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| 三亚三种海草可培养共附生放线菌多样性及其生物活性初探 | |
| 其他题名 | A Preliminary Study on the Actinobacteria Resource Associated with Seagrasses and Their Potential of Bioactivities Product |
| 陈文 | |
| 出版年 | 2012 |
| 学位类型 | 硕士 |
| 导师 | 吴后波 ; 李翔 |
| 学位授予单位 | 中国科学院大学 |
| 中文摘要 | 放线菌广泛分布于陆地、海洋、沙漠以及相关动植物宿主，是抗生素和其他活性次级代谢产物的重要来源。据统计，抗生素数据库中超过一半的活性化合物来自放线菌。随着陆源放线菌活性物质重复发现率的升高，人们开始将研究目光投向海洋来源放线菌。目前对于海洋放线菌资源的研究主要集中在海水、沉积物、以及各种海洋动植物中。\n海草是海洋生态系统的重要组成部分。近年来国内外对海草的研究主要集中在海草的分类与分布、生态功能、生物量与初级生产力、共附生细菌和真菌等领域，尚无关于海草可培养共附生放线菌的相关报道。\n本论文运用微生物学和分子生物学手段，对采自中国陵水新村港的泰莱藻(Thalassia hemprichii)、海神草(Cymodocea rotundata)和海菖蒲(Enhalus acoroides)等三种海草可培养共附生放线菌种质资源进行了研究，并就这些放线菌的生物活性和产生活性化合物的潜力进行了初步研究，取得了以下几方面的主要成果：\n(1)采用七种不同的培养基对三种海草进行共附生可培养放线菌分离、培养，总共获得放线菌纯培养293株，经16S rDNA鉴定它们属于链霉菌属(Streptomyces)、微杆菌属(Microbacterium)、分支杆菌属(Mycobacterium)，小单胞菌属(Micromonospora)、(Actinomycetospora)菌属、野野村胞菌属(Nonomuraea)、疣孢菌属(Verrucosispora)、拟诺卡氏菌属(Nocardiopsis)、糖霉菌属(Glycomyces)、(Isoptericola)菌属、(Prauserella)菌属、克氏库克菌属(Kocuria)、糖单胞菌属(Saccharomonospora)、纤维素菌属(Cellulomonas)以及纤维单胞菌属(Cellulosimicrobium)等15个属,其中疣孢菌属(Verrucosispora)、纤维单胞菌属(Cellulosimicrobium)和(Prauserella)菌属为首次从植物中分离得到。 在所得菌株中，放线菌M8D17、GXG、GXE、GX52、HCZ4、M1X63、M8Z38、M1Z45和G2Z20为潜在新种。\n同时，通过对不同培养基获得的微生物多样性比较发现，培养基组分和浓度对微生物的生长具有重要影响，部分微生物对宿主成分具有依赖性。\n(2)通过功能基因(聚酮合成酶(PKSⅠ、PKSⅡ)和非核糖体多肽合成酶(NRPS))的筛选，大部分菌株的PKSⅡ和NRPS基因筛选结果为阳性，PKSⅠ基因筛选结果阳性率相对较低。菌株GZ38和HCD20三种功能基因筛选结果都为阴性。该研究结果展现了海草共附生放线菌在合成聚酮类及非核糖体肽类物质方面具有较好的潜力。\n(3)琼脂块法抗菌实验结果显示：泰莱藻中进行抗菌实验的35株放线菌中有20株放线菌有一种以上抗菌活性，占约51.7%；海神草中进行抗菌实验的24株放线菌中有7株放线菌有一种以上抗菌活性，占约29.2%；海菖蒲中进行抗菌实验的14株放线菌中有7株放线菌有一种以上抗菌活性，占约50%。该研究结果表明海草共附生放线菌在提供生物活性物质方面具有很大的潜力。\n综上所述，海草中蕴含着极其丰富的放线菌资源，不同海草所孕育的放线菌资源存在显著的宿主差异。这些放线菌资源是巨大的潜在的活性天然产物来源。 |
| 英文摘要 | As a great important source of antibiotics and bioactive molecules, actinobacteria almost produce over half of the bioactive compounds in the Antibiotic Literature Database.It is well known that the diversity of actinobacteria is profuse both on terrestrial and marine environment. As the increasingly high rediscovery rate of known bioactive compounds from terrestrial actinobacteria, researchers began to shift the focus on the marine actinobacteria.The study about marine actinobacteria mostly concentrated on sea water,marine sediment and marine flora and fauna,and so far much more research attention has been paid to the actinobacteria associated with marine sponges.\nSeagrass played a pivotal role in marine ecosystem. The important ecological functions of seagrass have attracted more and more scientists’ interests. Recent studies are focus on the distribution and classification of seagrass, its ecological functions, biomass and primary production and so on. Nevertheless, studies on the diversity of seagrass-associated culturable actinobacteria have an expansive prospect. \nIn this study, classic microbiology and molecular techniques were employed to study the diversity of seagrass associated actinobacteria from three different kinds of seagrasses. At the same time, the potential of bioactivites from culturable actinobacteria associated with seagrasses were also been have a preliminary study. The research efforts finally led to some important findings from the following four aspects.\n(1)A total of 293 actinomycetes were isolated using seven different media. According to the phylogenetic analysis, the dominate actinomycete in these three species of seagrasses were all Streptomyces. Another fourteen different less frequently recovered actinobacteria genera found in these three seagrasses were as follows: Micromonospora, Mycobacterium, Verrucosispora, Microbacterium, Glycomyces, Kocuria, Actinomycetospora, Nonomuraea, Cellulosimicrobium, Cellulomonas, Isoptericola, Saccharomonospora, Prauserella and Nocardiopsis. To our knowledge, this is the first time that so many different genera of actinobacteria isolated from the plant, and the genera of Verrucosispora, Cellulosimicrobium and Isoptericola were first recorded to be isolated from the plant. Eight potential representatives of new actinobacteria taxa were isolated from three kinds of seagrasses. By sequence alignment and phylogenetic tree construction indicated that isolates M8D17, GXG, GXE, GX52, HCZ4, M1X63, M8Z38, M1Z45 and G2Z20 might be candidates of novel species(GX52 and HCZ4 might be the same species). This study gives a comparative study of actinobacterial diversity in the seagrasses and a preliminary exploration of the potential natural product diversity.\n We also could conclude that different nutrients in culture medium have a certain influence on the growth of microorganism in some samples, this also provided a new way of thinking for how to design medium.\n(3)Three sets of degenerate primers targeting genes encoding polyketide synthases (PKS-I and PKS-II) and nonribosomal peptide synthetase (NRPS) were used to identify the diversity of the potential bioactive compounds among these Actinobacteria isolated from seagrasses. PKS-II and NRPS sequence were amplified in almost all of the isolates, PKS-I was just detected in only 36.8% of the isolates. None of the target sequences was found in the strain HCD20 and strain GZ38.These results gave a glimpse of the great bioactive agent production ability of seagrass associated actinobacteia.\n(4)The result of antimicrobial activity test showed that seagrass has a great potential of producing bioactivities compound, with 20 of 35 isolates from Thalassia hemprichii, 7 of 24 isolates from Cymodocea rotundata and 7 of 14 isolates from Enhalus acoroides have at least one antibiotic activity.\nIn conclusion, although the differences were exist both in quantity of culturable actinomycetes and the species among the three kinds of seagrasses, but common confirmed seagrass contains extremely rich actinomycetes resources, including many rare actinomycetes and potential representatives of new actinobacteria taxa. The result of antimicrobial activity test also confirmed that these strains isolated from seagrass have great potential in production of bioactive natural compounds. |
| 中文关键词 | 海草 ; 共附生放线菌 ; 多样性 ; 生物活性 |
| 英文关键词 | Seagrass Actinobacteria associated with seagrass Diversity Bioactivities |
| 语种 | 中文 |
| 国家 | 中国 |
| 来源学科分类 | 海洋生物学 |
| 来源机构 | 中国科学院南海海洋研究所 |
| 资源类型 | 学位论文 |
| 条目标识符 | http://119.78.100.177/qdio/handle/2XILL650/287112 |
| 推荐引用方式 GB/T 7714 | 陈文. 三亚三种海草可培养共附生放线菌多样性及其生物活性初探[D]. 中国科学院大学,2012. |
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