Thermophilic Digestion Research Articles

Taking Advantage of Waste Heat Resource from Vinasses for Anaerobic Co-digestion of Waste Activated Sludge under the Thermophilic Condition: Energy Balance and Kinetic Analysis.

Vinasses are not only an easily biodegradable substrate but also a heat energy resource. In this study, the energy balance and kinetic model of anaerobic co-digestion of waste activated sludge (WAS) with vinasses have been investigated in semicontinuous reactor experiments at 55 °C. Herein, the maximum energy balance value, the ratio of energy to mass, and the kinetic constants μmax and K of anaerobic digestion of WAS were −33.44 kJ·day–1, −5.72 kJ·VS–1·day–1, and 0.0894 day–1 and 0.7294, respectively, at an organic loading rate (OLR) of 1.17 VS·L–3·day–1; when the mixture ratio of WAS to vinasses was 2:1 (dry VS) for co-digestion, the maximum energy balance value, the maximum ratio of energy to mass, and the kinetic constants μmax and K of anaerobic co-digestion of WAS and vinasses were +39.73 kJ·day–1, 8.1 kJ·VS–1·day–1, and 0.2619 day–1 and 1.9583, respectively, at an OLR of 1.73 VS·L–3·day–1. The positive energy balance was obtained for two reasons: one is for making the best use of the high-temperature heat energy resource of vinasses and the other is for enhancing the amount of biogas yield. The bottleneck of the negative energy balance of thermophilic digestion of WAS can be broken by anaerobic co-digestion of WAS and vinasses. The results indicate a promising future in the application of anaerobic thermophilic co-digestion of WAS and vinasses. Methane production from digestion and co-digestion was also predicted by the Chen–Hashimoto kinetic model.

Conventional and Innovative Hygienization of Feedstock for Biogas Production: Resistance of Indicator Bacteria to Thermal Pasteurization, Pulsed Electric Field Treatment, and Anaerobic Digestion

Animal by-products (ABP) can be valorized via anaerobic digestion (AD) for biogas energy generation. The digestate issued from AD process is usually used to fertilize farming land for agricultural activities, which may cause potential sanitary risk to the environment. The European Union (EU) requires that certain ABP be thermally pasteurized in order to minimize this sanitary risk. This process is called hygienization, which can be replaced by alternative nonthermal technologies like pulsed electric field (PEF). In the present study, Enterococcus faecalis ATCC 19433 and Escherichia coli ATCC 25922 were used as indicator bacteria. Their resistance to thermal pasteurization and PEF treatment were characterized. Results show that Ent. faecalis and E. coli are reduced by 5 log10 in less than 1 min during thermal pasteurization at 70 °C. The critical electric field strength was estimated at 18 kV∙cm−1 for Ent. faecalis and 1 kV∙cm−1 for E. coli. “G+” bacteria Ent. faecalis are generally more resistant than “G−” bacteria E. coli. AD process also plays an important role in pathogens inactivation, whose performance depends on the microorganisms considered, digestion temperature, residence time, and type of feedstock. Thermophilic digestion is usually more efficient in pathogens removal than mesophilic digestion.

Vivianite scaling in wastewater treatment plants: Occurrence, formation mechanisms and mitigation solutions

The presence of soluble iron and phosphorus in wastewater sludge can lead to vivianite scaling. This problem is not often reported in literature, most likely due to the difficult identification and quantification of this mineral. It is usually present as a hard and blue deposit that can also be brown or black depending on its composition and location. From samples and information gathered in 14 wastewater treatment plants worldwide, it became clear that vivianite scaling is common and can cause operational issues. Vivianite scaling mainly occurred in 3 zones, for which formation hypotheses were discussed. Firstly, iron reduction seems to be the trigger for scaling in anaerobic zones like sludge pipes, mainly after sludge thickening. Secondly, pH increase was evaluated to be the major cause for the formation of a mixed scaling (a majority of oxidized vivianite with some iron hydroxides) around dewatering centrifuges of undigested sludge. Thirdly, the temperature dependence of vivianite solubility appears to be the driver for vivianite deposition in heat exchanger around mesophilic digesters (37 °C), while higher temperatures potentially aggravate the phenomenon, for instance in thermophilic digesters. Mitigation solutions like the use of buffer tanks or steam injections are discussed. Finally, best practices for safe mixing of sludges with each other are proposed, since poor admixing can contribute to scaling aggravation. The relevance of this study lays in the occurrence of ironphosphate scaling, while the use of iron coagulants will probably increase in the future to meet more stringent phosphorus discharge limits.

Thermophilic rather than mesophilic sludge anaerobic digesters possess lower antibiotic resistant genes abundance

For exploring the impact of temperature on antibiotic resistant genes (ARGs) during sludge anaerobic digestion (AD), the dynamic variations of sludge ARGs, plasmid ARGs, and cell-free ARGs in mesophilic (25 °C and 35 °C) and thermophilic (55 °C) digesters were investigated. The study revealed that the abundance of sludge ARGs and plasmid ARGs in thermophilic sludge AD was significantly lower than that in mesophilic digesters, while the cell-free ARGs abundance of the thermophilic digesters was similar to mesophilic digesters. Higher archaea abundance, lower bacteria abundance, and different microbial community were found in thermophilic digesters compared to that of mesophilic ones. Firmicutes might be a main group of potential hosts of ARGs in sludge AD. The distinct microbial community was the main contributor to the low ARGs abundance in thermophilic sludge AD. Thermophilic operation at 55 °C rather than mesophilic operation is more conducive to control ARGs in sludge anaerobic digestion.

The screening of early warning indicators and microbial community of chicken manure thermophilic digestion at high organic loading rate

Anaerobic digestion of chicken manure (CM) with high nitrogen content is easily suffered inhibition of ammonia, especially under thermophilic and high organic loading rate (OLR) condition. To screen the early warning indicators for instability, thermophilic digestion of CM was conducted in a bench-scale reactor (working volume 55 L) under OLR from 1.0 to 6.0 g VS/(L·d). Microbial community analysis was used to reveal the mechanism of instability at high OLR. The system collapsed at an OLR of 6.0 g VS/(L·d). The initial ammonia inhibition evoked serious acidification, subsequently, the dual suppression of free volatile fatty acids and free ammonia caused the final collapse. The most likely mechanism of iso-butyrate degradation was propionate pathway. The ratio of n-butyrate to iso-butyrate was screened as an early warning indicator with the threshold of above 0.45. The high OLR of 6.0 g VS/(L·d) broke the balance of syntrophic oxidation bacteria (norank_o__MBA03, Gelria, and norank_o__D8A-2), hydrogenotrophic methanogens (Methanothermobacter) and fermentative and acid-producing bacteria (Ruminiclostridium_1, Bacteroides, Defluviitalea, norank_o__Clostridia, and norank_o__M55-D21). The non-functional acid tolerant bacteria (Defluviitoga, norank_f__Family_XI, Corynebacterrium_1, Caldicoprobacter, Lactobacillus, and Erysipelothrix) became the dominant after the system collapsed.

Comparison of microbial community structures between mesophilic and thermophilic anaerobic digestion of vegetable waste.

The anaerobic digestion performance correlates with the functional microbial community. Mesophilic and thermophilic digestions of vegetable waste were conducted, and dynamics of the microbial community were investigated. The mesophilic and thermophilic collapsed stages occurred at organic loading rates of 1.5 and 2.0g VS/(Ld) due to the accumulation of volatile fatty acids with final concentrations of 2276 and 6476mg/L, respectively. A high concentration of volatile fatty acids caused the severe inhibition of methanogens, which finally led to the imbalance between acetogenesis and methanogenesis. The mesophilic digestion exhibited a higher microbial diversity and richness than the thermophilic digestion. Syntrophic acetate-oxidizing coupled with hydrogenotrophic methanogenesis was the dominant pathway in the thermophilic stable system, and acetoclastic methanogenesis in the mesophilic stable system. The dominant acidogens, syntrophus, and methanogens were unclassified_f__Anaerolineaceae (8.68%), Candidatus_Cloacamonas (19.70%), Methanosaeta (6.10%), and Methanosarcina (4.08%) in the mesophilic stable stage, and Anaerobaculum (12.59%), Syntrophaceticus (4.84%), Methanosarcina (30.58%), and Methanothermobacter (3.17%) in thermophilic stable stage. Spirochaetae and Thermotogae phyla were the characteristic microorganisms in the mesophilic and thermophilic collapsed stages, respectively. These findings provided valuable information for the deep understanding of the difference of the microbial community and methane-producing mechanism between mesophilic and thermophilic digestion of vegetable waste.

Development of a novel reactor for simultaneous production of biogas from oil-palm empty fruit bunches (EFB) and palm oil mill effluents (POME)

A novel reactor was developed in this research to simultaneously produce biogas from solid- and liquid-state anaerobic digestions in one container for the first time. Excess heat from the outer liquid-state anaerobic digestion (OL-AD) of POME was transferred to the inner solid-state anaerobic digestion (IS-AD) of EFB for promoting thermophilic digestions (55 °C and 70 °C). Continuous operations at various hydraulic retention times were performed. Bacteria and archaea communities were identified. The results showed that the maximum methane yields were 507.07 and 125.84 mL-CH4/g-VS for OL-AD and IS-AD, respectively. In OL-AD, raising temperature increased the methane yield by 6.4%. In IS-AD, the yield was not significantly affected by elevated temperature. However, the pretreatment of EFB with NaOH increased the methane yield by 14.1%. The main bacteria and archaea communities in OL-AD and IS-AD were found similar. The novel reactor could save energy, reduce costs and digester space, and increase the methane yield.

Evolution of the microbial community structure in biogas reactors inoculated with seeds from different origin

The aim of this work was to provide solid proofs regarding the achievement of “steady-state conditions”, which means that the performance of the anaerobic digester is representative of the applied environmental conditions. For this reason, we investigated how, starting from different inoculum sources (i.e., municipal wastewater treatment, bio-waste treatment, and agricultural waste biogas plant), the microbial community adapted to the operational parameters and led to stable biogas production in thermophilic digesters treating the same influent feedstock. The results revealed that the different system achieved similar process performance and microbial community structure after a period that was equal to four hydraulic retention times, approved by a constant pH of 7.89 ± 0.08, 7.92 ± 0.05 and 7.85 ± 0.08, respectively, and stable TAN concentration of 1500 mg/L. Moreover, it was found that the microbial composition of the inocula was a key factor for the speed of achieving stable process performance; thus, a pre-adapted to the influent feedstock inoculum can shorten the stabilization process. On the contrary, after long term reactor operation, the microbial structure was shaped according to the chemical composition of the influent feedstock. The results of the study can also be used as a guide in future researches on anaerobic degradation, particularly in determining the time interval of an experiment to reflect changes in the microbial community of anaerobic digester.

Characterizing the growing microorganisms at species level in 46 anaerobic digesters at Danish wastewater treatment plants: A six-year survey on microbial community structure and key drivers

Anaerobic digestion (AD) is a key technology at many wastewater treatment plants (WWTPs) for converting primary and surplus activated sludge to methane-rich biogas. However, the limited number of surveys and the lack of comprehensive datasets have hindered a deeper understanding of the characteristics and associations between key variables and the microbial community composition. Here, we present a six-year survey of 46 anaerobic digesters, located at 22 WWTPs in Denmark, which is the first and largest known study of the microbial ecology of AD at WWTPs at a regional scale. For three types of AD (mesophilic, mesophilic with thermal hydrolysis pretreatment, and thermophilic), we present the typical value range of 12 key parameters including operational variables and performance parameters. High-resolution bacterial and archaeal community analyses were carried out at species level using amplicon sequencing of >1,000 samples and the new ecosystem-specific MiDAS 3 reference database. We detected 42 phyla, 1,600 genera, and 3,584 species in the bacterial community, where 70% of the genera and 93% of the species represented environmental taxa that were only classified based on MiDAS 3 de novo placeholder taxonomy. More than 40% of the bacterial species were found not to grow in the mesophilic and thermophilic digesters and were only present due to immigration with the feed sludge. Ammonium concentration was the main driver shaping the bacterial community while temperature and pH were main drivers for the archaea in the three types of ADs. Sub-setting for the growing microbes improved significantly the correlation analyses and revealed the main drivers for the presence of specific species. Within mesophilic digesters, feed sludge composition and other key parameters (organic loading rate, biogas yield, and ammonium concentration) correlated with specific growing species. This survey provides a comprehensive insight into community structure at species level, providing a foundation for future studies of the ecological significance/characteristics and function of the many novel or poorly described taxa.

Co-digestion of Dairy Cattle Waste in a Pilot-Scale Thermophilic Digester Adapted to Poultry Litter Feedstock: Stress, Recovery, and Microbiome Response

The adaptability of a thermophilic anaerobic digester stabilized long-term with poultry litter as sole feedstock was measured with regard to co-digestion with dairy cattle manure. Both feedstocks have low C/N ratios but differ in carbohydrates, protein, and fiber content. The digester was a pilot-scale (40 m3) sCSTR (semi-continuously stirred tank reactor) with a 20-day hydraulic retention time (HRT). Dairy manure was added as three step-wise increases in concentration: 20, 40, and 80%. At 20% dairy manure, methane production was unchanged (17–19 m3 day−1). But as the fraction of dairy manure increased, methane, ammonia, sCOD, acetate, and propionate decreased. At 80% dairy manure, methane volume dropped by 60%. Acetate concentration decreased from an initial 878 ± 209 mg L−1 during poultry litter mono-digestion to 285 ± 169 mg L−1 with 80% dairy manure. Upon returning to mono-digestion, methane production was on a trajectory of recovery although acetate remained low. Bacterial community structure shifted during co-digestion, but community resilience was also underway following return to the original feedstock for one HRT. The dominant bacterial phylotypes belonged to the groups Thermotogales, Synergistales, Clostridiales, and Halanaerobiales, and three uncharacterized orders (MBA08, SHA-98, OPB54) in class Clostridia. Co-digestion led to a shift in dominance with reductions in Thermotogales and OPB54, and an increase in MBA08. This experiment provides guidance for digester operators who wish to mix livestock and poultry manures for thermophilic co-digestion.

Entomological approach to the impact of ionophore-feed additives on greenhouse gas emissions from pasture land in cattle.

The suppressive effect of monensin as an ionophore-feed additive on enteric methane (CH4) emission and renewable methanogenesis were evaluated. To clarify the suppressive effect of monensin a respiratory trial with head cage was performed using Holstein-Friesian steers. Steers were offered high concentrate diets (80% concentrate and 20% hay) ad libitum with or without monensin, galacto-oligosaccharides (GOS) or L-cysteine. Steers that received monensin containing diet had significantly (p < 0.01) lower enteric CH4 emissions as well as those that received GOS containing diet (p < 0.05) compared to steers fed control diets. Thermophilic digesters at 55°C that received manure from steers fed on monensin diets had a delay in the initial CH4 production. Monensin is a strong inhibitor of enteric methanogenesis, but has a negative impact on biogas energy production at short retention times. Effects of the activity of coprophagous insects on CH4 and nitrous oxide (N2O) emissions from cattle dung pats were assessed in anaerobic in vitro continuous gas quantification system modified to aerobic quantification device. The CH4 emission from dungs with adults of Caccobius jessoensis Harold (dung beetle) and the larvae of the fly Neomyia cornicina (Fabricius) were compared with that from control dung without insect. The cumulative CH4 emission rate from dung with dung insects decreased at 42.2% in dung beetles and 77.8% in fly larvae compared to that from control dung without insects. However, the cumulative N2O emission rate increased 23.4% in dung beetles even though it reduced 88.6% in fly larvae compared to dung without coprophagous insects. It was suggested that the antibacterial efficacy of ionophores supplemented as a growth promoter still continued even in the digested slurry, consequently, possible environmental contamination with the antibiotics might be active to put the negative impact to land ecosystem involved in greenhouse gas mitigation when the digested slurry was applied to the fields as liquid manure.

Enhancement of organic household waste anaerobic digestion performances in a thermophilic pilot digester

Performances of organic household waste anaerobic digestion was investigated in a new horizontal thermophilic pilot digester of 50 L total capacity. The operation was performed under favorable conditions without any chemical additives. The presence of predigested cattle manure in the digester provided an interesting buffer capacity that was able to balance the produced fatty acids during the digestion process.The biomethane production reached 0.44 m3 CH4/Kg VS while achieving a COD removal of 70%.The operating time and the anaerobic digestion steps were shortened under thermophilic wet digestion. Only 4 days were sufficient to reach the daily maximum methane production (0.084 m3 CH4/(kgVS.day)) and 10 days to reach 80% of the overall biomethane potential. The process efficiency in removing volatile solid was found to be more than 35% within 5 days.Therefore, wet anaerobic digestion under thermophilic temperature is suitable for industrial scale anaerobic digestion of biowaste regarding the high biomethane yield, the large COD and solid removing and the shortness of process operating time.

Optimisation of methane fermentation as a valorisation method for food waste products

The aim of the study was to optimize methane fermentation of food waste products based on the composition of substrates mixture and technical parameters of anaerobic digestion to obtain the highest biogas and methane production. The study was conducted in the laboratory scale in the mesophilic (37°C) and thermophilic (55°C) conditions. Also the effects of the organic loading rates (OLRs) on the biogas productivity and composition were analysed. The highest biogas yield was obtained in mesophilic digestion of the substrates mixture containing 50% of meat, 40% dairy products, 10% fruit and vegetables. It amounted to 740.4 ± 19.9 cm3·g ODM−1 of biogas with 68.6 ± 1.8% of methane. Biogas potential in thermophilic digestion was from 2.1 to 2.8 times lower for analogical feedstock. It was caused by high concentration of volatile fatty acids (VFAs) in the reactor chamber. The ratio of VFAs to alkalinity was on the level from 0.5 ± 0.1 to 0.8 ± 0.1. In continuous experiments, the average biogas production rate was from 2646.9 ± 117.3 cm3·d−1 for OLR of 2 g ODM·dm−3·d−1 to 9893.4 ± 362.0 cm3·d−1 for OLR amounting to 8 g ODM·dm−3·d−1. Methane content in biogas was between 65.6 ± 2.5 and 65.5 ± 1.9%, respectively. It was confirmed that applying OLR above 6 g ODM·dm−3·d−1 causes inhibition of methane fermentation.

Production of Biosolids by Autothermal Thermophilic Aerobic Digestion (ATAD) from a Municipal Sewage Sludge: The Polish Case Study

This manuscript analyzed the process of autothermal thermophilic aerobic digestion (ATAD) used in installations of municipal sewage treatment plants in Poland. Additionally, solutions for sludge management and the parameters of operating installations were presented. Attention was also put to the energy consumption of the process, where the energy consumption for 1 m3 of sludge treated was between 18.4 and 27.79 kWh. The amount of sewage flowing into the analyzed plants was between 1500 and 14,000 m3/d. On the basis of research carried out in the years 2003–2019 in the selected plants, the characteristics of sludge after the ATAD process were presented. The parameters that determine the usefulness of the sludge as an organic fertilizer were indicated above all. The content of total nitrogen, which was from 2.4 to 8.1% of dry matter, ammonium nitrogen, which was from 0.8 to 1.8% of dry matter, and total phosphorus, which was from 1.1 to 4.2% of dry matter, recommended using sludge for fertilization. It was also pointed out that sewage sludge should be regularly tested for the dynamics of changes in chemical composition and biological parameters. These are the factors that increase risk and limit the use of sludge for fertilization.

The transition temperature (42 °C) from mesophilic to thermophilic micro-organisms enhances biomethane potential of corn stover

Mesophilic and thermophilic digestion has long been considered as preferred temperature ranges for anaerobic digestion. However, in this study, the effects of temperatures (37, 42, 47, and 55 °C) on the biomethane potential of corn stover were conducted with batch experiments, and the highest biomethane potential was at 42 °C. It was inferred that the change of feed materials, e.g., pretreatment caused by acidification (pH 6.0) during the lag time (4 days), was the main driver for higher biomethane potential. The natural pretreatment stimulated by a slight digestive temperature increase to 42 °C can enhance the biomethane potential of corn stover without adding extra acid. Meanwhile, metabolic pathways of methanogens changed from acetoclastic to mixotrophic and hydrogenotrophic methanogenesis. Based on these results, the transition temperature (42 °C) from mesophilic to thermophilic micro-organisms could be a promising option for corn stover anaerobic digestion.

Stress induced by crude glycerol in a thermophilic digester: microbial community divergence and resilience, but slow process recovery.

Recovery from stress is an important property for anaerobic digestion (AD). Although AD is quite adaptable with regard to waste composition, new substrates added to stable systems may cause process decline. We tested whether crude glycerol would cause stress to a thermophilic AD microbiome previously stabilized long-term on a low C/N ratio feedstock. Three-percent (v/v) crude glycerol was added to the basal substrate (poultry litter) for two hydraulic retention time (HRT) periods. This caused stress where biogas volume and methane percentage dramatically decreased and VFA levels increased. When the basal substrate was resumed, secondary inhibition occurred, resulting in even greater stress (biogas production ceased, methane 3.6%). Unassisted recovery of system processes required eight HRT periods. In contrast, crude glycerol applied at a lower organic loading rate did not cause inhibition. Crude glycerol caused changes in dominance in the microbial community (16S rRNA pyrotags). Although process resilience was slow, the recovery of digester functions occurred in conjunction with the recovery of community structure, particularly putative syntrophic acetate-oxidizing bacteria. KEY POINTS: • Crude glycerol caused stress in thermophilic co-digestion with poultry litter. • Unassisted resilience of digester functions (methane) required 8 HRT. • Syntrophic acetate-oxidizing bacteria implicated for keystone resilience functions. Graphical abstract.

Post-treatment of dewatered digested sewage sludge by thermophilic high-solid digestion for pasteurization with positive energy output

This study investigated the possibility to use thermophilic anaerobic high solid digestion of dewatered digested sewage sludge (DDS) at a wastewater treatment plant (WWTP) as a measure to increase total methane yield, achieve pasteurization and reduce risk for methane emissions during storage of the digestate. A pilot-scale plug-flow reactor was used to mimic thermophilic post-treatment of DDS from a WWTP in Linköping, Sweden. Process operation was evaluated with respect to biogas process performance, using both chemical and microbiological parameters. Initially, the process showed disturbance, with low methane yields and high volatile fatty acid (VFA) accumulation. However, after initiation of digestate recirculation performance improved and the specific methane production reached 46 mL CH4/g VS. Plug flow conditions were assessed with lithium chloride and the hydraulic retention time (HRT) was determined to be 19–29 days, sufficient to reach successful pasteurization. Degradation rate of raw protein was high and resulted in ammonia-nitrogen levels of up to 2.0 g/L and a 30% lower protein content in the digestate as compared to DDS. Microbial analysis suggested a shift in the methane producing pathway, with dominance of syntrophic acetate oxidation and the candidate methanogen family WSA2 by the end of the experiment. Energy balance calculations based on annual DDS production of 10000 ton/year showed that introduction of high-solid digestion as a post-treatment and pasteurization method would result in a positive energy output of 340 MWh/year. Post-digestion of DDS also decreased residual methane potential (RMP) by>96% compared with fresh DDS.

Mechanism of Organic Odor Generation from Thermophilically Digested Biosolids

In anticipation to generate more stabilized biosolids, thermophilic anaerobic digestion systems are widely used to destroy greater sludge organics thus making them less odorous. In this study, single-stage thermophilic (at 55 ℃) and mesophilic (at 37 ℃) anaerobic digesters were studied to compare organic removal efficiencies and sulfur-based odor generation from their biosolids. Although the thermophilic system removed about 9% more volatile solids than the mesophilic system, about 55 times more odorous organic sulfur compounds were measured from dewatered thermophilic biosolids. Different methanogenic species were found to be responsible for malodorous dewatered biosolids from the thermophilic anaerobic digester.

Characterization for the necessity of thermophilic biogas digester of tea waste and cooked waste for biogas production

&lt;p&gt;&lt;span&gt;Characterization of tea waste, cooked waste has been done by various authors but for the first time it has been done for understanding the necessity of thermophilic digestion. And for this kind of digestion takes place in thermophilic digester for efficient biogas production. Detailed morphological analysis of feedstock has been determined. In the present study, thermo gravimetric analysis carried out For easy and fast digestion of cooked waste, a novel design of thermophilic digester is proposed and tested.&lt;/span&gt;&lt;/p&gt;

Performance of Anaerobic Digestion of Acidified Palm Oil Mill Effluent under Various Organic Loading Rates and Temperatures

This study compared the performance of thermophilic and mesophilic digesters of an anaerobic digestion system from palm oil mill effluent (POME), in which temperature is a key parameter that can greatly affect the performance of anaerobic digestion. The digesters were incubated at two distinct temperatures of 55 and 37 °C, and operated with varying organic loading rates (OLRs) of 2.4, 3.2, and 4.0 g COD/L.d by altering the chemical oxygen demand (COD) of acidified POME during feeding. The results indicated that the performance of anaerobic digestion increased as the OLR increased from 2.4 to 4.0 g COD/L.d. At the OLR of 4.0 g COD/L.d, the thermophilic condition showed the highest methane yield of 0.31 ± 0.01 L/g COD, accompanied by the highest COD removal and volatile solid reduction, which were found to be higher than the mesophilic condition. Microbial community analysis via denaturing gradient gel electrophoresis (DGGE) revealed that Methanothermobacter sp. emerges as the dominant microbe, which is known to utilize the carbon dioxide pathway with hydrogen acting as an electron donor for methane formation