Screening Of Cellulose-degrading Bacteria And Its Application In Agricultural And Forestry Waste Recycling

Anaerobic digestion of organic waste to produce clean energy-methane is one of the effective ways to deal with environmental pollution and energy crisis.However,the refractory characterization of lignocellulosic material,which is the most commonly feedstock used for anaerobic digestion,limits the application of biogas fermentation technology.Using cellulose-degrading strain or enzymes to promote the degradation of lignocellulosic material is considered to be one of the effective ways to improve the efficiency of anaerobic digestion.In this paper,samples were collected from different environments of Tibet to isolate and purify cellulose-degrading strains,and the growth conditions and enzyme-producing properties of the strains were analyzed.Besides,the effects of the cellulose-degrading strain or commercial cellulase,hemicellulase and laccase on anaerobic digestion of organic waste was studied.Meanwhile,the effect of anaerobic digestion residue on seed germination was studied.The main conclusions are as follows:1.Five cellulose-degrading strains were isolated from different habitats of Tibet,and the degradation performance of carboxymethyl cellulase,filter paper enzyme,hemicellulase and laccase production of the five strains were analyzed by semi-quantitatively.The strain with best performance was identified using morphological and molecular biological method.The strain belongs to white rot fungus,is highly affiliated to Trametes sp.4267,and named as Trametes sp.W-4.2.Conditions for enzyme production of strain W-4 were optimized using single factor method.The effects of growth temperature,nitrogen source and fermentation time on the activity of carboxymethyl cellulase(CMCase)were analyzed.The activities of filter paper enzyme,xylanase and laccase of strain W-4 were determined under the optimal conditions.The results showed that the optimal growth temperature of strain W-4 was30℃,and peptone was the best nitrogen source.Under the optimal growth temperature and nitrogen source conditions,CMCase activity reached the highest value of 3.05 U/m L at 30 h.The analysis to the enzyme reaction conditions of CMCase showed that the optimal temperature was 50℃ and the optimal p H was 4.0.Under these conditions,the activity of filter paper enzyme,xylanase and laccase of the strain W-4 were 0.41 U/m L,0.29U/m L and 1.11 U/m L,respectively.3.The organic wastes including straw,cow dung and pig dung,were pretreated with strain W-4 for 0 day(control group),3 days,7 days,and 14 days to explore the effects of strain pretreatment on the anaerobic digestion.The results showed that under sterilization condition,the maximum biogas yield of 248.29 m L/g-VS(standard condition)was obtained from the pretreatment of 3 days,which was 75.29% higher than the control group.The cumulative biogas yield of the control group was the highest when the substrate was not sterilized,which was 182.27 m L/g-VS.In addition,whether the substrate was sterilized or not,the TS,VS,COD and ammonia nitrogen of the substrate decreased in different degrees after both the pretreatment and anaerobic fermentation,while the p H increased.4.To investigate the influence of the added enzymes on biogas fermentation,different combinations and concentrations of the commercial cellulase,hemicellulose and laccase were directly added into the anaerobic fermentation system using straw and fresh cow dung as feedstock.The results showed that when the enzymes were free,the maximum biogas production of 177.71 m L/g-VS was obtained from the group ASE2,which was 59.57% higher than that of CK group.When the enzymes were immobilized,the cumulative biogas production could be increased to251.67 m L/g-VS,which was 125.98% higher than that of CK group.5.The influence of anaerobic fermentation residue on the seed germination of C.angustifolium subsp.circumvagu was detected.The results showed that the anaerobic fermentation residue has a positive effect on the germination no matter which treatment was conducted.Among them,the highest germination rate of pretreatment group,free enzymes group and immobilized enzymes group could reach to 82.67%,86.00% and 84.00%respectively.