Obtainment of lignocellulose degradation microbial community: the effect of acid-base combination after restrictive enrichment.

Microbial communities for bioconversion of lignocellulose have received widespread attention. Many cellulose-degrading microbial communities have been enriched from different sources. Combining two microbial communities with acidic and basic properties (acid-base combination) is a technique used alongside restricted enrichment culturing. Understanding how changes to microbial communities result in community's structure and function is important for mechanistic reconstruction of microbiomes. In this study, we analyzed changes in microbial community structure to elucidate determination of the mechanisms of acid-base combination. We found that after restricted enrichment, the bacteria that were primarily retaining included not only those that decompose and utilize lignocellulose, such as Clostridium and Pseudomonas, but also synergistic microbiota such as Alkalobacillaceae. When the proportion of these two types of bacteria was unbalanced, the degradative ability of the microbial community was low, or pH changes of it did not compound regular changes, which may lead to the failure of restricted enrichment. Microbial communities were re-constituted by acid-base combination, whereby the degrading and synergistic strains were adjusted to a more appropriate proportion. The acid-base combination fixed the instability of microbial communities caused by the randomness of restrictive screening enrichment; it provided an effective method for obtaining high-quality lignocellulose-degrading microbial community.

Bioaugmentation with methanogens cultured in a micro-aerobic microbial community for overloaded anaerobic digestion recovery.

Anaerobic digestion (AD) is widely used for conversion of waste materials into biogas, but inhibition of methane production caused by overloading can be a major problem. The micro-aerobic microbial community MC1 was used to successfully culture methanogens, Methanosarcina acetivorans C2A and Methanosaeta thermophila NBRC 101360. The maximum 16S rRNA gene concentrations of Methanosarcina acetivorans C2A and Methanosaeta thermophila NBRC101360 were 1.06 × 106 and 1.35 × 103 copies/mL, respectively. The five key bacteria in MC1 were quantified to assess the effect of inoculation on the abundances of the bacteria in the mixed culture. The original MC1 total 16S rRNA gene concentration was 1.93 × 108 copies/mL, and the total 16S rRNA gene concentration had increased to 4.79 × 109 copies/mL on day 9 (p < 0.05). The proportions of the key strains in MC1+MST had changed by day 9. Cells were harvested and used to bioaugment and increase the pH values of the high- and medium-temperature anaerobic systems. After bioaugmentation, thermophilic AD recovered well. The cumulative amounts of gas produced were 44.78% and 28.28% higher in the MC1+MST and MC1 groups, respectively, than the sterilized control. The MC1+MST group gave better results than the chemical addition control group (CaCO3). There was no clear effect of bioaugmentation in mesophilic AD. When compared with traditional pure culture of methanogens as inoculants, methanogen cultivation in MC1 was simple and there was no need to separate and purify the target strains. This simplified methanogenic bioaugmentation agent was useful to study the mechanism of bioaugmentation for the recovery from low pH inhibition, showing the potential for practical application.

Composted biogas residue and spent mushroom substrate as a growth medium for tomato and pepper seedlings.

A composted material derived from biogas production residues, spent mushroom substrate (SMS) and pig manure was evaluated as a partial or total replacement for peat in growth medium for tomato and pepper seedlings. Five different substrates were tested: T1, compost + perlite (5:1, v:v); T2, compost + peat + perlite (4:1:1, v:v:v); T3, compost + peat + perlite (2.5:2.5:1, v:v:v); T4, compost + peat + perlite (1:4:1, v:v:v); and CK, a commercial peat + perlite (5:1, v:v). The physical-chemical characteristics of the various media were analyzed, and the germination rate and morphological growth were also measured. Real-time Quantitative PCR (qPCR) was used to quantify Fusarium concentrations. The addition of compost to peat-based growth medium increased the pH, electrical conductivity, air porosity, bulk density, and nutrition (NPK), and decreased the water holding capacity and total porosity. The use of compost did not affect the percent germination at day 15 of the tomato and pepper seedlings. The addition of compost resulted in better or comparable seedling quality compared with CK and fertilized CK. The best growth parameters were seen in tomato and pepper seedlings grown in T1 and T2, with higher morphological growth in comparison with CK and fertilized CK. However, T2 showed the highest Fusarium concentration compared to compost and all growth media. Fusarium concentrations in T1, T3, and T4 did not differ significantly from those in CK for tomato seedlings, and those in T1 and T4 were also similar to those in CK for pepper seedlings. The results suggest that biogas residues and SMS compost is a good alternative to peat, allowing 100% replacement, and that 20-50% replacement produces tomato and pepper seedlings with higher morphological growth and lower Fusarium concentrations.

[Composition diversity and metabolic characters of lactic acid bacteria community SGL].

OBJECTIVE: We aimed to select a stable lactic acid bacteria community from switchgrass silage, that was efficient in lactic acid production. METHODS: We obtained the community by continuous restricted subcultivation in MRS broth, and analysed the composition diversity and stability of the community by 16S rRNA gene-based pyrosequencing and Denaturing Gradient Gel Electrophoresis (DGGE), respectively. In addition, we studied the effect of different nitrogen sources on growth and lactic acid production of the community, through adding different concentrations of yeast extraction, different nitrogen sources [yeast extract, peptone, urea and (NH4) 2SO4] and different proportions of (NH4)2SO4 and yeast extract leveled with elemental nitrogen 1.8 g/L. RESULTS: The microbial composition of SGL became stable from the 8th generation according to the results of DGGE. The pH value of the MRS inoculated with SGL dropped to 3.7, and the concentration of lactic acid reached 26 g/L after 24 h cultivation. The result of the pyrosequencing showed that the major composition of SGL were Lactobacillus nantensis (78.78%), Lactobacillus plantarum (7.92%), Lactobacillus pantheris (5.27%), Bacillus coagulans (4.41%) and Lactococcus lactics (3.31%). The best supplementation of yeast extraction for SGL was 20 g/L. When the elemental nitrogen ratio of (NH4) 2SO4 to yeast extract was 1:4, the growth and lactic acid production were no significant difference with 0:5 (P < 0.05). CONCLUSION: SGL had a great potential of application, as an efficient inoculant for ensilage or lactic acid production. This study would offer theoretical basis for cultivate and application of SGL in production.

Food waste impact on dry anaerobic digestion of straw in a novel reactor: Biogas yield, stability, and hydrolysis-methanogenesis processes.

Gradient anaerobic digestion reactor (GADR) can improve substrate utilization efficiency by solving the problem of the "short circuit" of materials. However, the substrate's composition significantly affects the reactor's performance. This study investigated the impact of food waste (FW) levels on corn straw's dry anaerobic digestion (AD) in a novel GADR. The results show that biomethane production can be improved by coupling urban and agricultural solid waste recycling. The mechanism is to increase the hydrolysis and acid production efficiency, and the abundance of enzymes related to methanogenesis. The maximum methane yield (494.2 mL CH4/g VS) and the highest anaerobic biodegradability (85.7 %) were obtained when the FW was added at 60 %. The co-digestion of FW and straw can improve the hydrolysis and acid production efficiency and methane yield, which improves the buffering capacity and stability of the system compared with the single digestion of FW.

Effect of bioaugmentation on gas production and microbial community during anaerobic digestion in a low-temperature fixed-bed reactor.

Low temperature is one of the limiting factors for anaerobic digestion in cold regions. To improve the efficiency of anaerobic digestion for methane production in stationary reactors under low-temperature conditions, and to improve the structure of the microbial community for anaerobic digestion at low temperatures. We investigated the effects of different concentrations of exogenous Methanomicrobium (10, 20, 30%) and different volumes of carbon fiber carriers (0, 10, 20%) on gas production and microbial communities to improve the performance of low-temperature anaerobic digestion systems. The results show that the addition of 30% exogenous microorganisms and a 10% volume of carbon fiber carrier led to the highest daily (128.15 mL/g VS) and cumulative (576.62 mL/g VS) methane production. This treatment effectively reduced the concentrations of COD and organic acid, in addition to stabilizing the pH of the system. High-throughput sequencing analysis revealed that the dominant bacteria under these conditions were Acidobacteria and Firmicutes and the dominant archaea were Candidatus_Udaeobacter and Methanobacterium. While the abundance of microorganisms that metabolize organic acids was reduced, the functional abundance of hydrogenophilic methanogenic microorganisms was increased. Therefore, the synergistic effect of Methanomicrobium bioaugmentation with carbon fiber carriers can significantly improve the performance and efficiency of low-temperature anaerobic fermentation systems.

Synergistic analysis of lignin degrading bacterial consortium and its application in rice straw fiber film.

Fiber film have received widespread attention due to its green friendliness. We can use microorganisms to degrade lignin in straw to obtain cellulose and make fiber films. Herein, a group of high-temperature (50 °C) lignin degrading bacterial consortium (LDH) was enriched and culture conditions for lignin degradation were optimized. Combined with high-throughput sequencing technology, the synergistic effect of LDH-composited bacteria was analyzed. Then LDH was used to treat rice straw for the bio-pulping experiment. The results showed that the lignin of rice straw was degraded 32.4 % by LDH at 50 °C for 10 d, and after the optimization of culture conditions, lignin degradation rate increased by 9.05 % (P < 0.001). The bacteria that compose in LDH can synergistically degrade lignin. Paenibacillus can encode all lignin-degrading enzymes present in the LDH. Preliminary tests of LDH in the pulping industry have been completed. This study is the first to use high temperature lignin degrading bacteria to fabricate fiber film.

Optimization of semi-continuous dry anaerobic digestion process and biogas yield of dry yellow corn straw: Based on "gradient anaerobic digestion reactor".

In China, the problem of low biogas yield of traditional biogas projects has become increasingly prominent. This study investigated the effects of different hydraulic retention times (HRTs) on the biogas production efficiency and microbial community under pilot conditions. The results show that the "Gradient anaerobic digestion reactor" can stably carry out semi-continuous dry anaerobic digestion and improve biogas yield. The highest volatile solids (VS) biogas yield (413.73 L/kg VS and 221.61 L CH4/kg VS) and VS degradation rate (48.41%) were observed at an HRT of 25 days. When the HRT was 15 days, the volumetric biogas yield was the highest (2.73 L/L/d, 1.43 L CH4/L/d), but the VS biogas yield and degradation rate were significantly decreased. Microbial analysis showed that HRT significantly affected microbial community. It provides basic data support for the development of a new anaerobic digestion process and the practical application of the straw biogas project in China.

Effect of temperature and storage methods on liquid digestate: Focusing on the stability, phytotoxicity, and microbial community.

Identifying the stability and phytotoxicity of liquid digestate (LD) is necessary for safe agricultural utilization. Storage temperature, method, and time are critical factors that affect the stability and phytotoxicity of LD. This study therefore aimed to explore the dynamics of stability, phytotoxicity, and microbial community of LD in cattle farms under different storage conditions. The results showed that the contents of solids, organic matter, nitrogen, and phosphorous decreased during storage and exhibited temperature dependency. Conversely, the seed germination index increased, which was negatively correlated with dissolved organic carbon and ammonium nitrogen and positively correlated with certain bacteria (Thermovirga and Fastidiosipila). Open storage and/or higher temperature were found to contribute to the stabilization efficiency and phytotoxicity disappearance of LD. Open storage of LD at 30 °C for 60 days and 20 °C for 90 days was safe for its agricultural utilization, while hermetic storage of LD at 30 °C for 120 days and 20 °C for 150 days was safe. However, for storage at 10 °C for 180 days, additional post-treatment is required.

The effects of adding exogenous lignocellulose degrading bacteria during straw incorporation in cold regions on degradation characteristics and soil indigenous bacteria communities.

Low temperature is one of the bottleneck factors that limits the degradation of straw during rice straw incorporation. Determining strategies to promote the efficient degradation of straw in cold regions has become a highly active research area. This study was to investigate the effect of rice straw incorporation by adding exogenous lignocellulose decomposition microbial consortiums at different soil depths in cold regions. The results showed that the lignocellulose was degraded the most efficiently during straw incorporation, which was in deep soil with the full addition of a high-temperature bacterial system. The composite bacterial systems changed the indigenous soil microbial community structure and diminished the effect of straw incorporation on soil pH, it also significantly increased rice yield and effectively enhanced the functional abundance of soil microorganisms. The predominant bacteria SJA-15, Gemmatimonadaceae, and Bradyrhizobium promoted straw degradation. The concentration of bacterial system and the depth of soil had significantly positive correlations on lignocellulose degradation. These results provide new insights and a theoretical basis for the changes in the soil microbial community and the application of lignocellulose-degrading composite microbial systems with straw incorporation in cold regions.

Biogas Residues Improved Microbial Diversity and Disease Suppression Function under Extent Indigenous Soil Microbial Biomass.

Indigenous soil microbial biomass (ISMB) plays a key role in maintaining essential functions and biodiversity of soil health. One of the critical unknowns is how the indigenous microorganisms respond to different fertilizers which is directly related to agricultural production. Therefore, we used Mi-Seq sequencing and network analyses to compare the response of ISMB to biogas residue and chemical fertilizers. The results showed that crop production was profoundly influenced by levels of ISMB present and is further dependent on the strategy of fertilizer application. Higher ISMB primarily manifests through retention of richer microbial abundance, a balanced community structure, and tightened co-occurrence within a certain proportion of Nitrospirae, Rhizophlyctidaceae, and Gemmatimonadetes. Compared to chemical fertilizer, biogas residue resulted in higher production with more strongly linked nodes such as Actinobacteria, Chloroflexi and Gemmatimonadetes. Under the same level of ISMB, the microbial diversity was richer and co-occurrence was tighter when biogas residues were applied compared with chemical fertilizer. In addition, the higher level of ISMB with biogas residue applied had a lower abundance of potential fungal pathogens in both bulk and rhizosphere soil compared with chemical fertilizer. This study provides critical data to understand the influence of ISMB and biogas residue on soil ecological system.

An innovative strategy to enhance the ensiling quality and methane production of excessively wilted wheat straw: Using acetic acid or hetero-fermentative lactic acid bacterial community as additives.

Ensiling is an effective storage strategy for agricultural biomass, especially for energy crops (mainly energy grasses and maize). However, the ensiling of excessively wilted crop straw is limited due to material characteristics, such as a high lignocellulosic content and low water-soluble carbohydrate and moisture contents. In this study, acetic acid or hetero-fermentative lactic acid bacterial community (hetero-fermentative LAB) were employed as silage additives to improve the ensiling process of excessively wilted wheat straw (EWS). The results showed that the additives inhibited the growth of Enterobacteriaceae and Clostridium_sensu_stricto_12, whose abundances decreased from 55.8% to 0.03-0.2%, respectively. The growth of Lactobacillus was accelerated, and the abundances increased from 1.3% to 80.1-98.4% during the ensiling process. Lactic acid fermentation was the dominant metabolic pathway in the no additive treatment. The additives increased acetic acid fermentation and preserved the hemicellulose and cellulose contents, increasing the methane yield by 17.7-23.9%. This study shows that ensiling with acetic acid or hetero-fermentative LAB is an effective preservation and storage strategy for efficient methane production from EWS.

Microbial population dynamics and changes in main nutrients during the acidification process of pig manures.

This study evaluated the impact of pig manure acidification on anaerobic treatment and composition of the fecal microbial community. According to the different chemical oxygen demand (COD) in the anaerobic treatment processes, pig manure was diluted 4 times (x4), 16 times (x16), or 64 times (x64) and subjected to acidification. During the acidification process, pH, soluble chemical oxygen demand (SCOD), volatile fatty acids (VFAs), nitrogen (N), phosphorus (P) and potassium (K) were determined along with microbial population dynamics. The pH of the three dilutions first declined, and then slowly increased. The total VFAs of x4 and x16 dilutions peaked on day 15 and 20, respectively. The content of acetic acid, propanoic acid, butanoic acid and valeric acid of the x4 dilution were 23.6, 11.4, 8.8 and 0.6 g/L respectively, and that of the x16 dilution was 5.6, 2.3, 0.9 and 0.2 g/L respectively. Only acetic acid was detected in the x64 dilution, and its level peaked on day 10. The results showed that the liquid pig manure was more suitable to enter the anaerobic methanogenic bioreactors after two weeks of acidification. During the acidification process, total P concentration increased during the first ten days, then dropped sharply, and rose again to a relatively high final concentration, while total N concentration rose initially and then declined. Based on the analysis of denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone library, we concluded that the acidification process reduced the number of pathogenic bacteria species in pig manure.

Characterization of the effective cellulose degrading strain CTL-6.

An efficient cellulose degrading bacteria exists in the thermophilic wheat straw-degrading community, WDC2. However, this strain cannot be isolated and cultured using conventional separation techniques under strict anaerobic conditions. We successfully isolated a strain of effective cellulose degrading bacteria CTL-6 using a wash, heat shock, and solid-liquid alternating process. Analysis of its properties revealed that, although the community containing the strain CTL-6 grew under aerobic conditions, the purified strain CTL-6 only grew under anaerobic culture conditions. The strain CTL-6 had a striking capability of degrading cellulose (80.9% weight loss after 9 days of culture). The highest efficiency value of the endocellulase (CMCase activity) was 0.404 micromol/(min mL), cellulose degradation efficiency by CTL-6 was remarkably high at 50-65 degrees C with the highest degradation efficiency observed at 60 degrees C. The 16S rRNA gene sequence analysis indicated that the closest relative to strain CTL-6 belonged to the genus Clostridium thermocellum. Strain CTL-6 was capable of utilizing cellulose, cellobiose, and glucose. Strain CTL-6 also grew with Sorbitol as the sole carbon source, whereas C. thermocellum is unable to do so.

[Microbial diversity of a community for ensiling rice straw at low temperature and fermentation dynamics].

OBJECTIVE: To accelerate the conversion of rice straw into feeds in the low-temperature region, a microbial community was constructed by continuous enrichment cultivation. Microbial diversity and dynamics during the fermentation at 10 degrees C was analyzed. METHODS: The community was selected at 5 degrees C under static condition. To analyze the inoculating effects, the community and commercial inoculant ( CI: composed of Lactobacillus plantarum, Enterococcus faecium, L. salivarilus, Pediococcus acidilactici) were respectively inoculated into the rice straw for 30 d fermentation at 10 degrees C. Fermented products were detected by gas chromatography - mass spectrometry (GC-MS). Composition microorganisms of the community were analyzed using cloning library. Microbial dynamics during the fermentation was detected by denatured gradient gel eletrophoresis (DGGE). Quantitative PCR was used for tracking the composition microorganisms of the community during the fermentation. RESULTS: The results from 16S rDNA cloning library showed that the community was mainly composed of Lactobacillus spp. and Leuconostoc spp. At 6d fermentation, the pH and the lactic acid bacterial colony forming units (LAB CFUs) in the fermented rice straw with the community amounted to 4.3 and 2.9 x 10(9) CFU/g fresh matter (FM), respectively. The pH and LAB CFUs with the CI were respectively 5.3 and 2.9 x 10(9) CFU/g FM. At 30 d fermentation, the lactic acid concentrations with the community and the CI were respectively 8.1 g/kg FM and 2.0 g/kg FM. From DGGE patterns, both L. sakei and Leuconostoc inhae of the community were detected at 6d fermentation and existed during the fermentation. For the treatment with the CI, the uncultured bacterium was detected at 6d fermentation besides the composition microorganisms of the CI. At 16d and 30d fermentation, only L. plantarum and E. faecium were detected. Quantitative PCR showed DNA mass of L. sakei amounted to 41.0% at 6d fermentation in the treatment with the community. At 16d, DNA mass of L. sakei was 65%. The highest value (5.5%) of DNA mass of Le inhae appeared at 6d of fermentation. CONCLUSION: The community could effectively colonize into the rice straw fermentation system and accelerate the fermentation process at low temperature. The dominating microorganism of the community was L. sakei at 10 degrees C.

Metagenomic Insight into Lignocellulose Degradation of the Thermophilic Microbial Consortium TMC7.

Biodegradation is the key process involved in natural lignocellulose biotransformation and utilization. Microbial consortia represent promising candidates for applications in lignocellulose conversion strategies for biofuel production; however, cooperation among the enzymes and the labor division of microbes in the microbial consortia remains unclear. In this study, metagenomic analysis was performed to reveal the community structure and extremozyme systems of a lignocellulolytic microbial consortium, TMC7. The taxonomic affiliation of TMC7 metagenome included members of the genera Ruminiclostridium (42.85%), Thermoanaerobacterium (18.41%), Geobacillus (10.44%), unclassified_f__Bacillaceae (7.48%), Aeribacillus (2.65%), Symbiobacterium (2.47%), Desulfotomaculum (2.33%), Caldibacillus (1.56%), Clostridium (1.26%), and others (10.55%). The carbohydrate-active enzyme annotation revealed that TMC7 encoded a broad array of enzymes responsible for cellulose and hemicellulose degradation. Ten glycoside hydrolases (GHs) endoglucanase, 4 GHs exoglucanase, and 6 GHs ß-glucosidase were identified for cellulose degradation; 6 GHs endo-ß-1,4-xylanase, 9 GHs ß-xylosidase, and 3 GHs ß-mannanase were identified for degradation of the hemicellulose main chain; 6 GHs arabinofuranosidase, 2 GHs α-mannosidase, 11 GHs galactosidase, 3 GHs α-rhamnosidase, and 4 GHs α-fucosidase were identified as xylan debranching enzymes. Furthermore, by introducing a factor named as the contribution coefficient, we found that Ruminiclostridium and Thermoanaerobacterium may be the dominant contributors, whereas Symbiobacterium and Desulfotomaculum may serve as "sugar cheaters" in lignocellulose degradation by TMC7. Our findings provide mechanistic profiles of an array of enzymes that degrade complex lignocellulosic biomass in the microbial consortium TMC7 and provide a promising approach for studying the potential contribution of microbes in microbial consortia.

Degradation of lignocelluloses in straw using AC-1, a thermophilic composite microbial system.

In composting, the degradation of lignocellulose in straw is problematic due to its complex structures such as lignin. A common solution to this problem is the addition of exogenous inoculants. AC-1, a stable thermophilic microbial composite, was isolated from high temperature compost samples that can decompose lignocellulose at 50-70 °C. AC-1 had a best degradation efficiency of rice straw at 60 °C (78.92%), of hemicellulose, cellulose and lignin were 82.49%, 97.20% and 20.12%, respectively. It showed degrad-ability on both simple (filter paper, absorbent cotton) and complex (rice straw) cellulose materials. It produced acetic and formic acid during decomposition process and the pH had a trend of first downward then upward. High throughput sequencing revealed the main bacterial components of AC-1 were Tepidimicrobium, Haloplasma, norank-f-Limnochordaceae, Ruminiclostridium and Rhodothermus which provides major theoretical basis for further application of AC-1.

The effects of C/N (10-25) on the relationship of substrates, metabolites, and microorganisms in "inhibited steady-state" of anaerobic digestion.

The carbon/nitrogen ratio (C/N) is a key parameter that affects the performance of anaerobic digestion (AD). Recent AD research has focused on optimizing the C/N of feedstock. The so-called "inhibited steady-state" refers to a special state of ammonia inhibition of AD that often occurs at low-C/N (below 25) when degradable nitrogen-rich substrates, such as livestock manure, are used as feedstock. However, the mechanism behind the "inhibited steady-state" is still unknown. In the current study, co-digestion and recirculation were used to create a C/N gradient in the influent to explore the relationship between substrates, metabolites, and microorganisms in the "inhibited steady-state." Data were collected at the macro, microbial, and genetic levels. Three CSTRs were successfully made run into the "inhibited steady-state" using influent C/Ns of 10-12. Digestion performance levels of R10-R12 were low and stable, transitioning from an aceticlastic methane-producing pathway to a hydrogenotrophic pathway as the C/N gradually decreased. As the abundance of the hydrogenophilic methanogens increased, the abundance of syntrophic acetate-oxidizing bacteria (SAOB) also increased. The succession between populations of Methanosaeta and Methanosarcina may be used as a microbiological indicator of ammonia inhibition. Under high-C/Ns, cooperation among bacteria was high, while under low-C/Ns, competition among bacteria was high. These results clarify the processes underlying the "inhibited steady-state," which is a condition often faced in actual large-scale biogas facilities that use degradable nitrogen-rich substrates. Moreover, practical guidelines for evaluating ammonia inhibition are provided, and strategies to alleviate ammonia suppression are developed.

Functional characteristics and diversity of a novel lignocelluloses degrading composite microbial system with high xylanase activity.

To obtain an efficient natural lignocellulolytic complex enzyme, we screened an efficient lignocellulose degrading composite microbial system (XDC-2) from composted agricultural and animal wastes amended soil following a long-term directed acclimation. The XDC-2 could not only degrade natural lignocelluloses, but could also secret extracellular xylanase efficiently in liquid culture under static conditions at room temperature. The XDC-2 degraded rice straw by 60.3% after fermentation for 15 days. Hemicelluloses were decomposed effectively, while the extracellular xylanase activity was dominant with an activity of 8.357 U ml-1 on day 6 of the fermentation period. The extracellular crude enzyme noticeably hydrolyzed natural lignocelluloses. The optimum temperature and pH for the xylanase activity were 40 degrees and 6.0. However, the xylanase was activated in a wide pH range of 3.0-10.0, and retained more than 80% of its activity at 25-35 degrees and pH 5.0-8.0 after three days of incubation in liquid culture under static conditions. PCR-DGGE analysis of successive subculture indicated that the XDC-2 was structurally stable over long-term restricted and directed cultivation. Analysis of 16S rRNA gene clone library showed that the XDC-2 was mainly composed of mesophilic bacteria related to the genera Clostridium, Bacteroides, Alcaligenes, Pseudomonas, etc. Our results offer a new approach to exploring efficient lignocellulolytic enzymes by constructing a high-performance composite microbial system with synergistic complex enzymes.

A comparison and evaluation of the effects of biochar on the anaerobic digestion of excess and anaerobic sludge.

The mechanisms and enhancing effects of different biochar loadings on the digesters receiving low and high excess (or anaerobic) sludge loadings were thoroughly examined in the present study. This was done to explore an efficient method for converting excess sludge to anaerobic sludge. Biochar had an obvious effect on the anaerobic digestion of excess sludge but not on the anaerobic sludge. When the amount of biochar added was equivalent to 100% of the sludge TS, the cumulative methane yields of anaerobic digestion inoculated with small and large amounts of excess sludge were respectively 30.2 and 1.7 times that of those without biochar. The number of methanogens in the digesters that received small and large inoculations of excess sludge with 100% biochar, were respectively 105.4% and 20.6% higher than those without biochar. The biochar enhanced the systems performance because it selectively enriched the Trichococcus and Methanomicrobiales tightly attach to it. This enhanced the synergy and overall activity of the system by promoting biofilm development. Ultimately, the integration of 100% biochar and excess sludge can be used as a substitute for anaerobic sludge as an inoculum by giving similar overall performance.