耐铅锌菌群的鉴定及其去除铅锌特性研究
发布时间：2018-10-31 14:10
【摘要】：长期生活在重金属污染环境的微生物部分已具备耐受重金属的能力,从环境中将其分离驯化,是目前获得高效微生物修复资源的有效途径。通常,单一的微生物易受环境影响,难以广泛应用。而经过长期适应的多种微生物构成的协同菌群具有一定的稳定性,同时兼具一定系统功能性,在强化植物修复方面颇具潜力。本文以铅锌尾矿区有机菌肥处理组与对照组获得的可培养根际菌群为研究对象,通过正交试验对比其去除铅锌的能力及特性的差异,优化培养条件以获得高效的耐性菌群。通过营养液-沙培试验分析耐性菌群对铅锌胁迫下蓖麻种子萌发、幼苗生长及重金属积累的影响机制,结合高通量测序技术对耐性菌群的结构及多样性进行分析鉴定。主要研究结论如下:1)在铅锌复合胁迫下,有机菌肥处理组与对照组获得的可培养根际菌群去除铅的差异大,去除锌的差异不显著。菌群LJ(加有机菌肥)的铅、锌去除率较菌群LJO(未加有机菌肥)有不同程度的增加,最大增量可达40.2%、12.7%。铅锌浓度、培养基类型等因素对菌群LJ的铅、锌去除率有一定影响,其中培养基类型的影响最大。综合而言,菌群LJ去除铅锌的最佳培养条件为:铅锌浓度50mg/L、接菌量1.0mL、培养时间6 d、牛肉膏蛋白胨培养基。2)优化培养条件获得的两组耐性菌群对蓖麻初期的生长发育产生了不同的影响。在种子萌发阶段,接种牛肉膏菌群(细菌)对发芽有一定促进作用,马丁氏菌群(真菌)则呈现抑制作用。在幼苗生长发育阶段,牛肉膏菌群有利于植物株高的伸长,马丁氏菌群则更益于幼苗根系的发育。蓖麻幼苗对锌的积累及转运能力均强于铅,重金属主要积累在根部。两组菌群对植物根系吸收锌的抑制作用较明显,对铅的吸收影响较小。在较高铅锌浓度(100 mg/L)胁迫下,接种两组菌群更有利于植物对铅锌的向上转移。3)蓖麻根际的耐性细菌类群丰富,优势细菌门为变形菌门,优势细菌纲为蓝细菌纲、芽孢杆菌纲,优势细菌属为剑菌属、贪铜菌属。100 mg/L铅锌胁迫下,耐性细菌群落的丰富度有所降低,优势菌的相对丰度也有下降趋势,但不改变其优势地位,少数类群出现了相对丰度增加的现象。总体上,耐性细菌群落具有一定的稳定性。由于测序深度不够,所检测的真菌类群十分单一。子囊菌门(Ascomycota)、粪壳菌纲(Sordariomycetes)、镰刀菌属(Fusarium)是群落在门、纲、属分类水平上的优势菌,占据了真菌总量的99.9%。100 mg/L铅锌胁迫对镰刀菌属的相对丰度有显著的降低作用,对曲霉属则有极显著的提高。
[Abstract]:The microbes living in the polluted environment of heavy metals for a long time already have the ability to tolerate heavy metals. It is an effective way to obtain efficient microbial remediation resources by separating and domesticating them from the environment. Usually, a single microorganism is easily affected by the environment and is difficult to be widely used. The synergistic microflora which has been adapted for a long time has a certain stability and a certain system function. It has great potential in strengthening the phytoremediation. In this paper, the culturable rhizosphere flora obtained from the organic bacterial fertilizer treatment group and the control group in the lead-zinc tail mining area was studied. The differences in the ability and characteristics of removing lead and zinc were compared by orthogonal experiment, and the culture conditions were optimized to obtain the efficient tolerant flora. The effects of tolerant microflora on seed germination, seedling growth and heavy metal accumulation of castor under lead and zinc stress were analyzed by nutrient liquid-sand culture test. The structure and diversity of tolerant flora were analyzed and identified by high-throughput sequencing technique. The main conclusions are as follows: 1) under the combined stress of lead and zinc, the differences of lead removal and zinc removal between the organic fertilizer treatment group and the control group were great. The removal rate of lead and zinc in LJ was higher than that in LJO, and the maximum increment was 40.2%. The concentration of lead and zinc and the type of culture medium had a certain influence on the removal rate of lead and zinc of LJ, and the type of culture medium had the most influence on the removal rate of lead and zinc. Generally speaking, the optimum culture conditions for removal of lead and zinc by LJ were as follows: lead and zinc concentration 50 mg / L, inoculation amount 1.0 mL, culture time 6 days. Beef extract peptone medium. 2) the two groups of resistant bacteria obtained by optimizing the culture conditions had different effects on the growth and development of castor. At the stage of seed germination, inoculation of beef paste bacteria (bacteria) promoted germination to some extent, while Martin's group (fungi) showed inhibitory effect. In the stage of seedling growth and development, the plant height elongation was facilitated by the beef extract flora, while the Martin's group was more beneficial to the development of the seedling root system. The accumulation and transport of zinc in castor seedlings were stronger than that of lead, and heavy metals mainly accumulated in roots. The inhibition of zinc uptake by the two groups was more obvious than that of lead absorption. Under the stress of high concentration of lead and zinc (100 mg/L), inoculation of the two groups of bacteria was more favorable to the upward transfer of lead and zinc. 3) the dominant bacteria phylum was Proteus and the dominant class was cyanobacterium in the rhizosphere of castor. Under lead and zinc stress of 100 mg/L, the abundance of tolerant bacteria community decreased, and the relative abundance of dominant bacteria decreased, but it did not change its dominant position. A few groups appeared the phenomenon of relative abundance increase. Generally speaking, the tolerant bacterial community has certain stability. Because the sequencing depth is not enough, the detected fungi group is very single. (Sordariomycetes), is the dominant species in the taxonomic level of phylum, class and taxonomy of Fusarium (Sordariomycetes),. The relative abundance of Fusarium decreased significantly under lead and zinc stress of 99.9. 100 mg/L and increased significantly on Aspergillus.
【学位授予单位】：中南林业科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X172;X753
[Abstract]:The microbes living in the polluted environment of heavy metals for a long time already have the ability to tolerate heavy metals. It is an effective way to obtain efficient microbial remediation resources by separating and domesticating them from the environment. Usually, a single microorganism is easily affected by the environment and is difficult to be widely used. The synergistic microflora which has been adapted for a long time has a certain stability and a certain system function. It has great potential in strengthening the phytoremediation. In this paper, the culturable rhizosphere flora obtained from the organic bacterial fertilizer treatment group and the control group in the lead-zinc tail mining area was studied. The differences in the ability and characteristics of removing lead and zinc were compared by orthogonal experiment, and the culture conditions were optimized to obtain the efficient tolerant flora. The effects of tolerant microflora on seed germination, seedling growth and heavy metal accumulation of castor under lead and zinc stress were analyzed by nutrient liquid-sand culture test. The structure and diversity of tolerant flora were analyzed and identified by high-throughput sequencing technique. The main conclusions are as follows: 1) under the combined stress of lead and zinc, the differences of lead removal and zinc removal between the organic fertilizer treatment group and the control group were great. The removal rate of lead and zinc in LJ was higher than that in LJO, and the maximum increment was 40.2%. The concentration of lead and zinc and the type of culture medium had a certain influence on the removal rate of lead and zinc of LJ, and the type of culture medium had the most influence on the removal rate of lead and zinc. Generally speaking, the optimum culture conditions for removal of lead and zinc by LJ were as follows: lead and zinc concentration 50 mg / L, inoculation amount 1.0 mL, culture time 6 days. Beef extract peptone medium. 2) the two groups of resistant bacteria obtained by optimizing the culture conditions had different effects on the growth and development of castor. At the stage of seed germination, inoculation of beef paste bacteria (bacteria) promoted germination to some extent, while Martin's group (fungi) showed inhibitory effect. In the stage of seedling growth and development, the plant height elongation was facilitated by the beef extract flora, while the Martin's group was more beneficial to the development of the seedling root system. The accumulation and transport of zinc in castor seedlings were stronger than that of lead, and heavy metals mainly accumulated in roots. The inhibition of zinc uptake by the two groups was more obvious than that of lead absorption. Under the stress of high concentration of lead and zinc (100 mg/L), inoculation of the two groups of bacteria was more favorable to the upward transfer of lead and zinc. 3) the dominant bacteria phylum was Proteus and the dominant class was cyanobacterium in the rhizosphere of castor. Under lead and zinc stress of 100 mg/L, the abundance of tolerant bacteria community decreased, and the relative abundance of dominant bacteria decreased, but it did not change its dominant position. A few groups appeared the phenomenon of relative abundance increase. Generally speaking, the tolerant bacterial community has certain stability. Because the sequencing depth is not enough, the detected fungi group is very single. (Sordariomycetes), is the dominant species in the taxonomic level of phylum, class and taxonomy of Fusarium (Sordariomycetes),. The relative abundance of Fusarium decreased significantly under lead and zinc stress of 99.9. 100 mg/L and increased significantly on Aspergillus.
【学位授予单位】：中南林业科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X172;X753
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