云南星云湖酵母菌多样性及产类胡萝卜素的评价
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摘要
【背景】星云湖属云南高原湖泊,对其酵母菌的多样性研究和产类胡萝卜素菌株的筛选有助于开发利用星云湖酵母菌资源。【目的】研究云南星云湖酵母菌群落结构多样性,筛选和评价产类胡萝卜素活性菌株。【方法】采用膜过滤平置培养对星云湖水样进行酵母菌分离;应用26S rRNA D1/D2区域序列分析和形态特征及生理生化指标相结合,对分离获得的酵母菌进行鉴定;运用软件BIO-DAP分析酵母菌类群的多样性;采用酸热法提取类胡萝卜素,紫外分光光度计测定类胡萝卜素含量。【结果】从星云湖湖水中分离得到797株酵母菌,鉴定为18个属37个种。胶红酵母(Rhodotorula mucilaginosa)和Naganishia albida是优势种,分别占总菌株数的36.64%和28.86%;有28株酵母菌类胡萝卜素产量可达200μg/g细胞干重以上;有4株类胡萝卜素产量可达300μg/g细胞干重以上;粘红酵母(Rhodotorula glutinis) 2株、胶红酵母(Rhodotorula mucilaginosa)和Rhodosporidiobolus fluviale各1株。【结论】云南高原湖泊星云湖的酵母菌具有丰富的多样性,湖泊的营养化程度以及地理差异影响酵母菌群落的结构;红冬孢酵母属(Rhodosporidiobolus)和红酵母属(Rhodotorula)是星云湖类胡萝卜素产量可达300μg/g细胞干重以上的主要类群。有进一步开发研究的价值。
[Background] Xingyun Lake is one of plateau lakes of Yunnan. The study of its diversity of yeasts and the screening of carotenoids-producing strains contribute to the development and utilization of the yeast resources of Xingyun Lake. [Objective] To study the diversity of community structure of yeasts in Xingyun Lake, and to screen the carotenoids-producing strains. [Methods] Yeast separation was carried to on the water samples of Xingyun Lake by membrane filtration flat culture. Identification was based on sequence analysis of D1/D2 domains of 26 S rRNA gene, and to combinate with morphological characteristics and physiological and biochemical indicators, and the diversity of yeast was analyzed by software BIO-DAP. The carotenoids were extracted by acid-heating method, and the content of carotenoids were determined by ultraviolet spectrophotometer. [Results] A total of 797 strains were isolated from the water samples of Xingyun Lake, and identified as 18 genera and 37 species. The Rhodotorula mucilaginosa and Naganishia albida are the dominant species, accounting for 36.64% and28.86% of the total strains, respectively. There were 28 strains of yeast carotenoids with a yield of more than 200 μg/g cell dry weight, and 4 strains of carotenoeds-producing were up to 300 μg/g cell dry weight:Rhodotorula glutinis(2 strains), Rhodotorula mucilaginosa(1 strain) and Rhodosporidiobolus fluviale(1 strain). [Conclusion] The yeasts of Xingyun Lake are rich in diversity, and the degree of lake nutrition and geographical difference affect the structure of yeast community. In Xingyun Lake, Rhodosporidiobolus and Rhodotorula are the major groups of carotenoids-producting, and carotenoids-producting were up to300 μg/g of cell dry weight. It has the value of further development research.
[Background] Xingyun Lake is one of plateau lakes of Yunnan. The study of its diversity of yeasts and the screening of carotenoids-producing strains contribute to the development and utilization of the yeast resources of Xingyun Lake. [Objective] To study the diversity of community structure of yeasts in Xingyun Lake, and to screen the carotenoids-producing strains. [Methods] Yeast separation was carried to on the water samples of Xingyun Lake by membrane filtration flat culture. Identification was based on sequence analysis of D1/D2 domains of 26 S rRNA gene, and to combinate with morphological characteristics and physiological and biochemical indicators, and the diversity of yeast was analyzed by software BIO-DAP. The carotenoids were extracted by acid-heating method, and the content of carotenoids were determined by ultraviolet spectrophotometer. [Results] A total of 797 strains were isolated from the water samples of Xingyun Lake, and identified as 18 genera and 37 species. The Rhodotorula mucilaginosa and Naganishia albida are the dominant species, accounting for 36.64% and28.86% of the total strains, respectively. There were 28 strains of yeast carotenoids with a yield of more than 200 μg/g cell dry weight, and 4 strains of carotenoeds-producing were up to 300 μg/g cell dry weight:Rhodotorula glutinis(2 strains), Rhodotorula mucilaginosa(1 strain) and Rhodosporidiobolus fluviale(1 strain). [Conclusion] The yeasts of Xingyun Lake are rich in diversity, and the degree of lake nutrition and geographical difference affect the structure of yeast community. In Xingyun Lake, Rhodosporidiobolus and Rhodotorula are the major groups of carotenoids-producting, and carotenoids-producting were up to300 μg/g of cell dry weight. It has the value of further development research.
引文
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[7]Dong MH,Li ZY,Zhou B,et al.Diversity of cultivable yeast in Qilu Lake in winter[J].Acta Microbiologica Sinica,2016,56(4):603-613(in Chinese)董明华,李治滢,周斌,等.云南高原湖泊杞麓湖冬季可培养酵母菌多样性分析[J].微生物学报,2016,56(4):603-613
[8]Guo XF,Li ZY,Dong MH,et al.Spatial dynamics of yeast community and its relationship to environmental factors in Lake Fuxian,Yunnan Province[J].Journal of Lake Sciences,2016,28(2):358-369(in Chinese)郭小芳,李治滢,董明华,等.云南高原湖泊抚仙湖酵母菌空间分布及其与环境因子的关系[J].湖泊科学,2016,28(2):358-369
[9]Li ZY,Zhou XL,Zhou B,et al.Diversity and extracellular enzymes of yeasts from Chenghai Lake in winter[J].Mycosystema,2017,36(2):177-185(in Chinese)李治滢,周新丽,周斌,等.云南程海湖冬季酵母菌多样性及胞外酶活性研究[J].菌物学报,2017,36(2):177-185
[10]Zaky AS,Tucker GA,Daw ZY,et al.Marine yeast isolation and industrial application[J].FEMS Yeast Research,2015,14(6):813-825
[11]Fan ZQ,Li ZY,Dong MH,et al.Carotenoid-producing yeasts resources in Fuxian Lake,Yunnan[J].Microbiology China,2017,44(2):296-304(in Chinese)樊竹青,李治滢,董明华,等.云南抚仙湖产类胡萝卜素酵母菌的资源调查[J].微生物学通报,2017,44(2):296-304
[12]Villarreal P,Carrasco M,Barahona S,et al.Tolerance to ultraviolet radiation of psychrotolerant yeasts and analysis of their carotenoid,mycosporine,and ergosterol content[J].Current Microbiology,2016,72(1):94-101
[13]Kurtzman CP,Fell JW,Boekhout T.The Yeast:A Taxonomic Study[M].5th ed.London:Elsevier Science,2011:293-2058
[14]Nisiotou AA,Spiropoulos AE,Nychas GJE.Yeast community structures and dynamics in healthy and Botrytis-affected grape must fermentations[J].Applied and Environment Microbiology,2007,73(21):6705-6713
[15]Kurtzman CP,Robnett CJ.Identification and phylogeny of ascomycetous yeasts from analysis of nuclear large subunit(26S)ribosomal DNA partial sequences[J].Antonie van Leeuwenhoek,1998,73(4):331-371
[16]Fell JW,Noekhout T,Fonseca A,et al.Biodiversity and systematics of basidiomycetous yeasts as determined by large-subunit rDNA D1/D2 domain sequence analysis[J].International Journal of Systematic and Evolutionary Microbiology,2000,50(3):1351-1371
[17]Saitou N,Nei M.The neighbor-joining method:a new method for reconstructing phylogenetic trees[J].Molecular Biology and Evolution,1987,4(4):406-425
[18]Larkin MA,Blackshields G,Brown NP,et al.Clustal W and Clustal X version 2.0[J].Bioinformatics,2007,23(21):2947-2948
[19]Tamura K,Peterson D,Peterson N,et al.MEGA5:molecular evolutionary genetics analysis using maximum likelihood,evolutionary distance,and maximum parsimony methods[J].Molecular Biology and Evolution,2011,28(10):2731-2739
[20]Liu XZ,Wang QM,G?ker M,et al.Towards an integrated phylogenetic classification of the Tremellomycetes[J].Studies in Mycology,2015,81:85-147
[21]Wang QM,Yurkov AM,G?ker M,et al.Phylogenetic classification of yeasts and related taxa within Pucciniomycotina[J].Studies in Mycology,2015,81:149-189
[22]Thomas G,Clay D,Magurran A.BIODAP-Ecological Diversity and Its Measurement[M].Alma,New Brunswick,Canada:Fundy National Park,2000
[23]Brand?o LR,Libkind D,Vaz ABM,et al.Yeasts from an oligotrophic lake in Patagonia(Argentina):diversity,distribution and synthesis of photoprotective compounds and extracellular enzymes[J].FEMS Microbiology Ecology,2011,76(1):1-13
[24]Wang W,Wu Q,Xu Y.Identification and analysis of yeast community structure in Chinese light-style liquor brewing process[J].Microbiology China,2012,39(9):1272-1279(in Chinese)王薇,吴群,徐岩.清香型白酒固态酿造过程中酵母种群结构和多样性分析[J].微生物学通报,2012,39(9):1272-1279
[25]Qing MJ,Bai M,Zhang Y,et al.Identification and biodiversity of yeasts from Qula in Tibet and milk cake in Yunnan of China[J].Acta Microbiologica Sinica,2010,50(9):1141-1146(in Chinese)卿蔓君,白梅,张勇,等.西藏曲拉和云南乳饼中酵母菌的鉴定及其生物多样性[J].微生物学报,2010,50(9):1141-1146
[26]Chen K,Li GB,Li DH,et al.Water quality improvement and algal inhabitation by Wolffia arrhiza,Schizosaccharomyces and Rhodospirillum[J].Ecological Science,2007,26(5):428-431(in Chinese).陈坤,李根保,李敦海,等.无根萍、酵母菌和红螺菌的水质净化和抑藻作用[J].生态科学,2007,26(5):428-431
[27]Tan JL,Li ZY,Zhou B,et al.Extracellular enzymatic activities of yeasts isolated from Fuxian Lake and Xingyun Lake in Yunnan Plateau[J].Microbiology China,2018,45(2):302-313(in Chinese)谭金连,李治滢,周斌,等.云南高原湖泊抚仙湖和星云湖的酵母菌胞外酶活性[J].微生物学通报,2018,45(2):302-313
[28]MolinéM,Regina Flores M,Libkind D,et al.Photoprotection by carotenoid pigments in the yeast Rhodotorula mucilaginosa:the role of torularhodin[J].Photochemical&Photobiological Sciences,2010,9(8):1145-1151
[29]Libkind D,MolinéM,Sampaio JP,et al.Yeasts from high-altitude lakes:influence of UV radiation[J].FEMSMicrobiology Ecology,2009,69(3):353-362
[30]Libkind D,Brizzio S,Ruffini A,et al.Molecular characterization of carotenogenic yeasts from aquatic environments in patagonia,argentina[J].Antonie van Leeuwenhoek,2003,84(4):313-322
[31]Mata-Gómez LC,Monta?ez JC,Méndez-Zavala A,et al.Biotechnological production of carotenoids by yeasts:an overview[J].Microbial Cell Factories,2014,13:12
[32]MolinéM,Libkind D,Diéguez MC,et al.Photoprotective role of carotenoids in yeasts:Response to UV-B of pigmented and naturally-occurring albino strains[J].Journal of Photochemistry and Photobiology B:Biology,2009,95(3):156-161
[33]Mannazzu I,Landolfo S,da Silva TL,et al.Red yeasts and carotenoid production:outlining a future for non-conventional yeasts of biotechnological interest[J].World Journal of Microbiology and Biotechnology,2015,31(11):1165-1173
[34]Xie FX,Zhang FF,Zhou K,et al.Isolation,screening and identification of yeast for aquaculture water purification[J].Acta Microbiologica Sinica,2015,55(5):635-642(in Chinese)谢凤行,张峰峰,周可,等.水质净化酵母菌的分离筛选及鉴定[J].微生物学报,2015,55(5):635-642
[2]Yang JL,Li J,Li JW,et al.Summarize on the research of the hydro-bios and aquatic environment in Fuxian Lake and Xingyun Lake[J].Yunnan Geographic Environment Research,2012,24(2):98-102,109(in Chinese)杨加林,李杰,李经纬,等.抚仙湖-星云湖水生生物与水环境研究综述[J].云南地理环境研究,2012,24(2):98-102,109
[3]Nagahama T.Yeast biodiversity in freshwater,marine and deep-sea environments[A]//Péter G,Rosa C.Biodiversity and Ecophysiology of Yeasts[M].Berlin Heidelberg:Springer,2006:241-262
[4]Baker BJ,Sheik CS,Taylor CA,et al.Community transcriptomic assembly reveals microbes that contribute to deep-sea carbon and nitrogen cycling[J].The ISME Journal,2013,7(10):1962-1973
[5]Zhou XL,Li ZY,Yang LY,et al.Identification of yeasts isolated from Chenghai Lake,a plateau lake in Yunnan province[J].Acta Microbiologica Sinica,2010,51(4):547-553(in Chinese)周新丽,李治滢,杨丽源,等.云南程海湖酵母菌多样性及应用[J].微生物学报,2010,51(4):547-553
[6]Yan YP,Li ZY,Dong MH,et al.Yeasts from Yangzonghai Lake in Yunnan(China):diversity and extracellular enzymes[J].Acta Microbiologica Sinica,2013,53(11):1205-1212(in Chinese)严亚萍,李治滢,董明华,等.云南阳宗海酵母菌种群结构及产胞外酶测试[J].微生物学报,2013,53(11):1205-1212
[7]Dong MH,Li ZY,Zhou B,et al.Diversity of cultivable yeast in Qilu Lake in winter[J].Acta Microbiologica Sinica,2016,56(4):603-613(in Chinese)董明华,李治滢,周斌,等.云南高原湖泊杞麓湖冬季可培养酵母菌多样性分析[J].微生物学报,2016,56(4):603-613
[8]Guo XF,Li ZY,Dong MH,et al.Spatial dynamics of yeast community and its relationship to environmental factors in Lake Fuxian,Yunnan Province[J].Journal of Lake Sciences,2016,28(2):358-369(in Chinese)郭小芳,李治滢,董明华,等.云南高原湖泊抚仙湖酵母菌空间分布及其与环境因子的关系[J].湖泊科学,2016,28(2):358-369
[9]Li ZY,Zhou XL,Zhou B,et al.Diversity and extracellular enzymes of yeasts from Chenghai Lake in winter[J].Mycosystema,2017,36(2):177-185(in Chinese)李治滢,周新丽,周斌,等.云南程海湖冬季酵母菌多样性及胞外酶活性研究[J].菌物学报,2017,36(2):177-185
[10]Zaky AS,Tucker GA,Daw ZY,et al.Marine yeast isolation and industrial application[J].FEMS Yeast Research,2015,14(6):813-825
[11]Fan ZQ,Li ZY,Dong MH,et al.Carotenoid-producing yeasts resources in Fuxian Lake,Yunnan[J].Microbiology China,2017,44(2):296-304(in Chinese)樊竹青,李治滢,董明华,等.云南抚仙湖产类胡萝卜素酵母菌的资源调查[J].微生物学通报,2017,44(2):296-304
[12]Villarreal P,Carrasco M,Barahona S,et al.Tolerance to ultraviolet radiation of psychrotolerant yeasts and analysis of their carotenoid,mycosporine,and ergosterol content[J].Current Microbiology,2016,72(1):94-101
[13]Kurtzman CP,Fell JW,Boekhout T.The Yeast:A Taxonomic Study[M].5th ed.London:Elsevier Science,2011:293-2058
[14]Nisiotou AA,Spiropoulos AE,Nychas GJE.Yeast community structures and dynamics in healthy and Botrytis-affected grape must fermentations[J].Applied and Environment Microbiology,2007,73(21):6705-6713
[15]Kurtzman CP,Robnett CJ.Identification and phylogeny of ascomycetous yeasts from analysis of nuclear large subunit(26S)ribosomal DNA partial sequences[J].Antonie van Leeuwenhoek,1998,73(4):331-371
[16]Fell JW,Noekhout T,Fonseca A,et al.Biodiversity and systematics of basidiomycetous yeasts as determined by large-subunit rDNA D1/D2 domain sequence analysis[J].International Journal of Systematic and Evolutionary Microbiology,2000,50(3):1351-1371
[17]Saitou N,Nei M.The neighbor-joining method:a new method for reconstructing phylogenetic trees[J].Molecular Biology and Evolution,1987,4(4):406-425
[18]Larkin MA,Blackshields G,Brown NP,et al.Clustal W and Clustal X version 2.0[J].Bioinformatics,2007,23(21):2947-2948
[19]Tamura K,Peterson D,Peterson N,et al.MEGA5:molecular evolutionary genetics analysis using maximum likelihood,evolutionary distance,and maximum parsimony methods[J].Molecular Biology and Evolution,2011,28(10):2731-2739
[20]Liu XZ,Wang QM,G?ker M,et al.Towards an integrated phylogenetic classification of the Tremellomycetes[J].Studies in Mycology,2015,81:85-147
[21]Wang QM,Yurkov AM,G?ker M,et al.Phylogenetic classification of yeasts and related taxa within Pucciniomycotina[J].Studies in Mycology,2015,81:149-189
[22]Thomas G,Clay D,Magurran A.BIODAP-Ecological Diversity and Its Measurement[M].Alma,New Brunswick,Canada:Fundy National Park,2000
[23]Brand?o LR,Libkind D,Vaz ABM,et al.Yeasts from an oligotrophic lake in Patagonia(Argentina):diversity,distribution and synthesis of photoprotective compounds and extracellular enzymes[J].FEMS Microbiology Ecology,2011,76(1):1-13
[24]Wang W,Wu Q,Xu Y.Identification and analysis of yeast community structure in Chinese light-style liquor brewing process[J].Microbiology China,2012,39(9):1272-1279(in Chinese)王薇,吴群,徐岩.清香型白酒固态酿造过程中酵母种群结构和多样性分析[J].微生物学通报,2012,39(9):1272-1279
[25]Qing MJ,Bai M,Zhang Y,et al.Identification and biodiversity of yeasts from Qula in Tibet and milk cake in Yunnan of China[J].Acta Microbiologica Sinica,2010,50(9):1141-1146(in Chinese)卿蔓君,白梅,张勇,等.西藏曲拉和云南乳饼中酵母菌的鉴定及其生物多样性[J].微生物学报,2010,50(9):1141-1146
[26]Chen K,Li GB,Li DH,et al.Water quality improvement and algal inhabitation by Wolffia arrhiza,Schizosaccharomyces and Rhodospirillum[J].Ecological Science,2007,26(5):428-431(in Chinese).陈坤,李根保,李敦海,等.无根萍、酵母菌和红螺菌的水质净化和抑藻作用[J].生态科学,2007,26(5):428-431
[27]Tan JL,Li ZY,Zhou B,et al.Extracellular enzymatic activities of yeasts isolated from Fuxian Lake and Xingyun Lake in Yunnan Plateau[J].Microbiology China,2018,45(2):302-313(in Chinese)谭金连,李治滢,周斌,等.云南高原湖泊抚仙湖和星云湖的酵母菌胞外酶活性[J].微生物学通报,2018,45(2):302-313
[28]MolinéM,Regina Flores M,Libkind D,et al.Photoprotection by carotenoid pigments in the yeast Rhodotorula mucilaginosa:the role of torularhodin[J].Photochemical&Photobiological Sciences,2010,9(8):1145-1151
[29]Libkind D,MolinéM,Sampaio JP,et al.Yeasts from high-altitude lakes:influence of UV radiation[J].FEMSMicrobiology Ecology,2009,69(3):353-362
[30]Libkind D,Brizzio S,Ruffini A,et al.Molecular characterization of carotenogenic yeasts from aquatic environments in patagonia,argentina[J].Antonie van Leeuwenhoek,2003,84(4):313-322
[31]Mata-Gómez LC,Monta?ez JC,Méndez-Zavala A,et al.Biotechnological production of carotenoids by yeasts:an overview[J].Microbial Cell Factories,2014,13:12
[32]MolinéM,Libkind D,Diéguez MC,et al.Photoprotective role of carotenoids in yeasts:Response to UV-B of pigmented and naturally-occurring albino strains[J].Journal of Photochemistry and Photobiology B:Biology,2009,95(3):156-161
[33]Mannazzu I,Landolfo S,da Silva TL,et al.Red yeasts and carotenoid production:outlining a future for non-conventional yeasts of biotechnological interest[J].World Journal of Microbiology and Biotechnology,2015,31(11):1165-1173
[34]Xie FX,Zhang FF,Zhou K,et al.Isolation,screening and identification of yeast for aquaculture water purification[J].Acta Microbiologica Sinica,2015,55(5):635-642(in Chinese)谢凤行,张峰峰,周可,等.水质净化酵母菌的分离筛选及鉴定[J].微生物学报,2015,55(5):635-642
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