南海4种珊瑚附生可培养真菌多样性分析
|
摘要
本研究利用从南海采集的4种珊瑚分离附生真菌,并通过菌株的分类鉴定及系统发育分析获得珊瑚附生可培养真菌的多样性信息.选用酵母麦芽糖琼脂培养基(YM)、马铃薯葡萄糖琼脂培养基(PDA)、马丁氏培养基(Martin)、察氏琼脂培养基(CDA)和孟加拉红培养基(RBM)等5种培养基,采用平板涂布法培养分离附生真菌,基于ITS-rRNA基因序列对真菌菌株进行鉴定和系统发育分析.从4种珊瑚上共分离获得10个属的120株真菌,分别是:青霉属(Penicillum)56株、曲霉属(Aspergillus)25株、枝顶孢属(Acremonium)2株、枝孢属(Cladosporium)5株、毕赤酵母属(Meyerozyma)14株、隔孢伏革属(Peniophora)2株、拟茎点霉属(Phomopsis)1株、裂褶菌属(Schizophyllum)1株、梗孢酵母属(Sterigmatomyces)13株、篮状菌属(Talaromyces)1株.通过对不同珊瑚品种分离获得的附生真菌的比较,发现小叶鹿角珊瑚(Acropora microphthalma)和花鹿角珊瑚(Acropora floride)上分离到的菌株多样性最高,包含6属10种.进一步的分析发现不同珊瑚样品上真菌种类分布有较大差异,隔孢伏革属和裂褶菌属真菌均分离自小叶鹿角珊瑚,梗孢酵母属和篮状菌属均分离自角孔珊瑚(Goniopora sp.),扁脑珊瑚(Platygyra sp.)上则分离到拟茎点霉属.同时还发现不同培养基分离真菌的能力不同,YM培养基分离的菌株最多,CDA培养基分离真菌的多样性最多.以上的研究结果表明,南海珊瑚上存在大量且丰富的真菌资源,可以为后续生物活性物质的开发利用提供良好的菌种资源.
The diversity of culturable fungi derived from four coral species from the South China Sea were investigated in this study. By spread plate method,YM,PDA,Martin,CDA,RBM media were used to isolate coralassociated fungi. Identification and phylogenetic analysis of coral-associated fungi were conducted based on ITS-rRNA gene sequencing. A total of 120 fungi strains were isolated from 4 corals( Platygyra sp.,Goniopora sp.,Acropora microphthalma and Acropora florid) and these fungi strains belong to 11 genera,including Penicillum( 56 strains),Aspergillus( 25 strains),Acremonium( 2 strains),Cladosporium( 5 strains),Meyerozyma( 14 strains),Peniophora( 2 strains),Phomopsis( 1 strain),Schizophyllum( 1 strain),Sterigmatomyces( 13 strains) and Talaromyces( 1 strain). The most diverse coral species was Acropora microphthalma and Acropora floride,both including 6 genus and 10 species. Fungal species distribution varied among different coral species.Peniophora sp. and Schizophyllum commune were only recovered from the coral species Acropora microphthalma,while Sterigmatomyces sp. and Talaromyces verruculosus were only present in the coral species Goniopora sp..Only one strain Phomopsis sp. was obtained from coral Platygyra sp. Total number and species of culturable fungal strains recovered from the five media. YM media recovered the highest amount of fungal strains,accounting for 29. 17 % of the total strains. CDA media recovered the most diverse fungal species. These results suggest that there are diverse fungal symbionts associated with corals collected from the South China Sea,which are important sources for marine bioactive compounds.
The diversity of culturable fungi derived from four coral species from the South China Sea were investigated in this study. By spread plate method,YM,PDA,Martin,CDA,RBM media were used to isolate coralassociated fungi. Identification and phylogenetic analysis of coral-associated fungi were conducted based on ITS-rRNA gene sequencing. A total of 120 fungi strains were isolated from 4 corals( Platygyra sp.,Goniopora sp.,Acropora microphthalma and Acropora florid) and these fungi strains belong to 11 genera,including Penicillum( 56 strains),Aspergillus( 25 strains),Acremonium( 2 strains),Cladosporium( 5 strains),Meyerozyma( 14 strains),Peniophora( 2 strains),Phomopsis( 1 strain),Schizophyllum( 1 strain),Sterigmatomyces( 13 strains) and Talaromyces( 1 strain). The most diverse coral species was Acropora microphthalma and Acropora floride,both including 6 genus and 10 species. Fungal species distribution varied among different coral species.Peniophora sp. and Schizophyllum commune were only recovered from the coral species Acropora microphthalma,while Sterigmatomyces sp. and Talaromyces verruculosus were only present in the coral species Goniopora sp..Only one strain Phomopsis sp. was obtained from coral Platygyra sp. Total number and species of culturable fungal strains recovered from the five media. YM media recovered the highest amount of fungal strains,accounting for 29. 17 % of the total strains. CDA media recovered the most diverse fungal species. These results suggest that there are diverse fungal symbionts associated with corals collected from the South China Sea,which are important sources for marine bioactive compounds.
引文
[1]周进,晋慧,蔡中华.微生物在珊瑚礁生态系统中的作用与功能[J].应用生态学报,2014,25(3):919-930.
[2]Nagelkerken I K,Smith G W,Bonair K.Widespread disease in Caribbean sea fans:II.Patterns of infection and tissue loss[J].Marine Ecology Progress Series,1997,160:255-263.
[3]Geiser D M,Taylor J W,Ritchie K B,et al.Cause of sea fan death in the West Indies[J].Nature,1998,6(394):137-138.
[4]Alker A P,Smith G W,Kiho K.Characterization of Aspergillus sydowii(Thom et Church),a fungal pathogen of Caribbean sea fan corals[J].Hydrobiologia,2001,460(1/3):105-111.
[5]Preedanon S,Phongpaichit S,Rungjindama N,et al.Aspergillus of the gorgonian sea fan Annella sp.after the 2004 tsunami at Mu Ko Similan National Park,Andaman Sea,Thailand[J].Coral Reefs,2006,25(2):296.
[6]Rypien K L,Baker D M.Isotopic labeling and antifungal resistance as tracers of gut passage of the sea fan pathogen Aspergillus sydowii[J].Diseases of Aquatic Organisms,2009,86(1):1-7.
[7]Wegley L,Edwards R,Rodriguez-Brito B,et al.Metagenomic analysis of the microbial community associated with the coral Poritesastreoides[J].Environmental Microbiology,2007,9(11):2 707-2 719.
[8]Putri D A,Radjasa O K,Pringgenies D.Effectiveness of marine fungal symbiont isolated from soft coral Sinularia sp.from Panjang Island as antifungal[J].Procedia Environmental Sciences,2015,23:351-357.
[9]Canosa B J,Duenas L F,Sanchez J A.Isolation of potential fungal pathogens in gorgonian corals at the Tropical Eastern Pacific[J].Coral Reefs,2013,32(1):35-41.
[10]Toledo-Hernández C,Zuluaga-Montero A,Bones-González A,et al.Fungi in healthy and diseased sea fans(Gorgonia ventalina):is Aspergillus sydowii,always the pathogen?[J].Coral Reefs,2008,27(3):707-714.
[11]Yarden O,Ainsworth T D,Roff J,et al.Increased prevalence of ubiquitous Ascomycetes in an acroporid coral(Acroporaformosa)exhibiting symptoms of brown band syndrome and skeletal eroding band diseases[J].Applied and Environmental Microbiology,2007,73(8):2 755-2 757.
[12]Ramosflores T.Lower marine fungus associated with black line disease in star corals(Montastrea annularis,E and S)[J].Biological Bulletin,1983,165(2):429-435.
[13]Raghukumar C,Raghukumar S.Fungal invasion of massive corals[J].Marine Ecology,1991,12(3):251-160.
[14]肖胜蓝,雷晓凌,徐佳,等.徐闻8种珊瑚附生真菌的分离及初步鉴定[J].广东海洋大学学报:自然科学版,2011,31(6):91-95.
[15]肖胜蓝.徐闻3种石珊瑚附生可培养真菌多样性及其抗菌活性的研究[D].湛江:广东海洋大学,2012.
[16]徐佳,陈彬,雷晓凌,等.丛生盔形珊瑚附生可培养真菌多样性分析[J].微生物学通报,2011,38(8):1 193-1 198.
[17]王亚楠.中国南海柳珊瑚附生微生物多样性与抗菌作用研究[D].青岛:中国海洋大学,2012.
[18]Zhang X Y,Bao J,Wang G H,et al.Diversity and antimicrobial activity of culturable fungi isolated from six species of the South China Sea gorgonians[J].Microbial Ecology,2012,64(3):617-627.
[19]Wang Y N,Shao C L,Zheng C J.Diversity and antibacterial activities of fungi derived from the Gorgonian Echinogorgia rebekka from the South China Sea[J].Marine Drugs,2011,(9):1 379-1 390.
[20]Li J,Zhong M,Lei X,et al.Diversity and antibacterial activities of culturable fungi associated with coral Porites pukoensis[J].World Journal of Microbiology Biotechnology,2014,30(10):2 551-2 558.
[21]Zhang X Y,Sun Y L,Bao J,et al.Phylogenetic survey and antimicrobial activity of culturable microorganisms associated with the South China Sea black coral Antipathes dichotoma[J].FEMS Microbiology Letters,2012,336(2):122-130.
[22]张丹,蒋群,李英新,等.中国南海海绵和珊瑚共附生真菌多样性及其抑菌活性研究[J].中国海洋药物,2015,34(2):1-9.
[23]周静,陈敏,李筠,等.一株网刺柳珊瑚Echinogorgia sp.共附生真菌Peniophora sp.次级代谢产物研究[J].中国海洋药物,2012,31(4):8-13.
[24]刘楠,李英新,李志勇,等.珊瑚来源真菌天然产物及其活性的研究进展[J].中国海洋药物,2014,33(4):73-84.
[25]Zhang D,Satake M,Fukuzawa S,et al.Two new indole alkaloids,2-(3,3-dimethylprop-1-ene)-costaclavine and 2-(3,3-dimethylprop-1-ene)-epicostaclavine,from the marine-derived fungus Aspergillus fumigatus[J].Journal of Natural Medicines,2012,66(1):222-226.
[26]Khamthong N,Rukachaisirikul V,Phongpaichit S,et al.Bioactive polyketides from the sea fan-derived fungus Penicillum citrinum PSU-F51[J].Tetrahedron,2012,68(39):8 245-8 250.
[27]Shao C L,Wu H X,Wang C Y,et al.Potent antifouling resorcylic acid lactones from the gorgonian-derived fungus Cochliobolus lunatus[J].Journal of Natural Products,2011,74(4):629-633.
[28]Bao J,Sun Y L,Zhang X Y,et al.Antifouling and antibacterial polyketides from marine gorgonian coral-associated fungus Pencillum sp.SCSGAF 0023[J].Journal of Antibiotics,2013,66(4):219-223.
[29]Bao J,Zhang X Y,Xu X Y,et al.New cyclic tetrapeptides and asteltoxins from gorgonian-derived fungus Aspergillus sp.SCSGAF 0076[J].Tetrahedron,2013,69(9):2 113-2 117.
[30]Zhuang Y,Teng X,Wang Y,et al.Cyclopeptides and polyketides from coral-associated fungus,Aspergillus versicolor LCJ-5-4[J].Tetrahedron,2011,67(37):7 085-7 089.
[31]Wei M Y,Li D,Shao C L,et al.(±)-Pestalachloride D,an antibacterial racemate of chlorinated benzophenone derivative from a soft coral-derived fungus Pestalotiopsis sp.[J].Marine Drugs,2013,11(4):1 050-1 060.
[32]Tseng W R,Huang C Y,Tsai Y Y,et al.New cytotoxic and anti-inflammatory steroids from the soft coral Klyxum flaccidum[J].Bioorganic&Medicinal Chemistry Letters,2016,26(14):3 253-3 257.
[33]Zhang Y J,Zhang S,Liu X Z,et al.A simple method of genomic DNA extraction suitable for analysis of bulk fungal strains[J].Letters in Applied Microbiology,2010,51(1):114-118.
[34]Zhang T,Wei X L,Wei Y Z,et al.Diversity and distribution of cultured endolichenic fungi in the Ny-Alesund Region,Svalbard(High Arctic)[J].Extremophiles,2016,20(4):461-470.
[35]Amended A S,Barshis D J,Oliver T A.Coral-associated marine fungi form novel lineages and heterogeneous assemblages[J].International Society for Microbial Ecology,2011,6(7):1 291-1 301.
[36]Toledo-Hernández C,Bones-González A,Ortiz-Vázquez O E,et al.Fungi in the sea fan Gorgonia ventalina:diversity and sampling strategies[J].Coral Reefs,2007,26(3):725-730.
[2]Nagelkerken I K,Smith G W,Bonair K.Widespread disease in Caribbean sea fans:II.Patterns of infection and tissue loss[J].Marine Ecology Progress Series,1997,160:255-263.
[3]Geiser D M,Taylor J W,Ritchie K B,et al.Cause of sea fan death in the West Indies[J].Nature,1998,6(394):137-138.
[4]Alker A P,Smith G W,Kiho K.Characterization of Aspergillus sydowii(Thom et Church),a fungal pathogen of Caribbean sea fan corals[J].Hydrobiologia,2001,460(1/3):105-111.
[5]Preedanon S,Phongpaichit S,Rungjindama N,et al.Aspergillus of the gorgonian sea fan Annella sp.after the 2004 tsunami at Mu Ko Similan National Park,Andaman Sea,Thailand[J].Coral Reefs,2006,25(2):296.
[6]Rypien K L,Baker D M.Isotopic labeling and antifungal resistance as tracers of gut passage of the sea fan pathogen Aspergillus sydowii[J].Diseases of Aquatic Organisms,2009,86(1):1-7.
[7]Wegley L,Edwards R,Rodriguez-Brito B,et al.Metagenomic analysis of the microbial community associated with the coral Poritesastreoides[J].Environmental Microbiology,2007,9(11):2 707-2 719.
[8]Putri D A,Radjasa O K,Pringgenies D.Effectiveness of marine fungal symbiont isolated from soft coral Sinularia sp.from Panjang Island as antifungal[J].Procedia Environmental Sciences,2015,23:351-357.
[9]Canosa B J,Duenas L F,Sanchez J A.Isolation of potential fungal pathogens in gorgonian corals at the Tropical Eastern Pacific[J].Coral Reefs,2013,32(1):35-41.
[10]Toledo-Hernández C,Zuluaga-Montero A,Bones-González A,et al.Fungi in healthy and diseased sea fans(Gorgonia ventalina):is Aspergillus sydowii,always the pathogen?[J].Coral Reefs,2008,27(3):707-714.
[11]Yarden O,Ainsworth T D,Roff J,et al.Increased prevalence of ubiquitous Ascomycetes in an acroporid coral(Acroporaformosa)exhibiting symptoms of brown band syndrome and skeletal eroding band diseases[J].Applied and Environmental Microbiology,2007,73(8):2 755-2 757.
[12]Ramosflores T.Lower marine fungus associated with black line disease in star corals(Montastrea annularis,E and S)[J].Biological Bulletin,1983,165(2):429-435.
[13]Raghukumar C,Raghukumar S.Fungal invasion of massive corals[J].Marine Ecology,1991,12(3):251-160.
[14]肖胜蓝,雷晓凌,徐佳,等.徐闻8种珊瑚附生真菌的分离及初步鉴定[J].广东海洋大学学报:自然科学版,2011,31(6):91-95.
[15]肖胜蓝.徐闻3种石珊瑚附生可培养真菌多样性及其抗菌活性的研究[D].湛江:广东海洋大学,2012.
[16]徐佳,陈彬,雷晓凌,等.丛生盔形珊瑚附生可培养真菌多样性分析[J].微生物学通报,2011,38(8):1 193-1 198.
[17]王亚楠.中国南海柳珊瑚附生微生物多样性与抗菌作用研究[D].青岛:中国海洋大学,2012.
[18]Zhang X Y,Bao J,Wang G H,et al.Diversity and antimicrobial activity of culturable fungi isolated from six species of the South China Sea gorgonians[J].Microbial Ecology,2012,64(3):617-627.
[19]Wang Y N,Shao C L,Zheng C J.Diversity and antibacterial activities of fungi derived from the Gorgonian Echinogorgia rebekka from the South China Sea[J].Marine Drugs,2011,(9):1 379-1 390.
[20]Li J,Zhong M,Lei X,et al.Diversity and antibacterial activities of culturable fungi associated with coral Porites pukoensis[J].World Journal of Microbiology Biotechnology,2014,30(10):2 551-2 558.
[21]Zhang X Y,Sun Y L,Bao J,et al.Phylogenetic survey and antimicrobial activity of culturable microorganisms associated with the South China Sea black coral Antipathes dichotoma[J].FEMS Microbiology Letters,2012,336(2):122-130.
[22]张丹,蒋群,李英新,等.中国南海海绵和珊瑚共附生真菌多样性及其抑菌活性研究[J].中国海洋药物,2015,34(2):1-9.
[23]周静,陈敏,李筠,等.一株网刺柳珊瑚Echinogorgia sp.共附生真菌Peniophora sp.次级代谢产物研究[J].中国海洋药物,2012,31(4):8-13.
[24]刘楠,李英新,李志勇,等.珊瑚来源真菌天然产物及其活性的研究进展[J].中国海洋药物,2014,33(4):73-84.
[25]Zhang D,Satake M,Fukuzawa S,et al.Two new indole alkaloids,2-(3,3-dimethylprop-1-ene)-costaclavine and 2-(3,3-dimethylprop-1-ene)-epicostaclavine,from the marine-derived fungus Aspergillus fumigatus[J].Journal of Natural Medicines,2012,66(1):222-226.
[26]Khamthong N,Rukachaisirikul V,Phongpaichit S,et al.Bioactive polyketides from the sea fan-derived fungus Penicillum citrinum PSU-F51[J].Tetrahedron,2012,68(39):8 245-8 250.
[27]Shao C L,Wu H X,Wang C Y,et al.Potent antifouling resorcylic acid lactones from the gorgonian-derived fungus Cochliobolus lunatus[J].Journal of Natural Products,2011,74(4):629-633.
[28]Bao J,Sun Y L,Zhang X Y,et al.Antifouling and antibacterial polyketides from marine gorgonian coral-associated fungus Pencillum sp.SCSGAF 0023[J].Journal of Antibiotics,2013,66(4):219-223.
[29]Bao J,Zhang X Y,Xu X Y,et al.New cyclic tetrapeptides and asteltoxins from gorgonian-derived fungus Aspergillus sp.SCSGAF 0076[J].Tetrahedron,2013,69(9):2 113-2 117.
[30]Zhuang Y,Teng X,Wang Y,et al.Cyclopeptides and polyketides from coral-associated fungus,Aspergillus versicolor LCJ-5-4[J].Tetrahedron,2011,67(37):7 085-7 089.
[31]Wei M Y,Li D,Shao C L,et al.(±)-Pestalachloride D,an antibacterial racemate of chlorinated benzophenone derivative from a soft coral-derived fungus Pestalotiopsis sp.[J].Marine Drugs,2013,11(4):1 050-1 060.
[32]Tseng W R,Huang C Y,Tsai Y Y,et al.New cytotoxic and anti-inflammatory steroids from the soft coral Klyxum flaccidum[J].Bioorganic&Medicinal Chemistry Letters,2016,26(14):3 253-3 257.
[33]Zhang Y J,Zhang S,Liu X Z,et al.A simple method of genomic DNA extraction suitable for analysis of bulk fungal strains[J].Letters in Applied Microbiology,2010,51(1):114-118.
[34]Zhang T,Wei X L,Wei Y Z,et al.Diversity and distribution of cultured endolichenic fungi in the Ny-Alesund Region,Svalbard(High Arctic)[J].Extremophiles,2016,20(4):461-470.
[35]Amended A S,Barshis D J,Oliver T A.Coral-associated marine fungi form novel lineages and heterogeneous assemblages[J].International Society for Microbial Ecology,2011,6(7):1 291-1 301.
[36]Toledo-Hernández C,Bones-González A,Ortiz-Vázquez O E,et al.Fungi in the sea fan Gorgonia ventalina:diversity and sampling strategies[J].Coral Reefs,2007,26(3):725-730.