胶州湾近岸污染与半知菌群体多样性关系的研究
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摘要
本文测试研究了冬、春、秋三个季节胶州湾近岸李村河入海口、海泊河入海口沉积物中氮、磷及重金属含量,以石老人近岸沉积物作为对照,利用传统生物学分离培养方法与现代分子生物学18S PCR—DGGE相结合的方法对其污染状况与该生境半知菌种群多样性进行了研究,结果表明,李村河入海口综合污染最为严重,为重污染区,海泊河综合污染较轻,为中度污染区。沉积物中可培养半知菌数量随着污染程度增加而减少,这种变化与重金属污染的程度相关性较强。而半知菌种类多样性变化趋势则不同,在轻度到中度污染的范围内,重金属综合污染指数增大时,半知菌多样性指数也相应增大,但达到重度污染时,综合污染指数增大半知菌多样性下降较快。总氮、总磷的变化只对半知菌部分种群的数量产生影响,对半知菌整体的数量以及多样性的影响不明显。试验选择了半知菌作为指示海洋环境污染的指示微生物,并根据历年研究报道,选择不同污染区域李村河入海口(重污染区)、海泊河(中污染区)以及石老人近岸(轻度污染对照区)不同站点的沉积物,通过对测试沉积物中的化学分析结果以及传统生物学培养方法所获得的沉积物中半知菌的种群和多样性的信息进行的统计、分析和归纳,并结合分子生物学18S PCR—DGGE的方法所获得的结果与上述结果进行比较,通过对三个季节半知菌种群和多样性的信息与重金属综合污染之间的相互关系进行统计分析和动态分析,表明了半知菌可以作为海洋污染的良好的指示生物;试验总结获得了半知菌种群数量以及种类多样性与重金属综合污染之间的相互关系,并据此初步设计了半知菌种群数量、多样性与海洋污染之间的关系模型,为日后在海洋污染监测的应用奠定了基础。
The contents of N, P, heavy metal in Kiaochow bay coast of Licun river and Haibo river estuaries sediments collected in winter, spring and autumn were measured with Shilaoren as control. Statistical analysis of contamination status and biodiversity of Deuteromycetes in these habitats were conducted using traditional methods combined with modern 18S PCR-DGGE methods. The results showed Licun river estuaries was seriously polluted while Haibo river estuaries was moderate polluted and the amount of culturable Deuteromycetes decreased as the increase of pollution degree and this tendency was obviously relevant with heavy metal pollution degree. However, the tendency of Deuteromycetes biodiversity was different: within slight to moderate pollution scope, the biodiversity index of Deuteromycetes increased as the increase of heavy metal index while the biodiversity index of Deuteromycetes decreased sharply within seriously pollution scope. The change of total N, P could only exert effect on some Deuteromycetes populations and had little effect on the amount and biodiversity of the integrity Deuteromycetes. The innovation point of this study lies in the fact that deuteromycete was chosen as marine environmental microorganism indicator to investigate a variety of polluted sites including Licun River estuary region (heavy polluted area), Haibo River estuary region(medium polluted area) with the Shilaoren beach bathe (clean area) as control. Molecular methods such as 18S-PCR denaturing gradient gel electrophoresis (18S PCR-DGGE) combined with traditional culture method were utilized to analysis the relevance between Deuteromycete population diversity and heavy metal pollution in three seasonsand the resultes strongly suggested that Deuteromycete could be used as a reliable marine pollution microorganism indicator. A relevance model between Deuteromycete population, Deuteromycete diversity and marine pollution was primarily designed on the basis of above obtained results and established foundation for further marine pollution monitoring.
The contents of N, P, heavy metal in Kiaochow bay coast of Licun river and Haibo river estuaries sediments collected in winter, spring and autumn were measured with Shilaoren as control. Statistical analysis of contamination status and biodiversity of Deuteromycetes in these habitats were conducted using traditional methods combined with modern 18S PCR-DGGE methods. The results showed Licun river estuaries was seriously polluted while Haibo river estuaries was moderate polluted and the amount of culturable Deuteromycetes decreased as the increase of pollution degree and this tendency was obviously relevant with heavy metal pollution degree. However, the tendency of Deuteromycetes biodiversity was different: within slight to moderate pollution scope, the biodiversity index of Deuteromycetes increased as the increase of heavy metal index while the biodiversity index of Deuteromycetes decreased sharply within seriously pollution scope. The change of total N, P could only exert effect on some Deuteromycetes populations and had little effect on the amount and biodiversity of the integrity Deuteromycetes. The innovation point of this study lies in the fact that deuteromycete was chosen as marine environmental microorganism indicator to investigate a variety of polluted sites including Licun River estuary region (heavy polluted area), Haibo River estuary region(medium polluted area) with the Shilaoren beach bathe (clean area) as control. Molecular methods such as 18S-PCR denaturing gradient gel electrophoresis (18S PCR-DGGE) combined with traditional culture method were utilized to analysis the relevance between Deuteromycete population diversity and heavy metal pollution in three seasonsand the resultes strongly suggested that Deuteromycete could be used as a reliable marine pollution microorganism indicator. A relevance model between Deuteromycete population, Deuteromycete diversity and marine pollution was primarily designed on the basis of above obtained results and established foundation for further marine pollution monitoring.
引文
[1]沈颖.海洋环境污染与环境保护[J] .环境科学进展,1997,5 (1) :67-75.
[2]王项南.海洋环境遥感监测[J].海洋技术,1996,15(4):18-24.
[3]李慧蓉,生物监测技术及其研究进展,江苏石油化工学院学报(2002) 02-0057-04.
[4]张志杰,张维平.环境污染生物监测与评价[M].北京:中国环境科学出版社, 1991,12-21
[5]王海黎,陶澍,生物标志物在水环境研究中的应用[J].中国环境科学,1999, 19(5):421 - 4241
[6]周元清,曾艳,周丽清等,生物监测在环境污染监测中的应用研究[J],云南环境科学,(2004)02-0057-03.
[7]孙铁珩,周启星,李培军.污染生态学[M].北京科学出版社,2000,123～1241
[8]常学秀,王焕校,文传浩.Cd2 +、Al3 +对蚕豆胚根根尖细胞遗传学毒性效应研究[J].农业环境保护, 1999,18(1):1～31
[9]赵贤四.大气颗粒有机提取对大鼠肝细胞DNA合成的影响[J].环境与健康杂志, 1996,13,(3);105～1071.
[10] Berg G, Opelt K, Schmidt S, Zachow C, Lottmann J, Gotz M,Costa R & Smalla K (2005a) The rhizosphere effect on bacteria antagonistic towards the pathogenic fungus Verticillium differs depending on plant species and site. FEMS Microbiol Ecol,in press.
[11] Rueff J, Chiapella C, et a1.Development and validation of alternative metabolic systems for mufagenicify testing in short - term assays , Mutation Research , 353 (1996) : 151～1761
[12] Stock J B, Stock A M, Mottonen J M, 1999, Signal transduction in bacteria1Nature, 344 (6265) : 395～3641
[13]周元清,曾艳,周丽清等,Fried J, Mayr G, Berger H.Monitoring protozoa and Metazoa bio-film communities for assessing wastewater quality impact and reactor up-scaling effect1Wat Sci Tech, 2000, 41(4 - 5):309～3161
[14] American Petroleum Institute. Bio-monitoring: Techniques for Measuring the Biological Effects of Liquid Effluents [M]. Washington,1987.
[15]沈韫芬,顾曼如,冯伟松.水污染的微型生物监测[J].生命科学, 1997,9(2): 81–821.
[16] Giller K E, Witter Ernst, Mcgrath S P.Toxicity of heavy metal to microorganisms and microbial process in agriculture soils: a review [J].Soil Biology & Biochemistry,1998,30 (10-11): 1389-1414.
[17] HIROKI M. Effects of heavy metal contamination on soil microbial population [J].Soil Sci Plant Nutr, 1992, 38:141-147.
[18] Delong E F. Diversity of naturally occurring prokaryotes[A]. Microbial Diversity in Time and Space [C]. New York: Plenum Press, 1996.
[19] LiYZ and Chen. The diversity of the marine microbiology resources[J].Advance in Biology Engineering,1998,18(4):34-40.李越中,陈琦.海洋微生物资源多样性[J].生物工程进展,1998,18(4):34-40.
[20] Amans R I, Luding E and Schleifer KH. Phylogenetic identifiction and in situ detection of individual microbial cellswithout cultivation[J].Microbial.,1995,66: 4547-4554.
[21] Woese C R and Fox G E. Phylogenetic structure of the prokaryotic domain, the primary kingdoms [J].Proceeding of the National Academy of Science,USA, 1977,74(11): 5088-5090.
[22] Tunlid A,White D C. Biochemical analysis of biomass,community structure,nutritional status,and metabolic activity of microbial community in soil, In: Stotzky G, Bollag J M, eds.Soil Bioehemistry.New York:Dekker,1991,229-262.
[23]曾润颖,赵晶.东太平洋沉积物深海嗜冷菌的分子鉴定分类[J].微生物学通报, 2002,29(6):12-16.
[24] Jain R K, Saylor G S, Wilson J T, et al. Maintenance and stability of introduced genotypes in ground water aquifer material[J].Applied and Environmental Microbiology,1987,53: 996-1002.
[25] Lobert B E, Rossello M R and Amann R. Microbial community composition of Wadden Sea sediment as revealed by fluorescence in situ hybridization [J].Applied and Environmental Microbiology, 1998,64(12): 2691-2696.
[26] Buckiey D H and Schmidt TM. The structure of microbial communities in soil and the lasting impact of cultivation [J].Microb Ecol., 2001,42: 11-21.
[27] Widmer F. A high selective PCR protocol for detecting 16SrRNA genes of the genus Pseudomonas(Sensu Stricto)in environmental sample[J].Applied and environmental Microbiology, 1998,64(7): 2345-2553.
[28] Sabine P, Stefanie K, Frank S,et al. Succession of microbial communities during hot composing as detected by PCR-single-strand-conformation polymorphism-based genetic profiles of small-subunit rRNA genes[J].Applied and Environmental Microbiology, 2000,66(3): 930-936.
[29] Beja O, Sazuki MT, Heideberg J E,et al. Unsuspected diversity among marine aerobic anoxygenic phototrophs [J].Nature,2002,415(6287): 630-633.
[30] Ritz K, Griffiths BS, TorsvikVL,et al. Analysisof soil land bacterio plankton community DNA by melting profiles and reassociation kinetics[J].FEMS Microbiology Letter, 1997,149: 151-156.
[31] Hosein S G,Millette D,Butler B J,et al. Catabolicgene probe analysis of an aquifer microbial community degrading corsatee related polycyclic aromatic and heterorcyclic compounds. Micorbiol Ecol,1997,34(2):81-89.
[32] Liu XL and Qian B C. Study of Japanese marine bioactive substance[J].Chinese Journal of Marine Drugs, 1997,16(1): 45-49.刘雪莉,钱伯初.日本海洋天然活性物质研究简况[J].中国海洋药物,1997,16(1): 45-49.
[33] Osawa R, et al. An investigation of aquatic bacteria capable of utilizing chitin asthe sole source of nutrients [J]. Lett. Appl. Microbiol., 1995,21(5):288-291.
[34] Jutta K, Oliver P, Martin H S,et al. Sulfate-reducing bacterial community response to carbon source amendments in contaminated aquifer microcosms[J].FEMS Microbiology Ecology, 2002,42: 109-118.
[35] Hayashi K,et al. Identification of the position of disulfide bonds of chitinase frommarine bacterium Alteromonssp. strain 0~7[J].Biosci. Biotechnol. Biochem., 1995,59(10): 1981-1982.
[36] Yazawa K. Production of eicosapentaenoic acid from marine bacteria[J].Lipids., 1996,31(1): 297-300.
[37] Delong E F,et al. Biochemical function and ecological significance of novel bacteria lipids in deep sea prokaryotes[J].Trends Biotechnol., 1997,15:203-207.
[38] Leuschner C, Antranikian G and World G. Xylanolytic enzymes from fungi and bacteria[J].Microbiol. Biotechnol., 1995,11: 95-114.
[39] Zheng T L, Zhuang T C, Cai L Z, et al. The role of microbes in bioremediation of marine polluted environment[J].Journal of Xia men University (Natural Science Edition), 2001,40(2): 524-534.郑天凌,庄铁诚,蔡立哲,等.微生物在海洋污染环境中的生物修复作用[J].厦门大学学报(自然科学版), 2001,40(2): 524-534.
[40] Zhang J Y, Li J, Xia W L, et al. Advances in bioremediation researches [J].Chin. J. Appl. Environ. Biol., 1996,2(2): 193-199.张甲耀,李静,夏威林,等.生物修复研究进展[J].应用与环境生物学报, 1996,2(2): 193.
[41] Bruggen AHC and Semenov A M. Research of biological indicators for soil health and disease suppression [J]. Appl. Soil. Ecol., 2000,15 (1): 13-24.
[42] Ravenschlag K, Sahm K, Pernthaler J,et al. High bacterial diversity in permanently cold marine sediments[J]. Applied andEnvironmentalMicrobiology,1999,65(9): 3982-3989.
[43] Glovanoni S J, Britschgi T B, Moyer C L,et al. Genetic diversity in Sargasso Sea bacterial plankton [J].Nature. 1990,45: 60-63.
[44] Dai X, Zhou H, Cai C H,et al. A preliminary study on the novel taxa of bacteria in marine sediments [J].Acta Scientiarum Naturalium Universitatis Sunyatseni., 2001,40(6):51-54.戴欣,周惠,蔡创华,等.海洋沉积物中特有细菌类群的初步探讨[J].中山大学学报(自然科学版), 2001,40(6): 51-54.
[45]海洋监测规GB17378.1-2007
[46]李学刚,宋金明,牛丽凤等,近海沉积物中氮磷的同时测定及其在胶州湾沉积物中的应用,岩矿测试,2007,02—0087—06.
[47]海洋沉积物质量标准GB 18668-2002。
[48]金岚,王振堂,朱秀丽,等.环境生态学.北京:高等教育出版社[M],2001:108-109.
[49]汪亚平,高抒,贾建军.胶州湾及邻近海域沉积物分布特征和运移趋势[J].地理学报,2000, 55 ( 4) :449 - 458.
[50]殷效彩,杨永亮,余季金,等.胶州湾表层沉积物重金属分布研究[J].青岛大学报,2001 ,14(1) :76- 80.
[51] Chun mei Xue, Tian Li, Zhi wei Deng et al. Janthinolide A-B,two new 2,5-piperrazinedione derivatives from the endophytic Penicillium janthinellum isolated from the soft coral Dendronephthya sp. 2006, Pharmazie, 61:1041-1044
[52] ZHAO Xiangwei , LUO Yongming , TENG Ying, et a1.Genetic diversity of microbial communities in farmland soils contaminated with mixed heavy metals [J].Aeta Scientiae Circumstantiae , 2005 ,25
[53] Sigler W V,Zeyer J.Microbial diversity and activity along the fore一fields of two receding glaciers[J].Microbial Ecology, 2002, 43(4) : 397—407
[54] Hopkins W A,Roe JH,Snodgrass,J W Nondestructive indices of trace elenlent exposure in saquanlata reptiles Environ Poll,2001,115:1～77
[55]杨济龙,祖艳群,洪常青,等,蔬菜沉积物微生物种群数量与沉积物重金属含量的关系[M]生态环境,2003.
[56] American Petroleum Institute . Biomonitoring : Techniques for Measuring the Biological Effects of Liquid Effluents [M] . Washington ,1987 .
[57] Wizard? Plus Minipreps DNA Purification System INSTRUCTIONS FOR USE OF PRODUCTS A7100, A7500, A7510, A7141 AND A7211.
[58]张瑞福,曹慧,崔中利等,土壤微生物总DNA的提取和纯化.微生物报, 2003,1006-6179.
[59] Instruction Manual of UltraCleanTM Soil DNA Isolation Kit.Catalog# 12800-50 50preps.
[60] Kowalchuk G A 1999 New perspectives towards analysing fungal communities in terrestrial environments. Current Opinion in Biotechnology 10, 247–251.
[61] Bosshard PP, Santini Y, Gruter D, Stettler R, Bachofen R (2000).Bacterial diversity and community composition in the chemocline Of the meromictic alpine Lake Cadagno as revealed by 16S rDNA analysis. FEMS Microbiol Ecol 31:173–182
[62]杨红,李颖,关国华,等.尖孢镰刀菌异核体及其不同核型分离子rDNA ITS区序列分析.农业生物技术学报, 2002, 10(4): 318-384.
[63] White, T.J., Bruns, T., Lee, S., and Taylor, J. 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In PCR protocols: a guide to methods and applications. Edited by M.A. Innis, D.H. Gelfand, J.J. Sninsky, and T.J. White. Academic Press, Inc., San Diego, Calif. pp. 315–324.
[64] Vainio EJI&Hantula I(2000) Direct analysis of wood-inhabiting fungi using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Mycol Res 19:6823-6831.
[65] Kjellerup B V,Veeh R H,Sumithraratne P,et a1.Monitoring of microbial souring in chemi c~ y treated,produced—water biofilm systems using molecular techniques[J].Journal of Industrial Microbiology and Biotechnology,2005,32(4):163—170.
[66] Lisa A. May, Brenda Smiley, and Michael G. Schmidt. Comparative denaturing gradient gel electrophoresis analysis of fungal communities associated with whole plant corn silage[J]Can. J. Microbiol. 2001,47: 829–841
[67] Zhou JZ, BrunsMA, Tiedje AM. 1996. DNA recovery from soils of diverse composition. Appl Environ M icrobiol, 62: 316～322
[68] HinoueM, Fukuda K, Wan Y, et al. 2004. An effective method for extracting DNA from contaminated soil due to industrialwaste. JUoeh, 26: 13～21
[69] Kreader CA. 1996. Relief of amp lification inhibition in PCR with bovine serum albumin or T4 gene 32 protein. Appl Environ Microbiol, 62: 1102～1106
[70]李钧敏,金则新.一种高效可直接用于PCR分析的土壤总微生物DNA抽提方法[J]应用生态学报,2006,17 (11) : 2107～2111
[2]王项南.海洋环境遥感监测[J].海洋技术,1996,15(4):18-24.
[3]李慧蓉,生物监测技术及其研究进展,江苏石油化工学院学报(2002) 02-0057-04.
[4]张志杰,张维平.环境污染生物监测与评价[M].北京:中国环境科学出版社, 1991,12-21
[5]王海黎,陶澍,生物标志物在水环境研究中的应用[J].中国环境科学,1999, 19(5):421 - 4241
[6]周元清,曾艳,周丽清等,生物监测在环境污染监测中的应用研究[J],云南环境科学,(2004)02-0057-03.
[7]孙铁珩,周启星,李培军.污染生态学[M].北京科学出版社,2000,123～1241
[8]常学秀,王焕校,文传浩.Cd2 +、Al3 +对蚕豆胚根根尖细胞遗传学毒性效应研究[J].农业环境保护, 1999,18(1):1～31
[9]赵贤四.大气颗粒有机提取对大鼠肝细胞DNA合成的影响[J].环境与健康杂志, 1996,13,(3);105～1071.
[10] Berg G, Opelt K, Schmidt S, Zachow C, Lottmann J, Gotz M,Costa R & Smalla K (2005a) The rhizosphere effect on bacteria antagonistic towards the pathogenic fungus Verticillium differs depending on plant species and site. FEMS Microbiol Ecol,in press.
[11] Rueff J, Chiapella C, et a1.Development and validation of alternative metabolic systems for mufagenicify testing in short - term assays , Mutation Research , 353 (1996) : 151～1761
[12] Stock J B, Stock A M, Mottonen J M, 1999, Signal transduction in bacteria1Nature, 344 (6265) : 395～3641
[13]周元清,曾艳,周丽清等,Fried J, Mayr G, Berger H.Monitoring protozoa and Metazoa bio-film communities for assessing wastewater quality impact and reactor up-scaling effect1Wat Sci Tech, 2000, 41(4 - 5):309～3161
[14] American Petroleum Institute. Bio-monitoring: Techniques for Measuring the Biological Effects of Liquid Effluents [M]. Washington,1987.
[15]沈韫芬,顾曼如,冯伟松.水污染的微型生物监测[J].生命科学, 1997,9(2): 81–821.
[16] Giller K E, Witter Ernst, Mcgrath S P.Toxicity of heavy metal to microorganisms and microbial process in agriculture soils: a review [J].Soil Biology & Biochemistry,1998,30 (10-11): 1389-1414.
[17] HIROKI M. Effects of heavy metal contamination on soil microbial population [J].Soil Sci Plant Nutr, 1992, 38:141-147.
[18] Delong E F. Diversity of naturally occurring prokaryotes[A]. Microbial Diversity in Time and Space [C]. New York: Plenum Press, 1996.
[19] LiYZ and Chen. The diversity of the marine microbiology resources[J].Advance in Biology Engineering,1998,18(4):34-40.李越中,陈琦.海洋微生物资源多样性[J].生物工程进展,1998,18(4):34-40.
[20] Amans R I, Luding E and Schleifer KH. Phylogenetic identifiction and in situ detection of individual microbial cellswithout cultivation[J].Microbial.,1995,66: 4547-4554.
[21] Woese C R and Fox G E. Phylogenetic structure of the prokaryotic domain, the primary kingdoms [J].Proceeding of the National Academy of Science,USA, 1977,74(11): 5088-5090.
[22] Tunlid A,White D C. Biochemical analysis of biomass,community structure,nutritional status,and metabolic activity of microbial community in soil, In: Stotzky G, Bollag J M, eds.Soil Bioehemistry.New York:Dekker,1991,229-262.
[23]曾润颖,赵晶.东太平洋沉积物深海嗜冷菌的分子鉴定分类[J].微生物学通报, 2002,29(6):12-16.
[24] Jain R K, Saylor G S, Wilson J T, et al. Maintenance and stability of introduced genotypes in ground water aquifer material[J].Applied and Environmental Microbiology,1987,53: 996-1002.
[25] Lobert B E, Rossello M R and Amann R. Microbial community composition of Wadden Sea sediment as revealed by fluorescence in situ hybridization [J].Applied and Environmental Microbiology, 1998,64(12): 2691-2696.
[26] Buckiey D H and Schmidt TM. The structure of microbial communities in soil and the lasting impact of cultivation [J].Microb Ecol., 2001,42: 11-21.
[27] Widmer F. A high selective PCR protocol for detecting 16SrRNA genes of the genus Pseudomonas(Sensu Stricto)in environmental sample[J].Applied and environmental Microbiology, 1998,64(7): 2345-2553.
[28] Sabine P, Stefanie K, Frank S,et al. Succession of microbial communities during hot composing as detected by PCR-single-strand-conformation polymorphism-based genetic profiles of small-subunit rRNA genes[J].Applied and Environmental Microbiology, 2000,66(3): 930-936.
[29] Beja O, Sazuki MT, Heideberg J E,et al. Unsuspected diversity among marine aerobic anoxygenic phototrophs [J].Nature,2002,415(6287): 630-633.
[30] Ritz K, Griffiths BS, TorsvikVL,et al. Analysisof soil land bacterio plankton community DNA by melting profiles and reassociation kinetics[J].FEMS Microbiology Letter, 1997,149: 151-156.
[31] Hosein S G,Millette D,Butler B J,et al. Catabolicgene probe analysis of an aquifer microbial community degrading corsatee related polycyclic aromatic and heterorcyclic compounds. Micorbiol Ecol,1997,34(2):81-89.
[32] Liu XL and Qian B C. Study of Japanese marine bioactive substance[J].Chinese Journal of Marine Drugs, 1997,16(1): 45-49.刘雪莉,钱伯初.日本海洋天然活性物质研究简况[J].中国海洋药物,1997,16(1): 45-49.
[33] Osawa R, et al. An investigation of aquatic bacteria capable of utilizing chitin asthe sole source of nutrients [J]. Lett. Appl. Microbiol., 1995,21(5):288-291.
[34] Jutta K, Oliver P, Martin H S,et al. Sulfate-reducing bacterial community response to carbon source amendments in contaminated aquifer microcosms[J].FEMS Microbiology Ecology, 2002,42: 109-118.
[35] Hayashi K,et al. Identification of the position of disulfide bonds of chitinase frommarine bacterium Alteromonssp. strain 0~7[J].Biosci. Biotechnol. Biochem., 1995,59(10): 1981-1982.
[36] Yazawa K. Production of eicosapentaenoic acid from marine bacteria[J].Lipids., 1996,31(1): 297-300.
[37] Delong E F,et al. Biochemical function and ecological significance of novel bacteria lipids in deep sea prokaryotes[J].Trends Biotechnol., 1997,15:203-207.
[38] Leuschner C, Antranikian G and World G. Xylanolytic enzymes from fungi and bacteria[J].Microbiol. Biotechnol., 1995,11: 95-114.
[39] Zheng T L, Zhuang T C, Cai L Z, et al. The role of microbes in bioremediation of marine polluted environment[J].Journal of Xia men University (Natural Science Edition), 2001,40(2): 524-534.郑天凌,庄铁诚,蔡立哲,等.微生物在海洋污染环境中的生物修复作用[J].厦门大学学报(自然科学版), 2001,40(2): 524-534.
[40] Zhang J Y, Li J, Xia W L, et al. Advances in bioremediation researches [J].Chin. J. Appl. Environ. Biol., 1996,2(2): 193-199.张甲耀,李静,夏威林,等.生物修复研究进展[J].应用与环境生物学报, 1996,2(2): 193.
[41] Bruggen AHC and Semenov A M. Research of biological indicators for soil health and disease suppression [J]. Appl. Soil. Ecol., 2000,15 (1): 13-24.
[42] Ravenschlag K, Sahm K, Pernthaler J,et al. High bacterial diversity in permanently cold marine sediments[J]. Applied andEnvironmentalMicrobiology,1999,65(9): 3982-3989.
[43] Glovanoni S J, Britschgi T B, Moyer C L,et al. Genetic diversity in Sargasso Sea bacterial plankton [J].Nature. 1990,45: 60-63.
[44] Dai X, Zhou H, Cai C H,et al. A preliminary study on the novel taxa of bacteria in marine sediments [J].Acta Scientiarum Naturalium Universitatis Sunyatseni., 2001,40(6):51-54.戴欣,周惠,蔡创华,等.海洋沉积物中特有细菌类群的初步探讨[J].中山大学学报(自然科学版), 2001,40(6): 51-54.
[45]海洋监测规GB17378.1-2007
[46]李学刚,宋金明,牛丽凤等,近海沉积物中氮磷的同时测定及其在胶州湾沉积物中的应用,岩矿测试,2007,02—0087—06.
[47]海洋沉积物质量标准GB 18668-2002。
[48]金岚,王振堂,朱秀丽,等.环境生态学.北京:高等教育出版社[M],2001:108-109.
[49]汪亚平,高抒,贾建军.胶州湾及邻近海域沉积物分布特征和运移趋势[J].地理学报,2000, 55 ( 4) :449 - 458.
[50]殷效彩,杨永亮,余季金,等.胶州湾表层沉积物重金属分布研究[J].青岛大学报,2001 ,14(1) :76- 80.
[51] Chun mei Xue, Tian Li, Zhi wei Deng et al. Janthinolide A-B,two new 2,5-piperrazinedione derivatives from the endophytic Penicillium janthinellum isolated from the soft coral Dendronephthya sp. 2006, Pharmazie, 61:1041-1044
[52] ZHAO Xiangwei , LUO Yongming , TENG Ying, et a1.Genetic diversity of microbial communities in farmland soils contaminated with mixed heavy metals [J].Aeta Scientiae Circumstantiae , 2005 ,25
[53] Sigler W V,Zeyer J.Microbial diversity and activity along the fore一fields of two receding glaciers[J].Microbial Ecology, 2002, 43(4) : 397—407
[54] Hopkins W A,Roe JH,Snodgrass,J W Nondestructive indices of trace elenlent exposure in saquanlata reptiles Environ Poll,2001,115:1～77
[55]杨济龙,祖艳群,洪常青,等,蔬菜沉积物微生物种群数量与沉积物重金属含量的关系[M]生态环境,2003.
[56] American Petroleum Institute . Biomonitoring : Techniques for Measuring the Biological Effects of Liquid Effluents [M] . Washington ,1987 .
[57] Wizard? Plus Minipreps DNA Purification System INSTRUCTIONS FOR USE OF PRODUCTS A7100, A7500, A7510, A7141 AND A7211.
[58]张瑞福,曹慧,崔中利等,土壤微生物总DNA的提取和纯化.微生物报, 2003,1006-6179.
[59] Instruction Manual of UltraCleanTM Soil DNA Isolation Kit.Catalog# 12800-50 50preps.
[60] Kowalchuk G A 1999 New perspectives towards analysing fungal communities in terrestrial environments. Current Opinion in Biotechnology 10, 247–251.
[61] Bosshard PP, Santini Y, Gruter D, Stettler R, Bachofen R (2000).Bacterial diversity and community composition in the chemocline Of the meromictic alpine Lake Cadagno as revealed by 16S rDNA analysis. FEMS Microbiol Ecol 31:173–182
[62]杨红,李颖,关国华,等.尖孢镰刀菌异核体及其不同核型分离子rDNA ITS区序列分析.农业生物技术学报, 2002, 10(4): 318-384.
[63] White, T.J., Bruns, T., Lee, S., and Taylor, J. 1990. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In PCR protocols: a guide to methods and applications. Edited by M.A. Innis, D.H. Gelfand, J.J. Sninsky, and T.J. White. Academic Press, Inc., San Diego, Calif. pp. 315–324.
[64] Vainio EJI&Hantula I(2000) Direct analysis of wood-inhabiting fungi using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Mycol Res 19:6823-6831.
[65] Kjellerup B V,Veeh R H,Sumithraratne P,et a1.Monitoring of microbial souring in chemi c~ y treated,produced—water biofilm systems using molecular techniques[J].Journal of Industrial Microbiology and Biotechnology,2005,32(4):163—170.
[66] Lisa A. May, Brenda Smiley, and Michael G. Schmidt. Comparative denaturing gradient gel electrophoresis analysis of fungal communities associated with whole plant corn silage[J]Can. J. Microbiol. 2001,47: 829–841
[67] Zhou JZ, BrunsMA, Tiedje AM. 1996. DNA recovery from soils of diverse composition. Appl Environ M icrobiol, 62: 316～322
[68] HinoueM, Fukuda K, Wan Y, et al. 2004. An effective method for extracting DNA from contaminated soil due to industrialwaste. JUoeh, 26: 13～21
[69] Kreader CA. 1996. Relief of amp lification inhibition in PCR with bovine serum albumin or T4 gene 32 protein. Appl Environ Microbiol, 62: 1102～1106
[70]李钧敏,金则新.一种高效可直接用于PCR分析的土壤总微生物DNA抽提方法[J]应用生态学报,2006,17 (11) : 2107～2111