Low Methane Yield Research Articles

Effects of Chlorella vulgaris, Nannochloropsis oceanica and Tetraselmis sp. supplementation levels on in vitro rumen fermentation

Microalgae constitute an underexploited feed resource for ruminant nutrition. The unique digestive tract of ruminants makes them best suited to utilise complex polysaccharides and non-protein nitrogen of eukaryotic microalgae, though contrasting effects on rumen fermentation have been reported. The current study aimed to assess the effects of microalgae species, inclusion level and basal substrates on in vitro rumen fermentation parameters. To this end, haylage and maize silage were supplemented with 2.5, 5 and 10% (w/w, dry matter basis) of Chlorella vulgaris, Nannochloropsis oceanica and Tetraselmis sp. and incubated for 24-h with rumen inocula non-adapted to microalgae. No significant interaction was observed among basal substrate, microalgae species and inclusion level for any of the parameters evaluated. Substrate and microalgae species interaction affected most fermentation parameters. The lowest gas and methane productions and yields, total volatile fatty acids (VFA) production and concentrations of acetic, butyric and most minor VFA observed when C. vulgaris was incubated with haylage suggest that rumen fermentation was inhibited to a greater extent by this microalgae, probably reflecting a low availability of rapidly fermentable carbohydrates due to differences in cell wall structure and composition. However, the opposite was observed when C. vulgaris was incubated with maize silage, suggesting an improvement in the synchrony between fermentable energy and degradable nitrogen. Microalgae species and inclusion level interaction mainly affected methanogenesis. The lowest methane yield with N. oceanica at 10% may be explained by its high content of polyunsaturated fatty acids. The interaction between basal substrate and microalgae inclusion level did not affect any parameter. Overall, the effects of the interactions between substrate and microalgae species and microalgae species and inclusion level, underpin the importance of identifying optimal microalgae species and inclusion levels according to the basal diet fed to ruminants.

Cow manure stabilizes anaerobic digestion of cocoa waste

Anaerobic digestion of a mono-feedstock often causes low methane yields and process instability. An effective strategy to overcome these barriers is co-digestion with animal manure. The obtained process improvement is often attributed to buffer capacity, nutrients, vitamins and trace metals, and microorganisms present in manure, but it remains unknown which factor plays the key role in digester performance. Here, we investigated anaerobic digestion of cocoa waste in four different treatments: mono-digestion, addition of synthetic nutrients, co-digestion with sterile cow manure, and co-digestion with raw cow manure. Co-digestion with raw manure resulted in the highest methane yield of 181 ± 39 L kg−1 VS (volatile solids), similar to the co-digestion with sterile manure, i.e., 162 ± 52 L kg−1 VS. The supplementation of synthetic nutrients to the anaerobic digestion of cocoa waste only temporarily increased methane yield, indicating that this will tackle a lack of nutrients in the short term, but has a limited long-term contribution to the stabilization of the process. Hence, because of the inability of synthetic nutrients to stabilize the digestion process and the similarity between the digesters fed sterile and raw manure, both at the physico-chemical and microbial level, the key contribution of manure co-digestion with cocoa seems to be the provision of buffering capacity.

Sustainable anaerobic digestion of euphorbiaceae waste for biogas production: Effects of feedstock variation

Anaerobic digestion (AD) refers to the biological process which can convert organic substrates to biogas in the absence of oxygen. The aim of this study was to determine the capability of feedstock to produce biogas and to quantify the biogas yield from different feedstocks. A co-digestion approach was carried out in a continuous stirred tank reactor operated under mesophilic conditions and at a constant organic loading rate of 0.0756 g COD/ L.day, with a hydraulic retention time of 25 days. For comparison, mono-digestion was also included in the experimental work. 2 L working volumes were used throughout the experimental work. The seed culture was obtained from composting as substrate digestion. When the feedstock was added to seeding, the biogas started to emit after three days of retention time. The highest volume of biogas was observed when the seeding volume used for 1000 mL. However, the lowest volume of biogas yield was obtained from both co-digestion reactors, with a value of 340 mL. For methane yield, the highest methane production rate was 0.16 L CH4/mg. The COD with yield was at 8.6% and the lowest was at 0.5%. The highest quantity of methane was obtained from a reactor of Euphorbiaceae peel with added seeding, while the lowest methane yield came from a reactor of Euphorbiaceae stems with added seeding. In this study, sodium bicarbonate (NaHCO3) was used as a buffering solution to correct the pH in the reactor if the reactor condition was found to be in a souring or acidic condition.

Anaerobic co-digestion of fruit and vegetable waste: Synergy and process stability analysis

ABSTRACT Anaerobic mono- and co-digestion of fruits and vegetable waste (FVW), slaughterhouse waste (SHW), and cattle manure (CM) under mesophilic conditions (35°C) were conducted through biochemical methane potential tests to investigate how the FVW in a co-substrate formulation improves the methane yield, the degradative synergy between substrates, and especially the stability of the process. The co-digestion of FVW:SHW and FVW:CM were evaluated with volatile solids (VS) ratios of 1:2, 1:1, and 2:1. The results indicated that the highest synergistic effect was found in the co-digestion FVW:CM at 1:1 VS ratio. However, the co-digestion FWV:SHW at 1:2 VS ratio increased the methane yield by 74.2% compared to the mono-digestion of FVW (776.3 mL CH4 g VSadded −1). As a critical condition in these processes, the stability was evaluated using the early warning indicator VFA/TA (volatile fatty acids/total alkalinity). The co-substrate SHW promotes greater stability in methane production as the soluble carbohydrate content in FVW increases. It was proposed that the high protein (49.04 ± 0.96% VS) and ammonia content (693 ± 3 mg L−1) of SHW leads to the formation of a dampening system known as a carbonate-acetic buffer. It was concluded that balanced distribution between carbohydrates, proteins, and lipids is crucial to increase methane yields, and the low methane productions were associated with low N-NH4 + concentrations (FVW:CM co-digestions). The results obtained in this study can serve as a basis to design full-scale digesters under similar operating conditions and with the same substrate:co-substrate relationships. Implications: The production of methane from the anaerobic digestion process of food, and lose waste presents a viable alternative of valorization and could help to mitigate environmental impacts. However, anaerobic digestion from these substrates carries high instabilities and low methane yields. The need to increase these yields and contribute to process stability must be considered in the selection of a co-substrate. In this context, this work aims to evaluate the best fruits and vegetable waste: co-substrateformulation, that promotes higher methane yield, a synergy between substrates, and to improve the AD process stability in the presence of perturbations in the substrate composition. We believe that our results could be helpful for the design processes for methane production from fruit and vegetable waste, to contribute to competitiveness with conventional energies and promote the sustainability of these processes.

Biochemical methane potential of Jatropha curcas fruit shell: comparative effect of mechanical, steam explosion and alkaline pretreatments

Jatropha curcas is a promising tropical and subtropical plant species for biodiesel production that can reduce the competition between food and energy production. Jatropha seed processing for oil extraction usually generates considerable amount of Jatropha curcas fruit shell (JCFS), which can be considered as a potential substrate for biogas production rather than being discarded as solid waste. However, the higher lignocellulosic constituents in JCFS potentially affect the biological degradation process. Thus, applying suitable pretreatment techniques in advance of anaerobic digestion could enhance the biodegradability and methane yield of JCFS. In this study, the effect of mechanical, steam explosion (SE), and alkaline pretreatments on the chemical composition and methane yield of JCFS was examined at various process conditions. As compared with the untreated sample, grinding the JCFS into a particle size of less than 1 mm increased the methane yield by 74.23%, while at the optimum SE pretreatment process (160 °C, 5 min), the methane yield was increased by 54.75%. The alkaline pretreatment was relatively less effective over the other pretreatments; 44.05% methane yield increment was achieved after soaking the JCFS with 7.32% NaOH at 36 °C for 54 h. The effect of SE on compositional change depends on the severity factor in which severe pretreatment conditions were adequate for solubilizing the hemicellulose but resulted in higher pseudo-lignin and lower methane yields. In conclusion, all pretreatments processes have significantly increased the methane yield of JCFS as compared to the untreated JCFS; however, mechanical pretreatment was more effective than SE and alkaline pretreatments.

The impact of single step ozonation of antibiotics-contaminated waste sludge to biogas production

Antibiotics contamination of the environment is a growing public health concern, due to their persistency, possible chronic effects and spread of antibiotics resistance genes. The aim of the study was to determine efficiency of ozonation of waste biological sludge, contaminated by various antibiotics (400 mg L−1 of Tiamulin, Amoxicillin and Levofloxacin) in terms of inreased biogas production potential. It was confirmed that contaminated waste sludge inhibits overall biogas production in further anaerobic stabilization for 10–30% resulting also in lower methane yield in the gas mixture (14–45%). Ozonation of waste biological sludge was accomplished in batch system for 10 (22–24 mgO3 gvss−1) or 20 (36–69 mgO3 gvss−1) minutes. The impact to biogas production potential was measured for untreated, contaminated, ozonated untreated and ozonated contaminated sludge after its addition to anaerobic sludge in closed system at 37 °C. Ozone at applied doses simultaneously removed antibiotics related inhibition of biogas production and in some cases enhances biogas production (13–18%) with improved methane yield (22–32%). The highest improvement in biogas production potential was determined for Tiamulin while ozonation of Levofloxacin contaminated sludge was less efficient. It was concluded, that proposed ozone doses resulted in removal of inhibition due to the antibiotics but did not lead to economically feasible increase of biogas production and methane yield.

Coupling granular activated carbon and exogenous hydrogen to enhance anaerobic digestion of phenol via predominant syntrophic acetate oxidation and hydrogenotrophic methanogenesis pathway

Anaerobic digestion is a promising biological method for treating phenol-containing wastewater. However, the low methane yield of phenol due to the biological toxicity limits its potential application. This study presents a novel method to enhance the conversion rate of phenol to methane by coupling of granular activated carbon and exogenous hydrogen (GAC/H2). The cumulative methane production in the GAC/H2, H2, GAC, and control groups were 408.2 ± 16.2, 336.5 ± 5.7, 287.2 ± 26. 2, and 258.1 ± 8.6 mL‬ CH4/g COD, respectively. Compared with the control group, the hydrogenotrophic methanogenic activity and electron transfer activity of GAC/H2 group were increased by 403.9 and 367.4%, respectively. The results of the 16SrRNA analysis indicated GAC enhanced the relative abundances of Syntrophus and Syntrophorhabdus, and hydrogen promoted the relative abundances of Cryptanaerobacter, Aminicenantes, and Methanobacterium. Therefore, the coupling of GAC and exogenous hydrogen promoted a dominate SAO-HM pathway to convert phenol to methane.

Invaded water flowback capability of high volatile bituminous coal after nitrogen displacing: from a perspective of fracture and seepage pore structural complexities

Water-based fracturing is applied for stimulating coal reservoir. However, invasion of water can result in low flowback efficiency and low coalbed methane (CBM) yield. Nitrogen injection is an effective method to enhance CBM production. In this study, nitrogen displacing water experiments using water saturated high volatile bituminous coal were conducted, during which magnetic resonance imaging (MRI) tests were applied to determine flowback capability. To research the relationship between flowback capability and complexities of fracture and seepage pore, characterising methods including micro-CT and mercury intrusion porosimetry (MIP) were used. The results show that flowback capabilities of complex structural seepage pores and fractures were low. Water is easily flooded into simple structural seepage pores and fractures, and water flowback capabilities of these simple spaces are relative higher, compared with those of the complex ones. Water phase trapping during nitrogen displacing water happens due to water imbibition and Jamin effect occurring in complex structural seepage pores and fractures. [Received: April 27, 2019; Accepted: August 7, 2020]

Invaded water flowback capability of high volatile bituminous coal after nitrogen displacing: from a perspective of fracture and seepage pore structural complexities

Water-based fracturing is applied for stimulating coal reservoir. However, invasion of water can result in low flowback efficiency and low coalbed methane (CBM) yield. Nitrogen injection is an effe...

Current status of biomethane production using aqueous liquid from pyrolysis and hydrothermal liquefaction of sewage sludge and similar biomass

Pyrolysis and hydrothermal liquefaction (HTL) are potential technologies for renewable energy production and waste valorization using municipal wastewater sewage sludge and other lignocellulosic biomass. However, the organic-rich aqueous pyrolysis liquid (APL) and HTL aqueous phase (HTL-AP) produced currently have no apparent use and are challenging to manage. Furthermore, the toxic organic compounds in them can be harmful to the environment. Anaerobic digestion (AD) may be a viable method to manage the liquids and recover energy in APL and HTL-AP in form of methane-rich biogas. Integrating thermochemical processes with AD could promote a circular economy by recovering resources and reducing environmental pollution. The challenge, however, is the presence of toxic compounds recalcitrant to anaerobic biodegradation such as phenols and nitrogen-containing organics that can inhibit methane-producing microbes. This review presents information on APL and HTL-AP characterization and biodegradability. Feedstock composition and process operational parameters are major factors affecting APL and HTL-AP composition, subsequent toxicity, and degradability. Feedstocks with high nitrogen content as well as increased thermochemical processing temperature and retention time result in a more toxic aqueous liquid and lower methane yield. Dilution and low AD organic loading are required to produce methane. More comprehensive APL and HTL-AP chemical characterization is needed to adopt suitable treatment strategies. Pretreatments such as overliming, air stripping, partial chemical oxidation, adsorption, and solvent extraction of toxic constituents as well as co-digestion and microbial acclimatization successfully reduce toxicity and increase methane yield.

Ranking of energy potentials of agro-industrial wastes: Bioconversion and thermo-conversion approach

The rising amount of organic wastes (biomasses) generated annually constitute serious environmental concerns but hold enormous potentials in solving the increasing global energy demand. When properly harnessed, they are good renewable energy sources with capabilities of bridging the energy demand gaps and ameliorating environmental concerns of developing climes. In this study, the mixed-method approach of anaerobic digestion and thermogravimetry were used in the ranking of eighteen selected agro-industrial biomasses based on their energy (biomethane or thermochemical) conversion potentials. The ultimate and proximate properties of the substrates were determined, then the experimental biomethane potentials (BMP) were carried out via anaerobic digestion of feedstocks at mesophilic conditions for 30 days. The biomasses were also degraded thermally within a temperature range of 50 and 800 °C, torrefaction and the pyrolysis char yields were determined. The calorific values and the theoretical biomethane potentials (TBMP) were calculated from the elemental compositions of substrates. From the results obtained, lignocellulosic plant materials and the animal manures recorded lower methane yields and biodegradability. Although the animal manures had appreciable high heating values (HHVs), their BMP yields were poor with biodegradability as low as 25.8%. Consistent with agro-based biomasses, most of the biomass had low H/Ceff, high H/C and O/C atomic ratios. Based on ranking criteria, waste samples from animal carcasses and oil crops were ranked topmost in both thermochemical and biochemical energy conversion. Co-digestion/degradation of high ranked biomass with others and enhancements of the substrate before energy conversion is recommended. Thermal upgrade of biomasses and co-pyrolysis with coal is also recommended.

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.

Effect of Temperature and Organic Load on the Performance of Anaerobic Bioreactors Treating Grasses

The organic residues generated in grasslands can be treated by adopting anaerobic digestion technology. This technology can enhance the efforts for sustainable waste management around the world. In the northern Netherlands, there is a vast amount of ditch clippings and canal grasses that can be used as a renewable source of energy; however, optimal bioenergy production from grasses is still under research and this study aims to evaluate biogas production from grassy residues at the local level in the context of a sustainable waste management scheme. Batch tests were facilitated to investigate the impact of temperature and organic load on the anaerobic digestion performance of grass mixtures (ditch clippings and canal grasses). The results showed that high temperature favors the degradation of high lignocellulosic materials like grasses. Specifically, bioreactors at 55 °C with an organic load of 30 g volatile solids (VS) L−1 reached 360.4 mL g VSsubstrate−1. Moreover, reactors with low organic loads resulted in a lower methane yield. The kinetics study also showed good fitting of the predicted and experimental values.

Suitability of cardoon (Cynara cardunculus L.) harvested at two stages of maturity to ensiling and methane production

A sustainable biogas production requires the use of a biomass that is not in competition with food production and does not require a high energy input for its production. Cardoon (Cynara cardunculus L.) was evaluated at ensiling to establish its suitability for methane production. The cardoon was harvested at full bloom (Stage I) and seed ripening (Stage II), and was then treated with lactic acid bacteria (LAB). Higher fermentative products (lactic acid and ethanol) and DM losses were determined at Stage I than at Stage II. The advanced stage of maturity (Stage II) determined a higher fiber content and lower methane yield per tonne of volatile solids than at Stage I. No difference was observed between the methane production at harvesting and that after ensiling, thus indicating the ability of the ensiling process to conserve most of the energy of the crop at harvesting.

Optimization of methane production in a swine manure–rice straw anaerobic co-digestion process with sycamore sawdust biochar application

Anaerobic co-digestion has been widely applied to re-utilize livestock manure; however, the low methane yield is still a key issue. Biochar could play an important role in anaerobic digestion. This study aims to produce and characterize sycamore sawdust biochar (SSB), determine the promoting effect on the swine manure–rice straw co-digestion system by applying the SSB, and obtain the optimal digesting conditions by using a Box–Behnken design of response surface methodology (RSM). The results showed that SSB had a porous structure composed of micropores and mesopores. The micropore surface area reached 162.87 m2/g, and the mesopore volume accounted for 43.67% of the total pore volume. SSB was also rich in C=O surface functional groups, and the BET specific surface area was up to 276.67 m2/g. Application of SSB could improve the pH value and the stability of the co-digestion system, and eventually improve the cumulative methane yield. The maximum cumulative methane yield in this test was 217.99 mL/(g VS) when 4% SSB was added. According to the response surface optimization analysis, the cumulative methane yield could be 247.20 mL/(g VS) under the optimal conditions: 94.54% swine manure, 8.78% TS and 5.49% SSB.

A critical review of microbial electrolysis cells coupled with anaerobic digester for enhanced biomethane recovery from high-strength feedstocks

Anaerobic digestion (AD) is considered as a traditional method for biomethane recovery from high-strength waste and wastewater. The limitations of conventional anaerobic digestion processes include sluggish kinetics, process instability, and low methane yield. Recently, microbial electrolysis cell (MEC) has been demonstrated as a promising approach for enhancing biomethane recovery from various organic feedstocks. In recent years, a variety of hybrid anaerobic bioreactors coupling MEC and AD were employed to improve biomethane recovery and anaerobic treatability of various high-strength feedstocks. The MEC-AD integrated systems also exhibited better process stability than traditional anaerobic digesters by alleviating the accumulation of short-chain volatile fatty acids at higher organic loading rates, as well as mitigating the inhibitory effects of various toxic and refractory compounds. Firstly, this article provides a comprehensive performance review of the MEC-AD systems treating various high-strength (>8 g COD/L) substrates. Secondly, the implications of process parameters, energy efficiencies, and microbial community characteristics are discussed. Finally, valuable operational insights and future research needs are highlighted.

A Novel Nitrogen Removal Technology Pre-Treating Chicken Manure, Prior to Anaerobic Digestion

Chicken manure is an agricultural by-product that is a problematic feedstock for anaerobic digestion due to its high nitrogen content inhibiting methane yields. This research examines a novel pilot-scale method of ammonia stripping, the nitrogen recovery process (NRP) developed by Alchemy Utilities Ltd. The NRP was designed to remove and recover nitrogen from chicken manure and two different operating conditions were examined. Both operating conditions demonstrated successful nitrogen removal and recovery. The biochemical methane potential assays were used to compare the digestibility of the NRP-treated chicken manures to that of a fresh chicken manure control. Overall, the biochemical methane potential assays demonstrated that some NRP-treated chicken manure treatments produced significantly more methane compared to untreated manure, with no inhibition occurring in relation to ammonium. However, some of the NRP-treated chicken manures produced similar or lower methane yields compared to fresh chicken manure. The NRP requires further development to improve the efficiency of the pilot-scale unit for commercial-scale operation and longer-term continuous anaerobic digestion trials are required to determine longer-term methane yield and ammonium inhibition effects. However, these initial results clearly demonstrate the technology’s potential and novel application for decentralised, on-farm nitrogen recovery and subsequent anaerobic digestion of chicken manure.

Anaerobic degradation of protein-rich biomass in an UASB reactor: Organic loading rate effect on product output and microbial communities dynamics

Anaerobic degradation of enzymatically pretreated Chlorella vulgaris was aimed in an upflow anaerobic sludge blanket reactor (UASB) to evaluate the organic loading rate (OLR) effect on biomass valorization. Low OLRs resulted in high methane yields (171 mL CH4/g CODin) at low hydraulic retention time (HRT of 6 days). Firmicutes (35–43%), Bacteroidetes (17–18%) and Euryarchaeota (11%) dominated at low OLRs, promoting methanogenic activity. On the contrary, the highest OLRs resulted in low methane yield (86 mL CH4/gCODin) with a concomitant short-chain fatty acids (SCFAs) accumulation of 37% SCFAs-COD/CODin. The highest OLR decreased UASB reactor biodiversity, hampering Euryarchaeota population development (2.5%) and boosting Firmicutes (55%) and Proteobacteria (14%). These results demonstrated the suitability of UASB reactor configuration to reach high bioprocess efficiency for both, biogas and SCFAs production, with lower energetic and area requirements than those normally needed in continuous stirred tank reactors.

In search of the optimal inoculum to substrate ratio during anaerobic co-digestion of spent coffee grounds and cow manure.

The inoculum to substrate (I:S) ratio is a crucial operating parameter during the start-up period of anaerobic digestion (AD) processes and this ratio shows high differentiation with respect to substrate composition. While spent coffee grounds (SCG) have started to gain attraction in AD as a co-substrate due to their vast production and promising methane potential, there is still not enough information on the operative environment of SCG-based biogas reactors. This study investigated the optimal I:S ratio during anaerobic co-digestion of SCG and cow manure. Biochemical methane potential tests were conducted at mesophilic conditions and the influence of I:S ratio on methane production and digestion stability was evaluated at a wide range of I:S ratios from 0.5:1 to 4:1 (volatile solids (VS) basis). Methane yields increased gradually starting from the I:S ratio of 0.5:1 up to 3:1 and the highest methane yield (225 mlCH4 gVS-1) was achieved at the I:S ratio of 3:1. Comparatively lower methane yields were obtained at the ratios of 3.5:1 and 4:1. Instable AD conditions were established at the lowest I:S ratio examined (0.5:1), which caused volatile fatty acid (VFA) accumulation. The results highlighted that anaerobic co-digestion of SCG and cow manure is a promising approach, while the I:S ratio should be well-maintained due to the high potential risk of rapid and/or excess VFA production of these feedstocks.

Process performance and microbial communities in response to selenite addition during anaerobic digestion of pig manure

The effects of selenite addition on process performance and microbial communities during the mesophilic anaerobic digestion of pig manure were investigated. At low selenium concentration (0.1 mg/L), methane yield and methane production rate increased by 121% and 113%, respectively. High selenium concentration (30–120 mg/L) decreased methane yield by 23.1–100% and methane production rate by 1.3–100% owing to the reduced hydrolytic enzyme and F420 activities at initial phase of anaerobic digestion. Meanwhile, the inhibited activity of methane production could be completely recovered through a successive supplementing organic matter for selenite reduction. Besides, selenite addition also altered the relative abundance of hydrolytic and acetogenic bacteria, and increased the relative abundance of acetoclastic methanogen. These results suggested that the inhibited methane production was owing to the fact that the selenite-reducing microorganism competes with methane-producing microorganism for H2 and acetate, but selenite might not directly inhibit methane-producing microorganism.