冻土细菌新种和冰缘植物内生细菌新种的特征和多相分类学研究
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摘要
冻土细菌是一类生活在低温、寡营养、液态水稀少和氧气稀缺环境中的极端微生物。它们的种类丰富多样,它们的代谢产物也可能是一类新资源,对于人类而言,蕴含着巨大的价值。这一切的前提就是得到更多的冻土细菌进行分门归类,并对它们与已有的种类进行比较后,从而找到其特殊的用途和特殊的代谢产物。
植物内生细菌定殖于健康的植物宿主体内,与植物建立了和谐联合的关系,对于宿主的健康并无威胁,还对植物宿主的生命活动有着积极的作用,研究植物内生细菌对宿主的作用可能是未来生态型农业发展的一条新途径。为了发展新资源,人们必须从不同环境生长的植物中分离到更多的内生细菌,并进行分门归类。
本工作主要针对从冻土微生物多样性研究和冰缘植物内生细菌多样性研究中得到的可能具有新分类地位的细菌菌株进行分类鉴定的工作,主要结果如下：
1.对分离于天山一号冰川采集的冻土样品中细菌进行筛选后,得到两株可能是新种的细菌菌株,记为TSBY 57T和TSBY 67T;对分离于冰缘植物火绒草的内生细菌进行筛选后,得到一株可能是新种的细菌菌株,记为Enf 54T,三株细菌的纯培养物的得到为进一步的研究提供了素材,这是首次对天山冻土中分离的细菌进行分类鉴定,也是首次对冰缘植物火绒草的内生细菌进行分类鉴定。
2. TSBY 57T的菌落呈黄色,细胞为杆状,大小为0.2-0.3μm×0.3-1.1μm; TSBY 67T的菌落呈橙黄色,Flexirubin-type色素反应为阳性,细胞为杆状,大小为0.3-0.5μm×0.6-1.6μm; Enf 54T的菌落呈淡黄色,细胞为短杆状,大小为0.3μm×0.9μm,它们的在形态方面的主要特征与各自所在属的特征基本相符,奠定了分类鉴定研究中形态学的基础。
3.通过培养特性研究,TSBY 57T能在NaCl浓度为0-1.5%,pH范围为6-8,、温度范围为0-30℃时生长；TSBY 67T能在NaCl浓度为0-2%,pH范围为6-8,温度范围为4-37℃时生长;Enf 54T能在NaCl浓度为0-4%,能在pH范围为6.0-8.5,温度范围为4-30℃时生长。从它们的培养特性可以看出,它们都具有一定的耐冷性,这是它们在长年低温的环境中生长对冷适应的一种外在表现。
4.通过生理生化特征的研究,发现TSBY 57T、TSBY 67T和Enf 54T的革兰氏染色、酶活、糖代谢、同化作用等方面都具有各自所在属的共性,与属内相近种的菌株进行细致比较后,发现它们与各自属已发表的种具有不同之处,为新种的分类鉴定提供了生理生化方面的证据。
5.通过细胞化学组分分析后,得出结论TSBY 57T、TSBY 67T和Enf 54T在脂肪酸的种类上与各自属的所有种的脂肪酸种类是一致的,而平行实验的结果显示,它们有几类脂肪酸的丰度与已发表的新种有很大的区别,为这三株细菌是各自属的一个新种提供了有力的证据。
6.通过分子生物学方法,对TSBY 57T、TSBY 67T和Enf 54T进行研究后,TSBY 57T、TSBY 67T和Enf 54T的G+C含量分别为34.1%,33.5%和66.8%,分别都在该属已报道的G+C含量的范围内,并且通过16S rRNA序列相似性比较,TSBY 57T与Epilithonimonas属的亲缘关系最近,TSBY 67T与Chryseobacterium属的亲缘关系最近,Enf 54T与Frondihabitans属的关系最近,进一步证明了它们分别是各自所在属的一个新种。
综上所述,确定TSBY 57T是Epilithonimonas的一个新种,命名为Epilithonimonas xinjiangense sp. nov.;确定TSBY 67T是Chryseobacterium属的一个新种,命名为Chryseobacterium xinjiangense sp. nov。初步判定Enf 54T是Frondihabitans属的一个新种,命名为Frondihabitans xinjiangense sp. nov。
Bacteria from alpine frozen soils live in the environment with low temperature, oligotrophic system, and the absence of liquid water and O2. So Bacteria from alpine frozen soils are kinds of extreme microorganism. Bacteria from alpine frozen soils have lots of species. At the same time, their metabolites are new types resources and have great benefit to people. To develop new types resources, people must isolate more bacteria from alpine frozen soils, and classify and identify them.
Endophytic bacteria residing within plant tissues without any harm to the plant play active roles in plant. It could be a new approach to explore ecosystem type agriculture. To develop new types resources, people must isolate more bacteria from plants living all kinds of environments, and classify and identify them.
My paper mainly classify and identify strains isolating from alpine frozen soils and subnival plant-Leontopodium leontopodoides, and confirm their new taxonomic positions.
1. Strain TSBY 57T and strain TSBY 67T were isolated from alpine permafrost. Strain Enf 54T was isolated from Leontopodium leontopodoides in Tianshan Mountains. Their pure cultures which we obtain provide good material of study. We have firstly performed systematic taxonomy of isolated strains from alpine permafrost and Leontopodium leontopodoides, in Tianshan Mountains.
2. Cells of strain TSBY 57T are non-spore-forming, non-motile rods,0.2-0.3μm in diameter and 0.3-1.1μm in length. Colonies are yellow; Cells of strain TSBY 67T are non-spore-forming, non-motile rods,0.3-0.5μm in diameter and 0.6-1.6μm in length. Colonies are smooth and shiny, orange-yellow. Flexirubin type pigments are produced; Cells of strain Enf 54T are non-endospore-forming, non-motile, irregular-shaped rods,0.3μmin diameter and 0.9μm in length. Colonies are pale yellow. Therefore, their morphological features are respectively in accordance with morphological features of their respective genus. These results lay the foundation for classification and identification in the aspect of morphology.
3. Strain TSBY 57T grew with 0-1.5% NaCl, at 0-30℃, and at pH 6-8; Strain TSBY 67T grew with 0-2% NaCl, at 4-37℃, and at pH 6-8; Strain Enf 54T grew with 0-4% NaCl, at 4-30℃, and at pH 6.0-8.5. As a conclusion, they have cold resistance to a certain extent. That is external expression that they adapt to low temperature all the year round.
4. In light of the physiological and chemical tests, Strains TSBY 57T, TSBY 67T and Enf 54T are respectively considered to have generality of their respective genus and have obviously distinct from other closely related members of their respective genus.
5. We conclude that the fatty acids'composition of TSBY 57T, TSBY 67T and Enf 54T were respectively consistent with those of the closest phylogenetic neighbours grown under the same conditions. However, the content of fatty acids differed from them by higher proportions by lower proportion. Hence, it is a powerful evidence to classify and identify.
6. Studying by means of molecular biology, The DNA G+C content of strain TSBY57T, TSBY67T and Enf 54T were respectively 34.1mol%,33.5 mol% and 66.8mol%, lying within the range reported for members of their respective genus. On the basis of the sequence similarity of 16S rRNA gene, strain TSBY 57T was closely related to members of the genus Epilithonimonas, strain TSBY 67T was closely related to members of the genus Chryseobacterium and strain Enf 54T was closely related to members of the genus Frondihabitans.
Therefore, based on polyphasic taxonomic analytical results, strain TSBY57T represents a novel species of the genus Epilithonimonas, which is named as Epilithonimonas xinjiangense sp. nov., strain TSBY 67T represents a novel species of the genus Chryseobacterium, which is named as Chryseobacterium xinjiangense sp. nov., and strain Enf 54T could be a novel species of the genus Frondihabitans, which is named as Frondihabitans xinjiangense sp. nov..
植物内生细菌定殖于健康的植物宿主体内,与植物建立了和谐联合的关系,对于宿主的健康并无威胁,还对植物宿主的生命活动有着积极的作用,研究植物内生细菌对宿主的作用可能是未来生态型农业发展的一条新途径。为了发展新资源,人们必须从不同环境生长的植物中分离到更多的内生细菌,并进行分门归类。
本工作主要针对从冻土微生物多样性研究和冰缘植物内生细菌多样性研究中得到的可能具有新分类地位的细菌菌株进行分类鉴定的工作,主要结果如下：
1.对分离于天山一号冰川采集的冻土样品中细菌进行筛选后,得到两株可能是新种的细菌菌株,记为TSBY 57T和TSBY 67T;对分离于冰缘植物火绒草的内生细菌进行筛选后,得到一株可能是新种的细菌菌株,记为Enf 54T,三株细菌的纯培养物的得到为进一步的研究提供了素材,这是首次对天山冻土中分离的细菌进行分类鉴定,也是首次对冰缘植物火绒草的内生细菌进行分类鉴定。
2. TSBY 57T的菌落呈黄色,细胞为杆状,大小为0.2-0.3μm×0.3-1.1μm; TSBY 67T的菌落呈橙黄色,Flexirubin-type色素反应为阳性,细胞为杆状,大小为0.3-0.5μm×0.6-1.6μm; Enf 54T的菌落呈淡黄色,细胞为短杆状,大小为0.3μm×0.9μm,它们的在形态方面的主要特征与各自所在属的特征基本相符,奠定了分类鉴定研究中形态学的基础。
3.通过培养特性研究,TSBY 57T能在NaCl浓度为0-1.5%,pH范围为6-8,、温度范围为0-30℃时生长；TSBY 67T能在NaCl浓度为0-2%,pH范围为6-8,温度范围为4-37℃时生长;Enf 54T能在NaCl浓度为0-4%,能在pH范围为6.0-8.5,温度范围为4-30℃时生长。从它们的培养特性可以看出,它们都具有一定的耐冷性,这是它们在长年低温的环境中生长对冷适应的一种外在表现。
4.通过生理生化特征的研究,发现TSBY 57T、TSBY 67T和Enf 54T的革兰氏染色、酶活、糖代谢、同化作用等方面都具有各自所在属的共性,与属内相近种的菌株进行细致比较后,发现它们与各自属已发表的种具有不同之处,为新种的分类鉴定提供了生理生化方面的证据。
5.通过细胞化学组分分析后,得出结论TSBY 57T、TSBY 67T和Enf 54T在脂肪酸的种类上与各自属的所有种的脂肪酸种类是一致的,而平行实验的结果显示,它们有几类脂肪酸的丰度与已发表的新种有很大的区别,为这三株细菌是各自属的一个新种提供了有力的证据。
6.通过分子生物学方法,对TSBY 57T、TSBY 67T和Enf 54T进行研究后,TSBY 57T、TSBY 67T和Enf 54T的G+C含量分别为34.1%,33.5%和66.8%,分别都在该属已报道的G+C含量的范围内,并且通过16S rRNA序列相似性比较,TSBY 57T与Epilithonimonas属的亲缘关系最近,TSBY 67T与Chryseobacterium属的亲缘关系最近,Enf 54T与Frondihabitans属的关系最近,进一步证明了它们分别是各自所在属的一个新种。
综上所述,确定TSBY 57T是Epilithonimonas的一个新种,命名为Epilithonimonas xinjiangense sp. nov.;确定TSBY 67T是Chryseobacterium属的一个新种,命名为Chryseobacterium xinjiangense sp. nov。初步判定Enf 54T是Frondihabitans属的一个新种,命名为Frondihabitans xinjiangense sp. nov。
Bacteria from alpine frozen soils live in the environment with low temperature, oligotrophic system, and the absence of liquid water and O2. So Bacteria from alpine frozen soils are kinds of extreme microorganism. Bacteria from alpine frozen soils have lots of species. At the same time, their metabolites are new types resources and have great benefit to people. To develop new types resources, people must isolate more bacteria from alpine frozen soils, and classify and identify them.
Endophytic bacteria residing within plant tissues without any harm to the plant play active roles in plant. It could be a new approach to explore ecosystem type agriculture. To develop new types resources, people must isolate more bacteria from plants living all kinds of environments, and classify and identify them.
My paper mainly classify and identify strains isolating from alpine frozen soils and subnival plant-Leontopodium leontopodoides, and confirm their new taxonomic positions.
1. Strain TSBY 57T and strain TSBY 67T were isolated from alpine permafrost. Strain Enf 54T was isolated from Leontopodium leontopodoides in Tianshan Mountains. Their pure cultures which we obtain provide good material of study. We have firstly performed systematic taxonomy of isolated strains from alpine permafrost and Leontopodium leontopodoides, in Tianshan Mountains.
2. Cells of strain TSBY 57T are non-spore-forming, non-motile rods,0.2-0.3μm in diameter and 0.3-1.1μm in length. Colonies are yellow; Cells of strain TSBY 67T are non-spore-forming, non-motile rods,0.3-0.5μm in diameter and 0.6-1.6μm in length. Colonies are smooth and shiny, orange-yellow. Flexirubin type pigments are produced; Cells of strain Enf 54T are non-endospore-forming, non-motile, irregular-shaped rods,0.3μmin diameter and 0.9μm in length. Colonies are pale yellow. Therefore, their morphological features are respectively in accordance with morphological features of their respective genus. These results lay the foundation for classification and identification in the aspect of morphology.
3. Strain TSBY 57T grew with 0-1.5% NaCl, at 0-30℃, and at pH 6-8; Strain TSBY 67T grew with 0-2% NaCl, at 4-37℃, and at pH 6-8; Strain Enf 54T grew with 0-4% NaCl, at 4-30℃, and at pH 6.0-8.5. As a conclusion, they have cold resistance to a certain extent. That is external expression that they adapt to low temperature all the year round.
4. In light of the physiological and chemical tests, Strains TSBY 57T, TSBY 67T and Enf 54T are respectively considered to have generality of their respective genus and have obviously distinct from other closely related members of their respective genus.
5. We conclude that the fatty acids'composition of TSBY 57T, TSBY 67T and Enf 54T were respectively consistent with those of the closest phylogenetic neighbours grown under the same conditions. However, the content of fatty acids differed from them by higher proportions by lower proportion. Hence, it is a powerful evidence to classify and identify.
6. Studying by means of molecular biology, The DNA G+C content of strain TSBY57T, TSBY67T and Enf 54T were respectively 34.1mol%,33.5 mol% and 66.8mol%, lying within the range reported for members of their respective genus. On the basis of the sequence similarity of 16S rRNA gene, strain TSBY 57T was closely related to members of the genus Epilithonimonas, strain TSBY 67T was closely related to members of the genus Chryseobacterium and strain Enf 54T was closely related to members of the genus Frondihabitans.
Therefore, based on polyphasic taxonomic analytical results, strain TSBY57T represents a novel species of the genus Epilithonimonas, which is named as Epilithonimonas xinjiangense sp. nov., strain TSBY 67T represents a novel species of the genus Chryseobacterium, which is named as Chryseobacterium xinjiangense sp. nov., and strain Enf 54T could be a novel species of the genus Frondihabitans, which is named as Frondihabitans xinjiangense sp. nov..
引文
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