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The characteristics of sludge retained in a down-flow hanging sponge reactor were investigated to provide a better understanding of the sewage treatment process in the reactor. The organic removal and sulfur oxidation conditions were found to differ between the first layer and the following three layers. It was found that 63% and 59% of the organic matter was removed in the first layer, even though the hydraulic retention time was only 0.2 h. It is thought that the organic removal resulted from aerobic and anaerobic biodegradation on the sponge medium. The sulfate concentration increased 1.5–1.9-fold in the first layer, with almost no subsequent change in the second to fourth layers. It was shown that oxidation of sulfide in the influent was completed in the first layer. The result of the oxygen uptake rate test with an ammonium nitrogen substrate suggested that the ammonium oxidation rate was affected by the condition of dissolved oxygen (DO) or oxidation–reduction potential (ORP).

This study investigated down-flow hanging sponge (DHS) technology as a promising trickling filter (TF) using sponge media as a biomass carrier with an emphasis on protection of the biomass against macrofauna overgrazing. A pilot-scale DHS reactor fed with low-strength municipal sewage was operated under ambient temperature conditions for 1 year at a sewage treatment plant in Bangkok, Thailand. The results showed that snails (macrofauna) were present on the surface of the sponge media, but could not enter into it, because the sponge media with smaller pores physically protected the biomass from the snails. As a result, the sponge media maintained a dense biomass, with an average value of 22.3 gVSS/L sponge (58.1 gTSS/L sponge) on day 370. The snails could graze biomass on the surface of the sponge media. The DHS reactor process performance was also successful. The DHS reactor requires neither chemical treatments nor specific operations such as flooding for snail control. Overall, the results of this study indicate that the DHS reactor is able to protect biomass from snail overgrazing.

An up-flow anaerobic sludge blanket (UASB) – down-flow hanging sponge (DHS) was applied to Japanese municipal sewage treatment, and its treatability, energy consumption, and sludge production were evaluated. The designed sewage load was 50 m3/d. The sewage typically had a chemical oxygen demand (COD) of 402 mg/L, a suspended solids (SS) content of 167 mg/L, and a temperature of 17–29 °C. The UASB and DHS exhibited theoretical hydraulic retention times of 9.7 and 2.5 h, respectively. The entire system was operated without temperature control. Operation was started with mesophilic anaerobic digested sludge for the UASB and various sponge media for the DHS. Continuous operational data suggest that although the cellulose decomposition and methanogenic process in the UASB are temperature sensitive, stable operation can be obtained by maintaining a satisfactory sludge volume index and sludge concentration. For the DHS, the cube-type medium G3-2 offers superior filling rates, biological preservation and operational execution. The SS derived from the DHS contaminated the effluent but could be removed by optional sand filtration. A comparison with conventional activated sludge (CAS) treatment confirmed that this system is adequate for municipal sewage treatment, with an estimated energy requirement and excess sludge production approximately 75 and 85% less than those of CAS, respectively.

The biodegradation characteristics of palm oil mill effluent (POME) and the related microbial community were studied in both actual sequential anaerobic ponds in Malaysia and enrichment cultures. The significant degradation of the POME was observed in the second pond, in which the temperature was 35–37 °C. In this pond, biodegradation of major long chain fatty acids (LCFA), such as palmitic acid (C16:0) and oleic acid (C18:1), was also confirmed. The enrichment culture experiment was conducted with different feeding substrates, i.e. POME, C16:0 and C18:1, at 35 °C. Good recovery of methane indicated biodegradation of feeds in the POME and C16:0 enrichments. The methane production rate of the C18:1 enrichment was slower than other substrates and inhibition of methanogenesis was frequently observed. Denaturing gradient gel electrophoresis (DGGE) analyses indicated the existence of LCFA-degrading bacteria, such as the genus Syntrophus and Syntorophomonas, in all enrichment cultures operated at 35 °C. Anaerobic degradation of the POME under mesophilic conditions was stably processed as compared with thermophilic conditions.

The microbial community structures in the thermophilic anaerobic digestion (TAD) of waste activated sludge (WAS) and WAS were analyzed with molecular biological techniques including real-time polymerase chain reaction (PCR) and cloning. The microbial community of TAD had less diversity than that of WAS, and the sequences obtained in WAS were not present in TAD by the cloning analysis. In the TAD bacterial clone library, 97.5% of total clones were affiliated with the phylum Firmicutes and 73.1% with the genus Coprothermobacter. Real-time PCR and cloning analysis revealed that the number of Methanosarcina thermophila, which is an acetoclastic methanogen, is larger than that of Methanoculleus thermophilus, which is a hydrogenotrophic methanogen, in terms of the numbers of copies of 16S ribosomal DNA (rDNA).

A 2.0 L volume of EGSB reactor was operated at 20 degrees C for more than 500 days with 0.3-0.4 g COD/L of sucrose base wastewater to investigate the influence of effluent-recirculation on the process performance. At the start up period, the reactor was operated in EGSB mode with 5 m/h upflow velocity by continuous effluent recirculation. The COD loading was set to 7.2-9.6 kg COD/m(3) day with HRT of 1 hour. However, in this mode, EGSB reactor exhibited insufficient COD removal efficiency, i.e., 50-60%. Therefore, UASB mode (without recirculation, 0.7 m/h upflow velocity) was used for 30 minutes in every 40 minutes cycle to increase the COD concentration in the sludge bed. As a result, an excellent process performance was shown. The COD removal efficiency increased from 65% to 91% and the reactor could maintain a good physical property of retained sludge (sludge concentration: 33.4 g VSS/L and SVI: 25 mL/g VSS). Furthermore, retained sludge possessed sufficient level of methanogenic activity at 20 degrees C.

In this study, a lab scale EGSB reactor was operated for 400 days to investigate the influence of temperature-decrease on the microbial characteristic of retained sludge. The EGSB reactor was started-up at 15 degrees C seeding with 20 degrees C-grown granular sludge. The influent COD of synthetic wastewater was set at 0.6-0.8 gCOD/L. The process-temperature was stepwise reduced from 15 degrees C to 5 degrees C during 400 days operation. Decrease of temperature of the reactor from 15 degrees C to 10 degrees C caused the decline of COD removal efficiency. However, continuous operation of the EGSB reactor led the efficient treatment of wastewater (70% of COD removal, 50% of methane recovery) at 10 degrees C. We confirmed that the both acetate-fed and hydrogen-fed methanogenic activities of retained sludge clearly increased under 15 to 20 degrees C. Changes of microbial profiles of methanogenic bacteria were analyzed by 16S rDNA-targeted DGGE analysis and cloning. It shows that genus Methanospirillum as hydrogen-utilizing methanogen proliferated due to low temperature operation of the reactor. On the other hand, genus Methanosaeta presented in abundance as acetoclastic-methanogen throughout the experiment.

Molecular approaches were applied to identify and enumerate denitrifying bacteria subsisting in a fluidized bed reactor (FBR). The FBR was continuously operated as a unit for the removal of nitrogen from the effluents of domestic sewage treatment plant, with an additional supply of methanol as a carbon source. By denaturing gradient gel electrophoresis (DGGE) and sequence analysis of 16S ribosomal RNA genes, Thauera group was found to be dominant among the denitrifying bacteria in the FBR sludge. Oligonucleotide probe THA155 for fluorescence in situ hybridization (FISH) was newly designed for specifically targeting the Thauera group. However, the THA155 signal obtained from the sludge was only 0.9–5.7% of the DAPI-stained total cells. The real-time polymerase chain reaction (PCR) targeting the sequences of nitrite reductase (NIR) gene, a key enzyme of denitrification processes, was performed to quantify the cells of denitrifying bacteria cells including the Thauera group in FBR sludge. An excellent correlation was obtained between the numbers of nirS genes and the activity of denitrifiers in the FBR sludge.

The objective of this research is to make a novel wastewater treatment process activated by a sulfur-redox cycle action of microbes in low temperature conditions. This action is carried out by sulfate-reducing bacteria (SRB) and sulfur-oxidizing bacteria (SOB). The process was comprised of a UASB reactor as pre-treatment and an aerobic downflow hanging sponge (DHS) reactor as post-treatment. As the results of reactor operation, the whole process achieved that over 90% of CODcr removal efficiency, less than 30 mgCODcr/L (less than 15 mgBOD/L) of final effluent, at 12 h of HRT and at 8 °C of UASB reactor temperature. Acetobacterium sp. was detected as the predominant species by PCR-DGGE method targeting 16SrDNA with band excision and sequence analysis. In the UASB reactor, various species of sulfate-reducing bacterium, Desulfobulbus sp., Desulfovibrio sp., and Desulfomicrobium sp., were found by cloning analysis. In the DHS reactor, Tetracoccus sp. presented as dominant. The proposed sulfur-redox action process was considered as an applicable process for low strength wastewater treatment in low temperature conditions.

A pilot-scale multi-staged UASB (MS-UASB) reactor with a working volume of 2.5 m3 was operated for thermophilic (55°C) treatment of an alcohol distillery wastewater for a period of over 600 days. The reactor steadily achieved a super-high rate COD removal, that is, 60 kgCOD m−3 d−1 with over 80% COD removal. However, when higher organic loading rates were further imposed upon the reactor, that is, above 90 kgCOD m−3 d−1 for barely-based alcohol distillery wastewater (ADWW) and above 100 kgCOD m−3 d−1 for sweet potato-based ADWW, the reactor performance somewhat deteriorated to 60 and 70% COD removal, respectively. Methanogenic activity (MA) of the retained sludge in the thermophilic MS-UASB reactor was assessed along the time course of continuous run by serum-vial test using different substrates as a vial sole substrate. With the elapsed time of continuous run, hydrogen-utilizing MA, acetate-utilizing MA and propionate-fed MA increased at maximum of 13.2, 1.95 and 0.263 kgCOD kgVSS−1 d−1, respectively, indicating that propionate-fed MA attained only 1/50 of hydrogen-utilizing MA and 1/7 of acetate-utilizing MA. Since the ADWW applied herewith is a typical seasonal campaign wastewater, the influence of shut-down upon the decline in sludge MA was also investigated. Hydrogen-utilizing MA and acetate-utilizing MA decreased slightly by 3/4, during a month of non-feeding period, whereas propionate-fed MA were decreased significantly by 1/5. Relatively low values of propionate-fed MA and its vulnerability to adverse conditions suggests that the propionate degradation step is the most critical bottleneck of overall anaerobic degradation of organic matters under thermophilic condition.

This paper presents an evaluation of the process performance of a pilot-scale “fourth generation” downflow hanging sponge (DHS) post-treatment system combined with a UASB pretreatment unit treating municipal wastewater. After the successful operation of the second- and third-generation DHS reactors, the fourth-generation DHS reactor was developed to overcome a few shortcomings of its predecessors. This reactor was designed to further enhance the treatment efficiency and simplify the construction process in real scale, especially for the application in developing countries. Configuration of the reactor was modified to enhance the dissolution of air into the wastewater and to avert the possible clogging of the reactor especially during sudden washout from the UASB reactor. The whole system was operated at a total hydraulic retention time (HRT) of 8 h (UASB: 6 h and DHS: 2 h) for a period of over 600 days.The combined system was able to remove 96% of unfiltered BOD with only 9 mg/L remaining in the final effluent. Likewise, F. coli were removed by 3.45 log with the final count of 103 to 104 MPN/100 ml. Nutrient removal by the system was also satisfactory.

The transcription level of amoA mRNA encoding a subunit of ammonia monooxygenase (AMO) in ammonia-oxidizing bacteria (AOB) was quantified by reverse transcription-polymerase chain reaction (RT-PCR) methods in combination with real-time PCR technology. The effects of ammonia concentration and dissolved oxygen (DO) on the transcription levels of amoA mRNA and 16S rRNA in AOB were evaluated in batch experiments with nitrifying sludge taken from a lab-scale reactor treating artificial wastewater. A batch incubation without ammonia resulted in a rapid decrease, within four hours, in the transcription level of amoA mRNA to as low as 1/10 of that at the beginning of the experiment, while the 16S rRNA level in AOB was almost constant. After subsequent incubation with 3 mM ammonia for eight hours, a small increase in the transcription level of amoA mRNA occurred, but ammonia oxidation proceeded in the interim. Copy numbers of amoA mRNA showed an almost fixed value for over eight hours in the absence of dissolved oxygen.

The microbial community structure of mesophilic (35°C) and thermophilic (55°C) methanogenic granular sludges was surveyed by using both cultivation-independent molecular approach and conventional cultivation technique in order to address the fundamental questions on the microbial populations, i.e. who are present, where they are located, and what they are doing there. To elucidate the microbial constituents within both sludges, we first constructed 16S ribosomal DNA clone libraries, and partial sequencing of the clones was conducted for phylogenetic analysis. In this experiment, we found a number of unidentifiable clones within the domain Bacteria as well as clones that were closely related with 16S rDNAs of cultured microbes. The unidentifiable clones accounted for approximately 60–70% of the total clones in both mesophilic and thermophilic libraries. 16S rRNA-targeted in situ hybridization combined with confocal laser scanning microscopy was subsequently employed to examine where the uncultured populations were located within sludge granules. Spatial organization of uncultured microbes was visualized in thin-sections of both types of granules using fluorescent oligonucleotide probes, which were designed based on the clone sequences of certain novel clusters. This resulted in the detection of two types of uncultured cells in specific locations inside the granules. Finally, the goal-directed conventional cultivation technique was employed to recover such uncultured anaerobes and uncover their physiology and functions. In this approach, a total of five new species of thermophilic microorganisms were isolated, including several types of syntrophs and a novel sugar-fermenting bacterium. In the previous molecular approaches, all of these isolates were suggested to be significant populations within thermophilic granular sludge, hence obtaining these isolates in pure culture decreased the fraction of unknown clones in the previous thermophilic clone library from 70% to 40%. In conclusion, these approaches successfully revealed biodiversity and spatial organization of microbes of interest in sludge granules, and enlarged the fundamental knowledge of microbial constituents functioning as significant populations in the UASB processes.

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

Whole cell fluorescence in-situ hybridization (FISH) with 16S rRNA targeted oligonucleotides was applied to reveal the microbial ecological structure of UASB-grown granular sludge. The FISH analysis indicated that the members of the domain Archaea accounted for 28 to 53% of the total cells in various granular sludge sources, while Methanosaeta and Methanobacteriaceae cells accounted for 13 to 38%, and 4 to 27%, respectively. Methanosaeta cell density and Methanobacteriaceae cell density were strongly correlated, respectively, with acetate-utilizing methane production activity and with hydrogen-utilizing methane production activity.

Fluorescence in-situ hybridization (FISH) and DO microelectrodes were applied to biofilms developed in a novel reactor named DHS (downflow hanging sponge-cubes), to investigate the mechanisms of simultaneous carbon removal and nitrification. The DHS reactor was employed as an aerobic post-treatment process after an UASB anaerobic pre-treatment process receiving a municipal sewage. The presence ratio of Nitrosomonas and Nitrobacter cells to total cells of the DHS biomass was estimated by FISH technique to be 1.4% and 0.18%, respectively. Cell concentrations of both nitrifying bacteria were in good agreement with the magnitudes of ammonia-oxidizing and nitrite-oxidizing activities evaluated from batch tests. The habitats of both nitrifiers were the interior space of sponge-cubes, rather than within the biofilms attached onto sponge-cube surfaces. DO microelectrodes verify that the sponge-cubes insides were kept aerobically throughout the whole reactor height excluding the inlet vicinity portion.

A novel methodology is proposed in this study to evaluate the tensile strength of biofilms by using a newly developed tensile test device. Three different types of biofilms were prepared for tensile test: Run 1 biofilm was aerobically cultivated on glucose, and Run 2 and Run 3 biofilms were denitrifying biofilms grown on glucose and on methanol, respectively, as a sole carbon source. Each of three biofilms was formed on the outer surfaces of tygon-made tubes (diameter 4.76 mm) submerged in a rectangular open-channel reactor. The results demonstrated that the tensile strengths of three different biofilms had a tendency to increase with the biofilm development over a period of 50 days of reactor operation and eventually reached a similar range, approximately 500 to 1000 Pa, independent of biofilm types or cultivation conditions. An elastic coefficient of the biofilm was determined by assuming that a biofilm behaves as an elastic material during the tensile test. A strong positive correlation was found between the elastic coefficient and the tensile strength. However, the behavior of extracellular biopolymer (ECP) content and the biofilm density were not dependent on the tensile strength. Scanning electron microscope (SEM) observations also suggested that the biofilm structural properties such as elastic coefficient are more determinative factors for the tensile strength than are the biofilm physiological properties such as ECP content.

A novel methodology is proposed in this study to evaluate biofilm adhesion strength in two different ways: by measuring detached biomass caused by tensile force and by shear force. Tensile force was provided by centrifuging biofilm-attached plates installed on rotary tables. Shear force was provided by colliding biofilm-attached plates by gravity. Test biofilms consisting of denitrifiers were formed on the flat surfaces of square (25 cm2) plates that had been submerged in a rectangular open-channel reactor. The detachment tests revealed that, although biofilm adhesion strength was relatively high at the earlier growth stage, it drastically decreased at the later stage. The most weakened location toward biofilm depth was observed at the substratum surface, at which the adhesion strength by tensile force dropped from a several Pa to below 1 Pa as biofilms became aged. The adhesion strength by shear force was all the time more than 100 times as large as that by tensile force, even though having a similar behavior. The proportion of cavity, i.e., biofilm-absent area at the biofilm/substratum interface, increased as biofilms became mature. Cavity formation was strongly responsible for lessening the adhesion strength. It is suggested that biofilm slough-off is caused by the decline of adhesion strength by tensile force rather than by shear force.

In this study the in situ behavior of biofilm development and detachment was continuously observed by a video camera. The model biofilms consisting of denitrifiers were formed on the flat surfaces of polyvinyl-chloride plates that were placed in a rectangular open-channel reactor. The effects of intrinsic biofilm properties on biofilm detachment, such as intrafilm gas-vacuoles formation, extracellular biopolymer (ECP) content, biofilm density and cavity formation at the biofilm/substratum interface were quantified using attached-biofilms onto flat substrata. The result indicated that there were three phenotypes of massive biofilm detachment (see Fig. 4) under the conditions of low shear stress of flow. The first type showed that a large fraction (approximately 1/4-1/3) of the biofilm was sloughed off as a whole layer from the substratum surface at a relatively early stage of biofilm formation. This type was primarily caused by the cavity formation and by the decrease of ECP content. Succeeding to the first type, the second type occurred as fragmentary detachments. The second phenomenon was attributable to the increase in biofilm buoyancy due to the gas-vacuoles formation within the biofilm. The third type took place as the combination of the first and the second types following the occurrence of the second type. In contrast, at a higher flow shear stress only the second type of detachment became prevalent throughout the progression of biofilm formation.

A cross-flow ultrafiltration (UF) membrane separation was applied to anaerobic process for treatment of a wastewater containing high proportion of particulate COD. A synthetic wastewater with the total strength of 5000 mg COD·1−1 consisting of soluble and particulate COD (cellulose) in the ratio of 1:1 was fed to the reactor. The reactor was operated for 190 days at two loading rates, i.e. 1.5 and 2.5 kg COD·m−3·d−1. More than 98% of COD removal was consistently achieved throughout the duration, and the system was furthermore likely to accommodate much higher loading. Although the permeate COD was always kept at less than 80 mg COD·1−1, soluble COD of reactor broth accumulated up to 1200 mg COD·1−1. The biomass concentration saturated around 15 000 mg ML VSS·1−1. There was no tendency for cellulose to accumulate in the reactor over the whole period, which constituted only 1-2% of the total solids retained in the reactor. The methanogenic activity of the sludge increased 3.4 times for H2/CO2 and 10 times for acetate after 40 days operation. Afterwards, however, a further cultivation caused declines to 50% of the respective peak values for both substrates, because of a low sludge loading. The methanogenic activity using cellulose as a test substrate also exhibited a similar tendency. The membrane permeate flux deteriorated significantly with the cultivation time, owing to the change in rheological properties of the reactor mixed liquor that had been caused not only by increment of MLSS but also by accumulation of soluble high molecular organics.

In this study a novel methodology is proposed to estimate the adhesion strength of biofilm developed in an attached-growth reactor. The time variation of adhesion strength was quantified in the course of formation of a biofilm consisting of denitrifiers. The model biofilm was formed on the surfaces of polyvinyl-chloride plates placed in a rectangular open-channel reactor. The result indicates that the adhesion strength was not uniformly distributed throughout biofilm, but had a tendency to increase with biofilm depth and with the progression of biofilm formation. For example on the 32nd day from start-up the adhesion strength near the substratum interface attained a level in the order of 102 dyne·cm−2, whereas that in the vicinity of biofilm outer surface was only in the order of 100 dyne·cm−2. As well the biofilm dry density did not remain constant throughout biofilm, having a tendency similar to that of the adhesion strength. A strong correlation was observed between the dry density and the extracellular biopolymer content per unit biofilm volume. It was suggested that the adhesion strength was significantly affected by the dry density.

A laboratory-scale UASB reactor was operated at 55°C over 600 days in order to investigate the granulation mechanism under thermophilic conditions. The reactor was seeded with a thermophilic digested sewage sludge, and fed with a synthetic wastewater composed of sucrose and volatile fatty acids. As a result of establishment of the whole granulated sludge bed, the reactor allowed ultimately a volumetric COD loading of 45 kgCOD m−3 d−1 with 90% COD removal. The maximum sludge loading achieved was 3.7 gCOD·gVSS−1 d−1, that is two to three times as large as that of mesophilically grown sludge. The behavior of SRT (sludge retention time) was closely correlated with the biogas evolution flux: excessive biogas evolution at the highest COD loading (45 kgCOD m−3 d−1) enhanced the sludge washout, and thus led to shortening of SRT into less than 7 days. Methanogenic activities of the retained sludge increased finally up to 110 times for acetate, 25 times for propionate, and 3.6 times for hydrogen as large as those of the seeded sludge.

After seeded with mesophilically-grown (35°C) granular sludge, a laboratory-scale UASB reactor was thermophilically (55°C) operated over 8 months by feeding with an alcohol distillery wastewater. Use of mesophilically-grown granules as a seed material proved to be more advantageous for rapid and stable start-up of thermophilic UASB process, compared with the use of suspended-growth sludge taken from a thermophilic anaerobic digester. The reactor accommodated successfully a COD loading 30 kgCOD·m−3·d−1, with a COD removal efficiency of 85%. However, during a period of 30 kgCOD·m−3·d−1, propionate accumulated in the effluent up to 300-600 mgCOD·l−1. Thermophilic cultivation caused a drastic increase of methanogenic activities (55°C) of the retained sludge: 4.4 times for acetate, 4.6 times for propionate, and 3.5 times for hydrogen as large as those of the seed sludge. A considerably low value of propionate-fed methanogenic activity, i.e. only 1/5 of acetate-fed activity and only 1/23 of hydrogen-fed activity, suggested that the propionate degradation is subject to be a rate-limiting step in thermophilic anaerobic processes.

A novel sewage treatment system was proposed, which consists of a UASB anaerobic pre-treatment unit and the following DHS (downflow hanging sponge-cubes) aerobic post-treatment unit, as a low-cost and easy-maintenance process for developing countries. Over six months experiment by feeding sewage our proposed system achieved 94% of total-COD removal, 81% of soluble-COD removal, and nearly perfect SS removal and total-BOD removal at the overall HRT of 8.3 hr (7 hr in UASB and 1.3 hr in DHS unit). Moreover, the DHS reactor was capable of performing high (73–78%) nitrification. Our whole system requires neither external aeration input nor withdrawal of excess sludge.

The treatability of raw sewage in a temperate climate (wintertime around 10–20°C) using an upflow anaerobic sludge blanket (UASB) reactor and the hanging sponge cubes process was evaluated. After being seeded with digested sewage sludge, a 47.1 L UASB reactor was continuously operated for more than 2 years by feeding raw sewage, which had average COD around 300 mg/L (41% soluble). During summertime at an HRT of 7 h, effluent COD approximately 70 mg/L total, 50 mg/L soluble and BOD5 20 mg/L total, 12 mg/L soluble was obtained. During wintertime also, treatment efficiency and process stability was good. With the hanging sponge cubes process using the effluent of the UASB reactor treating raw sewage, the following results were obtained. The ammonia oxidation rates of 1.9 and 3.5 g NH4-N·m−2·d−1 in a downflow hanging sponge cubes biofilter, under natural air intake only were obtained during wintertime and summertime, respectively. With post-denitrification and an external carbon source, 84% in average N (NO3+NO2) was removed with an HRT of less than 1 hour and in the temperature range of 13 to 30°C using an upflow submerged hanging sponge bed bioreactor, under anaerobic conditions. The overall system using a UASB reactor and the hanging sponge cubes process could be quite an attractive treatment alternative.