A compound enzyme as an additive to a continuous anaerobic dynamic membrane bioreactor for enhanced lignocellulose removal from codigestion: Performance, membrane characteristics and microorganisms.

To explore the enzyme-enhanced strategy of a continuous anaerobic dynamic membrane reactor (AnDMBR), the anaerobic codigestion system of food waste and corn straw was first operated stably, and then the best combination of compound enzymes (laccase, endo-ß-1,4-glucanase, xylanase) was determined via a series of batch trials. The results showed that the methane yield (186.8 ± 19.9 mL/g VS) with enzyme addition was 12.2 % higher than that without enzyme addition. Furthermore, the removal rates of cellulose, hemicellulose and lignin increased by 31 %, 36 % and 78 %, respectively. In addition, dynamic membranes can form faster and more stably with enzyme addition. The addition of enzymes changed the structure of microbial communities while maintaining sufficient hydrolysis bacteria (Bacteroidetes), promoting the proliferation of Proteobacteria as a dominant strain and bringing stronger acetylation ability. In summary, the compound enzyme strengthening strategy successfully improved the methane production, dynamic membrane effect, and degradation rate of lignocellulose in AnDMBR.

A new substrate equalization method for optimizing the influent conditions and fluid flow patterns of a multifed upflow anaerobic sludge blanket reactor with mature anammox granules.

To improve nitrogen removal efficiency (NRE) and achieve homogenous distribution of anammox sludge and substrate, a new substrate equalization theory and a cumulative overload index was proposed for multifed upflow anaerobic sludge bed (MUASB) reactors with mature anammox granules. The performance and flow patterns of MUASB reactors were investigated under various influent conditions. The results showed that the nitrogen removal performance and stability of MUASB reactors could be optimized by minimizing the cumulative load. The NRE gradually increased from 83.3 ± 2.2 %, 86.8 ± 4.2 % to 89.3 ± 4.1 % and 89.7 ± 1.6 % in feeding flow tests and feeding port tests, respectively. Furthermore, the flow patterns were compared based on residence time distribution and computational fluid dynamics, indicating that a better equilibrium distribution of microorganisms and substrates could be achieved in the MUASB reactors under the lowest cumulative load. Therefore, substrate equalization theory can be used to optimize the nitrogen removal performance of MUASB reactors with low-carbon footprints.

Long-term in-situ starvation and reactivation of co-digestion with food waste and corn straw in a continuous AnDMBR: Performance, sludge characteristics, and microorganism community.

To explore the effects of in-situ starvation and reactivation in a continuous anaerobic dynamic membrane reactor (AnDMBR), the anaerobic co-digestion system of food waste and corn straw was firstly start-up and stability operated, and then stopped feeding substrate approximately 70 days. After long-term in-situ starvation, the continuous AnDMBR was reactivated using the same operation conditions and organic loading rate as the continuous AnDMBR used before in-situ starvation. Results shown that the anaerobic co-digestion of corn straw and food waste in the continuous AnDMBR can resume stable operation within 5 days, and the corresponding methane production of 1.38 ± 0.26 L/L/d was completely returned to the methane production before in-situ starvation (1.32 ± 0.10 L/L/d). Through analysis of the specific methanogenic activity and key enzyme activity of the digestate sludge, only the acetic acid degradation activity of methanogenic archaea can be partially recovered, however, the activities of lignocellulose enzyme (lignin peroxidase, laccase, and endoglucanase), hydrolase (α-glucosidase) and acidogenic enzyme (acetate kinas, butyrate kinase, and CoA-transferase) can be fully recovered. Analysis of microorganism community structure using metagenomic sequencing technology showed that starvation decreased the abundance of hydrolytic bacteria (Bacteroidetes and Firmicutes) and increased the abundance of small molecule-utilizing bacteria (Proteobacteria and Chloroflexi) due to lack of substrate during the long-term in-situ starvation stage. Furthermore, the microbial community structure and key functional microorganism still maintained and similar with that of starvation final stage even after long-term continuous reactivation. The reactor performance and sludge enzymes activity in the continuous AnDMBR co-digestion of food waste and corn straw can be well reactivated after long-term in-situ starvation, even though the microbial community structure can not be recovered to the initiating stage.

Comparative study of the effect of loading increments on the mesophilic codigestion of waste activated sludge and food waste: Reactor performance, stability analysis, and microbial community.

The performance and stability of mesophilic codigestion of waste activated sludge (WAS) and food waste (FW) were compared in two parallel, continuously stirred tank reactors using high- and low-magnitude loading increments for the loading regimes. The results indicated that a high methane (CH4) production of 6.98 L L-1·d-1 was realized without volatile fatty acid accumulation via low-magnitude loading increments at a high loading of 26.5 g-COD·L-1·d-1, and this system was more stable and achieved a higher efficiency than the codigestion system that used high-magnitude loading increments at similar loading and operating conditions. Furthermore, higher CH4 yields of 258-334 mL-CH4·g-COD-1, TCOD removal efficiencies of 64-79%, conversion ratios of 62-88%, and methanogenic activities of 0.37-0.40 g-CH4-COD·g-VS-1·d-1 were consistently maintained via the low-magnitude loading increments during the high-rate period. High abundances of the phyla Firmicutes (63.3%) and genus Methanosarcina (94.5%) contributed to the high rates and stable operating conditions of the mesophilic system for WAS and FW codigestion using low-magnitude loading increments.

Effects of long-term acclimatization on the optimum substrate mixture ratio and substrate to inoculum ratio in anaerobic codigestion of food waste and cow manure.

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.

A comparative study of artificial cow and sheep rumen fermentation of corn straw and food waste: Batch and continuous operation.

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.

Acclimatization of anaerobic sludge with cow manure and realization of high-rate food waste digestion for biogas production.

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.

Stable and high-rate anaerobic co-digestion of food waste and cow manure: Optimisation of start-up conditions.

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.

Cosubstrate strategy for enhancing lignocellulose degradation during rumen fermentation in vitro: Characteristics and microorganism composition.

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.

Persistent action of cow rumen microorganisms in enhancing biodegradation of wheat straw by rumen fermentation.

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 as additive to a DMBR for stable and high-rate digestion of food waste: Performance and microbial community.

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.

Interaction and coexistence characteristics of dissolved organic matter with toxic metals and pesticides in shallow groundwater.

The long-term and large-scale utilization of fertilizers and pesticides in facility agriculture leads to groundwater pollution. However, the coexistence and interactions between organic fertilizers (i.e., organic matter), toxic metals, and pesticides in shallow groundwater have seldom been studied. Thus, the study sought to characterize said interactions via fluorescence, ultraviolet-visible spectroscopy (UV-Vis), and Fourier-transform infrared spectroscopy coupled with two-dimensional correlation spectroscopy and chemometric techniques. The results indicated that groundwater DOM was comprised of protein-, polysaccharide-, and lignin-like substances derived from organic fertilizers. Protein-like substances accounted for the binding of Co, Ni, and Fe, while polysaccharide- and lignin-like substances were mainly responsible for Cr and Mo complexation. Moreover, lignin- and polysaccharide-like substances played a key role in the binding of pesticides (i.e., dichlorodiphenyltrichloroethane [DDT], endosulfan, γ-hexachlorocyclohexane [γ-HCH], monocrotophos, chlorpyrifos, and chlorfenvinphos), rendering the conversion of γ-HCH to ß-hexachlorocyclohexane (ß-HCH) and the degradation of DDT to dichlorobenzene dichloroethylene (DDE) ineffective. However, the presence of protein-like substances in groundwater benefited the degradation and conversion of γ-HCH and α-endosulfan. Redundancy analyses showed that lignin- and polysaccharide-like matter had the most impacts on the coexistence of DOM with toxic metals and pesticides.

The defense system for Bidens pilosa root exudate treatments in Pteris multifida gametophyte.

According to the novel weapons hypothesis, root exudates are the inhibition factors for native species growth and development through invasive plants. It is hypothesized that antioxidant system (AOS) presents an effective role in plant defense system. The allelopathy indexes of P. multifida gametophyte biomass and sporogonium conversions rates turn negative with the dose and time effects, and the synthetical allelopathic effect index was -55.07% at 100% treatments under root exudates treatments. Under transmission electron microscopy, the cell structures turn burry. Next, AOS and programmed cell death (PCD) were tested in this study. In AOS, strong activities of superoxide dismutase, catalase, glutathione reductase and glutathione S-transferase (GST) were identified in gametophyte cells under the treatments, as well as the contents of glutathione, ascorbic acid and reduced ascorbate, while GPX activity decreased. Based on the input (SOD activity) and the output (GST activity) of antioxidant system, and the decreasing system control would be a reason leading gametophyte death under root exudates. At day 10, PCD would get its peak of 46.93% at 100% root exudates. We found a dynamic balance of PCD and AOS under the exudates treatments. We detected hexadecanoic acid, ethylene glycol and undecane are three major chemicals in root exudates. Our results provide a reference of AOS and PCD working under root exudates treatments in plants and offer novel strategy for the native species protection and invasion plants control in environment science.

[Influence of Sludge Retention Time on the Performance and Stability of Mesophilic Anaerobic Co-digestion of Food Waste with Waste Activated Sludge].

Two parallel digestion systems of food waste (FW) and waste-activated sludge (WAS) were successfully initiated using a continuous stirred-tank reactor (CSTR), and the effect of different reduction extents of sludge retention time (SRT) on the co-digestion of FW and WAS was investigated. SRT Reduction extents longer than 8.3 d were not conducive to the stable operation of the co-digestion system when the organic load rate (OLR) was increased. The reduction extent of SRT should be reduced gradually from 5 d to 0.9 d to achieve high load and stable operation of the co-digestion of FW and WAS. After a long-term operation (approximately 282 d), the co-digestion reached stable operation at SRT of 9.1 d and OLR (calculated by COD) of (12.9±1.5) g·(L·d)-1. The corresponding methane production, methane yield (calculated by COD), pH, and volatile fatty acid (VFA, calculated by COD) were 3.94-4.25 L·(L·d)-1, 288-302 mL·g-1, 7.80-7.83, and 0.32-0.39 g·L-1, respectively. Additionly, the sludge characteristics of the co-digestion of FW and WAS under a high loading rate were also investigated. The results showed that the primary pathway of methane conversion was through acetic acid during the co-digestion of FW and WAS. Meanwhile, higher methanogenic activity of acetic acid, propionic acid, butyric acid, valeric acid, and coenzyme F420 concentration were also measured.

Inhibitory effects of heavy metals and antibiotics on nitrifying bacterial activities in mature partial nitritation.

To facilitate the use of partial nitritation (PN) in nitrogen removal processes for livestock wastewater, this work investigated the inhibitory effects of heavy metals and antibiotics on the nitrifying bacterial activities of the PN processes. Biomass was collected from a continuous-flow internal-loop airlift reactor and cultured with fixed ammonium concentrations of 921 mg N L-1. Batch activity tests were conducted to determine the specific oxygen uptake rate. The individual and interactive inhibitory effects of Zn2+, Cu2+, oxytetracycline (OTC) and sulfamethazine (SMZ) were evaluated using an orthogonal test. The results showed that the half maximal inhibitory concentration (IC50) values of Zn2+, Cu2+, OTC and SMZ on PN sludge were 50.1, 35.4, 447 and 1890 mg L-1, respectively. The joint toxicities of heavy metals (Zn2+ and Cu2+) and antibiotics (OTC and SMZ) in the PN mixed culture were generally synergistic, except for between Zn2+ and Cu2+, which was antagonistic. In joint toxicity tests, the significance of the inhibitory effect of Zn2+ (15.3-164.3 mg L-1), Cu2+ (13.8-90.9 mg L-1), OTC (27.0-866.5 mg L-1) and SMZ (290-3490 mg L-1) on the nitrifying bacterial activity can be ranked in the following order: SMZ > Cu2+ > Zn2+ > OTC. Additionally, different exposure times (1 h, 3 h and 24 h) with or without aeration were also comparatively studied. These results show that a greater PN sludge activity loss than when exposure without aeration.

Influence of preservation temperature on the characteristics of anaerobic ammonium oxidation (anammox) granular sludge.

Preserving active anaerobic ammonium oxidation (anammox) biomass is a potential method for securing sufficient seeding biomass for the rapid start-up of full-scale anammox processes. In this study, anammox granules were cultured in an upflow anaerobic sludge blanket (UASB) reactor (R0), and then the enriched anammox granules were preserved at 35, 20, 4, and -30 °C. The subsequent reactivation characteristics of the granules were evaluated in four UASB reactors (denoted R1, R2, R3, and R4, respectively) to investigate the effect of preservation temperature on the characteristics of anammox granules and their reactivation performance. The results demonstrated that 4 °C was the optimal preservation temperature for maintaining the biomass, activity, settleability, and integrity of the anammox granules and their cellular structures. During the preservation period, a first-order exponential decay model may be used to simulate the decay of anammox biomass and activity. The protein-to-polysaccharide ratio in the extracellular polymeric substances and the heme c content could not effectively indicate the changes in settleability and activity of the anammox granules, respectively, and a loss of bioactivity was positively associated with the degree of anaerobic ammonium-oxidizing bacteria cell lysis. After 42 days of storage, the anammox granules preserved at 4 °C (R3) exhibited a better recovery performance than those preserved at 20 °C (R2), -30 °C (R4), and 35 °C (R1). The comprehensive comparison indicated that 4 °C is the optimal storage temperature for anammox granular sludge because it promotes improved maintenance and recovery performance properties.

[Variations and Influencing Factors of Oral Bioaccessibility of Polybrominated Diphenyl Ethers in Soils Using an In-vitro Gastrointestinal Model].

Intake of contaminated soils is considered as an important exposure pathway of polybrominated diphenyl ethers (PBDEs) to humans, especially for children during their outdoor hand-to-mouth activities. Oral bioaccessibility is an essential tool to quantitatively assess the exposure risk of pollutants. In this study, we employed an in vitro digestion model to mimic the gastrointestinal digestion of typical PBDEs (BDE-28, BDE-47, BDE-99 and BDE-153 at a series of initial concentrations) in three natural soil samples with different TOC contents and to verify a previous hypothesis that the sorption of PBDE fraction mobilized from soil into digestive fluid on the surface of residual solid phase may lead to an underestimation of bioaccessibility of PBDEs. In addition, a method with multiple fluid-to-solid ratios was applied to calibrate the underestimation. The results indicated that the calibrated digestibility values were commonly higher than those without correction. For the different soil samples, the averaged increasing rates of PBDE digestibility at different initial concentrations ranged from 14.3% to 42.3%, from 11.1% to 32.1%, from 4.9% to 12.3% and from 0.0% to 7.7% for BDE-28, BDE-47, BDE-99 and BDE-153, respectively. Therefore, the bioaccessibility of PBDEs in gastrointestinal gut would be significantly underestimated without calibration, especially for tri- and tetra-BDEs and soil samples with low TOC contents or high PBDEs concentrations. The corrected digestibility values of BDE-28, BDE-47, BDE-99 and BDE-153 were 21.9%-54.7%, 18.8%-43.1%, 13.4%-27.2% and 9.3%-19.9%, respectively. The results indicated that the PBDEs digestibility was negatively correlated with lgK(ow); whereas there was no significant correlation of PBDE bioaccessibility with TOC contents in soils or with initial concentrations of PBDEs, particularly for the highly brominated components.