Anammox bacteria enrichment in upflow anaerobic sludge blanket (UASB) reactor

We investigated the anaerobic ammonium oxidation (anammox) reaction in a labscale upflow anaerobic sludge blanket (UASB) reactor. Our aim was to detect and enrich the organisms responsible for the anammox reaction using a synthetic medium that contained low concentrations of substrates (ammonium and nitrite). The reactor was inoculated with granular sludge collected from a full-scale anaerobic digestor used for treating brewery wastewater. The experiment was performed during 260 days under conditions of constant ammonium concentration (50 mg NH₄/⁺-N/L) and different nitrite concentrations (50∼150 mg NO₂-N/L). After 200 days, anammox activity was observed in the system. The microorganisms involved in this anammox reaction were identified as CandidatusB. Anammoxidans andK. Stuttgartiensis using fluorescencein situ hybridization (FISH) method.     

Granular sludge formation in upflow anaerobic sludge blanket (UASB) reactors

The state of the art for upflow anaerobic sludge blanket (UASB) reactors is discussed, focusing on the microbiology of immobilized anaerobic bacteria and the mechanism of granule formation. The development of granular sludge is the key factor for successful operation of the UASB reactors. Criteria for determining if granular sludge has developed in a UASB reactor is given based on the densities and diameters of the granular sludge. The shape and composition of granular sludge can vary significantly. Granules typically have a spherical form with a diameter from 0.14 to 5 mm. The inorganic mineral content varies from 10 to 90% of the dry weight of the granules, depending on the wastewater composition etc. The main components of the ash are calcium, potassium, and iron. The extracellular polymers in the granular sludge are important for the structure and maintenance of granules, while the inorganic composition seems to be of less importance. The extracellular polymer content varies between 0.6 and 20% of the volatile suspended solids and consists mainly of protein and polysaccharides. Both Methanosaeta spp. (formerly Methanothrix) and Methanosarcina spp. have been identified as important aceticlastic methanogens for the initial granulation and development of granular sludge. Immunological methods have been used to identify other methanogens in the granules. The results have showed that, besides the aceticlastic methanogens Methanosaeta spp. and Methanosarcina spp., hydrogen and formate utilizing bacteria are also present, e.g., Methanobacterium formicicum, Methanobacterium thermoautotrophicum, and Methanobrevibacter spp. Microcolonies of syntrophic bacteria are often observed in the granules, and the significant electron transfer in these microcolonies occurs through interspecies hydrogen transfer. The internal organization of the various groups of bacteria in the granules depends on the wastewater composition and the dominating metabolic pathways in the granules. Internal organization is observed in granules where such an arrangement is beneficial for an optimal degradation of the wastewater. A four-step model is given for the initial development of granular sludge. (c) 1996 John Wiley & Sons, Inc.     

Granulation in thermophilic upflow anaerobic sludge blanket (UASB) reactors

The state of the art for thermophilic UASB reactors is discussed focusing on the start-up of UASB reactors, the influence of the waste water composition and temperature on the development and maintenance of thermophilic granules, and the microbial composition and structure of thermophilic granules.     

Performance of upflow anaerobic sludge blanket (UASB) reactor treating wastewaters containing carbon tetrachloride

The anaerobic biodegradation of carbon tetrachloride (CT) was investigated during the granulation process by reducing the hydraulic retention time, increasing the chemical oxygen demand (COD) and CT loadings in a 2l laboratory-scale upflow anaerobic sludge blanket (UASB) reactor. Anaerobic unacclimated sludge and glucose were used as seed and primary substrate, respectively. Granules were developed 4 weeks after start-up, which grew at an accelerated rate for 8 months, and then became fully grown. The effect of operational parameters such as influent CT concentrations, COD, CT loading, food to biomass ratio and specific methanogenic activity (SMA) were also considered during granulation. The granular sludge cultivated had a maximum diameter of 2.1 mm and SMA of 1.6 g COD/g total suspended solid (TSS) day. COD and CT removal efficiencies of 92 and 88% were achieved when the reactor was firstly operating at CT and COD loading rates of 17.5 mg/l day and 12.5 g/l day, respectively. This corresponds to hydraulic retention time of 0.28 day and food to biomass ratio of 0.5 g COD/g TSS day. Kinetic coefficients of maximum specific substrate utilization rate, half velocity coefficient, growth yield coefficient and decay coefficient were determined to be 2.4 × 10^–3 mg CT/TSS day^–1, 1.37 mg CT/l, 0.69 mg TSS/mg CT and 0.046 day^–1, respectively for CT biotransformation during granulation.     

Treatment of catechol bearing wastewater in an upflow anaerobic sludge blanket (UASB) reactor: sludge characteristics

This paper deals with the characteristics of anaerobic microbial granules grown in an UASB reactor treating catechol bearing synthetic wastewater (SWW). The specific methanogenic activity of the sludge showed an increase in trend with an increase in the organic loading rate and the catechol concentration in the SWW. The settling velocity of individual granules in the size range of 0.5-2.5mm was found to be in the range of 30-75mh(-1). The ash content in the sludge was 11.7% with a sludge volume index of 18-20mlg(-1). The inorganic elemental distribution within the granules showed a decrease except that for phosphorous and cobalt, which increased by approximately 12% and 18%, respectively, after the treatment of SWW. Scanning electron microscopy (SEM) coupled with electron disperse X-ray analysis showed an increase in the sulphur content by approximately 300% after the treatment of SWW. Surface mineral composition of the granules determined by XRD analysis indicated the existence of vuagnatite (CaAlSiO(4)(OH)). SEM observation of the granules showed the predominance of Methanosaeta and Methanobacterium type of species on the surface along with a variety of other species.     

Biosorption and biodegradation of pentachlorophenol (PCP) in an upflow anaerobic sludge blanket (UASB) reactor

In order to understand the fate of PCP in upflow anaerobic sludge blanket reactor (UASB) more completely, the sorption and biodegradation of pentachlorophenol (PCP) by anaerobic sludge granules were investigated. The anaerobic granular sludge degrading PCP was formed in UASB reactor, which was seeded with anaerobic sludge acclimated by chlorophenols. At the hydraulic retention time (HRT) of 20–22 h, and PCP loading rate of 200–220 mg l⁻¹ d⁻¹, UASB reactor exhibited good performance in treating wastewater which containing 170–180 mg l⁻¹ PCP and the PCP removal rate of 99.5% was achieved. Sequential appearance of tetra-, tri-, di-, and mono-chlorophenol was observed in the reactor effluent after 20 mg l⁻¹ PCP introduction. Sorption and desorption of PCP on the anaerobic sludge granules were all fitted to the Freundlich isotherm equation. Sorption of PCP was partly irreversible. The Freundlich equation could describe the behavior of PCP amount sorbed by granular sludge in anaerobic reactor reasonably well. The results demonstrated that the main mechanism leading to removal of PCP on anaerobic granular sludge was biodegradation, not sorption or volatization.     

Anaerobic granule development for removal of pentachlorophenol in an upflow anaerobic sludge blanket (UASB) reactor

The granulation process using synthetic wastewater containing pentachlorophenol (PCP) in four 1.1 l laboratory scale upflow anaerobic sludge blanket (UASB) reactors was studied, and the anaerobic biotransformation of PCP during the granulation process investigated. After 110 days granular sludge was developed and up to 160 and 180 mg/l of PCP was added into the reactors R1 and R2, respectively, when they were inoculated with acclimated anaerobic sludge from an anaerobic digester of a citric acid plant. The inoculum was predominately composed of bacilli and filamentous bacteria. Granulation did not occur in reactors R3 and R4 which were inoculated with acclimated anaerobic sludge from aerobic sludge of the municipal sewage treatment plant which consisted mainly of cocci. Despite similar bacilli in the granule, the filamentous bacteria from reactor R1 were thicker than those of reactor R2. The granular sludge had a maximum diameter of 2.5 and 2.2 mm, and SMA of 1.44 and 1.32 gCOD/gTVS per day for reactors R1 and R2, respectively. Over 98% chemical oxygen demand (COD) removal rate and 99% of PCP removal rate were achieved when reactors R1 and R2 were operated at PCP and COD loading rates of 150 and 7.5 g/l per day, respectively. H₂-producing acetogens were the dominant anaerobes in the granular sludge.     

Characteristics of methanogenic granules grown on propionate in an upflow anaerobic sludge blanket (UASB) reactor

Granulation of a propionate-degrading consortia was performed with a mesophilic propionate-acclimatized sludge in an upflow anaerobic sludge blanket (UASB) reactor. The granules formed were relatively small, ranging mainly from 0.3 to 0.6 mm in diameter, but had an excellent sedimentation velocity due to a high specific gravity of 1.355 g/cm³ (ash content, 48.2%). The ash consisted mainly of calcium (30.2%), phosphorus (19.7%), and magnesium (3.95%) forming plate crystals in the granules. The populations of three bacterial trophic groups present in the granules, propionate-degraders, hydrogenotrophic and aceticlastic methanogens were 5.6 × 10⁸, 1.6 × 10¹⁰, and 2 × 10⁹ (in most probable number/g mixed-liquor volatile suspended solids [MLVSS]), respectively, while the specific utilization rates of propionate, hydrogen, and acetate of the granules were 9.4, 850, and 20.9 (mmol/g MLVSS·d), respectively. Electron microscopic analysis showed that Methanothrix spp. appeared dominant over the granules. Total granular sludge concentration retained in the UASB reactor during 178 d of operation was 80.0 g mixed-liquor suspended solids (MLSS)/l-reactor, corresponding to 41.4 g MLVSS/l-reactor, which realized a high-rate methanogenic fermentation of propionate of 85 g chemical oxygen demand (COD)/l-reactor·d.     

Performance and granulation in an upflow anaerobic sludge blanket (UASB) reactor treating saline sulfate wastewater

An upflow anaerobic sludge blanket reactor was employed to treat saline sulfate wastewater. Mesophilic operation (35 ± 0.5 °C) was performed with hydraulic retention time fixed at 16 h. When the salinity was 28 g L^?1, the chemical oxygen demand and sulfate removal efficiencies were 52 and 67 %, respectively. The salinity effect on sulfate removal was less than that on organics removal. The methane productions were 887 and 329 cm³ L^?1 corresponding to the NaCl concentrations of 12 and 28 g L^?1, respectively. High salinity could stimulate microbes to produce more extracellular polymeric substances (EPSs) and granulation could be performed better. Besides, with the high saline surroundings, a great deal of Na⁺ compressed the colloidal electrical double-layer, neutralized the negative charge of the sludge particles and decreased their electrostatic repulsion. The repulsion barrier disappeared and coagulation took place. The maximum size of granules was 5 mm, which resulted from the coupled triggering forces of high EPSs and Na⁺ contents. Sulfate-reducing bacteria (SRB) were dominant in the high saline surroundings while the methane-producing archaea dominated in the low saline surroundings. The SRB were affected least by the salinity.     

Methane fermentation of coastal mud sediment by a two-stage upflow anaerobic sludge blanket (UASB) reactor system

The removal of organic matter from a coastal mud sediment was carried out by a methane fermentation process under anaerobic conditions. In a batch acidogenic fermentation, the addition of vitamins containing thiamine, nicotinic acid and biotin dramatically enhanced acetate production from the mud sediment (200 g wet wt l(-1) artificial sea water), yielding 77 mM acetate after 6 days, which corresponded to 77% of the organic matter in the mud sediment, measured on the basis of chemical oxygen demand. Thereafter, the two-fold diluted, post-acidogenic fermentation liquor (PAF liquor) was continuously treated at 2.4x original dilution rate day(-1) for 30 days, using an upflow anaerobic sludge blanket methanogenic reactor containing the acclimated methanogenic sludge from the mud sediment. Acetate, 42 mM in the PAF liquor, was converted to methane at a maximum methane production rate of 96 mmol l(-1) day(-1); and 87.5% of the acetate and 88.7% of the total organic carbon in the PAF liquor were removed. Moreover, an efficient treatment of the mud sediment was carried out by a semi-continuous, two-stage reactor system, where the culture broth was circulated between acidogenic and methanogenic reactors. This two-stage reactor system gave a stable operation at 4-day intervals for one treatment period, yielding 112 mmol methane from the wet mud in the PAF liquor (278 g l(-1)).     

Extracellular polymers in granular sludge from different upflow anaerobic sludge blanket (UASB) reactors

Thermal extraction was used to quantify extracellular polymers (ECP) in granules from anaerobic upflow reactors. The optimal time for extraction was determined as the time needed before the intracellular material gives a significant contribution to the extracted extracellular material due to cell lysis. ECP contents of 41 to 92 mg · g^–1 volatile suspended solids of granules were found depending on the type of granular sludge examined. The content of polysaccharides, protein and lipids in the extracted ECP was quantified. Furthermore, the different methyl esters of the lipids were determined and quantified. Lower amounts of polysaccharides and proteins were found in the extracellular material from granules grown on methanogenic and acetogenic substrates compared to granules grown on more complex substrates. In contrast, the lipid content was lower on complex substrates. Changing the feed of an upflow anaerobic sludge blanket reactor from a sugar-containing waste-water to a synthetic waste-water containing acetate, propionate and butyrate resulted in a decrease in both the protein and polysaccharide content and an increase in the lipid content of the extracellular material. Furthermore, the amount of protein and polysaccharides in the ECP found under mesophilic conditions was significantly higher than under thermophilic conditions, while the lipid content was lower.     

Hydraulics of laboratory and full-scale upflow anaerobic sludge blanket (UASB) reactors

Laboratory-scale upflow anaerobic sludge blanket (UASB) reactors are often used as test platforms to evaluate full-scale applications. However, for a given volume specific hydraulic loading rate and geometry, the gas and liquid flows increase proportionally with the cube root of volume. In this communication, we demonstrate that a laboratory-scale reactor had plug-flow hydraulics, while a full-scale reactor had mixed flow hydraulics. The laboratory-scale reactor could be modeled using an existing biochemical model, and parameters identified, but because of computational speed with plug-flow hydraulics, mixed systems are instead recommended for parameter identification studies. Because of the scaling issues identified, operational data should not be directly projected from laboratory-scale results to the full-scale design.     

Improved performance of upflow anaerobic sludge blanket (UASB) reactors by operating alternatives

Summary The efficient operation of UASB reactors treating complex soluble wastewater containing high protein and lipid content was attempted by mixing in different modes. Higher superficial flow rate increased COD removal efficiency, sludge retainment, and methane content in biogas not only in the start-up period but also at high volumetric loading rates. However, formation of sludge particles in larger size was hindered by increased upflow liquid velocity.     

Start-up of a thermophilic upflow anaerobic sludge bed (UASB) reactor with mesophilic granular sludge

Summary Fast start-up of thermophilic upflow anaerobic sludge bed (UASB) reactors was achieved at process temperatures of 46, 55 and 64° C, using mesophilic granular sludge as inoculum and fatty acid mixtures as feed. The start-up was brought about by increasing the temperature of mesophilic UASB reactors in a single step, which initially led to a sharp drop in the methane production rate. Thereafter, stable thermophilic methanogenesis was achieved within a period of 1 or 2 weeks depending on the temperature of operation. Mesophilic granules functioned initially as effective carrier material for thermophilic organisms. However, long-term operation led to disintegration of the granules, resulting in wash-out of thermophilic biomass. The temperature optima for acetotrophic methanogenic activity of the sludges cultivated at 46, 55 and 64° C, were similar, but differed significantly from the temperature optimum of the mesophilic inoculum. All the sludges examined were dominated by Methanothrix-like rods. These could be distinguished by antigenic fingerprinting into two subpopulations, one predominant at 36° C and the other predominant at 46° C and above. Offprint requests to: J. B. van Lier     

Anaerobic biodegradation of linear alkylbenzene sulfonate (LAS) in upflow anaerobic sludge blanket (UASB) reactors

The anaerobic biodegradation of Linear Alkylbenzene Sulfonate (LAS) was studied in Upflow Anaerobic Sludge Blanket Reactors (UASB). One reactor was fed with easily degradable substrates and commercial LAS solution during a period of 3 months (Reactor 1), meanwhile a second reactor was fed with a commercial LAS solution without co-substrate (Reactor 2) during 4 months. Both reactors were operated with an organic loading rate of 4–5 mg-LAS/l*day and a hydraulic retention time of one day.The LAS biodegradation was determined by full mass balance. LAS was analysed by HPLC in the liquid phase (influent and effluent streams of the reactors) as well as in the solid phase (granular sludge used as biomass). The results indicate a high level of removal (primary biodegradation: 64–85%). Biodegradation was higher in the absence of external co-substrates than in the presence of additional sources of carbon. This indicates that the surfactant can be partially used as carbon and energy source by anaerobic bacteria. Under the operating conditions used, inhibition of the methanogenic activity or any other negative effects on the biomass due to the presence of LAS were not observed. The methanogenic activity remained high and stable throughout the experiment.     

Continuous dechlorination of tetrachloroethene in an upflow anaerobic sludge blanket reactor

Influences of hydraulic retention time (HRT) on dechlorination of tetrachloroethene (PCE) were investigated in an upflow anaerobic sludge blanket (UASB) reactor inoculated with anaerobic granular sludge non-pre-exposed to chlorinated compounds. PCE was introduced into the reactor at a loading rate of 3 mg/l d. PCE removal increased from 51 +/- 5% to 87 +/- 3% when HRT increased from 1 to 4 d, corresponding to an increase in the PCE biotransformation rate from 10.5 +/- 2.3 to 21.3 +/- 3.7 mumol/d. A higher ethene production rate, 0.9 +/- 0.2 mumol/d, was attained without accumulation of dichloroethenes at the HRT of 4 d. Dehalococcoides-like species were detected in sludge granules by fluorescence in situ hybridization, with signal strength in proportion to the extent of PCE dechlorination.     

Assessment of active methanogenic archaea in a methanol-fed upflow anaerobic sludge blanket reactor

Methanogenic archaea enrichment of a granular sludge was undertaken in an upflow anaerobic sludge blanket (UASB) reactor fed with methanol in order to enrich methylotrophic and hydrogenotrophic methanogenic populations. A microbial community assessment, in terms of microbial composition and activity—throughout the different stages of the feeding process with methanol and acetate—was performed using specific methanogenic activity (SMA) assays, quantitative real-time polymerase chain reaction (qPCR), and high-throughput sequencing of 16S ribosomal RNA (rRNA) genes from DNA and complementary DNA (cDNA). Distinct methanogenic enrichment was revealed by qPCR of mcrA gene in the methanol-fed community, being two orders of magnitude higher with respect to the initial inoculum, achieving a final mcrA/16S rRNA ratio of 0.25. High-throughput sequencing analysis revealed that the resulting methanogenic population was mainly composed by methylotrophic archaea (Methanomethylovorans and Methanolobus genus), being also highly active according to the RNA-based assessment. SMA confirmed that the methylotrophic pathway, with a direct conversion of methanol to CH₄, was the main step of methanol degradation in the UASB. The biomass from the UASB, enriched in methanogenic archaea, may bear great potential as additional inoculum for bioreactors to carry out biogas production and other related processes.     

Poultry slaughter wastewater treatment with an up-flow anaerobic sludge blanket (UASB) reactor

Removal of organic material from poultry slaughter wastewater as determined by changes in biological oxygen demand (BOD5) was investigated by adding three different types of inoculum combining cow manure, yeast extract or hydraulic residence time as variables with response vector of reduction of BOD5. In a 3-l reactors, a 95% removal of BOD5 from poultry slaughter wastewater was obtained with organic loading rates up to 31 kg BOD5 m(-3) d(-1) without loss of stability. This 95% removal was obtained between 25 and 39 degrees C with a hydraulic residence time between 3.5 and 4.5 h. The growth of the consortium of micro-organisms in the reactor followed a first-order kinetic with a constant specific growth rate of 0.054 h(-1). It was concluded that an inoculum from cow manure added with nutrients and yeast extract allowed a 95% removal of BOD5 from poultry slaughter wastewater at ambient temperatures within a hydraulic residence time of 4 h, sharply reducing possible environmental hazards.     

Toluene mineralization by denitrification in an up flow anaerobic sludge blanket (UASB) reactor

In order to examine the effect of easily degradable substrate such as acetate on toluene mineralization by denitrification, an upflow anaerobic sludge blanket (UASB) reactor in steady state was set up. The experimentation was carried out in two stages. Initially, the reactor was fed with a carbon loading rate of 250 mg acetate-C L-1 d-1 as electron source. Nitrate loading rate (mg ) was adjusted to obtain a constant C/N ratio of 1.4. In the second stage, five toluene-C loading rates (TLR, mg toluene-C L-1 d-1), 25, 50, 75, 100 and 125, were assessed while total carbon loading rate and C/N were maintained constant at 250 mg C L-1 d-1 and 1.4, respectively. In so doing, acetate-C loading rate (mg acetate-C L-1 d-1) was gradually substituted by toluene-C. When acetate-C was the only electron source a dissimilative denitrifying process resulted as indicated by bicarbonate yield YHCO3, mg produced/mg carbon consumed) of 0.74 +/- 0.005 and denitrifying yield (YN2, mg N2 produced/mg consumed) of 0.89 +/- 0.042. The addition of different TLR did not affect the biological process as consumption carbon efficiency (CCE) values remained up to 95% +/- 3.5 and YHCO3 and YN2 values were higher than 0.71 +/- 0.03 and 0.88 +/- 0.01, respectively. Toluene mineralization by denitrification in continuous culture was successfully achieved. A simple UASB denitrifying reactor system has promising applications for complete conversion of nitrate, toluene and acetate into N2 and CO2 with a minimal sludge production.     

The settling characteristics and mean settling velocity of granular sludge in upflow anaerobic sludge blanket (UASB)-like reactors

Mean settling velocity of granular sludge in full-scale UASB (upflow anaerobic sludge blanket) and EGSB (expanded granular sludge bed) reactors was evaluated by settling column tests, and a settling velocity model based on the experimental results and available literature data was developed. It is concluded that the settling velocity should be calculated by the Allen formula, because the settling process of the granules is in the category of intermediate flow regime rather than in the laminar flow one. The comparison between calculated and measured values of the settling velocity shows an excellent agreement, with an average relative error of 4.04%. A simple but reliable mathematical method to determine the settling velocity is therefore proposed.