威海近海沉积物细菌富集分离、新菌鉴定及B210捕食现象探究
发布时间：2018-07-15 13:14
【摘要】：近海沉积物中微生物多样性丰富,蕴藏大量的潜在新菌资源。本研究通过细菌富集分离手段,对不同富集时期的近海沉积物样品进行新菌资源挖掘,并通过454FLX+平台宏基因组测序探究了不同富集时期富集培养液中细菌区系演替规律。本研究发现了大量潜在新菌,并对7株海洋新菌进行了多相分类。本研究发现慢生单胞菌新菌B210具有捕食现象,并对其进行了初步探究。通过富集分离,获得326株细菌,包括49株潜在新菌。通过宏基因组测序分析得出:不同富集时期,富集培养液中有其独特的细菌区系,为以后有目的性地挖掘海洋新菌提供指导;物种丰富度会随着富集时间的增加而降低,而至第30天时,呈上升趋势;某些丰富度极低的细菌门类通过富集,数量得到了剧增,如酸杆菌门细菌。菌株Z1~T和JL1分别分离自石花菜和真江蓠,可降解琼胶。呼吸醌为Q-8。菌株 Z1~T 的主要脂肪酸是 C_(18:1)ωa7c、C_(16:0) 和 summed feature 3(C_(16:1)ω7c and/or iso-C_(15:0)2-OH)。主要极性脂是磷脂酰乙醇胺、磷脂酰甘油和1种未知的氨基脂。DNA G+C含量是45.1 mol%。与最相近物种的16S rDNA序列相似性低于92.3%。根据多相分类,菌株Z1~T和JL1代表Gammaproteobacteriaa的一个新属新种,命名为 Marinagarivorans algicola gen.nov.,sp.nov。模式菌株为 Z1~T(=ATCC BAA-2617~T=CICC 10859~T)。菌株NC2-42~T分离自威海近海沉积物,革兰氏染色可变。呼吸醌是MK-7。细胞壁含二氨基庚二酸。极性脂组分包括磷脂酰甘油、磷脂酰乙醇胺、2种未知的糖脂、1种未知的磷脂、1种未知的氨基磷脂、1种未知的脂质。主要脂肪酸是anteiso-C_(15:0)、iso-C_(16:0)和C_(16:0)。基因组DNAG+C含量58.1mol%。菌株NC2-42~T与Paenibacillus profundus有最高16SrDNA序列相似性(92.32%)。通过多相分类,NC2-42~T为 Paenibacillaceae 的一个新属新种,命名为 Marinicrinis sediminis gen.nov.,sp.nov。模式菌株为 NC2-42~T(=KCTC 33676~T =MCCC 1K01238~T)。菌株XDB06~T分离于威海近海沉积物,革兰氏染色阴性。呼吸醌是Q-8。主要脂肪酸是iso-C_(15:0)和C_(16:0)。极性脂成分为双磷脂酰甘油、磷脂酰甘油、2种糖脂和 4 种磷脂。基因组 DNA G+C 含量 65.0 mol%。与Wenzhouxiangella marina的16SrDNA相似性最高,为96.5%。通过多相分类,菌株XDB06~T是WenzhouxiangeHa 的一个新物种,命名为Wenzhouxiangella sediminis sp.nov。模式菌株为 XDB06~T(=KCTC 52041~T= MCCC 1K02285~T)。菌株0W14~T分离于文登海洋盐田,革兰氏染色阳性。MK-7是唯一呼吸醌。肽聚糖类型是A4βL-Orn-D-Glu。主要脂肪酸组分是anteiso-C_(15:0)。极性脂组分是双磷脂酰甘油、磷脂酰甘油、2种未知的糖脂和4种未知的磷脂。基因组DNA G+C 含量 44.6mol%。菌株 0W14T与Virgibacillus litoralis具有最高 16SrDNA 相似性(96.0%)。根据多相分类,菌株0W14T代表Bacillaceae的一个新属新种,命名为 Marinisalinussediminis gen.nov.,sp.nov。模式菌株为 0W14T(=KCTC 33835~T=MCCC 1H00171~T)。菌株NC2-31~T分离于威海近海沉积物,革兰氏染色阳性。呼吸醌为MK-7,肽聚糖中含二氨基庚二酸。主要极性脂是双磷脂酰甘油、磷脂酰甘油和磷脂酰乙醇胺。基因组DNA G+C含量46.3mol%。主要脂肪酸为iso-C_(15:0)、anteiso-C_(15:0)、iso-C_(16:0)和 SummedFeature3(C_(16:1)ω7cand/or iso-C_(15:0)2-OH)。基于 16SrDNA 序列的系统发育分析表明菌株NC2-31T在Bacillus内形成独立分支。根据多相分类,菌株NC2-31~T代表Baacillus的一个新物种,命名为Bacillusmarinisedimentorum sp.nov。模式菌株为 NC2-31~T(=KCTC33721~T = MCCC1K01239~T)。菌株WDN1C137~T分离于文登海洋盐田,革兰氏染色阴性。Q-10是唯一醌型。主要脂肪酸成分为C_(18:1)ω7c、cycloC_(19:0)ω8c、C_(16:0)。主要极性脂为磷脂酰甘油、磷酸糖脂、1种未知的糖酯、1种未知的脂质、2种未知的氨基脂和3种未知的磷脂。基因组DNA G+C含量71.4 mol%。菌株WDN1C137T与Roseivivax jejudonensi.有最高16S rDNA相似性(94.5%)。根据多相分类,菌株WDN1C137T代表Rhodobacteracea 的一个新属新种,Rhodosalinus sediminis gen.nov.,sp.nov。模式菌株为 WDN1C137~T(=KCTC52478~T = MCCC 1H00170~T)。菌株B210T分离于威海小石岛近海沉积物,革兰氏染色阴性。呼吸醌是MK-7。极性脂组分是双磷脂酰甘油、磷脂酰甘油、磷脂酰乙醇胺、1种未知的磷脂、1种未知的脂质。主要脂肪酸是iso-C_(15:0)。基因组DNA G+C含量是64.7 mol%。系菌株 B210T 与 Bradymonas sediminis FA350~T有最高 16S rDNA 序列相似性(89.2%)。通过多相分类,B210~T为Bradymonadaless的一个新属新种。模式菌株为 B210~T(=KCTC42951~T)。对鉴定的慢生单胞菌新菌B210捕食现象进行了初步探究。其捕食谱广泛,能捕食革兰氏染色阳性和阴性细菌,这同黄色黏球菌的捕食谱相似,而不同于只能捕食革兰氏阴性菌的蛭弧菌;通过扫描电子显微镜观察、捕食接触性试验推断其捕食需要细胞间接触。通过对菌株B210发酵液不同组分的抑菌作用的探究,发现其发酵上清液和胞内产物都不具有抑菌作用,这进一步佐证了其捕食行为需借助与猎物的细胞接触。
[Abstract]:In the offshore sediments, the microbial diversity is abundant and contains a large number of potential new bacteria resources. Through the method of bacterial enrichment and separation, the new bacteria resources are excavated in the sediment samples of different enrichment period, and the succession law of the bacterial flora in the enrichment culture medium of different enrichment period is explored through the 454FLX+ platform macrogenome sequencing. This study found a large number of potential new bacteria and a multiphase classification of 7 new marine bacteria. This study found that B210 has a predatory phenomenon and has carried out preliminary exploration. Through enrichment and separation, 326 strains of bacteria, including 49 potential new bacteria, were obtained by Acer sequence analysis: enrichment culture and enrichment culture. There is a unique bacterial flora in the nutrient solution, which provides guidance for the future purpose of mining the new marine bacteria. The species richness will decrease with the increase of the enrichment time, and up to thirtieth days. Some kinds of bacteria with extremely low richness have been enriched, such as bacilli bacteria, bacteria Z1~T and JL1, respectively. The main fatty acids of the Q-8. strain Z1~T are C_ (18:1) Omega a7c, C_ (16:0) and summed feature 3 (C_ (16:1) Omega 7C). The main polar lipids are phosphatidylethanolamine, phosphatidyl glycerol and 1 kinds of unknown amino groups, which are 45.1 and the closest species The similarity of 16S rDNA sequence is lower than 92.3%. based on Polyphasic classification. Strain Z1~T and JL1 represent a new genus and new species of Gammaproteobacteriaa, named Marinagarivorans algicola gen.nov., sp.nov. model strain is Z1~T (=ATCC). Quinone is MK-7. cell wall containing two amino heptane diacid. Polar lipid components include phosphatidyl glycerol, phosphatidyl ethanolamine, 2 kinds of unknown glycolipids, 1 unknown phospholipids, 1 unknown amino phospholipids, 1 kinds of unknown lipids. The main fatty acids are anteiso-C_ (15:0), iso-C_ (16:0) and C_ (16:0). 58.1mol%. strains NC2-42~T and Paenibaci in genomic DNAG+C Llus profundus has the highest 16SrDNA sequence similarity (92.32%). Through multiphase classification, NC2-42~T is a new genus and new species of Paenibacillaceae, named Marinicrinis sediminis gen.nov., sp.nov. mode strain is NC2-42~T (=KCTC 33676~T). The strain is separated from the sediments of Weihai and gram-negative. The main fatty acids of Q-8. are iso-C_ (15:0) and C_ (16:0). The polar lipids are diphosphatidyl glycerol, phosphatidyl glycerol, 2 kinds of glycolipids and 4 kinds of phospholipids. The genomic DNA G+C content is the highest similarity between 65 mol%. and Wenzhouxiangella marina. The strain of Wenzhouxiangella sediminis sp.nov. model is XDB06~T (=KCTC 52041~T= MCCC 1K02285~T). Strain 0W14~T is separated from the marine salt field in Wendeng, and gram-positive.MK-7 is the only respiratory quinone. Lipoyl glycerol, 2 unknown glycols and 4 kinds of unknown phospholipids. Genomic DNA G+C content 44.6mol%. strain 0W14T and Virgibacillus litoralis have the highest 16SrDNA similarity (96%). According to multiphase classification, strain 0W14T represents a new species of Bacillaceae, named Marinisalinussediminis gen.nov., sp.nov. mode strain is 0 W14T (=KCTC 33835~T=MCCC 1H00171~T). Strain NC2-31~T was isolated from Weihai offshore sediments, gram-positive. Respiratory quinone was MK-7, and peptidoglycan contained two amino heptanic acid. The main polar lipids were diphosphatidyl glycerol, phosphatidyl glycerol and phosphatidyl ethanolamine. Genomic DNA G+C content 46.3mol%. main fatty acids were iso-C_ (15:0), anteiso-C_. (15:0), iso-C_ (16:0) and SummedFeature3 (C_ (16:1) Omega 7cand/or iso-C_ (15:0) 2-OH). Phylogenetic analysis based on 16SrDNA sequence shows that strain NC2-31T forms an independent branch. 31~T (=KCTC33721~T = MCCC1K01239~T). Strain WDN1C137~T was isolated from the Wendeng ocean salt field. Gram-negative.Q-10 was the only quinone. The main fatty acids were C_ (18:1) Omega 7C, cycloC_ (19:0) Omega 8C, C_. 1 kinds of unknown sugar esters, 1 unknown lipids, 2 unknown amino fat and 3 were the main polar lipids. Genomic DNA G+C content 71.4 mol%. strain WDN1C137T and Roseivivax jejudonensi. have the highest 16S rDNA similarity (94.5%). According to the multiphase classification, the strain WDN1C137T represents a new species of Rhodobacteracea, Rhodosalinus sediminis. Strain B210T is separated from the sediments of Weihai small Shidao offshore, gram-negative. Respiratory quinone is a MK-7. polar lipid component which is diphosphatidyl glycerol, phosphatidyl glycerol, phosphatidyl ethanolamine, 1 kinds of unknown phospholipids, 1 kinds of unknown lipids. The main fatty acids are iso-C_ (15:0). The DNA G+C content of the group is 64.7 mol%. strain B210T and Bradymona S sediminis FA350~T has the highest 16S rDNA sequence similarity (89.2%). Through multiphase classification, B210~T is a new genus and new species of Bradymonadaless. The pattern strain is B210~T (=KCTC42951~T). The preliminary exploration of the identification of the new bacteria B210 of the new bacteria of the bacteria is preliminarily explored. It is widely used to capture Gram-positive and negative bacteria. This was similar to that of the Staphylococcus aureus, but different from the vermiculorvibrio that only preyed on the Gram-negative bacteria. Through the scanning electron microscope, the predatory contact test concluded that the predation needed intercellular contact. By investigating the Bacteriostasis of different components of the strain B210, the fermentation supernatant and the intracellular products were not found. It has bacteriostatic effect, which further confirms its predatory behavior by means of contact with prey cells.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q93
本文编号：2124190
[Abstract]:In the offshore sediments, the microbial diversity is abundant and contains a large number of potential new bacteria resources. Through the method of bacterial enrichment and separation, the new bacteria resources are excavated in the sediment samples of different enrichment period, and the succession law of the bacterial flora in the enrichment culture medium of different enrichment period is explored through the 454FLX+ platform macrogenome sequencing. This study found a large number of potential new bacteria and a multiphase classification of 7 new marine bacteria. This study found that B210 has a predatory phenomenon and has carried out preliminary exploration. Through enrichment and separation, 326 strains of bacteria, including 49 potential new bacteria, were obtained by Acer sequence analysis: enrichment culture and enrichment culture. There is a unique bacterial flora in the nutrient solution, which provides guidance for the future purpose of mining the new marine bacteria. The species richness will decrease with the increase of the enrichment time, and up to thirtieth days. Some kinds of bacteria with extremely low richness have been enriched, such as bacilli bacteria, bacteria Z1~T and JL1, respectively. The main fatty acids of the Q-8. strain Z1~T are C_ (18:1) Omega a7c, C_ (16:0) and summed feature 3 (C_ (16:1) Omega 7C). The main polar lipids are phosphatidylethanolamine, phosphatidyl glycerol and 1 kinds of unknown amino groups, which are 45.1 and the closest species The similarity of 16S rDNA sequence is lower than 92.3%. based on Polyphasic classification. Strain Z1~T and JL1 represent a new genus and new species of Gammaproteobacteriaa, named Marinagarivorans algicola gen.nov., sp.nov. model strain is Z1~T (=ATCC). Quinone is MK-7. cell wall containing two amino heptane diacid. Polar lipid components include phosphatidyl glycerol, phosphatidyl ethanolamine, 2 kinds of unknown glycolipids, 1 unknown phospholipids, 1 unknown amino phospholipids, 1 kinds of unknown lipids. The main fatty acids are anteiso-C_ (15:0), iso-C_ (16:0) and C_ (16:0). 58.1mol%. strains NC2-42~T and Paenibaci in genomic DNAG+C Llus profundus has the highest 16SrDNA sequence similarity (92.32%). Through multiphase classification, NC2-42~T is a new genus and new species of Paenibacillaceae, named Marinicrinis sediminis gen.nov., sp.nov. mode strain is NC2-42~T (=KCTC 33676~T). The strain is separated from the sediments of Weihai and gram-negative. The main fatty acids of Q-8. are iso-C_ (15:0) and C_ (16:0). The polar lipids are diphosphatidyl glycerol, phosphatidyl glycerol, 2 kinds of glycolipids and 4 kinds of phospholipids. The genomic DNA G+C content is the highest similarity between 65 mol%. and Wenzhouxiangella marina. The strain of Wenzhouxiangella sediminis sp.nov. model is XDB06~T (=KCTC 52041~T= MCCC 1K02285~T). Strain 0W14~T is separated from the marine salt field in Wendeng, and gram-positive.MK-7 is the only respiratory quinone. Lipoyl glycerol, 2 unknown glycols and 4 kinds of unknown phospholipids. Genomic DNA G+C content 44.6mol%. strain 0W14T and Virgibacillus litoralis have the highest 16SrDNA similarity (96%). According to multiphase classification, strain 0W14T represents a new species of Bacillaceae, named Marinisalinussediminis gen.nov., sp.nov. mode strain is 0 W14T (=KCTC 33835~T=MCCC 1H00171~T). Strain NC2-31~T was isolated from Weihai offshore sediments, gram-positive. Respiratory quinone was MK-7, and peptidoglycan contained two amino heptanic acid. The main polar lipids were diphosphatidyl glycerol, phosphatidyl glycerol and phosphatidyl ethanolamine. Genomic DNA G+C content 46.3mol%. main fatty acids were iso-C_ (15:0), anteiso-C_. (15:0), iso-C_ (16:0) and SummedFeature3 (C_ (16:1) Omega 7cand/or iso-C_ (15:0) 2-OH). Phylogenetic analysis based on 16SrDNA sequence shows that strain NC2-31T forms an independent branch. 31~T (=KCTC33721~T = MCCC1K01239~T). Strain WDN1C137~T was isolated from the Wendeng ocean salt field. Gram-negative.Q-10 was the only quinone. The main fatty acids were C_ (18:1) Omega 7C, cycloC_ (19:0) Omega 8C, C_. 1 kinds of unknown sugar esters, 1 unknown lipids, 2 unknown amino fat and 3 were the main polar lipids. Genomic DNA G+C content 71.4 mol%. strain WDN1C137T and Roseivivax jejudonensi. have the highest 16S rDNA similarity (94.5%). According to the multiphase classification, the strain WDN1C137T represents a new species of Rhodobacteracea, Rhodosalinus sediminis. Strain B210T is separated from the sediments of Weihai small Shidao offshore, gram-negative. Respiratory quinone is a MK-7. polar lipid component which is diphosphatidyl glycerol, phosphatidyl glycerol, phosphatidyl ethanolamine, 1 kinds of unknown phospholipids, 1 kinds of unknown lipids. The main fatty acids are iso-C_ (15:0). The DNA G+C content of the group is 64.7 mol%. strain B210T and Bradymona S sediminis FA350~T has the highest 16S rDNA sequence similarity (89.2%). Through multiphase classification, B210~T is a new genus and new species of Bradymonadaless. The pattern strain is B210~T (=KCTC42951~T). The preliminary exploration of the identification of the new bacteria B210 of the new bacteria of the bacteria is preliminarily explored. It is widely used to capture Gram-positive and negative bacteria. This was similar to that of the Staphylococcus aureus, but different from the vermiculorvibrio that only preyed on the Gram-negative bacteria. Through the scanning electron microscope, the predatory contact test concluded that the predation needed intercellular contact. By investigating the Bacteriostasis of different components of the strain B210, the fermentation supernatant and the intracellular products were not found. It has bacteriostatic effect, which further confirms its predatory behavior by means of contact with prey cells.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q93
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