Gram Of Volatile Suspended Solids Research Articles

Microbial reduction of antimony under anaerobic conditions by inocula from different sources

Antimony (Sb) is a toxic and carcinogenic metalloid that can be present in contaminated water generated by mining operations and other industrial activities. Dissimilatory Sb(V) reduction to Sb(III) under anaerobic conditions by prokaryotic microbial communities has been reported; however, the fate of antimony in anaerobic wastewater treatment systems is poorly understood. This study investigated the potential of six different inoculum sources to reduce Sb(V) under anaerobic conditions and correlated the Sb(V) reduction rate with the microbial community composition. Microbial reduction of Sb(V) to Sb(III) by all the inoculum sources tested was observed in anaerobic incubations using hydrogen as an electron donor after a month. The highest reduction rate observed was 12.5 mg Sb per gram of volatile suspended solids per day. In some inocula, not only Sb reduction was observed, but also a decrease in the total Sb concentration, suggesting the precipitation of Sb(III). Metagenomic analysis revealed high variability in the community diversity associated with the origin of the inoculum. For the family taxonomic category, the three inocula that showed the highest Sb reduction potential were characterized by the presence of the Bacillaceae family.

Influence of salt addition to stimulating biopolymers production in aerobic granular sludge systems

The influence of salt addition to stimulating biopolymers production in aerobic granular sludge (AGS) systems was evaluated. The control systems (R1: acetate and R2: propionate) initially obtained less accumulation of mixed liquor volatile suspended solids (MLVSS), indicating that the osmotic pressure in the salt-supplemented systems (R3: acetate and R4: propionate) contributed to biomass growth. However, the salt-supplemented systems collapsed between days 110 and 130 of operation. R3 and R4 showed better performance regarding nutrients removal due to the greater abundance of nitrifying and denitrifying bacteria and phosphate-accumulating organisms. Salt also contributed to the higher production of biopolymers such as alginate-like exopolymers (ALE) per gram of volatile suspended solids (VSS) (R1: 397 mgALE∙gVSS−1, R2: 140 mgALE∙gVSS−1, R3: 483 mgALE∙gVSS−1, R4: 311 mgALE∙gVSS−1). Amino acids like tyrosine and tryptophan were better identified in extracellular polymeric substances extract from salt-operated reactors. This study brings important results in the context of resource recovery by treating saline effluents.

Carbon source affects the resource recovery in aerobic granular sludge systems treating wastewater

This study evaluated the influence of carbon sources on alginate-like exopolymers (ALE) and tryptophan (Trp) biosynthesis in the aerobic granular sludge (AGS). With acetate, the highest biopolymers levels, per gram of volatile suspended solids (VSS) (418.7 mgALE∙g−1 and 4.1 mgTrp∙gVSS−1), were found likely due to biomass loss throughout the operation, which resulted in lower sludge age (4–7 days) and shorter famine period. During granulation, encouraging results on ALE production were obtained with propionate (>250 mgALE∙gVSS−1), significantly higher than those found with glycerol, glucose, and sucrose. Regarding tryptophan production, propionate and glycerol proved to be good substrates, although the content was still lower than acetate (1.6 mgTrp∙gVSS−1). Granules fed with glucose showed the worst results compared to the other substrates (38.5 mgALE∙VSS−1 and 0.6 mgTrp∙gVSS−1) due to the filamentous microorganisms’ abundance found. Therefore, this study provides insights to value the production of compounds of industrial interest in AGS systems.

Impact of solids retention time on the biological performance of an AnMBR treating lipid-rich synthetic dairy wastewater

ABSTRACT In this study, the impact of applied solids retention time (SRT) on the biological performance of an anaerobic membrane bioreactor (AnMBR) treating synthetic dairy wastewater with high lipid content was assessed. Two side-stream AnMBR systems were operated at an SRT of 20 and 40 days (R20 and R40, respectively), equipped with an inside-out tubular membrane operated in cross-flow mode under full-scale operational conditions, i.e. crossflow velocity, transmembrane pressure, membrane flux. Successful operation was achieved and removal efficiencies of both reactors were up to 99% applying an organic loading rate (OLR) of 4.7 g COD L−1 d−1. No precipitation of lipids was observed throughout the operational period, keeping the lipids available for the anaerobic degradation. Long chain fatty acid (LCFA) accumulation was very modest and amounted 148 and 115 mg LCFA-COD per gram of volatile suspended solids (VSS) for R20 and R40, respectively. At an SRT of 40 days, a slightly better biological conversion was obtained. Periodically performed specific methanogenic activity (SMA) tests showed stabilization of the SMA for R40 sludge, whereas for R20 sludge the SMA continued to decrease. This study revealed a more stable reactor performance operating the AnMBR at an SRT of 40 days compared to 20 days.

Sludge granulation in an UASB–moving bed biofilm hybrid reactor for efficient organic matter removal and nitrogen removal in biofilm reactor

ABSTRACTA hybrid upflow anaerobic sludge blanket (UASB)–moving bed biofilm (MBB) and rope bed biofilm (RBB) reactor was designed for treatment of sewage. Possibility of enhancing granulation in an UASB reactor using moving media to improve sludge retention was explored while treating low-strength wastewater. The presence of moving media in the top portion of the UASB reactor allowed a high solid retention time even at very short hydraulic retention times and helped in maintaining selection pressure in the sludge bed to promote formation of different sized sludge granules with an average settling velocity of 67 m/h. These granules were also found to contain plenty of extracellular polymeric substance (EPS) such as 58 mg of polysaccharides (PS) per gram of volatile suspended solids (VSS) and protein (PN) content of 37 mg/g VSS. Enriched sludge of nitrogen-removing bacteria forming a porous biofilm on the media in RBB was also observed in a concentration of around 894 g/m2. The nitrogen removing sludge also had a high EPS content of around 22 mg PS/g VSS and 28 mg PN/g VSS. This hybrid UASB-MBB-RBB reactor with enhanced anaerobic granular sludge treating both carbonaceous and nitrogenous matter may be a sustainable solution for decentralized sewage treatment.

Effect of intermittent aeration on microbial diversity in an intermittently aerated IFAS reactor treating municipal wastewater: A field study

ABSTRACTIn the present study, the effect of three intermittent aeration (IA) cycles on treatment performance and microbial diversity was investigated in an integrated fixed film activated sludge (IFAS) reactor treating municipal wastewater. The results showed that IA strategies were able to achieve efficient removal of organics and nitrogen ranging between 90 and 95% and 70 and 80%, respectively, however the phosphorus removal was found to be inversely proportional to the duration of aeration off time in each IA cycle. The microscopic analysis revealed that the suspended and attached biomass had compact morphology and open floc structure, respectively. For each gram of volatile suspended solids, 165 and 148 mg of extracellular polymeric substances (EPS) were extracted from attached and suspended biomass, respectively, constituting carbohydrates (∼24%), proteins (∼31%), humic acids (∼28%), DNA (∼2%) and unknown substances (∼12%). The microbial diversities of suspended biomass in IFAS reactor were investigated using culture-dependent approach, which confirmed the presence of Clostridium spp., Pseudomonas spp., Bacillus spp., Escherichia coli spp., Nitrosococcus spp., Streptococcus spp., Acinetobacter spp., Betaproteobacteria outliers, Klebsiella pneumoniae, Klebsiella aerogenes, Serratia marcescens, Micrococcus, Proteus vulgaris spp., Actinomycetes spp., and Actinobacteria including Micromonospora spp. and Streptomyces spp. Molecular tools for diversity analyses were used for ammonia and nitrite oxidizer identification, such as Nitrospira and Nitrosococcus species. Denitrifiers include the species of Pseudomonas, Betaproteobacteria and Flavobacterium. Acinetobacter, Betaproteobacteria and Gammaproteobacteria were responsible for the phosphorus removal in the present system. Overall, the system performed efficiently showing Proteobacteria (59%), Acinetobacter (12%) and Bacteroidetes (11%) as the dominant bacterial groups. However, the dominance of the bacterial diversity varied with the IA cycle time numerating the maximum percentage of bacterial species during IA1 phase i.e. 2.5 h aeration/0.5 h non-aeration.

A new biological process for short-chain fatty acid generation from waste activated sludge improved by Clostridiales enhancement.

Short-chain fatty acids (SCFAs), the carbon source of biological nutrient removal, can be produced by waste activated sludge (WAS) anaerobic fermentation. To get more SCFAs from sludge, most studies in literature focused on the mechanical process control or the structure of microbial community; little attention has been paid to the key microorganisms and their function related to SCFA generation. In this study, a different sludge pretreated method, i.e., pretreating sludge by proteinase K for 2days followed by pretreating at pH 10 for 4days, is reported, by which the proportion of Clostridiales was increased and SCFA generation was enhanced. First, the effects of different proteinase K concentrations and initial pH on sludge hydrolysis and SCFA generation were investigated. The optimal conditions showed the highest SCFA generation (352.91mg COD per gram of volatile suspended solids), which was 2.89-fold of the blank (un-pretreated). Further, the new biological pretreatment process led to the conversion of other SCFAs to acetic acid. Acetic acid accounted for 60.8% of total SCFAs with the new biological pretreatment process compared with 44.9% in the blank test. Then, the investigation on the key microorganisms related to SCFA production with 16S rRNA gene clone library and fluorescence in situ hybridization (FISH) indicated that there were much greater active Clostridiales when SCFAs were generated with the proteinase K and pH 10 pretreated sludge. Further, the mechanisms for the optimal conditions significantly enhancing SCFA generation were investigated. It was found that pretreating sludge by proteinase K and pH 10 caused the greatest key enzyme activities, organic consumption, and inhibition of methane generation. Graphical abstract A new biological process for short-chain fatty acid generation from waste activated sludge improved by Clostridiales enhancement.

Development of a down-flow hanging sponge reactor for the treatment of low strength sewage

The process performance of a down-flow hanging sponge (DHS) reactor for treating low strength sewage (biochemical oxygen demand (BOD) 20-50 mg/L) was investigated in Bangkok, Thailand. The hydraulic retention time (HRT) was set at 4 h during the start-up period and was reduced to 1.5 h in a stepwise manner. Throughout the 300-day operational period, the DHS reactor shows high performance with respect to the removal of total suspended solid (>90% total suspended solid removal efficiency). No clogging of sponge media was observed in response to the self-digestion phenomena of the biofilm. At a HRT of 1.5 h, the BOD removal efficiency was sufficiently high (about 85%). The pathogen Escherichia coli and other coliform bacteria were removed almost completely as well (removal was 99.4% and 98.1%, respectively). Regarding the retained sludge activity measurement, the nitrite oxidation rate was higher than the ammonium oxidation rate (0.031 and 0.022 gram of nitrogen per gram of volatile suspended solids per day, respectively). In the 300 days of operation, the amount of excess sludge production was negligible. Thus, no sludge treatment system is required. Introduction of the DHS system in developing countries is recommended because this system requires a relatively small area, and has low electricity consumption and operation costs.

Dynamics of Intracellular Polymers in Enhanced Biological Phosphorus Removal Processes under Different Organic Carbon Concentrations

Enhanced biological phosphorus removal (EBPR) may deteriorate or fail during low organic carbon loading periods. Polyphosphate accumulating organisms (PAOs) in EBPR were acclimated under both high and low organic carbon conditions, and then dynamics of polymers in typical cycles, anaerobic conditions with excess organic carbons, and endogenous respiration conditions were examined. After long-term acclimation, it was found that organic loading rates did not affect the yield of PAOs and the applied low organic carbon concentrations were advantageous for the enrichment of PAOs. A low influent organic carbon concentration induced a high production of extracellular carbohydrate. During both anaerobic and aerobic endogenous respirations, when glycogen decreased to around 80 ± 10 mg C per gram of volatile suspended solids, PAOs began to utilize polyphosphate significantly. Regressed by the first-order reaction model, glycogen possessed the highest degradation rate and then was followed by polyphosphate, while biomass decay had the lowest degradation rate.

Evaluation of biodegradation of soluble and particulate matter produced during sewage sludge ozonation by a combination of chemical and microbial approaches

In this study a batch aerobic reactor fed with the soluble and particulate matter produced during sludge ozonation (reactor 1) is proposed and its performance was evaluated in comparison to a reactor fed with municipal wastewater (reactor 2) to investigate the biodegradation properties of the ozone-treated sludge. During long-term operation of nearly 7 months, the efficiency of reactor 1 was preserved, although there was a 5% decrease in total COD elimination, compared with reactor 2. The released proteins and polysaccharides during sludge ozonation were effectively decomposed. The volatile suspended solid (VSS)/total suspended solid (TSS) ratio and the oxygen uptake rate in reactor 1 were lower than those in reactor 2 by 19% and 27%, respectively. However, the protease activity per gram of volatile suspended solids and the ammonium oxidation rate in reactor 1 were higher. The yields of sludge in reactor 1 fed with ozonated sludge were lower which implied that the cryptic growth using the organic autochthonous substrate might produce reduced biomass yields, which would decrease sludge production. The debris-like substances observed from scanning electron microscope (SEM) images and the functional groups associated with calcite were only found in reactor 1 by FT-IR analysis, which implies that some of the inert or inorganic substances from sludge ozonation were accumulated in the reactor.

Sulfate reduction at pH 4.0 for treatment of process and wastewaters

Acidic industrial process and wastewaters often contain high sulfate and metal concentrations and their direct biological treatment is thus far not possible as biological processes at pH < 5 have been neglected. Sulfate-reducing bacteria convert sulfate to sulfide that can subsequently be used to recover metals as metal-sulfides precipitate. This study reports on high-rate sulfate reduction with a mixed microbial community at pH 4.0 and 4.5 with hydrogen and/or formate as electron donors. The maximum sulfate reducing activity at pH 4.0 was sustained for over 40 days with a specific activity 500-fold greater than previously reported values: 151 mmol sulfate reduced/L reactor liquid per day with a maximum specific activity of 84 mmol sulfate per gram of volatile suspended solids per day. The biomass yield gradually decreased from 38 to 0.4 g volatile suspended solids per kilogram of sulfate when decreasing the reactor pH from pH 6 to 4. The microorganisms had a high maintenance requirement probably due maintaining pH homeostasis and the toxicity of sulfide at low pH. The microbial community diversity in the pH 4.0 membrane bioreactor decreased over time, while the diversity of the sulfate reducing community increased. Thus, a specialized microbial community containing a lower proportion of microorganisms capable of activity at pH 4 developed in the reactor compared with those present at the start of the experiment. The 16S rRNA genes identified from the pH 4.0 grown mixed culture were most similar to those of Desulfovibrio species and Desulfosporosinus sp. M1.

Enhancement of Waste Activated Sludge Protein Conversion and Volatile Fatty Acids Accumulation during Waste Activated Sludge Anaerobic Fermentation by Carbohydrate Substrate Addition: The Effect of pH

Volatile fatty acids (VFAs), the carbon source of biological nutrients removal, can be produced by waste activated sludge (WAS) anaerobic fermentation. However, because of high protein content and low carbon to nitrogen mass ratio (C/N) of WAS, the production of VFAs, especially propionic acid, a more preferred VFA than acetic acid for enhanced biological phosphorus removal (EBPR), is limited. After the addition of carbohydrate (rice was used as the model matter) to the WAS anaerobic fermentation system to balance the C/N ratio, the effect of pH on WAS protein conversion and VFAs production was investigated in this paper. Experimental results showed that the addition of carbohydrate matter caused a remarkable enhancement of WAS protein conversion and protease activity, and an apparent synergistic effect between WAS and carbohydrate matter was observed. The study of pH effect revealed that pH influenced not only the total VFAs production but the percentage of individual VFA. The maximal VFAs production (520.1 mg COD per gram of volatile suspended solids (VSS)) occurred at pH 8.0 and a fermentation time of 8 d, which was more than three times that at uncontrolled pH (150.2 mg COD/g VSS). The analysis of the composition of VFAs showed that propionic acid ranked first at pH 6.0-9.0 (around 50%) whereas acetic acid was the dominant product at other pHs investigated. Thus, the suitable conditions for propionic acid-enriched VFAs production were pH 8.0 and a time of 8 d. Further investigation showed that as there was more fermentation substrate consumption with lower biogas generation at pH 8.0, improved VFAs production was observed. Also, the key enzymes relevant to propionic acid formation exhibited the highest activities at pH 8.0, which resulted in the greatest propionic acid content in the fermentative VFAs. The 16S rRNA gene clone library demonstrated that Clostridia, beta-Proteobacteria, and Bacteroidetes were the dominant microbial community when the current anaerobic fermentation system was operated at pH 8.0. With the fermentative VFAs as the additional carbon source of municipal wastewater, the EBPR performance was significantly increased.

The effects of the ratio of propionate to acetate on the transformation and composition of polyhydroxyalkanoates with enriched cultures of glycogen‐accumulating organisms

Studies on the synthesis of polyhydroxyalkanoates (PHA) mainly focus on the enriched cultures of phosphorus‐accumulating (PAO) or glycogen‐accumulating organisms (GAO) with acetate or propionate as the sole carbon source. However, PHA production with a mixture of propionate and acetate by enriched cultures of GAO has not been reported. It is well known that the physical and mechanical properties of PHA can be improved by incorporating 3‐hydroxyvalerate (3HV) into the polymer. To synthesize PHA with a high 3HV percentage, five laboratory‐scale sequencing batch reactors were operated with the enriched cultures of GAO using different ratios of propionate and acetate (P/A) as the carbon sources in this study. The effects of P/A ratio on the anaerobic and aerobic transformations, composition and synthesis rate of PHA were studied. The results showed that with the increase in the P/A ratio, the anaerobic yields of total PHA declined from 4.226 to 2.469 mmol‐C per gram of volatile suspended solids (VSS), and the 3‐hydroxybutyrate (3HB) fraction in PHA decreased from approximately 70 to 10%, along with an increase in 3HV and 3‐hydroxy‐2‐methylvalerate (3H2MV) percentages. When there were equal quantities (mmol‐C L−1) of propionate and acetate, 3HB was produced more quickly than 3H2MV. By increasing P/A ratio from 1/10 to 10/1, the initial synthesis rate of 3H2MV rose from 0.002 to 0.029, while 3HB and the total PHA declined from 0.083 to 0.007 and 0.110 to 0.071 mmol‐C g‐VSS−1 min−1, respectively. Also, it was observed that more 3HV was incorporated into PHA with increasing P/A ratio. This study suggested that when the enriched cultures of GAO were applied to the synthesis of PHA, the composition, yield and synthesis rate of PHA could be manipulated by varying the feed composition.

Improved Bioproduction of Short-Chain Fatty Acids (SCFAs) from Excess Sludge under Alkaline Conditions

The production of short-chain fatty acids (SCFAs) from excess sludge was conducted in batch fermentation tests at different pH values ranging from 4.0 to 11.0. Experimental results of the impacts of different pHs on SCFAs production showed that during the first 8-day fermentation time the total SCFAs production at either pH 9.0 or pH 10.0 was much greater than that at acidic or neutral pH, and the maximal yield of 256.2 mg SCFAs-COD per gram of volatile suspended solids (VSS) was at pH 10.0, which was, respectively, over3 and 4times that at pH 5.0 and uncontrolled pH. Clearly, SCFAs production from excess sludge could be significantly improved and maintained stable by controlling the fermentation pH at 10.0. The composition of SCFAs and the percent distribution of individual SCFAs accounting for total SCFAs at pH 10.0 were analyzed. The SCFAs consisted of acetic, propionic, iso-butyric, n-butyric, iso-valeric, and n-valeric acids, and acetic acid was the most prevalent product with a fraction of 40-55%. Because the results of this study were differentfrom those of previous studies of SCFAs production, the mechanism of increased SCFAs production under alkaline conditions was investigated. Results showed that as soluble COD increased, more soluble protein was provided as the substrate for producing SCFAs. In addition, less or even no SCFAs were consumed by methanogens at alkaline pH, so the SCFAs production was therefore remarkably improved. Further investigation revealed thatthe formation of SCFA at pH 10.0 was dominated by biological effects rather than by chemical hydrolysis.

Thermophilic H2 production from a cellulose-containing wastewater.

Cellulose in wastewater was converted into H2 by a mixed culture in batch experiments at 55 degrees C with various wastewaters pH (5.5-8.5) and cellulose concentrations (10-40 g l(-1)). At the optimal pH of 6.5, the maximum H2 yield was 102 ml g(-1) cellulose and the maximum production rate was 287 ml d(-1) for each gram of volatile suspended solids (VSS). Analysis of 16S rDNA sequences showed that the cellulose-degrading mixed culture was composed of microbes closely affiliated to genus Thermoanaerobacterium.

Microbial diversity of a mesophilic hydrogen-producing sludge

A hydrogen-producing sludge degraded 99% of glucose at 36 degrees C and pH 5.5, producing a methane-free biogas (comprising 64% hydrogen) and an effluent comprising mostly butyrate, acetate, and ethanol. The yield was 0.26 l H2 g(-1) glucose and the production rate per gram of volatile suspended solids was 4.6 1 H2 day(-1). A 16S rDNA library was constructed from the sludge for microbial species determination. A total of 96 clones were selected for plasmids recovery, screened by denaturing gradient gel electrophoresis, and sequenced for rDNA. Based on the phylogenetic analysis of the rDNA sequences, 64.6% of all the clones were affiliated with three Clostridium species (Clostridiaceae), 18.8% with Enterobacteriaceae, and 3.1% with Streptococcus bovis (Streptococcaceae). The remaining 13.5% belonged to eight operational taxonomic units, the affiliations of which were not identified.

Detection and quantification of Desulforhabdus amnigenus in anaerobic granular sludge by dot blot hybridization and PCR amplification

A species-specific 16S rRNA oligonucleotide probe (ASRB1) was developed for the detection of Desulforhabdus amnigenus in anaerobic granular sludge. The presence of nucleic acids from cells of D. amnigenus in granular sludge was determined using ASRBI as a specific primer for polymerase chain reaction (PCR) amplification or as a probe for dot blot hybridizations. The detection threshold and the reproducibility of these two methods were determined with sludge amended with 10 4 -10 10 D. amnigenus cells per gram of volatile suspended solids (VSS), For D. amnigenus cells with a ribosomal RNA content of 15 fg cell -1 , the lowest number of target cells detected by hybridization was 1 x 10 8 cells g -1 VSS. With the PCR amplification method the lowest number of target cells which could be detected was 1 x 10 7 g -1 VSS. This corresponds to a threshold level for hybridization of 0.1-0.001‰ of the total bacterial sludge population, while the threshold level obtained with the PCR approach amounted to 0.01-0.0001‰. The rRNA content of D. amnigenus was found to be affected by the growth rate and the growth phase, and it ranged from 19 fg cell -1 in slow-growing cultures to 90 fg cell -1 in fast-growing cultures. Therefore, the detection threshold of the dot blot hybridization method for fast-growing cells is lower than for slow-growing cells.

Impact of the reactor hydrodynamics and organic loading on the size and activity of anaerobic granules

Wastewater treatment processes based on the upflow anaerobic sludge bed design are strongly dependent on the aggregation of biomass into macroscopic granules (1–3 mm) which settle well. The reactor hydrodynamics is of importance in the granulation process. The effect of the liquid upflow velocity υ UP associated with the operating time on the mean granule size and on the hydrogen, formate, acetate, propionate and glucose specific activities was studied, at various specific loading rates, in upflow sludge bed and filter reactors of 13 l fed with sugar wastewater. Reactors which were operated at 0.9 m h −1 behaved as fixed beds while those run at 2.2, 4.4 and 6.6 m h −1 were fluidized, because an immediate spatial gradient of the sludge particle sizes was induced. The υ UP had a significant positive effect on mean granule size. A specific loading rate increase from 0.5 to 1.5 g chemical oxygen demand per gram of volatile suspended solids per day raised proportionally the biomass growth rate, but had no positive effect on the granule development in size. Moreover, the υ UP had little effect on the specific wash-out rate of the smaller particles. Henceforth the resulting final size of granules is essentially a function of the hydrodynamic regime. Major impact on granule net steady size is attributed to several mechanisms related to fluidization: improved penetration of substrates into biofilm; insignificance of liquid shear relative to the shear of gas; reduction of particle friction and attrition with the bed voidage. Acidogenic (glucotrophic) activity decreased with υ UP increased yielding minimum values at intermediate υ UP, between 2 and 5 m h −1. Postacidogenic activities (propionate, acetate, formate, H 2) were positively influenced by υ UP to a slight extent. Glucose activity gradient within the granule bed was highly and inversely correlated to the granule size, while for acetate activity gradients, the correlation was direct, although less strong. These observations are discussed in detail with regard to an ordered distribution of the consortium populations within the granule spatial structure.

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.