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The effects of long-term acclimatization on the optimum food waste to cow manure ratio (FW/CM) and substrate to inoculum ratio (S/I) in anaerobic codigestion with FW and CM were investigated by batch trials. For the unacclimated sludge, the highest CH4 yields of 646.6 and 653.4 mL/g VS were achieved under the optimum FW/CM (2.5 VS/VS) and S/I (0.07 VS/VS) ratios, respectively. After more than 550 days of acclimatization, the optimum FW/CM and S/I of the acclimated sludge were improved to 3.4 and 0.68 VS/VS with more anaerobic digestion enzymes and lignocellulose, respectively. Based on high-throughput sequencing analysis, the microbial community structures of bacteria, fungi, and archaea were changed, which was the main reason for the change in the optimum FW/CM and S/I. Therefore, the FW/CM and S/I should be periodically optimized during the long-term operation of codigestion to improve the codigestion efficiency for biogas production.

To optimize the artificial rumen microorganism sources and develop a stable artificial rumen system, batch and continuous operation were investigated with corn straw and food waste as substrates. The batch trials evaluated the volatile fatty acid (VFA) yield, biogas production, and lignocellulose degradation efficiency. The continuous test evaluated the performance of the artificial cow and sheep rumen systems using a dynamic membrane bioreactor (DMBR) with a stepwise organic loading rate at mesophilic temperature. The anaerobic digestion (AD) of the lignocellulose biomass after rumen fermentation pretreatment and of the permeate from the artificial rumen system were also evaluated for CH4 production. The results indicated that the cow rumen microorganisms were more suitable than sheep rumen microorganisms for lignocellulosic biomass pretreatment and maximized the CH4 yield through the AD process without inhibition. After approximately four months of continuous operation, a stable and continuous artificial rumen system for lignocellulosic biomass degradation was achieved with cow rumen fluid as inoculum. Based on analysis of the core lignocellulose-degrading enzyme levels and gel filtration chromatography, the cow rumen microorganisms could secrete more extracellular multienzyme complexes to hydrolyze lignocellulosic biomass than the sheep rumen microorganisms in vitro. During the batch and continuous operations, a high diversity and similar richness of bacteria and fungi demonstrated that the cow rumen microorganisms can be used as a preferred inoculum for the artificial rumen system. The use of an artificial cow rumen system with a DMBR is a promising way to construct a stable and continuous artificial rumen system to biodegrade lignocellulosic biomass for biogas production.

CO2 bioelectromethanosynthesis represents a promising strategy for the capture and utilization of CO2. In such process, the continuous generation of electron (e-) and proton (H+) in anodic oxidation are of prime importance for the efficient cathodic CO2 electroreduction and process stability. Proton transfer, however, is very easy to be hindered due to the fouling of proton exchange membrane (PEM). In this study, an artificial channel in microbial electrolysis cell (MEC) was proposed to strengthen the transport of protons from anodic to cathodic compartment, and H+-rich anolyte was semi-continuously circulated to the cathodic chamber to provide protons for CO2 electroreduction. The results indicated that the daily CH4 yield in cathode with anolyte circulation (18.5 mL/d·L-reactor) was 5.4-fold higher than that without circulation (2.9 mL/d·L-reactor). Meanwhile, efficient anodic biodegradation of organic components was observed with COD, proteins and polysaccharides removal of up to 95.6 ± 1.9%, 96.3 ± 3.7% and 99.1 ± 0.2% respectively, which supplied a continuous e-/H+ donor for CO2 electroconversion. 16S rRNA gene pyrosequencing analysis identified a large number of proteins-utilizing Bacteroidetes (14.11%) and polysaccharides-consuming Thermotogae (18.49%) in anodic biofilm, conductive to the biodegradation of organic components. Moreover, a high abundance of Methanobacterium (81.07%) was detected to prevail in cathodic biofilm, demonstrating the occurrence of highly enhanced CH4 bioelectrosysthesis. The syntrophic and symbiotic relationship was established in the dual-bioelectrode system, creating a beneficial environment and an energy-efficient approach for biowaste refinery and CO2 electromethanosynthesis.

Bioeletromethanogenesis, as a cutting-edge option to capture CO2 and produce multi-carbon biofuels, has received extensive attraction. However, how electroactive biofilm (EAB) as the biocatalyst drives CO2 electromethanogenesis is still not well recognized. In this study, a two-chamber bioelectrochemical cell equipped with a hybrid skirt-shaped cathode was constructed and the electrocatalytic performance of EAB and the electron shuttling mechanisms involved in extracellular electron transfer (EET) were systematically studied. The EAB colonizing on biocathode showed an excellent cathodic electrocatalytic activity and the minimum charge transfer resistance. The CH4 production rate of 298.0 ± 46.7 mL/L/d was obtained at the cathodic potential of −1.0 V vs. Ag/AgCl with the highest Coulombic efficiency of 75.8 ± 9.9%. The gel-like extracellular polymeric substances, secreted by EAB, facilitated the adhesion/aggregation of microbes and EAB development. Further analysis suggested that CO2 electromethanogenesis exhibited a positive association with Methanobacterium (54.4%) in EAB. Moreover, metagenome analysis confirmed the presence of direct EET-related genes (i.e., hdrA, ehaA, and ehbC), which accelerated the formation of corresponding functional protein complexes (particularly heterodisulfide reductase A, HdrA) and electron exchange. The mechanism for electron shuttling process in catalyzing CO2 electromethanogenesis was further proposed. This study provides a new insight into direct extracellular electron transfer (DEET) mechanisms of CO2 electromethanogenesis, and is useful for promoting EAB electrocatalytic activities and CO2 emission reduction and reuse.

The potential of disordered mesoporous carbon (DMC) as catalyst of peroxydisulfate (PDS) to improve sludge solubilization and methane production was investigated. Results showed that DMC activated PDS (DMC/PDS) to produce sulfate radicals (SO4 −), facilitating cells rupture and sludge matrix dissociation by degrading the carbonyl and amide groups in organic biopolymers (especially proteins, polysaccharides and humus). At the optimal DMC/PDS dosage of 0.04/1.2 g-mmol/g-VS, SCOD was increased from initial 294.0 to 681.5 mg/L, with the methane production rate of 12.6 mL/g-VS/day. Moreover, DMC could serve as electron mediator to accelerate electron transfer of microorganisms, building a more robust anaerobic metabolic environment. Modelling analysis further demonstrated the crucial role of DMC/PDS pretreatment in biological degradation and methane productivity. This study indicated that DMC/PDS pretreatment can prominently enhance the release of soluble substances and methane production, aiding the utilization of PDS oxidation technology for improving anaerobic bioconversion of sludge.

Microbial electrolytic cell (MEC) and magnetite (M) have shown excellent performance in promoting anaerobic digestion (AD) of biowastes. In this study, four types of anaerobic systems (i.e. single AD, M-AD, MEC-AD, and M-MEC-AD) were developed to comprehensively investigate the potential effects of magnetite-enhanced bioelectrochemical stimulation on the biodegradation of waste activated sludge (WAS) and methane (CH4) production. Results showed that M-MEC-AD system produced the highest cumulative CH4 yield, 9.4% higher than that observed in MEC-AD system. Bioelectrochemical stimulation enriched electroactive Geobacter, and classical methanogens (Methanosaeta and Methanobacterium), and the proliferation was further promoted when coupling with magnetite. The relative abundance of Geobacter (6.9%), Methanosaeta (0.3%), and Methanobacterium (12.6%) in M-MEC-AD system was about 10.8, 1.2, and 1.2 times of MEC-AD system, respectively. The integration of magnetite could serve as the conductive materials, and promote inherent indirect electron transfer (IET) and emerging direct electron transfer (DET) between methanogens and fermentative bacteria, building a more energy-efficient route for interspecies electron transfer and methane productivity. This study demonstrated the positive promotion of the coupled bioelectrochemical regulation and magnetite on organic biodegradation, process stability and CH4 productivity, providing some references for the integrated technology in sludge treatment and bioenergy recovery.

Food waste (FW) and cow manure (CM) were co-digested to achieve a stable and high-rate of methane production. The start-up conditions (substrate mixing (FW/CM) ratio, substrate to inoculum ratio, and initial pH) were optimised, and the optimised parameters were experimentally confirmed by batch operation under mesophilic temperatures. To further verify the effects of start-up conditions on the long-term co-digestion process, a semi-continuous dynamic membrane bioreactor was operated for over 300 days with an FW/CM ratio of 2.5. Following the optimised operation scheme, the organic loading rate gradually increased to 11.9 g COD/L/d. Thus, stable anaerobic co-digestion was maintained at FW/CM ratio of 2.5, and a high CH4 production of 2.71 L CH4/L/d and CH4 yield of 441 mL CH4/g VS was achieved. In the long-term operation, the digestate pH was stable at approximately 8.4, which indicated a very favourable anaerobic reaction condition without volatile fatty acid accumulation.

Rumen fermentation is known to be effective for lignocellulosic-wastes biodegradation to certain extent but it is still unclear if there exists a termination of the microorganisms' action to further degrade the bio-refractory fractions. In order to illuminate the related microbiological characteristics, experiments were conducted in a prolonged duration of rumen fermentation of mechanically ruptured wheat straw, with inoculation of cow rumen microorganisms in vitro. Although the organic wastes could not be biodegraded quickly, continuous conversion of the lignocellulosic contents to volatile fatty acids and biogas proceeded in the duration of more than three months, resulting in 96-97% cellulose and hemicellulose decomposition, and 42% lignin decomposition. X-ray diffraction and Fourier transform infrared spectroscopy further demonstrated the characteristics of lignocellulosic structure decomposition. Under the actions of cow rumen microorganisms, stable pH was maintained in the fermentation liquid, along with a steady NH4+-N, volatile fatty acids accumulation, and a large buffering ability. It was identified by enzyme analysis and Illumina MiSeq sequencing that the rich core lignocellulolytic enzymes secreted by the abundant and diverse rumen bacteria and fungi contributed to the persistent degradation of lignocellulosic wastes. Members of the Clostridiales order and Basidiomycota phylum were found to be the dominant lignocellulolytic bacteria and fungi, respectively. It could thus be inferred that the main lignocellulose degradation processes were a series of catalytic reactions under the actions of lignocellulolytic enzymes secreted from bacteria and fungi. The dominant hydrogenotrophic methanogens (Methanomassiliicoccus, Methanobrevibacter, Methanosphaera, and Methanoculleus) in the rumen could also assist CH4 production if the rumen fermentation was followed with anaerobic digestion.

Cow manure (CM) was added to a dynamic membrane bioreactor (DMBR) operated under anaerobic condition for enhancing food waste (FW) digestion for over 300 days with stepwise increase of organic loading rates (OLRs) from 1.07 to 11.9 g COD/L/day. At a FW/CM ratio of 3.5:1 (based on volatile solids), the mixed liquor pH was always above 8.0 and no apparent volatile fatty acids (VFAs) accumulation occurred even at the highest OLR of 11.9 g COD/L/day (hydraulic retention time as 10 days and solid retention time as 15.5 days, correspondingly), indicating a very stable operation condition which resulted in an average CH4 yield as high as 250 mL/g COD and CH4 production as high as 2.71 L CH4/L/day. The hardly biodegradable organic components, such as cellulose, hemicellulose, and lignin, were effectively degraded by 78.3%, 58.8%, and 47.5%, respectively. Significantly high anaerobic digestion reaction ratios, especially the hydrolysis ratio which is usually the limiting factor, were calculated based on experimental results. Furthermore, the high lignocellulase contents and coenzyme F420 levels, along with the decrease of cellulose crystallinity from 72.6% to 16.4% in the feedstock, provided strong evidence of an enhanced biological activity by CM addition. By high-throughput sequencing analysis, more abundant and diverse bacterial, archaeal, and fungal genera were identified from the DMBR sludge. With CM addition, the biodegradation of lignocellulose might have produced sufficient H2 and CO2 for the hydrogenotrophic methanogens such as Methanoculleus, Methanomassiliicoccus, and Methanobacterium, which were highly tolerant to ammonium inhibition, and then the elevated ammonium level would have provided high buffering capacity in the DMBR thus ensuring a stable condition for high rate FW digestion and CH4 production.

The stabilization and disposal of the large amounts of sewage sludge pose a worldwide challenging problem. To solve this problem, the technical feasibility of coupled alkali-microwave–H2O2–oxidation pretreatment with in-situ bioelectrochemical regulation was used for enhancing sludge biodegradability and methane productivity. The optimum condition of the combined pretreatment was pH 10.0 ± 0.1, microwave 700 W and H2O2 0.4 g/g TS. In this case, soluble chemical oxygen demand (SCOD) of pretreated sludge was increased from initial 330.9 ± 10.0 to 3328.8 ± 49.6 mg/L. The highest accumulative methane yield of 234.3 mL CH4/g VS was obtained for pretreated sludge at the cathodic potential of −0.8 V vs. Ag/AgCl, increasing by 4.3-, and 1.9-fold compared with the raw and pretreated sludge, respectively. The microbial community analysis further provided a compelling evidence that bioelectrochemical regulation stimulated the growth of the functional microorganisms, especially in protein-degrading (Firmicutes), polysaccharides-utilizing (Chloroflexi), electroactive (Geobacter, and Desulfomicrobium) and methane-producing (Methanobacterium) microorganisms. In addition, pH adjustment of pretreated sludge by addition of H2SO4 could further enrich the abundance of microbial community and build a strong syntrophic interaction, accordingly provoking the hydrolysis and subsequent methane production. The result of this study will contribute to the establishment of an efficient sludge stabilization and bioenergy recovery strategy.

Long-term acclimatization of anaerobic sludge was conducted by operating a mesophilic continuously stirred anaerobic reactor (CSTR) with continuous feeding of food wastes (FW) and cow manure (CM). During the long-term acclimatization, continued increase of enzyme activity was revealed, while the microbial structure tended stable as shown by the Shannon index and microbial community. By biomethane potential analysis, the acclimated sludge had a methane yield about 13 times higher than the initial anaerobic sludge. The acclimated sludge was subsequently used for FW digestion with stepwise organic loading rate increase without CM addition. The functional phyla of Bacteroidetes and Proteobacteria, which originated from CM but not very abundant, were significantly enriched not only during sludge acclimatization with CM addition but also in the process of FW digestion without CM addition. A microbe coexistence network was proposed to support an explanation of the metabolic pathways of FW digestion using the acclimated sludge.

To enhance the degradation of wheat straw (WS) and corn straw (CS) in rumen fermentation, characterization of degradation and ruminal microorganisms of monosubstrate (WS/CS) groups and a cosubstrate strategy with food waste (FW) group was performed. The cellulose, hemicellulose, and lignin degradation efficiency of WS and CS; soluble chemical oxygen demand; volatile fatty acid yields; and activity of ligninolytic, cellulolytic, and hemicellulolytic enzymes for the cosubstrate group were improved compared with those for the corresponding monosubstrate groups. An accurate and a good of fit of the Weibull kinetic model, decreased crystallinity index values, and characteristic absorbance bands in the Fourier transform-infrared spectra further confirmed that cosubstrate addition with FW decreased the resistance of cellulose and hemicellulose to biodegradation. High-throughput sequencing results suggested that the bacterial diversity in CS rumen fermentation and fungal diversity and richness in WS rumen fermentation were promoted with FW as a cosubstrate. The cosubstrate addition with FW significantly affected the composition of the ruminal bacteria and fungi in rumen fermentation. The relative abundances (RAs) of rumen bacteria were increased in the cosubstrate CS/WS and FW fermentation conditions, and the enhancement of CS degradation with FW supplementation was stronger than that of WS rumen fermentation with FW supplementation. The RAs of the ruminal fungal genera Ustilago and Fusarium were promoted in CS and WS fermentation with FW, respectively. Moreover, the fermentation properties and rumen flora in the FW rumen fermentation also provided some evidence to suggest an enhancement of the cosubstrate strategy compared with the monosubstrate strategy.