蚂蚁内共生放线菌的鉴定及抗菌活性产物研究
发布时间：2018-08-09 20:03
【摘要】：近年来科学家们在探寻新型抗生素的筛选途径时,发现昆虫内生菌能够产生许多种类多样的次生代谢产物,尤其那些能够帮助宿主抵御病原菌入侵的放线菌,其中一些具有新颖的结构并表现出较好的生物活性。由于这些化合物能够存在于真核宿主体内,且不危害宿主,所以它们很有可能对人类也没有毒副作用。因此这些昆虫内生放线菌具有很大产生人类药用的抗生素潜力,所以近年来昆虫内共生放线菌受到越来越多科学家的关注,昆虫内生放线菌也将是持续地发现新抗生素的重要领域之一,这对于对抗日益增多的抗药病原菌和新型疾病具有重要意义。本研究以实验室筛选得到的30株蚂蚁内生菌为材料,通过培养特征观察、抗菌活性测试和16S rRNA序列测序结果分析,发现两株潜在的放线菌新种3H-GS17~T和2H-TWYE14~T和一株具有良好抗菌活性放线菌1H-GS2,对它们分别进行了系统的分类学鉴定和活性物质的分离鉴定,结果如下:(1)16S rRNA分析结果显示菌株3H-GS17~T是珊瑚放线菌属的一株菌,与Actinocorallia glomerata JCM 9376T(98.13%)和Actinocorallia longicatena JCM 9377~T(97.64%)的16S rRNA序列相似性最高。化学分类学特征也与Actinocorallia的特征一致。而DNA DNA杂交结果和表型分析结果显示菌株3H-GS17~T明显不同于与它相近的菌株,进一步的实验证明菌株3H-GS17~T和珊瑚放线菌属的其他菌株都不相同。因此确定菌株3H-GS17~T是珊瑚放线菌属的一个新种,命名为Actinocorallia lasicapitis,典型菌株是3H-GS17~T(=DSM 100595T=CGMCC4.7282T)。(2)16S rRNA分析结果显示2H-TWYE14~T是链霉菌属的一株菌,与Streptomyces niveus NRRL 2466T(98.84%)的16S rRNA序列相似性最高。gyr B基因分析结果同样显示2H-TWYE14~T属于链霉菌属。化学分类学特征也与链霉菌属的其他菌株相一致。而2H-TWYE14~T与S.niveus NRRL 2466T之间的DNA DNA杂交结果和表型分析结果显示2H-TWYE14~T明显不同于S.niveus NRRL 2466T。因此确定菌株2H-TWYE14~T是链霉菌菌属的一个新种,命名为Streptomyces camponoticapitis,典型菌株是2H-TWYE14~T(=DSM 100523T=CGMCC 4.7275T)。(3)活体生物活性检测显示菌株1H-GS2对部分测试真菌具有较好的生物活性,将菌株进行发酵,发酵产物同样显示良好的真菌抑制活性。对发酵产物中的化合物进行分离纯化后,利用现代光谱学进行结构分析鉴定,最终得到化合物白诺菌素(Albonoursin)。
[Abstract]:In recent years, while searching for new antibiotic screening pathways, scientists have found that insect endophytes can produce a variety of secondary metabolites, especially actinomycetes, which can help their hosts withstand the invasion of pathogens. Some of them have novel structures and exhibit good biological activity. Since these compounds are present in eukaryotic hosts and do not harm the host, they are likely to have no adverse effects on humans. Therefore, these insect endophytic actinomycetes have great antibiotic potential to produce human medicine, so in recent years, more and more scientists pay attention to insect endophytic actinomycetes. Insect endophytic actinomycetes will also be one of the important fields for the continuous discovery of new antibiotics, which is of great significance against the increasing number of drug-resistant pathogens and new diseases. In this study, 30 strains of ant endophytic bacteria screened in laboratory were used as materials, and the results of antibacterial activity test and 16s rRNA sequencing were analyzed through observation of culture characteristics. Two potential new actinomycetes, 3H-GS17~T and 2H-TWYE14~T, and one actinomycetes with good antibacterial activity, 1H-GS2, were found. The results were as follows: (1) the results of 16s rRNA analysis showed that the strain 3H-GS17~T was a strain of actinomycetes, and had the highest similarity with the 16s rRNA sequences of Actinocorallia glomerata JCM 9376T (98.13%) and Actinocorallia longicatena JCM 9377T (97.64%). The characteristics of chemotaxonomy are also consistent with those of Actinocorallia. The results of DNA DNA hybridization and phenotypic analysis showed that 3H-GS17~T was different from other strains of actinomycetes. Further experiments showed that 3H-GS17~T and other strains of actinomycetes were different. Therefore, 3H-GS17~T is a new species of actinomycetes, named Actinocorallia lasicapitis. the typical strain is 3H-GS17~T (DSM 100595T=CGMCC4.7282T). (2) 16s rRNA. The results show that 2H-TWYE14~T is a strain of Streptomyces. The analysis of 16s rRNA sequence with Streptomyces niveus NRRL 2466T (98.84%) also showed that 2H-TWYE14~T belongs to Streptomyces. The chemotaxonomic characteristics are also consistent with other strains of Streptomyces. The results of DNA DNA hybridization and phenotypic analysis between 2H-TWYE14~T and S.niveus NRRL 2466T showed that 2H-TWYE14~T was significantly different from S.niveus NRRL 2466T. Therefore, the strain 2H-TWYE14~T is a new species of Streptomyces, named Streptomyces camponticapitis. the typical strain is 2H-TWYE14~T (2H-TWYE14~T 100523T=CGMCC 4.7275T). (3), which shows that 1H-GS2 has good bioactivity to some tested fungi. The fermentation products also showed good fungal inhibitory activity. After the separation and purification of the compounds in the fermentation products, the structure analysis and identification of the compounds were carried out by modern spectroscopy. Finally, the compound (Albonoursin). Was obtained.
【学位授予单位】：东北农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R915
本文编号：2175167
[Abstract]:In recent years, while searching for new antibiotic screening pathways, scientists have found that insect endophytes can produce a variety of secondary metabolites, especially actinomycetes, which can help their hosts withstand the invasion of pathogens. Some of them have novel structures and exhibit good biological activity. Since these compounds are present in eukaryotic hosts and do not harm the host, they are likely to have no adverse effects on humans. Therefore, these insect endophytic actinomycetes have great antibiotic potential to produce human medicine, so in recent years, more and more scientists pay attention to insect endophytic actinomycetes. Insect endophytic actinomycetes will also be one of the important fields for the continuous discovery of new antibiotics, which is of great significance against the increasing number of drug-resistant pathogens and new diseases. In this study, 30 strains of ant endophytic bacteria screened in laboratory were used as materials, and the results of antibacterial activity test and 16s rRNA sequencing were analyzed through observation of culture characteristics. Two potential new actinomycetes, 3H-GS17~T and 2H-TWYE14~T, and one actinomycetes with good antibacterial activity, 1H-GS2, were found. The results were as follows: (1) the results of 16s rRNA analysis showed that the strain 3H-GS17~T was a strain of actinomycetes, and had the highest similarity with the 16s rRNA sequences of Actinocorallia glomerata JCM 9376T (98.13%) and Actinocorallia longicatena JCM 9377T (97.64%). The characteristics of chemotaxonomy are also consistent with those of Actinocorallia. The results of DNA DNA hybridization and phenotypic analysis showed that 3H-GS17~T was different from other strains of actinomycetes. Further experiments showed that 3H-GS17~T and other strains of actinomycetes were different. Therefore, 3H-GS17~T is a new species of actinomycetes, named Actinocorallia lasicapitis. the typical strain is 3H-GS17~T (DSM 100595T=CGMCC4.7282T). (2) 16s rRNA. The results show that 2H-TWYE14~T is a strain of Streptomyces. The analysis of 16s rRNA sequence with Streptomyces niveus NRRL 2466T (98.84%) also showed that 2H-TWYE14~T belongs to Streptomyces. The chemotaxonomic characteristics are also consistent with other strains of Streptomyces. The results of DNA DNA hybridization and phenotypic analysis between 2H-TWYE14~T and S.niveus NRRL 2466T showed that 2H-TWYE14~T was significantly different from S.niveus NRRL 2466T. Therefore, the strain 2H-TWYE14~T is a new species of Streptomyces, named Streptomyces camponticapitis. the typical strain is 2H-TWYE14~T (2H-TWYE14~T 100523T=CGMCC 4.7275T). (3), which shows that 1H-GS2 has good bioactivity to some tested fungi. The fermentation products also showed good fungal inhibitory activity. After the separation and purification of the compounds in the fermentation products, the structure analysis and identification of the compounds were carried out by modern spectroscopy. Finally, the compound (Albonoursin). Was obtained.
【学位授予单位】：东北农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R915
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