纤维素降解菌的筛选及其在果树枝条腐解中的应用

发布时间：2018-03-23 12:50

  本文选题：纤维素降解菌　切入点：复合菌剂　出处：《西北农林科技大学》2015年硕士论文

【摘要】：随着果品产业的大面积扩展,每年修剪果树枝条的量逐年增加,果树枝条堆放占用大面积的土地,焚烧造成环境污染,还浪费了宝贵的生物资源。本研究从堆肥产物和秦岭山区的土壤样品中分离纯化高效纤维素降解菌株,将其组合培养,构建筛选出一组高效的纤维素混合菌群。并制成复合微生物菌剂应用于果树枝条和猪粪堆肥过程中,研究复合微生物菌剂对果树枝条和猪粪堆肥过程中理化性质、酶活性、微生物功能多样性以及微生物遗传多样性的影响。主要结果表现在以下几个方面:1、纤维素降解菌的筛选、鉴定及混合菌群的构建通过刚果红染色法初筛,再以内切酶(CMC)、纤维素全酶(FPA)2种酶的六天酶活性为依据进行复筛,得到5株酶活性较高的纤维素降解菌。经生理生化测定和分子生物学鉴定,菌株2-3、10-2、3-5、6-2和10-4分别为罗尔斯通菌(Ralstoinia sp.)、青霉菌(Penicillium sp.)、黄灰青霉菌(Penicillium aurantioqriseum)、支顶孢菌(Acremonium alternatum)和毛壳霉菌(Chaetomium piluliferum)。对5株菌进行组合培养,得到最优组合为10-2/3-5/2-3/6-2,其CMC、FPA和木聚糖酶活性分别为16.3、58.6和11083.7 U/m L,均比单菌株有一定程度提高。2、复合微生物菌剂对果树枝条堆肥过程中酶活性的影响接种复合微生物菌剂在堆肥前期可以提高堆肥温度,比对照处理高温期(高于55℃)持续时间多3d,促进堆料的腐熟。接种菌剂还能有效地提高果树枝条堆肥过程中酶活性,脲酶、多酚氧化酶、纤维素酶、漆酶、锰过氧化物酶和木质素过氧化物酶活性在堆肥过程中分别比CK处理提高2.3%~71.4%、0.3%~28.4%、15.0%~19.8%、1.0%~11.0%、4.1%~26.8%、4.0%~22.2%。3、复合微生物菌剂对果树枝条堆肥过程中微生物多样性的影响Biolog微平板法测定结果表明,接种复合菌剂显著提高了堆肥中微生物的平均每孔颜色变化率(average well color development),并改变了高温期和降温期微生物对6大类碳源的利用;主成分分析表明,复合菌剂主要在高温期发挥作用,对堆料中微生物起分异作用的碳源主要为糖类和氨基酸类。测序结果表明,好氧堆肥过程中优势细菌主要为芽孢杆菌属(Bacillus sp.PML14)、不动杆菌属(Pseudomonas sp.)、厚壁菌门属(Firmicutes bacterium)、假单孢菌属(Pseudomonas sp.)、席藻菌属(Phormidium sp)和大量的用传统方法不能培养但在堆肥过程中发挥了重要作用的细菌,其中Bacillus sp.、Uncultured bacterium isolate DGGE gel band、Phormidium sp.和Uncultured beta proteobacterium isolate DGGE gel band在堆肥的高温期发挥了主要作用。接种菌剂处理能够显著提高堆肥过程的细菌群落多样性,促进细菌群落结构的演替,加快堆肥的腐熟进程。
[Abstract]:With the extensive expansion of fruit industry, the amount of pruning fruit tree branches increases year by year, and fruit tree branches pile up and occupy a large area of land, causing environmental pollution caused by incineration. High efficient cellulose degrading strains were isolated from compost products and soil samples from Qinling Mountain area and cultured in combination. A group of efficient cellulosic mixed bacteria was constructed and selected, and the compound microbial fungicides were used in the process of fruit tree branch and pig manure composting. The physicochemical properties and enzyme activity of the compound microbial fungicides on fruit tree branch and pig manure composting were studied. Effects of microbial functional diversity and microbial genetic diversity. The main results were as follows: 1. Screening and identification of cellulosic degrading bacteria and construction of mixed bacteria by Congo red staining. On the basis of six days enzyme activity of two enzymes, CMCX and FPAN, five strains of cellulose degrading bacteria with high enzyme activity were obtained, which were identified by physiological and biochemical tests and molecular biology. Strains 2-3, 10-2, 3-5, 6-2 and 10-4 were Ralstoinia sp., Penicillium sp., Penicillium aurantioqriseumum, Acremonium alternatum and Chaetomium piluliferum, respectively. The optimum combination was 10-2 / 3-5 / 2-3-3 / 6-2, and the CMCFPA and xylanase activities were 16.3U / 58.6 and 11083.7 U / mL, respectively, which were higher than that of single strain to some extent. The effect of compound microbial agents on enzyme activity in composting process of fruit trees was studied. At the early stage of composting, the composting temperature can be raised. Compared with the control treatment, the duration of high temperature period (above 55 鈩，

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