可降解吡啶的全食副球菌B21-3的筛选鉴定及降解特性
|
摘要
【背景】吡啶作为一种难降解的有机污染物普遍存在于焦化、炼油、皮革和制药等行业的废水中,并对环境造成危害。【目的】治理废水中残留的有机污染物吡啶,筛选高效降解菌。【方法】采用富集培养和选择培养,以石家庄某污水处理厂的活性污泥为材料进行吡啶降解菌的筛选,通过形态特征、生理生化特性、(G+C)mol%测定及16S rRNA基因序列系统发育分析对筛选到的降解菌进行鉴定,并分析其对吡啶的降解特性。【结果】分离筛选到一株能以吡啶为唯一碳源和氮源生长代谢的降解菌B21-3,经鉴定该菌株为全食副球菌(Paracoccuspantotrophus)。菌株B21-3对吡啶的最适降解温度为32°C,最适降解pH为7.0,吡啶浓度为100mg/L时降解率为48.50%±0.02%;通过逐步提高吡啶初始浓度对菌株进行驯化,驯化后菌株可耐受较高浓度吡啶且吡啶降解率显著增加,吡啶浓度为100 mg/L时驯化后菌株B21-3对吡啶的降解率为90.26%±1.70%。驯化后菌株在含吡啶的无机盐平板上传代培养15代后,对吡啶的降解率为89.39%±2.03%。【结论】菌株B21-3具有较强的吡啶降解能力及降解稳定性,该菌株可作为吡啶污染水体生物修复的潜在资源。
[Background] As a refractory organic pollutant, pyridine is commonly found in wastewater from coking, refining, leather and pharmaceutical industries, causing harm to the environment.[Objective] To treat pyridine contaminated wastewater, bacteria capable of degrading pyridine were screened. [Methods] Pyridine degrading bacteria were isolated from the activated sludge of a wastewater plant in Shijiazhuang by enrichment and selective medium. Bacterium B21-3 was identified by morphological, physiological and biochemical characteristics,(G+C)mol% assay, and 16 S rRNA gene phylogenic analysis. The pyridine degradation characteristics were analyzed. [Results] A bacterial strain B21-3 that used pyridine as the sole carbon and energy source was isolated and identified as Paracoccus pantotrophus. The optimal pH and temperature for pyridine degradation were 7.0 and 32 °C, respectively.When the initial concentration of pyridine was 100 mg/L, the degradation percentage of pyridine was48.50%±0.02%. After acclimation by increasing the initial concentration of pyridine, strain B21-3 tolerated higher concentrations of pyridine and the degradation of pyridine increased significantly. Under the pyridine concentration of 100 mg/L, the degradation percentage of pyridine by domesticated strain B21-3 was 90.26%±1.70%. After the acclimatized strain B21-3 was subcultured on mineral salt plates supplemented with pyridine for 15 generations, the degradation percentage of pyridine was89.39%±2.03%. [Conclusion] Strain B21-3 had strong pyridine degradation ability with potential for bioremediation of pyridine contaminated wastewater.
[Background] As a refractory organic pollutant, pyridine is commonly found in wastewater from coking, refining, leather and pharmaceutical industries, causing harm to the environment.[Objective] To treat pyridine contaminated wastewater, bacteria capable of degrading pyridine were screened. [Methods] Pyridine degrading bacteria were isolated from the activated sludge of a wastewater plant in Shijiazhuang by enrichment and selective medium. Bacterium B21-3 was identified by morphological, physiological and biochemical characteristics,(G+C)mol% assay, and 16 S rRNA gene phylogenic analysis. The pyridine degradation characteristics were analyzed. [Results] A bacterial strain B21-3 that used pyridine as the sole carbon and energy source was isolated and identified as Paracoccus pantotrophus. The optimal pH and temperature for pyridine degradation were 7.0 and 32 °C, respectively.When the initial concentration of pyridine was 100 mg/L, the degradation percentage of pyridine was48.50%±0.02%. After acclimation by increasing the initial concentration of pyridine, strain B21-3 tolerated higher concentrations of pyridine and the degradation of pyridine increased significantly. Under the pyridine concentration of 100 mg/L, the degradation percentage of pyridine by domesticated strain B21-3 was 90.26%±1.70%. After the acclimatized strain B21-3 was subcultured on mineral salt plates supplemented with pyridine for 15 generations, the degradation percentage of pyridine was89.39%±2.03%. [Conclusion] Strain B21-3 had strong pyridine degradation ability with potential for bioremediation of pyridine contaminated wastewater.
引文
[1]Zhao QX,Lin Z,Zhang T.Strategies for treatment of bio-refractory organic pollutants[J].Environmental Protection of Chemical Industry,1995,15(5):276-279(in Chinese)赵庆祥,林喆,张彤.生物难降解有机污染物的治理对策[J].化工环保,1995,15(5):276-279
[2]Zhuo M,He CD,Liu WH.Purification of refractory pyridine waste gas by anoxic denitrification[J].Chinese Journal of Environmental Engineering,2017,11(12):6345-6350(in Chinese)卓猛,何成达,刘伟慧.利用反硝化法净化难降解吡啶废气[J].环境工程学报,2017,11(12):6345-6350
[3]Jin TT,Ren JH,Zhang H,et al.Identification and characterization of a pyridine degrading bacterium[J].Ecology and Environmental Sciences,2016,25(7):1217-1224(in Chinese)晋婷婷,任嘉红,张晖,等.一株吡啶高效降解菌的鉴定及其降解特性[J].生态环境学报,2016,25(7):1217-1224
[4]Yi X,Zhong J.Biodegradation of pyridine and quinoline by two Pseudomonas strains[J].Acta Microbiologica Sinica,2011,51(8):1087-1097
[5]Liu JJ,Chen LJ,Xiong RL,et al.Physiological and biochemical characteristics of a pyridine-degrading bacterial strain[J].Acta Scientiarum Naturalium Universitatis Pekinensis,2008,44(6):983-986(in Chinese)刘江江,陈吕军,熊瑞林,等.一株吡啶降解菌的生理生化特性研究[J].北京大学学报:自然科学版,2008,44(6):983-986
[6]Chen P,Yan JB,Yu YD.Screening of a pyridine-degrading strain and its biodegradation capability[J].Environmental Protection of Chemical Industry,2015,35(6):566-570(in Chinese)陈佩,颜家保,余永登.一株吡啶降解菌的筛选及其降解性能[J].化工环保,2015,35(6):566-570
[7]Sun JQ,Xu L,Tang YQ,et al.Degradation of pyridine by one Rhodococcus strain in the presence of chromium(VI)or phenol[J].Journal of Hazardous Materials,2011,191(1/3):62-68
[8]Padoley KV,Mudliar SN,Pandey RA.Microbial degradation of pyridine andα-picoline using a strain of the genera Pseudomonas and Nocardia sp.[J].Bioprocess and Biosystems Engineering,2009,32(4):501-510
[9]Deng XQ,Wei CH,Ren Y,et al.Isolation and identification of Achromobacter sp.DN-06 and evaluation of its pyridine degradation kinetics[J].Water,Air,&Soil Pollution,2011,221(1/4):365-375
[10]Shen JY,Zhang X,Chen D,et al.Characteristics of pyridine biodegradation by a novel bacterial strain,Rhizobium sp.NJUST18[J].Desalination and Water Treatment,2015,53(7):2005-2013
[11]Zefirov NS,Agapova SR,Terentiev PB,et al.Degradation of pyridine by Arthrobacter crystallopoietes and Rhodococcus opacus strains[J].FEMS Microbiology Letters,1994,118(1/2):71-74
[12]Sun JQ,Xu L,Tang YQ,et al.Degradation characteristics of pyridine and phenol by Rhodococcus sp.Chr-9[J].Chinese Journal of Applied&Environmental Biology,2012,18(4):647-650(in Chinese)孙纪全,徐莲,汤岳琴,等.红球菌Chr-9降解吡啶和苯酚的特性[J].应用与环境生物学报,2012,18(4):647-650
[13]Mohan SV,Sistla S,Guru RK,et al.Microbial degradation of pyridine using Pseudomonas sp.and isolation of plasmid responsible for degradation[J].Waste Management,2003,23(2):167-171
[14]Ma YF,Wang L,Shao ZZ.Pseudomonas,the dominant polycyclic aromatic hydrocarbon-degrading bacteria isolated from Antarctic soils and the role of large plasmids in horizontal gene transfer[J].Envrionmental Microbiology,2006,8(3):455-465
[15]Bai YH.Microbial degradation of pyridine and quinolne and bioaugmentation for their removal:characteristics and mechanisms[D].Beijing:Doctoral Dissertation of Peking University,2009(in Chinese)柏耀辉.吡啶、喹啉微生物降解及生物强化去除的特性与机理[D].北京:北京大学博士学位论文,2009
[16]Li YX,Zhao CH.Determination of main pollutants in coking wastewater by ultraviolet spectrophotometry[J].China Water&Wastewater,2001,17(1):54-56(in Chinese)李亚新,赵晨红.紫外分光光度法测定焦化废水的主要污染物[J].中国给水排水,2001,17(1):54-56
[17]Edwards U,Rogall T,Bl?cker H,et al.Isolation and direct complete nucleotide determination of entire genes.Characterization of a gene coding for 16S ribosomal RNA[J].Nucleic Acids Research,1989,17(19):7843-7853
[18]Lane DJ.16S/23S r RNA sequencing[A]//Stackebrandt E,Goodfellow M.Nucleic Acid Techniques in bacterial Systematics[M].New York:John Wiley&Sons,1991:115-175
[19]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
[20]Dong XZ,Cai MY.Common Bacteria System Classification and Appraisal Method[M].Beijing:Science Press,2001:62-63,349-398(in Chinese)东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社,2001:62-63,349-398
[21]Mesbah M,Premachandran U,Whitman WB.Precise measurement of the G+C content of deoxyribonucleic acid by high-performance liquid chromatography[J].International Journal of Systematic and Evolutionary Microbiology,1989,39:159-167
[22]Santal AR,Singh NP,Saharan BS.A novel application of Paracoccus pantotrophus for the decolorization of melanoidins from distillery effluent under static conditions[J].Journal of Environmental Management,2016,169:78-83
[23]Faridha BI,Mohankumar R,Jeevan M,et al.GC-MS analysis of bio-active molecules derived from Paracoccus pantotrophus FMR19 and the antimicrobial activity against bacterial pathogens and MDROs[J].Indian Journal of Microbiology,2016,56(4):426-432
[24]Jiang JD,Zhang RF,He J,et al.Bacterial chemotaxis to environmental pollutants and it’s significance in bioremediation[J].Acta Ecologica Sinica,2005,25(7):1764-1771(in Chinese)蒋建东,张瑞福,何健,等.细菌对环境污染物的趋化性及其在生物修复中的作用[J].生态学报,2005,25(7):1764-1771
[25]Wang SP,Tuo BH,Ma XF,et al.Study on isolation and degradation kinetics of pyridine-degrading bacterium[J].Fuel&Chemical Processes,2013,44(6):41-45(in Chinese)王书萍,庹保华,马雄风,等.吡啶降解菌的分离及降解动力学研究[J].燃料与化工,2013,44(6):41-45
[26]Zhou M,Wang XH,Wei SB,et al.Strain isolation from the coking effluent and the degradation condition of pyridine by the strain[J].Fuel&Chemical Processes,2011,42(2):44-46,49(in Chinese)周敏,王小虎,魏松波,等.焦化废水吡啶降解菌的筛选及降解条件[J].燃料与化工,2011,42(2):44-46,49
[2]Zhuo M,He CD,Liu WH.Purification of refractory pyridine waste gas by anoxic denitrification[J].Chinese Journal of Environmental Engineering,2017,11(12):6345-6350(in Chinese)卓猛,何成达,刘伟慧.利用反硝化法净化难降解吡啶废气[J].环境工程学报,2017,11(12):6345-6350
[3]Jin TT,Ren JH,Zhang H,et al.Identification and characterization of a pyridine degrading bacterium[J].Ecology and Environmental Sciences,2016,25(7):1217-1224(in Chinese)晋婷婷,任嘉红,张晖,等.一株吡啶高效降解菌的鉴定及其降解特性[J].生态环境学报,2016,25(7):1217-1224
[4]Yi X,Zhong J.Biodegradation of pyridine and quinoline by two Pseudomonas strains[J].Acta Microbiologica Sinica,2011,51(8):1087-1097
[5]Liu JJ,Chen LJ,Xiong RL,et al.Physiological and biochemical characteristics of a pyridine-degrading bacterial strain[J].Acta Scientiarum Naturalium Universitatis Pekinensis,2008,44(6):983-986(in Chinese)刘江江,陈吕军,熊瑞林,等.一株吡啶降解菌的生理生化特性研究[J].北京大学学报:自然科学版,2008,44(6):983-986
[6]Chen P,Yan JB,Yu YD.Screening of a pyridine-degrading strain and its biodegradation capability[J].Environmental Protection of Chemical Industry,2015,35(6):566-570(in Chinese)陈佩,颜家保,余永登.一株吡啶降解菌的筛选及其降解性能[J].化工环保,2015,35(6):566-570
[7]Sun JQ,Xu L,Tang YQ,et al.Degradation of pyridine by one Rhodococcus strain in the presence of chromium(VI)or phenol[J].Journal of Hazardous Materials,2011,191(1/3):62-68
[8]Padoley KV,Mudliar SN,Pandey RA.Microbial degradation of pyridine andα-picoline using a strain of the genera Pseudomonas and Nocardia sp.[J].Bioprocess and Biosystems Engineering,2009,32(4):501-510
[9]Deng XQ,Wei CH,Ren Y,et al.Isolation and identification of Achromobacter sp.DN-06 and evaluation of its pyridine degradation kinetics[J].Water,Air,&Soil Pollution,2011,221(1/4):365-375
[10]Shen JY,Zhang X,Chen D,et al.Characteristics of pyridine biodegradation by a novel bacterial strain,Rhizobium sp.NJUST18[J].Desalination and Water Treatment,2015,53(7):2005-2013
[11]Zefirov NS,Agapova SR,Terentiev PB,et al.Degradation of pyridine by Arthrobacter crystallopoietes and Rhodococcus opacus strains[J].FEMS Microbiology Letters,1994,118(1/2):71-74
[12]Sun JQ,Xu L,Tang YQ,et al.Degradation characteristics of pyridine and phenol by Rhodococcus sp.Chr-9[J].Chinese Journal of Applied&Environmental Biology,2012,18(4):647-650(in Chinese)孙纪全,徐莲,汤岳琴,等.红球菌Chr-9降解吡啶和苯酚的特性[J].应用与环境生物学报,2012,18(4):647-650
[13]Mohan SV,Sistla S,Guru RK,et al.Microbial degradation of pyridine using Pseudomonas sp.and isolation of plasmid responsible for degradation[J].Waste Management,2003,23(2):167-171
[14]Ma YF,Wang L,Shao ZZ.Pseudomonas,the dominant polycyclic aromatic hydrocarbon-degrading bacteria isolated from Antarctic soils and the role of large plasmids in horizontal gene transfer[J].Envrionmental Microbiology,2006,8(3):455-465
[15]Bai YH.Microbial degradation of pyridine and quinolne and bioaugmentation for their removal:characteristics and mechanisms[D].Beijing:Doctoral Dissertation of Peking University,2009(in Chinese)柏耀辉.吡啶、喹啉微生物降解及生物强化去除的特性与机理[D].北京:北京大学博士学位论文,2009
[16]Li YX,Zhao CH.Determination of main pollutants in coking wastewater by ultraviolet spectrophotometry[J].China Water&Wastewater,2001,17(1):54-56(in Chinese)李亚新,赵晨红.紫外分光光度法测定焦化废水的主要污染物[J].中国给水排水,2001,17(1):54-56
[17]Edwards U,Rogall T,Bl?cker H,et al.Isolation and direct complete nucleotide determination of entire genes.Characterization of a gene coding for 16S ribosomal RNA[J].Nucleic Acids Research,1989,17(19):7843-7853
[18]Lane DJ.16S/23S r RNA sequencing[A]//Stackebrandt E,Goodfellow M.Nucleic Acid Techniques in bacterial Systematics[M].New York:John Wiley&Sons,1991:115-175
[19]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
[20]Dong XZ,Cai MY.Common Bacteria System Classification and Appraisal Method[M].Beijing:Science Press,2001:62-63,349-398(in Chinese)东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京:科学出版社,2001:62-63,349-398
[21]Mesbah M,Premachandran U,Whitman WB.Precise measurement of the G+C content of deoxyribonucleic acid by high-performance liquid chromatography[J].International Journal of Systematic and Evolutionary Microbiology,1989,39:159-167
[22]Santal AR,Singh NP,Saharan BS.A novel application of Paracoccus pantotrophus for the decolorization of melanoidins from distillery effluent under static conditions[J].Journal of Environmental Management,2016,169:78-83
[23]Faridha BI,Mohankumar R,Jeevan M,et al.GC-MS analysis of bio-active molecules derived from Paracoccus pantotrophus FMR19 and the antimicrobial activity against bacterial pathogens and MDROs[J].Indian Journal of Microbiology,2016,56(4):426-432
[24]Jiang JD,Zhang RF,He J,et al.Bacterial chemotaxis to environmental pollutants and it’s significance in bioremediation[J].Acta Ecologica Sinica,2005,25(7):1764-1771(in Chinese)蒋建东,张瑞福,何健,等.细菌对环境污染物的趋化性及其在生物修复中的作用[J].生态学报,2005,25(7):1764-1771
[25]Wang SP,Tuo BH,Ma XF,et al.Study on isolation and degradation kinetics of pyridine-degrading bacterium[J].Fuel&Chemical Processes,2013,44(6):41-45(in Chinese)王书萍,庹保华,马雄风,等.吡啶降解菌的分离及降解动力学研究[J].燃料与化工,2013,44(6):41-45
[26]Zhou M,Wang XH,Wei SB,et al.Strain isolation from the coking effluent and the degradation condition of pyridine by the strain[J].Fuel&Chemical Processes,2011,42(2):44-46,49(in Chinese)周敏,王小虎,魏松波,等.焦化废水吡啶降解菌的筛选及降解条件[J].燃料与化工,2011,42(2):44-46,49