3株红树林土壤来源菲降解菌的鉴定及其降解特性
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摘要
根据液体发酵浑浊程度判断,共分离出能利用菲作为碳源生长的菌种共9株,其中3株具有较强生长活性和降解活力,通过菌体菌落形态学观察、生理生化及16S rRNA基因序列系统发育分析,确定菌株UMBR 0019、UMBR 0020和UMBR 0197分别为Micromonospora palomenae NEAU-CX1T(相似性为99.64%)、Rhodococcus ruber DSM 43338T(相似性为100%)和Novosphingobium mathurense SM117T(相似性为99.24%)。研究结果表明,在以菲为唯一碳源的海水培养基中, 30℃,180 r/min培养条件下,UMBR 0019,UMBR 0020和UMBR 0197对菲的降解效率分别为48.80%、75%和63.54%,而通过菌株协同作用后能显著提高菲的降解效率,达到99.07%。红树林蕴含丰富的多环芳烃降解微生物资源,能够为海洋环境污染生物修复提供重要途径。
According to the muddy of fermentation liquid, 9 stains were found with phenanthrene as their sources of carbon,in which 3 stains grow rapidly and highly efficient in degrading bacteria. Through morphological observation, physiological and chemotaxonomic characteristics and 16S rRNA gene phylogenetic tree analysis, UMBR 0019, UMBR 0020 and UMBR 0197 were respectively identified as Micromonospora palomenae NEAU-CX1T(pairwise similarity 99.64%), Rhodococcus ruber DSM 43338T(pairwise similarity 100%) and Novosphingobium mathurense SM117T(pairwise similarity 99.24%). Degration percentage of phenanthrene by strain UMBR 0019, UMBR 0020 and UMBR 0197 were 48.80%, 75% and 63.54% at 30 ℃(180 r/min), while phenanthrene degradation efficiency was increased to 99.07% through synergistic effect in co-culture fermentation. There were abundant and diverse PAHs-degrading microbial resources in the mangrove sediments, which would be a main approach for bioremediation of PAHs-contaminated environment.
According to the muddy of fermentation liquid, 9 stains were found with phenanthrene as their sources of carbon,in which 3 stains grow rapidly and highly efficient in degrading bacteria. Through morphological observation, physiological and chemotaxonomic characteristics and 16S rRNA gene phylogenetic tree analysis, UMBR 0019, UMBR 0020 and UMBR 0197 were respectively identified as Micromonospora palomenae NEAU-CX1T(pairwise similarity 99.64%), Rhodococcus ruber DSM 43338T(pairwise similarity 100%) and Novosphingobium mathurense SM117T(pairwise similarity 99.24%). Degration percentage of phenanthrene by strain UMBR 0019, UMBR 0020 and UMBR 0197 were 48.80%, 75% and 63.54% at 30 ℃(180 r/min), while phenanthrene degradation efficiency was increased to 99.07% through synergistic effect in co-culture fermentation. There were abundant and diverse PAHs-degrading microbial resources in the mangrove sediments, which would be a main approach for bioremediation of PAHs-contaminated environment.
引文
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[2]Wilcke,Wolfgang.SYNOPSIS Polycyclic Aromatic Hydrocarbons(PAHs)in soil:a review[J].Journal of Plant Nutrition and Soil Science,2015,163(3):229-248.
[3]杨跃志,张海丽,李正炎,等.北部湾沿岸表层沉积物中酚类内分泌干扰物与多环芳烃的污染特征及生态风险评价[J].中国海洋大学学报:自然科学版,2013,43(6):87-92.Yang Yuezhi,Zhang Haili,Li Zhengyan,et al.Pollution characteristics and ecological risk assessment of phenolic endocrine disruptors and polycyclic aromatic hydrocarbons in surface sediments of Beibu Gulf Coast[J].The Journal of China Ocean University:Natural Science Edition,2013,43(6):87-92.
[4]刘慧杰.红树林湿地微生物对典型有机物污染的响应及其在生物修复中的作用研究[D].厦门:厦门大学,2008.Liu Huijie.Response of Microorganisms in Mangrove Wetlands to Typical Organic Pollutants and Their Role in Bioremediation[D]Xiamen:Xiamen University,2008.
[5]Chen J L,Wong Y S,Tam N F.Static and dynamic sorption of phenanthrene in mangrove sediment slurry[J].Journal of Hazardous Materials,2009,168(2/3):1422-1429.
[6]Mostafa A R,Wade T L,Sweet S T,et al.Distribution and characteristics of polycyclic aromatic hydrocarbons(PAHs)in sediments of Hadhramout coastal area,Gulf of Aden,Yemen[J].Journal of Marine Systems,2009,78(1):1-8.
[7]周双清,黄小龙,黄东益,等.Chelex-100快速提取放线菌DNA作为PCR扩增模板[J].生物技术通报,2010(2):123-125.Zhou Shuangqing,Huang Xiaolong,Huang Dongyi,et al.Chelex-100 rapid extraction of actinomycete DNA as a PCRamplification template[J].Biotechnology Bulletin,2010(2):123-125.
[8]Kim OS,Cho YJ,Lee K,et al.Introducing EzTaxon-e:a prokaryotic 16S rRNA gene sequence database with phylotypes that represent uncultured species[J].Int J Syst Evol Microbiol,2012,62(Pt 3):716-721.
[9]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&Evolution,2011,28(10):2731.
[10]Joseph Felsenstein.Confidence limits on phylogenies:an approach using the bootstrap[J].Evolution,1985,39(4):783-791.
[11]Ivshina I,Kostina L,Krivoruchko A,et al.Removal of polycyclic aromatic hydrocarbons in soil spiked with model mixtures of petroleum hydrocarbons and heterocycles using biosurfactants from Rhodococcus ruber IEGM 231[J].J Hazard Mater,2016:8-17.
[12]Song X,Xu Y,Li G,et al.Isolation,characterization of Rhodococcus sp.P14 capable of degrading high-molecularweight polycyclic aromatic hydrocarbons and aliphatic hydrocarbons[J].Marine Pollution Bulletin,2011,62(10):2122-2128.
[13]Li C,Zhang C,Song G,et al.Characterization of a protocatechuate catabolic gene cluster in Rhodococcus ruber OA1involved in naphthalene degradation[J].Annals of Microbiology,2015,66(1):469-478.
[14]Leigh MB,Prouzova P,Mackova M,et al.Polychlorinated biphenyl(PCB)-degrading bacteria associated with trees in a PCB-contaminated site[J].Appl Environ Microbiol,2006,72(4):2331-2342.
[15]Muangchinda C,Yamazoe A,Polrit D,et al.Biodegradation of high concentrations of mixed polycyclic aromatic hydrocarbons by indigenous bacteria from a river sediment:a microcosm study and bacterial community analysis[J].Environ Sci Pollut Res Int,2017,24(5):1-12.
[16]Kuyukina MS,Ivshina IB,Korshunova IO,et al.Diverse effects of a biosurfactant from Rhodococcus ruber IEGM 231on the adhesion of resting and growing bacteria to polystyrene[J].AMB Express,2016,6(1):14.
[17]Gupta SK,Lal D,Lal R.Novosphingobium panipatense sp.nov.and Novosphingobium mathurense sp.nov.from oilcontaminated soil[J].International Journal of Systematic&Evolutionary Microbiology,2009,59(1):156-161.
[18]Niharika N,Moskalikova H,Kaur J,et al.Novosphingobium barchaimii sp.nov.isolated from hexachlorocyclohexanecontaminated soil[J].International Journal of Systematic&Evolutionary Microbiology,2013,63(2):667-672.
[19]Anjali Saxena,Shailly Anand,Ankita Dua,et al.Novosphingobium lindaniclasticum sp.nov.a hexachlorocyclohexane(HCH)-degrading bacterium isolated from an HCH dumpsite[J].International Journal of Systematic and Evolutionary Microbiology,2009,63(Pt 12):3140-3144.
[20]Gan HM,Hudson AO,Rahman AY A,et al.Comparative genomic analysis of six bacteria belonging to the genus Novosphingobium:insights into marine adaptation,cell-cell signaling and bioremediation[J].BMC Genomics,2013,14(1):431.
[21]Margaret F Romine,Lisa C Stillwell,KwongKwok Wong,et al.Complete sequence of a 184-kilobase catabolic plasmid from Sphingomonas aromaticivorans F199[J].Journal of Bacteriology,1999,181(5):1585-1602.
[22]Idemudia,Nosagie O A,Omorede O.Comparative assessment of degradation potentials of bacteria and actinomycetes in soil contaminated with motorcycle spent oil[J].Asian Journal of Science and Technology,2014,5(8):482-487.
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[28]Machín Ramírez C,Morales D,Martínez Morales F,et al.Benzo[a]pyrene removal by axenic-and co-cultures of some bacterial and fungal strains[J].International Biodeterioration&Biodegradation,2010,64(7):538-544.