白蚁肠道细菌的多样性和生物活性研究

发布时间：2018-01-05 23:16

本文关键词：白蚁肠道细菌的多样性和生物活性研究　出处：《郑州大学》2017年硕士论文　论文类型：学位论文

【摘要】：白蚁肠道是一个比较封闭的微环境,白蚁肠道内的共生微生物也具有其独特的性质,它们可以帮助白蚁分解难消化的木质纤维素,产生白蚁生长所必需的刺激代谢产物等。本研究从白蚁肠道内微生物的多样性出发,利用免培养的高通量测序技术和纯培养的方法研究白蚁肠道细菌的多样性,筛选具有木质纤维素分解能力和抗菌活性的菌株。为以后的研究提供一定的实验依据。本研究通过高通量测序技术,对白蚁肠道样品中的细菌进行16SrDNA基因的V4区进行测序,最终获得62404条有效的序列,总共630个OTUs。鉴定出厚壁菌门(Firmicutes)、拟杆菌门(Bacteroidetes)、变形菌门(Proteobacteria)、放线菌门(Actinobacteria)、螺旋体门(Spirochaetes)等13个门的细菌,其中放线菌门占16.95%。本研究通过不同的培养基和分离方法分离白蚁肠道内的微生物。一共挑取1458株菌,最后得到放线菌327株,其中包括一个放线菌目(Actinobacterales),放线菌目下的5个亚目,分别为:微球菌亚目(Micrococcineae)的4个科;链霉菌亚目(Streptomycineae)的1个科;棒杆菌亚目(Corynebacterineae)的2个科;丙酸杆菌亚目(Propionibacterineae)的1个科;小单孢菌亚目(Micromonosporineae)的1个科,总共有10个科,18个属。并且分离到一株新分类单元,通过初步分析,鉴定YIM 110061为明串珠菌科(Leuconostocaceae)下的一个新属。本研究对分离得到的菌株筛选长势良好的菌株进行纤维素酶活和木质素酶活的测定。最终测量四株菌株的纤维素酶活随时间的变化曲线,比较发现YIM110167分解纤维素的活性整体最好,并在第五天酶活达到最大值为23.69/ml。本研究测量菌株的三种木质素酶活,分别是:漆酶、过氧化物酶、猛过氧化物酶。其中发现YIM 110063的过氧化物酶活性最高,这株菌株还检测到其他两种酶的活性,后续对这株菌株进行滤纸片的筛选,发现五天后滤纸片断裂,并且液体培养基变浑浊。本研究对分离到的327株放线菌进行抗菌活性和功能基因的筛选。最终得到11株抗大肠杆菌,22株抗枯草芽孢杆菌,30株抗金黄色葡萄球菌,32株抗白色链球菌,48株抗黑曲霉的菌株。并检测327株放线菌的六种功能基因。白蚁肠道是一个非常特殊的环境,存在大量值得研究的菌株,利用好这类资源一定会为人类的发展做出很大贡献。
[Abstract]:The intestinal tract of termites is a relatively closed microenvironment, and the symbiotic microorganisms in the intestinal tract of termites also have their unique properties, which can help termite to decompose the indigestible lignocellulose. This study based on the diversity of microbes in the intestinal tract of termites, using high-throughput sequencing technology and pure culture methods to study the diversity of termites intestinal bacteria. Screening the strains with lignocellulose decomposition ability and antimicrobial activity to provide a certain experimental basis for future research. This study through high-throughput sequencing technology. The V4 region of 16s rDNA gene was sequenced from the bacteria in the intestinal tract of termites and 62404 valid sequences were obtained. A total of 630 OTUs. were identified as Firmicutesus, Bacteroidetes. Proteobacteria, Actinobacteria, Spirochaetes. Actinomycetes accounted for 16.95. In this study, the bacteria in termites intestines were isolated by different culture medium and isolation methods. A total of 1458 strains of bacteria were selected, and 327 strains of actinomycetes were obtained. It includes an actinobacterium actinobacterium and five suborders of actinobacter, which are four families of Micrococcineaeae. A family of Streptomycineae; Two families of Corynebacterium Corynebacter. A family of Propionibacterium propioniae; A family of Micromonosporineae, consisting of 10 families and 18 genera, was isolated and a new taxonomic unit was isolated and analyzed preliminarily. YIM 110061 was identified as Leuconostocaceaeae. In this study, the cellulase activity and ligninase activity of the strains with good growth were determined. Finally, the cellulase activity curves of four strains were measured. It was found that the activity of cellulose decomposed by YIM110167 was the best on the whole and reached the maximum value of 23.69 / ml on 5th days. The three kinds of lignin enzyme activity of the strain were measured in this study. They were laccase, peroxidase and peroxidase. The highest peroxidase activity was found in YIM 110063, and the other two enzymes were also detected by this strain. The strain was screened for filter paper and found that the filter paper broke five days later. In this study, 327 strains of actinomycetes were screened for antimicrobial activity and functional genes. Finally, 11 strains resistant to Escherichia coli and 22 strains of Bacillus subtilis resistant to Bacillus subtilis were obtained. Thirty three strains of staphylococcus aureus were resistant to Streptococcus albicans and 48 strains were resistant to Aspergillus Niger. Six functional genes of 327 strains of actinomycetes were detected. The intestinal tract of termites is a very special environment. There are a lot of strains worth studying, and making good use of these resources will make a great contribution to human development.
【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q93

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