Mesophilic Digestion Research Articles

Enhancing Methane Production in a Two-Stage Anaerobic Digestion of Spent Mushroom Substrate and Chicken Manure via Activation of Sludge, Optimization of Temperature, and C/N Ratio

Management of spent mushroom substrate (SMS) is causing a global environmental concern due to tremendous increase in mushroom production globally. Therefore, in this research, the performance of a two-stage anaerobic co-digestion (TS-AD) of spent mushroom substrate and chicken manure was evaluated in terms of methane and biogas production and process stability with respect to single stage anaerobic digestion (SS-AD). Activation of anaerobic sludge using aeration or heat treatment in the first stage at mesophilic temperature followed by thermophilic co-digestion with chicken manure in the second stage was investigated. TS-AD exhibited better performance and enhanced methane generation over SS-AD. The optimal temperatures were determined as 35°C and 50°C for the first and the second stage of TS-AD, respectively. C/N ratio of 10 was the most suitable for biogas and methane production. TS-AD with C/N ratio of 10 and mesophilic digestion of SMS and sludge for 3 days at 35°C followed by co-digestion of the first stage effluent with chicken manure at 50°C was the optimized state producing 1359 mL of biogas of which 614.42 mL was methane, showing an increment by 59.44% in methane production as compared to SS-AD. TS-AD might be promising approach for utilization of SMS as feed stocks for biogas and methane production.

Methane production and microbial community acclimation of five manure inocula during psychrophilic anaerobic digestion of swine manure

For small-scale farms, the development of rustic and cheap psychrophilic anaerobic digestion systems appears as an opportunity to treat manure, mitigate gaseous emissions and promote decentralized renewable energy production. However, the development of such processes is limited by our understanding of their start-up. In this research, we tested the ability of one mesophilic digestate and four different manure to be used as inoculum for the start-up of psychrophilic anaerobic digestion of swine slurry at 13 °C. The most efficient inoculum appeared to be a swine manure that had been stored for 2 months in a pit. After 9 months of acclimation, the corresponding reactor produced a maximum methane yield of 42L of CH4/kg Volatile Solidesubstrate/day and a CH4 volume of 125L of CH4/kg-Chemical Oxygen Demandadded. The maximum methane production expressed at 13 °C was between 55% and 68% of that obtained at 37 °C. Monitoring of the microbial community dynamics by high throughput 16S rDNA sequencing showed the smooth adaptation of manure microbial species, underlining the transient dominance of the acetogen syntroph candidatus Cloacimonas during acclimation and the enrichment in the Methanosaeta and Methanosarcina methanogens for an efficient methane production.

Estimation of kinetic constants in high-density polyethylene bead degradation using hydrolytic enzymes

Microplastic beads are an emerging contaminant that can cause serious environmental and public health problems. Potential bypass of microplastic beads from wastewater to sludge treatment systems is a key challenge in the conventional wastewater treatment process. Moreover, there are no systematic studies on microplastic bead degradation by hydrolytic enzymes that are rich in concentration within wastewater and sludge treatment processes (e.g., anaerobic digestion (AD)). In this study, lab-scale experiments were conducted to investigate the degradation of high-density polyethylene beads by hydrolytic enzymes (e.g., lipase) under various experimental conditions (e.g., temperature). In a 3-day batch experiment, protease was most effective in polyethylene bead degradation as 4.0% of the initial bead mass was removed at an enzyme concentration of 88 mg/L under thermophilic temperature (55 °C). It was also found that the increasing enzyme concentration and high temperature enhanced the polyethylene bead degradation. In a separate 7-day experiment with repeated doses of protease, 23.3% of the initial mass of beads was removed at thermophilic temperature, indicating that AD with a long retention time (e.g., 20 days) and heated temperature has a significant potential for polyethylene bead degradation. A mathematical model was developed and calibrated using the experimental results to estimate the kinetic constant of the high-density polyethylene bead reduction by an enzyme (k1,i) and enzyme self-decay constant (k2,ii). The calibrated k1,i ranged from 5.0 to 8.1× 10−4 L/mg/hr while k2,ii was 0.44–1.10 L/mg/hr. Using the calibrated model, degradation of polyethylene beads using a mixture of cellulase and protease was simulated, considering an interactive-decay reaction between the two enzymes. The calibrated model was used to simulate the polyethylene bead degradation in AD where 70–95% of the initial bead mass was removed at typical retention time under mesophilic digestion (37.5 °C). Based on the experimental and simulation results, it can be concluded that hydrolytic enzymes can be an efficient technology for large-scale high-density polyethylene bead removal applications.

Anaerobic Mesophilic Digestion of Typha Latifolia Suspensions Using the Fermented Waste of Yeast Production as Fermentation Inoculum

Anaerobic Mesophilic Digestion of Typha Latifolia Suspensions Using the Fermented Waste of Yeast Production as Fermentation Inoculum

Microbial Community Successional Changes in a Full-Scale Mesophilic Anaerobic Digester from the Start-Up to the Steady-State Conditions.

Anaerobic digestion is a widely used technology for sewage sludge stabilization and biogas production. Although the structure and composition of the microbial communities responsible for the process in full-scale anaerobic digesters have been investigated, little is known about the microbial successional dynamics during the start-up phase and the response to variations occurring in such systems under real operating conditions. In this study, bacterial and archaeal population dynamics of a full-scale mesophilic digester treating activated sludge were investigated for the first time from the start-up, performed without adding external inoculum, to steady-state operation. High-throughput 16S rRNA gene sequencing was used to describe the microbiome evolution. The large majority of the reads were affiliated to fermentative bacteria. Bacteroidetes increased over time, reaching 22% of the total sequences. Furthermore, Methanosaeta represented the most abundant methanogenic component. The specific quantitative data generated by real-time PCR indicated an enrichment of bacteria and methanogens once the steady state was reached. The analysis allowed evaluation of the microbial components more susceptible to the shift from aerobic to anaerobic conditions and estimation of the microbial components growing or declining in the system. Additionally, activated sludge was investigated to evaluate the microbial core selected by the WWTP operative conditions.

The interaction between lipids and ammoniacal nitrogen mitigates inhibition in mesophilic anaerobic digestion

Ammoniacal nitrogen and long chain fatty acids (LCFA) are common inhibitors of the anaerobic digestion process. However, the interaction between these inhibitors has received little attention. Understanding the interaction between these inhibitors is important to optimise the operation of anaerobic digesters treating slaughterhouse waste or using fat, oil and grease (FOG) as co-substrate among others. To study the interaction between ammoniacal nitrogen and LCFA inhibition, 20 different conditions were trialled in mesophilic batch tests. Experimental conditions included 5 mixtures between slaughterhouse wastewater and LCFA (100:0, 75:25, 50:50, 20:80, 0:100 on a VS basis), each one tested at 4 different ammoniacal nitrogen concentrations (0, 1, 3, 6 gNadded·L-1). Experimental and modelling results showed that ammoniacal nitrogen inhibition was less severe in LCFA-rich mixtures, indicating that LCFA mitigated ammoniacal nitrogen inhibition to a certain extent. However, the positive interaction between inhibitors did not only depend on the LCFA concentration. A protective LCFA coat that limited the diffusion of free ammonia into the cell and/or provided a localised lower pH in the vicinity of the microbial cell could explain the experimental results. However, ammoniacal nitrogen and LCFA inhibition comprise up to 6 different but interrelated inhibitors (i.e. NH3, NH4+, LCFA, VFA, H2 and pH) and therefore the specific mechanism could not be elucidated. Nonetheless, these results suggest that LCFA do not exacerbate TAN-related inhibition and that LCFA-rich substrates can be utilised as co-substrates in mesophilic N-rich digesters.

Dry anaerobic digestion of the fine particle fraction of mechanically-sorted organic fraction of municipal solid waste in laboratory and pilot reactor.

High-solid anaerobic digestion of the very small particle fraction of mechanically-sorted organic fraction of municipal solid waste (OFMSW) was examined in mesophilic digestion tests in a conventional laboratory (0.013m3) and a pilot (0.300m3) reactor. The non-biodegradable and recalcitrant molecules together with the low protein and starch contents of the small-particles of OFMSW limited the methane generation potential of substrate. In the conventional AD system, methane yields remained low at 0.139 m3kgVS-1 due to formation of a non-reacting layer on digestate surface, which restricted utilization of the available in OFMSW digestible organics. The absence of surface solid crust in the pilot unit favoured consumption of a greater proportion of volatile solids of the OFMSW. Dry AD was remarkably stable over the entire period and negligibly effected by the toxic H2S yields. Methane generation (0.167 m3kgVS-1) was increased 1.2-fold compared to the conventional system due to a better mixing of substrate and microorganisms achieved inside the pilot reactor, which led to an increase of the digested volatile organics. Digestate presented low stability and high heavy metal content, both of which restrain its implementation as soil conditioner or fertilizer in agriculture. A secondary co-digestion treatment may be required for the neutralization of digestate.

Stimulatory effects of biochar addition on dry anaerobic co-digestion of pig manure and food waste under mesophilic conditions.

The stimulatory effect of biochar addition on dry anaerobic digestion (AD) has been rarely investigated. In this study, the effects of commonly used biochars (bamboo, rice husk, and pecan shell) on dry co-AD were investigated using mesophilic batch digesters fed with pig manure and food waste as substrates. The results show that the specific methane yield was mildly elevated with the addition of biochars by 7.9%, 9.4%, and 12.0% for bamboo, rice husk, and pecan shell-derived biochar additions, respectively. Biochar did facilitate the degradation of poorly biodegradable organics. In comparison, there was no significant effect on the peak methane production rate by the supplementation of the selected biochars. Among the three mechanisms of enhancing methanogenesis by biochar (buffering, providing supporting surface, and enhancing electron transfer), the first two mechanisms did not function significantly in dry co-AD, while the third mechanism (i.e., enhancing electron transfer) might play an important part in dry AD process. It is recommended that the utilization of biochar for the enhancement of biomethanation in dry AD should be more focused on mono digestion in future studies.

Biogas Production by Pilot-Scale Anaerobic Co-Digestion and Life Cycle Assessment Using a Real Scale Scenario: Independent Parameters and Co-Substrates Influence

This study evaluates the performance of different agricultural by-products to identify the potential effect of independent variables, using as the dependent variable the biogas production. A Box–Behnken experimental design was carried out in a pilot-scale plant of four stirred stainless-steel digesters under mesophilic semi-continuous digestion. The results obtained support the creation of a technical framework to scale up the process and further evaluation of the potential environmental impacts through life cycle assessment (LCA) methodology. A stable behaviour was achieved in 12 of the 13 experiments proposed. The highest value of daily biogas production was 2200.15 mL day−1 with a stabilization time of 14 days, an organic loading rate of 4 g VS feed daily, low C/N ratio and a 1:1 relation of nitrogen providers. The concentrations of CH4 remained stable after the production stabilization and an average biogas composition of 60.6% CH4, 40.1% CO2 and 0.3% O2 was obtained for the conditions mentioned above. Therefore, the real scale plant was estimated to manage 2.67 tonnes of residual biomass per day, generating 369.69 kWh day−1 of electricity. The LCA analysis confirms that the co-digestion process evaluated is a feasible and environmentally sustainable option for the diversification of the Colombian energy matrix and the development of the agro-industrial sector.

Mesophilic aerobic digestion: An efficient and inexpensive biological pretreatment to improve biogas production from highly-recalcitrant pinewood

Biological pretreatments and their combination with chemical and physical pretreatments are among the inexpensive methods for the improvement of biogas production from lignocellulosic biomass. In this study, mesophilic aerobic digestion was studied as a biological pretreatment to improve biogas production from pinewood. Its effects in combination with other pretreatments, i.e., liquid hot water (LHW), sodium hydroxide (NaOH), and ultrasonic pretreatments, were also investigated. LHW and Alkali (using 8% w/w NaOH) pretreatments were performed at less severe conditions (100 °C for 10 min) to evaluate the synergistic effect of these thermochemical pretreatments on biogas production. The ultrasonic pretreatment was performed at 40 KHz for 30 min at 40 °C. Biological pretreatment was carried out using aerobic sludge at 37 °C for 10 days. The aerobic digestion was the only pretreatment that could individually improve the methane yield from pinewood by 7.3 folds. However, the highest methane yield (57.7 mL/g VS of pretreated pinewood) was obtained by the combination of NaOH, biological, and ultrasonic pretreatments, which was 11.2 folds higher than that of the untreated substrate. It was concluded that the pretreatment with aerobic digestion could significantly improve the biogas production yield, especially in combination with mild thermo-chemical pretreatments.

Mass Balance of C, Nutrients, and Mineralization of Nitrogen during Anaerobic Co-Digestion of Rice Straw with Cow Manure

Anaerobic co-digestion technology (AcoD) can be used to process rice straw (RS) and cow manure (CoM) to produce energy and a digestate rich in nutrients, while the improper disposal of RS and CoM causes environmental problems. The overall effectiveness of the anaerobic digestion technology can be improved by utilizing the nutrients available in the digestate. It is also a way to reduce the usage of mineral fertilizer by recycling the nutrients available in the digestate. The co-digestion of RS with CoM was performed in a newly developed digester (F1) and in a mesophilic digester (F2) used as a reference. The mass balance of C, macronutrients (N, P, K, Ca, Mg, and S), and their distribution into a liquid digestate (LD) and a solid digestate (SD) was investigated in both digesters. The mass balance was used to evaluate the carbon available in the biogas and in the digestate. It was also used to investigate the recovery potential of the macronutrients after the AD process. Moreover, the assessment of the resulting digestate was carried out to suggest its potential use in agriculture. The amount of C measured in the biogas was the same in both digesters (41.0% and 38.0% of the initial C). Moreover, the conversion efficiency of C from the substrate into methane was 23.4% for F1 and 21.0% for F2. The Ca, Mg, K, and P were conserved in the digestate because their recovery rates (RR) were close to 100%. However, a relatively low RR was observed for N (84.1% in F1 and 86.8% in F2) and S (87.1% in F1 and 86.5% in F2) in both the digesters. After separation n of the SD, from 79.1 to 83.4% (in F1) and 75.0 to 82.4% (in F2) of the final nutrients were available in the LD. The assessment of the SD suggested its use in agriculture not only for soil amendment but also as a K-providing organic fertilizer.

The biological and abiotic effects of powdered activated carbon on the anaerobic digestion performance of cornstalk

To comprehensively evaluate the biological and abiotic influence of powdered activated carbon (PAC) on the anaerobic digestion of cornstalk, mesophilic and thermophilic digestion were conducted. Adding PAC (10 g/L) under thermophilic system obtained the maximum cellulose degradation rate and methane yield (MY), which were 57.47% and 128.19 L/kg VS. However, adding same dose of PAC at mesophilic system decreased the MY by 8.16% while increased the cellulose degradation rate and methane production rate by 6.48% and 17.92%. Under mesophilic conditions, the enhancement of PAC was owing to the enrichment of cellulolytic microorganisms, improvement of the syntrophic process and direct interspecies electron transfer. The lower methane yield was attributed to the adsorption of carbon source by PAC and CH4 consumption by Norank_c_Bathyarchaeia. The good performance of thermophilic system was owing to the lower adsorption capability of PAC, absence of Norank_c_Bathyarchaeia, and concentrated carbon flow to methane.

The dosage-effect of biochar on anaerobic digestion under the suppression of oily sludge: Performance variation, microbial community succession and potential detoxification mechanisms

This study investigated the dosage-effect of biochar on the suppressed mesophilic digestion of oily sludge (OS) containing naphthalene (recalcitrant compound) and starch (easily bioavailable substrate). Methanogenesis was inhibited in control with OS, where biomethane yield (63.33 mL/gVS) was obviously lower than theoretical yield (260.55 mL/gVS). With adding optimal dose of biochar (0.60 g/gVS OS), the highest CH4 yield (138.41 mL/gVS) was 2.19 times of control. Meanwhile, the efficiencies of hydrolysis, acidogenesis and acetogenesis were significantly enhanced. However, excessive biochar (4.80 g/gVS OS) caused negative effects with methanogenic efficiency diminished by 32.5% and lag phase prolonged by 5.72 h. Dissolved organic matter (DOM) analysis showed that humic acid-like and fulvic acid-like components percentages of fluorescence regional integration were decreased because of the adsorption of biochar. In addition, biochar mediating interspecies electron transfer selectively enriched electroactive fermentation bacteria (Clostridium and Bacteroides) and acetoclastic Methanosaeta, which was responsible for promoting mesophilic digestion performance. The functional genes related to metabolism and environmental information processing were potentially activated by biochar. Above results indicate that moderate biochar application may mitigate the bio-toxicity suppression of OS, which help to provide a promising pathway for reinforcing oily wastes bio-treatment.

The occurrence and fate of microplastics in a mesophilic anaerobic digester receiving sewage sludge, grease, and fatty slurries

The mesophilic digester of a wastewater treatment plant serving 790,000 inhabitants was analyzed for incoming and outgoing microplastics (MPs). The annual MP load on the digester was 7326 kg y−1 and 1.20 × 1014 N y−1, while the digested sludge contained 4885 kg y−1 and 0.85 × 1014 N y−1. The corresponding mean reduction of approximately 30% was though within the variability of the analyzed samples, and size distributions and polymer composition before and after the digester could similarly neither confirm nor deny if MPs were lost in the digester. The load on the digester corresponded to a per capita load of 0.025 g capita−1 day−1 or 4.14 × 105 N capita−1 day−1. In terms of MP numbers, the values were high compared to most previous studies, which most likely was due to differences in analytical methods, where the present study applied FPA-μFTIR hyperspectral imaging with automatized MP recognition in the size range 10–500 μm and ATR-FTIR on all particles in the range 500–5000 μm. The polymer composition was quite diverse, with 15 identified polymer types, of which polyethylene and polyester were the most common in the sludge, while the variation in composition was larger in the grease that came from the plant's grease trap and the fatty slurries which came from grease traps in the drainage area. The load on the digester was finally used to demonstrate how especially the determined mass loads can be used to set upper boundaries for specific sources, for example human excretion.

Elucidating the key factors in semicontinuous anaerobic digestion of urban biowaste: The crucial role of sludge addition in process stability, microbial community enrichment and methane production

Anaerobic digestion, a virtuous process to reduce and reuse biowaste to obtain renewable bioenergy, is often dramatically subject to process failure during long timescale operation. This study aimed to evaluate the process robustness and reliability by elucidating the composition of microbial populations and the link with process parameters under a wide range of different operating conditions in long-term semicontinuous reactors. Mesophilic semipilot digesters were fed with real urban biowaste, namely, food waste alone or food waste with activated sludge, to explore the impact of feedstock composition and organic loading rate (OLR: 0.8–3.5 kgVSfed m−3d−1) on volatile fatty acid (VFA) composition and pattern, biomethane production, and core microbiome dynamics. The major bacterial phyla were Bacteroidetes (basically more abundant when the feedstock was only food waste), followed by Chloroflexi and Firmicutes, while Euryarcheota hydrogenotrophic methanogens, mainly represented by members of the Methanomicrobiales family, prevailed in all systems. During monodigestion of food waste, however, independent of the OLR, methanogenic conversion of fermentative end-products decreased progressively after 1 HRT. In particular, the observed propionate and butyrate accumulations seemed to derive from thermodynamic limitations due to the increase in hydrogen levels, indicative of progressively lower H2 consumption rates. However, the induced process recovery assured a large degradation of fatty acids with higher carbon chain lengths together with methane production, suggesting the upswing of hydrogenotrophic methanogen activity emerging after stressful conditions. The addition of a minimal amount of sludge boosted the methane production rates (up to 0.29 ± 0.1 Nm3CH4 kg−1VSfed) along with excellent process stability, also at a high OLR, creating a robust methanogen community shaped by high biodiversity. Methanospirillum and Candidatus Methanofastidiosum, strictly related to the codigestion systems, have a large H2 consumption capacity, thus preventing thermodynamic bottlenecks and process failure.

The role of oxidation-reduction potential as an early warning indicator, and a microbial instability mechanism in a pilot-scale anaerobic mesophilic digestion of chicken manure

To identify an early warning indicator for the anaerobic digestion of chicken manure and reveal a microbial mechanism of instability, a bench-scale digester with a working volume of 55 L was utilized by raising the organic loading rates (OLRs). Three characteristics were identified and posited as early warning indicators, namely: one-way changes, a longer early response time, and sudden variations. The core early warning indicator set was constructed, which included the oxidation-reduction potential (ORP), individual volatile fatty acids, and CH4/CO2. Considering the convenience, economic nature, and reliability of industrial biogas plants, the ORP was recommended as a final early warning indicator with an inhibition threshold of > −540 mV. The initial instability was caused by the accumulation of ammonia, which suppressed methanogenic activity, although it stimulated the growth of Methanosarcina. The accumulation of free volatile fatty acids, which inhibited both the growth and methanogenic activity of Methanosarcina, was responsible for the final collapse.

Anaerobic Biodegradation of Biodiesel Industry Wastewater in Mesophilic and Thermophilic Fluidized Bed Reactors: Enhancing Treatment and Methane Recovery.

In the past few years, the extraction of value-added compounds from the anaerobic digestion of glycerol has been an option to add value to this waste because biodiesel production is increasing worldwide. The evolution of research on glycerol valorization by anaerobic digestion has reached the use of high-rate reactors. However, no study has evaluated glycerol digestion in an anaerobic fluidized bed reactor (AFBR), a configuration with potential advantages in methane production. Still, the best operating temperature for high-rate glycerol digestion remains unclear. To clarify these gaps, the present study aimed to compare glycerol digestion in mesophilic AFBR (30 °C) and thermophilic AFBR (55 °C). In both reactors, glycerol concentration was increased from 1.0 to 7.0 g L-1 at a fixed hydraulic retention time of 24 h, resulting in an increase at the organic loading rate from 1.2 to 7.6 kg COD m-3 day-1. Thermophilic digestion of glycerol achieved superior removals of organic matter (67.7-94.2%) and methane yield (330.8 mL CH4 g-1 COD) than the mesophilic digestion (48.6-93.0% and 266.6 mL CH4 g-1 COD). Additionally, the application of the kinetic model of substrate utilization (modified Stover-Kincannon model) indicated a higher substrate utilization coefficient in the thermophilic AFBR (23.09 g L-1 day-1) than the mesophilic AFBR (7.14 g L-1 day-1). Therefore, the application of glycerol concentrations higher than 7.0 g L-1 in thermophilic AFBR should be further investigated. Also, given only operational results, the application of the AFBR in the two-stage anaerobic digestion of glycerol is recommended.

Flexible Energy from Biogas: Use of Secondary Digesters for Heat Storage—Results of Fermentation Tests

AbstractA process for a more efficient and flexible use of the produced heat in biogas plants is tested under real operational conditions. The process uses a mesophilic digester and a thermophilic secondary digester, and the heat demand of the plant is covered by using the heat stored in the secondary digester. The efficient heat use is achieved by a controlled variation of temperature in combination with a suitable process monitoring, without affecting the biogas production or disturbing the biological process. The concept was previously tested in ideal laboratory conditions. Here, the validation tests, in preparation for a planned large‐scale trial, comprising one‐year semicontinuous fermentation tests in the laboratory (close to real operational conditions) are presented. These real conditions are characterized by a high organic load of the reactors as well as by a more manifold substrate mix. During the experimental phase, it is determined that the maximum temperature, at high variation rates between −4 and +2 K per day, reaches 54 °C, allowing a stable process. These experiments confirm that the adaptability to temperature changes depends on the boundary conditions of the fermentation and case‐specific experiments are required before significant changes are made to the temperature management of anaerobic digestion systems.

Anaerobic Digestion of Food Waste, Brewery Waste, and Agricultural Residues in an Off-Grid Continuous Reactor

Small-scale anaerobic digestion (AD) can be an effective organic waste management system that also provides energy for small businesses and rural communities. This study measured fuel production from digestions of single and mixed feedstocks using an unheated, 2 m3 digester operated continuously in a temperate climate for over three years. Using local food waste, brewery waste, grease waste, and agricultural residues, this study determined that small-scale AD co-digestions were almost always higher yielding than single feedstocks during psychrophilic operation and seasonal temperature transitions. Agricultural residues from Miscanthus x giganteus had the greatest impact on biomethane production during co-digestion (4.7-fold greater average biogas %CH4), while mesophilic digestion of brewery waste alone produced the most biogas (0.76 gCH4 gVS−1 d−1). Biogas production during the transition from mesophilic to psychrophilic was temporarily maintained at levels similar to mesophilic digestions, particularly during co-digestions, but biogas quality declined during these temperature shifts. Full-time operation of small-scale, unheated AD systems could be feasible in temperate climates if feedstock is intentionally amended to stabilize carbon content.

Effects of bentonite on antibiotic resistance genes in biogas slurry and residue from thermophilic and mesophilic anaerobic digestion of food waste

Anaerobic digestion (AD) processes of food waste (FW) have potential risk on environments due to the prevalence and dissemination of antibiotic resistance genes (ARGs). This study investigated the effect of bentonite on methane production and the abundance of ARGs in biogas slurry and residue during AD of FW. Results showed that methane production increased by 68.52% and 56.79% with 3 g/L and 5 g/L of bentonite in mesophilic and thermophilic digestion, respectively. Adding 5 g/L of bentonite effectively reduced the genes of ermB, ermF, tetQ, tetX, sul1, sul2 and intI1 with a range of 80.82% - 100.00% in biogas residue under mesophilic reactor. The abundance of ARGs and intI1 in biogas residue were lower than in slurry under both temperatures with 5 g/L of bentonite. Statistical analysis indicated that bentonite and temperature were main driver factors which could impact ARGs by influencing the abundance and structure of microbial communities.