteristic of Methanobrevibacter spp. In six enrichments derived from three patients, the common methanogen was antigenically similar to Methanobrevibacter smithii. The same was true for the three methanogenic isolates obtained in axenic culture from a fourth patient. The six enrichments and two of the three isolates were antigenically closer to strain ALI than to PS. Two of the enrichments also had subpopulations with weak antigenic similarity to Methanosphaera stadtmanae. The data indicate that methanogens in the oral cavity of humans are antigenically close to those found in the intestinal tract.     

Hydrogenotrophic methanogenesis by moderately acid-tolerant methanogens of a methane-emitting acidic peat

The emission of methane (1.3 mmol of CH(4) m(-2) day(-1)), precursors of methanogenesis, and the methanogenic microorganisms of acidic bog peat (pH 4.4) from a moderately reduced forest site were investigated by in situ measurements, microcosm incubations, and cultivation methods, respectively. Bog peat produced CH(4) (0.4 to 1.7 micro mol g [dry wt] of soil(-1) day(-1)) under anoxic conditions. At in situ pH, supplemental H(2)-CO(2), ethanol, and 1-propanol all increased CH(4) production rates while formate, acetate, propionate, and butyrate inhibited the production of CH(4); methanol had no effect. H(2)-dependent acetogenesis occurred in H(2)-CO(2)-supplemented bog peat only after extended incubation periods. Nonsupplemented bog peat initially produced small amounts of H(2) that were subsequently consumed. The accumulation of H(2) was stimulated by ethanol and 1-propanol or by inhibiting methanogenesis with bromoethanesulfonate, and the consumption of ethanol was inhibited by large amounts of H(2); these results collectively indicated that ethanol- or 1-propanol-utilizing bacteria were trophically associated with H(2)-utilizing methanogens. A total of 10(9) anaerobes and 10(7) hydrogenotrophic methanogens per g (dry weight) of bog peat were enumerated by cultivation techniques. A stable methanogenic enrichment was obtained with an acidic, H(2)-CO(2)-supplemented, fatty acid-enriched defined medium. CH(4) production rates by the enrichment were similar at pH 4.5 and 6.5, and acetate inhibited methanogenesis at pH 4.5 but not at pH 6.5. A total of 27 different archaeal 16S rRNA gene sequences indicative of Methanobacteriaceae, Methanomicrobiales, and Methanosarcinaceae were retrieved from the highest CH(4)-positive serial dilutions of bog peat and methanogenic enrichments. A total of 10 bacterial 16S rRNA gene sequences were also retrieved from the same dilutions and enrichments and were indicative of bacteria that might be responsible for the production of H(2) that could be used by hydrogenotrophic methanogens. These results indicated that in this acidic bog peat, (i) H(2) is an important substrate for acid-tolerant methanogens, (ii) interspecies hydrogen transfer is involved in the degradation of organic carbon, (iii) the accumulation of protonated volatile fatty acids inhibits methanogenesis, and (iv) methanogenesis might be due to the activities of methanogens that are phylogenetic members of the Methanobacteriaceae, Methanomicrobiales, and Methanosarcinaceae.     

Influence of hydrogen-consuming bacteria on cellulose degradation by anaerobic fungi

The presence of methanogens Methanobacterium arboriphilus, Methanobacterium bryantii, or Methanobrevibacter smithii increased the level of cellulose fermentation by 5 to 10% in cultures of several genera of anaerobic fungi. When Neocallimastix sp. strain L2 was grown in coculture with methanogens the rate of cellulose fermentation also increased relative to that for pure cultures of the fungus. Methanogens caused a shift in the fermentation products to more acetate and less lactate, succinate, and ethanol. Formate transfer in cocultures of anaerobic fungi and M. smithii did not result in further stimulation of cellulolysis above the level caused by H2 transfer. When Selenomonas ruminatium was used as a H2-consuming organism in coculture with Neocallimastix sp. strain L2, both the rate and level of cellulolysis increased. The observed influence of the presence of methanogens is interpreted to indicate a shift of electrons from the formation of electron sink carbon products to H2 via reduced pyridine nucleotides, favoring the production of additional acetate and probably ATP. It is not known how S. ruminantium exerts its influence. It might result from a lowered production of electron sink products by the fungus, from consumption of electron sink products or H2 by S. ruminantium, or from competition for free sugars which in pure culture could exert an inhibiting effect on cellulolysis.     

Influence of hydrogen-consuming bacteria on cellulose degradation by anaerobic fungi.

The presence of methanogens Methanobacterium arboriphilus, Methanobacterium bryantii, or Methanobrevibacter smithii increased the level of cellulose fermentation by 5 to 10% in cultures of several genera of anaerobic fungi. When Neocallimastix sp. strain L2 was grown in coculture with methanogens the rate of cellulose fermentation also increased relative to that for pure cultures of the fungus. Methanogens caused a shift in the fermentation products to more acetate and less lactate, succinate, and ethanol. Formate transfer in cocultures of anaerobic fungi and M. smithii did not result in further stimulation of cellulolysis above the level caused by H2 transfer. When Selenomonas ruminatium was used as a H2-consuming organism in coculture with Neocallimastix sp. strain L2, both the rate and level of cellulolysis increased. The observed influence of the presence of methanogens is interpreted to indicate a shift of electrons from the formation of electron sink carbon products to H2 via reduced pyridine nucleotides, favoring the production of additional acetate and probably ATP. It is not known how S. ruminantium exerts its influence. It might result from a lowered production of electron sink products by the fungus, from consumption of electron sink products or H2 by S. ruminantium, or from competition for free sugars which in pure culture could exert an inhibiting effect on cellulolysis.     

Interaction between methanogenic and sulfate-reducing microorganisms during dechlorination of a high concentration of tetrachloroethylene

A methanogenic and sulfate-reducing consortium, which was enriched on medium containing tetrachloroethylene (PCE), had the ability to dechlorinate high concentrations of PCE. Dehalogenation was due to the direct activity of methanogens. However, interactions between methanogenic and sulfate-reducing bacteria involved modification of the dechlorination process according to culture conditions. In the absence of sulfate, the relative percentage of electrons used in PCE dehalogenation increased after an addition of lactate in batch conditions. The sulfate reducers would produce further reductant from lactate catabolism. This reductant might be used by methanogenic bacteria in PCE dechlorination. A mutualistic interaction was observed in the absence of sulfate. However in the presence of sulfate, methanogenesis and dechlorination decreased because of interspecific competition, probably between the H(2)-oxydizing methanogenic and sulfate-reducing bacteria in batch conditions. In the semicontinuous fixed-bed reactor, the presence of sulfate did not affect dechlorination and methanogenesis. The sulfate-reducing bacteria may not be competitors of H(2)-consuming methanogens in the reactor because of the existence of microbial biofilm. The presence of the fixed film may be an advantage for bioremediation and industrial treatment of effluent charged in sulfate and PCE. This is the first report on the microbial ecology of a methanogenic and sulfate-reducing PCE-enrichment consortium.     

Effects of hydrogen and formate on the degradation of propionate and butyrate in thermophilic granules from an upflow anaerobic sludge blanket reactor.

Degradation of propionate and butyrate in whole and disintegrated granules from a thermophilic (55 degrees C) upflow anaerobic sludge blanket reactor fed with acetate, propionate, and butyrate as substrates was examined. The propionate and butyrate degradation rates in whole granules were 1.16 and 4.0 mumol/min/g of volatile solids, respectively, and the rates decreased 35 and 25%, respectively, after disintegration of the granules. The effect of adding different hydrogen-oxidizing bacteria (both sulfate reducers and methanogens), some of which used formate in addition to hydrogen, to disintegrated granules was tested. Addition of either Methanobacterium thermoautotrophicum delta H, a hydrogen-utilizing methanogen that does not use formate, or Methanobacterium sp. strain CB12, a hydrogen- and formate-utilizing methanogen, to disintegrated granules increased the degradation rate of both propionate and butyrate. Furthermore, addition of a thermophilic sulfate-reducing bacterium (a Desulfotomaculum sp. isolated in our laboratory) to disintegrated granules improved the degradation of both substrates even more than the addition of methanogens. By monitoring the hydrogen partial pressure in the cultures, a correlation between the hydrogen partial pressure and the degradation rate of propionate and butyrate was observed, showing a decrease in the degradation rate with increased hydrogen partial pressure. No significant differences in the stimulation of the degradation rates were observed when the disintegrated granules were supplied with methanogens that utilized hydrogen only or hydrogen and formate. This indicated that interspecies formate transfer was not important for stimulation of propionate and butyrate degradation.     

Pure-culture growth of fermentative bacteria, facilitated by H2 removal: bioenergetics and H2 production

We used an H2-purging culture vessel to replace an H2-consuming syntrophic partner, allowing the growth of pure cultures of Syntrophothermus lipocalidus on butyrate and Aminobacterium colombiense on alanine. By decoupling the syntrophic association, it was possible to manipulate and monitor the single organism's growth environment and determine the change in Gibbs free energy yield (DeltaG) in response to changes in the concentrations of reactants and products, the purging rate, and the temperature. In each of these situations, H2 production changed such that DeltaG remained nearly constant for each organism (-11.1 +/- 1.4 kJ mol butyrate(-1) for S. lipocalidus and -58.2 +/- 1.0 kJ mol alanine(-1) for A. colombiense). The cellular maintenance energy, determined from the DeltaG value and the hydrogen production rate at the point where the cell number was constant, was 4.6 x 10(-13) kJ cell(-1) day(-1) for S. lipocalidus at 55 degrees C and 6.2 x 10(-13) kJ cell(-1) day(-1) for A. colombiense at 37 degrees C. S. lipocalidus, in particular, seems adapted to thrive under conditions of low energy availability.     

Isolation of key methanogens for global methane emission from rice paddy fields: a novel isolate affiliated with the clone cluster rice cluster I

Despite the fact that rice paddy fields (RPFs) are contributing 10 to 25% of global methane emissions, the organisms responsible for methane production in RPFs have remained uncultivated and thus uncharacterized. Here we report the isolation of a methanogen (strain SANAE) belonging to an abundant and ubiquitous group of methanogens called rice cluster I (RC-I) previously identified as an ecologically important microbial component via culture-independent analyses. To enrich the RC-I methanogens from rice paddy samples, we attempted to mimic the in situ conditions of RC-I on the basis of the idea that methanogens in such ecosystems should thrive by receiving low concentrations of substrate (H(2)) continuously provided by heterotrophic H(2)-producing bacteria. For this purpose, we developed a coculture method using an indirect substrate (propionate) in defined medium and a propionate-oxidizing, H(2)-producing syntroph, Syntrophobacter fumaroxidans, as the H(2) supplier. By doing so, we significantly enriched the RC-I methanogens and eventually obtained a methanogen within the RC-I group in pure culture. This is the first report on the isolation of a methanogen within RC-I.     

Spore-forming thermophilic sulfate-reducing bacteria isolated from north sea oil field waters

Thermophilic sulfate-reducing bacteria were isolated from oil field waters from oil production platforms in the Norwegian sector of the North Sea. Spore-forming rods dominated in the enrichments when lactate, propionate, butyrate, or a mixture of aliphatic fatty acids (C(4) through C(6)) was added as a carbon source and electron donor. Representative strains were isolated and characterized. The isolates grew autotrophically on H(2)-CO(2) and heterotrophically on fatty acids such as formate, propionate, butyrate, caproate, valerate, pyruvate, and lactate and on alcohols such as methanol, ethanol, and propanol. Sulfate, sulfite, and thiosulfate but not nitrate could be used as an electron acceptor. The temperature range for growth was 43 to 78 degrees C; the spores were extremely heat resistant and survived 131 degrees C for 20 min. The optimum pH was 7.0. The isolates grew well in salt concentrations ranging from 0 to 800 mmol of NaCl per liter. Sulfite reductase P582 was present, but cytochrome c and desulfoviridin were not found. Electron micrographs revealed a gram-positive cell organization. The isolates were classified as a Desulfotomaculum sp. on the basis of spore formation, general physiological characteristics, and submicroscopic organization. To detect thermophilic spore-forming sulfate-reducing bacteria in oil field water, polyvalent antisera raised against antigens from two isolates were used. These bacteria were shown to be widespread in oil field water from different platforms. The origin of thermophilic sulfate-reducing bacteria in the pore water of oil reservoirs is discussed.     

CO2-dependent fermentation of phenol to acetate, butyrate and benzoate by an anaerobic, pasteurised culture

Fermentative degradation of phenol was studied using a non-methanogenic, pasteurised enrichment culture containing two morphologically different bacteria. Phenol was fermented to benzoate, acetate and butyrate and their relative occurrence depended on the concentration of hydrogen. Proportionately more benzoate was formed with high initial levels of H2. The influence of PH2 on the fermentation pattern was studied both in dense cell suspensions and in growing cultures by addition of hydrogen. An increase in growth yield (OD578) was observed, compared to controls, as a consequence of phenol degradation; however, the increase was less in H2-amended treatments, in which most of the phenol ended up as benzoate. The degradation of phenol in the dense cell suspension experiments was dependent on CO2. Benzoate was not degraded when added as a substrate to the growing culture. This is, to our knowledge, the first report concerning the fermentative degradation of phenol to nonaromatic products.     

Acetate as a carbon source for hydrogen production by photosynthetic bacteria

Hydrogen is a clean energy alternative to fossil fuels. Photosynthetic bacteria produce hydrogen from organic compounds by an anaerobic light-dependent electron transfer process. In the present study hydrogen production by three photosynthetic bacterial strains (Rhodopseudomonas sp., Rhodopseudomonas palustris and a non-identified strain), from four different short-chain organic acids (lactate, malate, acetate and butyrate) was investigated. The effect of light intensity on hydrogen production was also studied by supplying two different light intensities, using acetate as the electron donor. Hydrogen production rates and light efficiencies were compared. Rhodopseudomonas sp. produced the highest volume of H2. This strain reached a maximum H2 production rate of 25 ml H2 l(-1) h(-1), under a light intensity of 680 micromol photons m(-2) s(-1), and a maximum light efficiency of 6.2% under a light intensity of 43 micromol photons m(-2) s(-1). Furthermore, a decrease in acetate concentration from 22 to 11 mM resulted in a decrease in the hydrogen evolved from 214 to 27 ml H2 per vessel.     

Role of formate and hydrogen in the degradation of propionate and butyrate by defined suspended cocultures of acetogenic and methanogenic bacteria

The butyrate-degradingSyntrophospora bryantii degrades butyrate and a propionate-degrading strain (MPOB) degrades propionate in coculture with the hydrogen- and formate-utilizingMethanospirillum hungatii orMethanobacterium formicicum. However, the substrates are not degraded in constructed cocultures with twoMethanobrevibacter arboriphilus strains which are only able to consume hydrogen. Pure cultures of the acetogenic bacteria form both hydrogen and formate during butyrate oxidation with pentenoate as electron acceptor and during propionate oxidation with fumarate as electron acceptor. Using the highest hydrogen and formate levels which can be reached by the acetogens and the lowest hydrogen and formate levels which can be maintained by the methanogens it appeared that the calculated formate diffusion rates are about 100 times higher than the calculated hydrogen diffusion rates.     

Interspecies hydrogen transfer in co-cultures of methanol-utilizing acidogens and sulfate-reducing or methanogenic bacteria

Abstract The metabolism of methanol by acidogenic bacteria ( Butyribacterium methylotrophicum, Sporomusa ovata and Acetobacterium woodii ) was studied in pure culture and in defined mixed cultures with sulfate-reducing bacteria ( Desulfovibrio vulgaris ) or methanogenic bacteria ( Methanobrevibacter arboriphilus strain AZ). In the mixed cultures, less acids (acetate and/or butyrate) were formed per unit methanol converted than in pure cultures. In these mixed cultures, a significant production of sulfide or methane was observed despite the inability of the sulfate reducer and the methanogen to use methanol as an energy substrate. These results are explained in terms of interspecies hydrogen transfer between the acidogens (converting part of the methanol to 1 CO₂ and 3 H₂) and the Desulfovibrio or Methanobrevibacter species. The bioenergetic aspects of this process and its ecological implications are discussed.     

Characterization of 3-chlorobenzoate degrading aerobic bacteria isolated under various environmental conditions

The rates of bacterial growth in nature are often restricted by low concentrations of oxygen or carbon substrates. In the present study the metabolic properties of 24 isolates that had been isolated using various concentrations of 3-chlorobenzoate, benzoate and oxygen as well as using continuous culture at high and low growth rates were determined to investigate the effects of these parameters on the metabolism of monoaromatic compounds. Bacteria were enriched from different sampling sites and subsequently isolated. In batch culture this was done both under low oxygen (2% O(2)) and air-saturated concentrations. Chemostat enrichments were performed under either oxygen or 3-chlorobenzoate limiting conditions. Bacteria metabolizing aromatics with gentisate or protocatechuate as intermediates (gp bacteria) as well as bacteria metabolizing aromatic compounds via catechols (cat bacteria) were isolated from batch cultures when either benzoate or 3CBA were used as C sources, regardless of the enrichment conditions applied. In contrast, enrichments performed in chemostats at low dilution rates resulted in gp-type organisms only, whereas at high dilution rates cat-type organisms were enriched, irrespective of the oxygen and 3-chlorobenzoate concentration used during enrichment. It is noteworthy that the gp-type of bacteria possessed relatively low μ(max) values on 3CBA and benzoate along with relatively high substrate and oxygen affinities for these compounds. This is in contrast with cat-type of bacteria, which seemed to be characterized by high maximum specific growth rates on the aromatic substrates and relatively high apparent half saturation constants. In contrast, bacteria degrading chlorobenzoate via gentisate or protocatechuate may possibly be better adapted to conditions leading to growth at reduced rates such as low oxygen and low substrate concentrations.     

Sulfate reduction in methanogenic bioreactors

Abstract: In the anaerobic treatment of sulfate-containing wastewater, sulfate reduction interferes with methanogenesis. Both mutualistic and competitive interactions between sulfate-reducing bacteria and methanogenic bacteria have been observed. Sulfate reducers will compete with methanogens for the common substrates hydrogen, formate and acetate. In general, sulfate reducers have better growth kinetic properties than methanogens, but additional factors which may be of importance in the competition are adherence properties, mixed substrate utilization, affinity for sulfate of sulfate reducers, relative numbers of bacteria, and reactor conditions such as pH, temperature and sulfide concentration. Sulfate reducers also compete with syntrophic methanogenic consortia involved in the degradation of substrates like propionate and butyrate. In the absence of sulfate these methanogenic consortia are very important, but in the presence of sulfate they are thought to be easily outcompeted by sulfate reducers. However, at relatively low sulfate concentrations, syntrophic degradation of propionate and butyrate coupled to HZ removal via sulfate reduction rather than via methanogenesis may become important. A remarkable feature of some sulfate reducers is their ability to grow fermentatively or to grow in syntrophic association with methanogens in the absence of sulfate.     

Metabolism of polyethylene glycol by two anaerobic bacteria, Desulfovibrio desulfuricans and a Bacteroides sp

Two anaerobic bacteria were isolated from polyethylene glycol (PEG)-degrading, methanogenic, enrichment cultures obtained from a municipal sludge digester. One isolate, identified as Desulfovibrio desulfuricans (strain DG2), metabolized oligomers ranging from ethylene glycol (EG) to tetraethylene glycol. The other isolate, identified as a Bacteroides sp. (strain PG1), metabolized diethylene glycol and polymers of PEG up to an average molecular mass of 20,000 g/mol [PEG 20000; HO-(CH2-CH2-O-)nH]. Both strains produced acetaldehyde as an intermediate, with acetate, ethanol, and hydrogen as end products. In coculture with a Methanobacterium sp., the end products were acetate and methane. Polypropylene glycol [HO-(CH2-CH2-CH2-O-)nH] was not metabolized by either bacterium, and methanogenic enrichments could not be obtained on this substrate. Cell extracts of both bacteria dehydrogenated EG, PEGs up to PEG 400 in size, acetaldehyde, and other mono- and dihydroxylated compounds. Extracts of Bacteroides strain PG1 could not dehydrogenate long polymers of PEG (greater than or equal to 1,000 g/mol), but the bacterium grew with PEG 1000 or PEG 20000 as a substrate and therefore possesses a mechanism for PEG depolymerization not present in cell extracts. In contrast, extracts of D. desulfuricans DG2 dehydrogenated long polymers of PEG, but whole cells did not grow with these polymers as substrates. This indicated that the bacterium could not convert PEG to a product suitable for uptake.     

Metabolism of polyethylene glycol by two anaerobic bacteria, Desulfovibrio desulfuricans and a Bacteroides sp.

Two anaerobic bacteria were isolated from polyethylene glycol (PEG)-degrading, methanogenic, enrichment cultures obtained from a municipal sludge digester. One isolate, identified as Desulfovibrio desulfuricans (strain DG2), metabolized oligomers ranging from ethylene glycol (EG) to tetraethylene glycol. The other isolate, identified as a Bacteroides sp. (strain PG1), metabolized diethylene glycol and polymers of PEG up to an average molecular mass of 20,000 g/mol [PEG 20000; HO-(CH2-CH2-O-)nH]. Both strains produced acetaldehyde as an intermediate, with acetate, ethanol, and hydrogen as end products. In coculture with a Methanobacterium sp., the end products were acetate and methane. Polypropylene glycol [HO-(CH2-CH2-CH2-O-)nH] was not metabolized by either bacterium, and methanogenic enrichments could not be obtained on this substrate. Cell extracts of both bacteria dehydrogenated EG, PEGs up to PEG 400 in size, acetaldehyde, and other mono- and dihydroxylated compounds. Extracts of Bacteroides strain PG1 could not dehydrogenate long polymers of PEG (greater than or equal to 1,000 g/mol), but the bacterium grew with PEG 1000 or PEG 20000 as a substrate and therefore possesses a mechanism for PEG depolymerization not present in cell extracts. In contrast, extracts of D. desulfuricans DG2 dehydrogenated long polymers of PEG, but whole cells did not grow with these polymers as substrates. This indicated that the bacterium could not convert PEG to a product suitable for uptake.     

Characterization of metabolic performance of methanogenic granules treating brewery wastewater: role of sulfate-reducing bacteria.

Granules from an upflow anaerobic sludge blanket system treating a brewery wastewater that contained mainly ethanol, propionate, and acetate as carbon sources and sulfate (0.6 to 1.0 mM) were characterized for their physical and chemical properties, metabolic performance on various substrates, and microbial composition. Transmission electron microscopic examination showed that at least three types of microcolonies existed inside the granules. One type consisted of Methanothrix-like rods with low levels of Methanobacterium-like rods; two other types appeared to be associations between syntrophic-like acetogens and Methanobacterium-like organisms. The granules were observed to be have numerous vents or channels on the surface that extended into the interior portions of the granules that may be involved in release of gas formed within the granules. The maximum substrate conversion rates (millimoles per gram of volatile suspended solids per day) at 35 degrees C in the absence of sulfate were 45.1, 8.04, 4.14, and 5.75 for ethanol, acetate, propionate, and glucose, respectively. The maximum methane production rates (millimoles per gram of volatile suspended solids per day) from H2-CO2 and formate were essentially equal for intact granules (13.7 and 13.5) and for physically disrupted granules (42 and 37). During syntrophic ethanol conversion, both hydrogen and formate were formed by the granules. The concentrations of these two intermediates were maintained at a thermodynamic equilibrium, indicating that both are intermediate metabolites in degradation. Formate accumulated and was then consumed during methanogenesis from H2-CO2. Higher concentrations of formate accumulated in the absence of sulfate than in the presence of sulfate. The addition of sulfate (8 to 9 mM) increased the maximum substrate degradation rates for propionate and ethanol by 27 and 12%, respectively. In the presence of this level of sulfate, sulfate-reducing bacteria did not play a significant role in the metabolism of H2, formate, and acetate, but ethanol and propionate were converted via sulfate reduction by approximately 28 and 60%, respectively. In the presence of 2.0 mM molybdate, syntrophic propionate and ethanol conversion by the granules was inhibited by 97 and 29%, respectively. The data show that in this granular microbial consortium, methanogens and sulfate-reducing bacteria did not compete for common substrates. Syntrophic propionate and ethanol conversion was likely performed primarily by sulfate-reducing bacteria, while H2, formate, and acetate were consumed primarily by methanogens.