一株耐Cd菌株的分离、鉴定及基本特性
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摘要
【目的】从活性污泥中筛选耐镉(Cd)菌株,并研究其生长特性及对溶液中Cd~(2+)吸附的最佳条件,以期为重金属Cd污染水体的微生物修复提供菌株资源和应用技术参考。【方法】采用平板划线法,从活性污泥中分离、筛选、驯化出耐Cd菌株,通过16SrRNA基因序列分析及溶血试验、蛋白质毒素结晶试验进行初步鉴定,并采用单因素实验优化菌株的培养条件,通过正交实验确定菌粉吸附溶液中Cd~(2+)的最佳条件,同时利用SEM-EDS及FTIR分析探讨菌粉吸附Cd~(2+)的机理。【结果】经分离、驯化得到1株耐Cd细菌菌株,命名为H6,初步鉴定为蜡样芽孢杆菌(Bacilluscereus),最大Cd~(2+)耐受浓度为350 mg/L。菌株H6的最佳生长条件为:pH 6.0–8.0,温度28°C,转速120–210 r/min,接种量1%–5%;菌株H6在生长过程中,培养液pH值先稍微下降然后不断上升。菌粉吸附Cd~(2+)的正交优化条件为:菌粉用量0.125 g/L,吸附时间2 h,pH 5.0,温度30°C,此条件下吸附量为205 mg/g。SEM-EDS分析和红外光谱(FTIR)分析表明,在吸附过程中主要作用基团有羟基、羧基、羰基、酰胺基和烷基,此外,Ca~(2+)与Cd~(2+)发生了离子交换。【结论】从活性污泥中分离出的菌株H6,初步鉴定为蜡样芽孢杆菌(Bacillus cereus),是1株具有较强Cd~(2+)吸附能力的细菌菌株。
[Objective] We isolated and identified a cadmium resistant bacterium from activated sludge. Furthermore, we studied its growth characteristics and optimal biosorption conditions of Cd~(2+). This study was expected to provide strain resources and applied reference for microbial remediation of cadmium contaminated water. [Methods] The bacterium was isolated by streak plate and identified based on its 16 S rRNA analysis, hemolysis testing and protein crystallization dying experiment. The growth conditions of the strain were determined by single factor test. The optimal biosorption conditions of bacterial powder were investigated by designed orthogonal experiment. Besides, the biosorption mechanisms were discussed by Scanning electron microscope(SEM), Energy Dispersive X-ray Scanning(EDS) and Fourier transform-infrared spectroscopy(FTIR) analysis. [Results] The strain was identified as Bacillus cereus and named as H6. The maximum cadmium tolerance concentration of strain H6 was 350 mg/L. Its culture conditions were: initial pH 6.0 to 8.0 at 28 °C, shaking speed of 120 to 210 r/min, inoculation 1% to 5%. During the growth of strain H6, the pH of culture medium first decreased slightly and then increased gradually. The optimal biosorption conditions of bacterial powder were: dosage of 0.125 g/L, contact time of 2 h, pH 5.0 and 30 °C, the biosorption capacity reached 205 mg/g. SEM-EDS and FTIR results revealed that ion exchange between Ca~(2+) and Cd~(2+) occurred and the functional group of hydroxyl, carboxyl, carbonyl, amido and alkyl played a principal role during the biosorption process. [Conclusion] Strain H6, isolated from the activated sludge, was identified as Bacillus cereus with good adsorption property for Cd~(2+).
[Objective] We isolated and identified a cadmium resistant bacterium from activated sludge. Furthermore, we studied its growth characteristics and optimal biosorption conditions of Cd~(2+). This study was expected to provide strain resources and applied reference for microbial remediation of cadmium contaminated water. [Methods] The bacterium was isolated by streak plate and identified based on its 16 S rRNA analysis, hemolysis testing and protein crystallization dying experiment. The growth conditions of the strain were determined by single factor test. The optimal biosorption conditions of bacterial powder were investigated by designed orthogonal experiment. Besides, the biosorption mechanisms were discussed by Scanning electron microscope(SEM), Energy Dispersive X-ray Scanning(EDS) and Fourier transform-infrared spectroscopy(FTIR) analysis. [Results] The strain was identified as Bacillus cereus and named as H6. The maximum cadmium tolerance concentration of strain H6 was 350 mg/L. Its culture conditions were: initial pH 6.0 to 8.0 at 28 °C, shaking speed of 120 to 210 r/min, inoculation 1% to 5%. During the growth of strain H6, the pH of culture medium first decreased slightly and then increased gradually. The optimal biosorption conditions of bacterial powder were: dosage of 0.125 g/L, contact time of 2 h, pH 5.0 and 30 °C, the biosorption capacity reached 205 mg/g. SEM-EDS and FTIR results revealed that ion exchange between Ca~(2+) and Cd~(2+) occurred and the functional group of hydroxyl, carboxyl, carbonyl, amido and alkyl played a principal role during the biosorption process. [Conclusion] Strain H6, isolated from the activated sludge, was identified as Bacillus cereus with good adsorption property for Cd~(2+).
引文
[1]Wen XF,Du CY,Yuan HY,Zhang JF,Chen H,Yu GL,Hu XY,Peng XX.Adsorption of Cd2+in wastewater through modified magnetic nanoparticles immobilizing endogenous bacterium Bacillus nealsonii.Acta Scientiae Circumstantiae,2016,36(12):4376-4383.(in Chinese)文晓凤,杜春艳,袁瀚宇,张金帆,陈宏,余关龙,胡旭跃,彭向训.改性磁性纳米颗粒固定内生菌Bacillus nealsonii吸附废水中Cd2+的特性研究.环境科学学报,2016,36(12):4376-4383.
[2]Dong JQ,Zhang HZ,Wang JN,Zhang TZ.Modeling of cadmium speciation in Longjiang River during an emergent environmental incident.China Environmental Science,2015,35(10):3046-3052.(in Chinese)董璟琦,张红振,王金南,张天柱.龙江河突发环境事件河流镉污染化学形态模拟.中国环境科学,2015,35(10):3046-3052.
[3]Zhang XJ,Lin PF,Chen C.Emergent water supply in environmental accident of cadmium,hallium and arsenic in Xiannvhu Lake in Xinyu City.China Water&Wastewater,2017,33(9):1-10.(in Chinese)张晓健,林朋飞,陈超.江西新余仙女湖镉铊砷突发环境事件应急供水.中国给水排水,2017,33(9):1-10.
[4]Tran TA,Popova LP.Functions and toxicity of cadmium in plants:recent advances and future prospects.Turkish Journal of Botany,2013,37(1):1-13.
[5]Godt J,Scheidig F,Grosse-Siestrup C,Esche V,Brandenburg P,Reich A,Groneberg DA.The toxicity of cadmium and resulting hazards for human health.Journal of Occupational Medicine and Toxicology,2006,1:22.
[6]Cao Z,Luo SL,Zeng GM,Xiao X,Wan Y,Su F.Removal of Cd2+by an endophytic bacteria SDE06 obtained from Solanum Nigrum L..Microbiology China,2009,36(3):328-333.(in Chinese)曹喆,罗胜联,曾光明,肖潇,万勇,苏峰.一株龙葵内生细菌SDE06去除Cd2+的实验.微生物学通报,2009,36(3):328-333.
[7]Fu FL,Wang Q.Removal of heavy metal ions from wastewaters:a review.Journal of Environmental Management,2011,92(3):407-418.
[8]Ye JJ,Xiao HL,Xiao BL,Xu WS,Gao LX,Lin G.Bioremediation of heavy metal contaminated aqueous solution by using red algae Porphyra leucosticta.Water Science&Technology,2015,72(9):1662-1666.
[9]Xiao CW,Luo XY,Tian Y,Lu XY.Research progress of bioremediation of heavy metal cadmium pollution.Chemistry&Bioengineering,2013,30(8):1-4.(in Chinese)肖春文,罗秀云,田云,卢向阳.重金属镉污染生物修复的研究进展.化学与生物工程,2013,30(8):1-4.
[10]Lin XY,Mou RX,Cao ZY,Zhu ZW,Chen MX.Isolation and cadmium adsorption mechanisms of cadmium-resistant bacteria strains.Journal of Agro-Environment Science,2015,34(9):1700-1706.(in Chinese)林晓燕,牟仁祥,曹赵云,朱智伟,陈铭学.耐镉细菌菌株的分离及其吸附镉机理研究.农业环境科学学报,2015,34(9):1700-1706.
[11]Wang JL,Chen C.Research advances in heavy metal removal by biosorption.Acta Scientiae Circumstantiae,2010,30(4):673-701.(in Chinese)王建龙,陈灿.生物吸附法去除重金属离子的研究进展.环境科学学报,2010,30(4):673-701.
[12]Xiao W,Zhang SY,Zhao Q,Wang YX,Lai YH,Li ZY,Cui XL.Diversity and heavy-metal tolerance of bacteria isolated from Gejiu tin mining area of Yunnan.Acta Microbiologica Sinica,2013,53(11):1158-1165.(in Chinese)肖炜,张仕颖,赵琴,王永霞,赖泳红,李治滢,崔晓龙.云南个旧锡矿区可培养细菌多样性及其重金属抗性.微生物学报,2013,53(11):1158-1165.
[13]Igwe JC,Abia AA.A bioseparation process for removing heavy metals from waste water using biosorbents.African Journal of Biotechnology,2006,5(12):1167-1179.
[14]Loukidou MX,Karapantsios TD,Zouboulis AI,Matis KA.Diffusion kinetic study of cadmium(II)biosorption by Aeromonas caviae.Journal of Chemical Technology&Biotechnology,2004,79(7):711-719.
[15]Ziagova M,Dimitriadis G,Aslanidou D,Papaioannou X,Litopoulou E,Liakopoulou-Kyriakides M.Comparative study of Cd(II)and Cr(VI)biosorption on Staphylococcus xylosus and Pseudomonas sp.in single and binary mixtures.Bioresource Technology,2007,98(15):2859-2865.
[16]Flouty R,Estephane G.Bioaccumulation and biosorption of copper and lead by a unicellular algae Chlamydomonas reinhardtii in single and binary metal systems:a comparative study.Journal of Environmental Management,2012,111:106-114.
[17]KadukováJ,Vir?íkováE.Comparison of differences between copper bioaccumulation and biosorption.Environment International,2005,31(2):227-232.
[18]黄飞.蜡状芽孢杆菌对水体中镉的吸附特性与机理研究.华南理工大学博士学位论文,2013.
[19]Li HF,Lin YB,Guan WM,Chang JL,Xu L,Guo JK,Wei GH.Biosorption of Zn(II)by live and dead cells of Streptomyces ciscaucasicus strain CCNWHX 72-14.Journal of Hazardous Materials,2010,179(1/3):151-159.
[20]Limcharoensuk T,Sooksawat N,Sumarnrote A,Awutpet T,Kruatrachue M,Pokethitiyook P,Auesukaree C.Bioaccumulation and biosorption of Cd2+and Zn2+by bacteria isolated from a zinc mine in Thailand.Ecotoxicology and Environmental Safety,2015,122:322-330.
[21]Tang KX,Niu ZX,Sun CX,Pang X.Research on growth characteristics of Bacillus cereus.Shandong Journal of Animal Science and Veterinary Medicine,2000(1):6-7.(in Chinese)唐珂心,牛钟相,孙存孝,庞昕.蜡样芽胞杆菌生长特性的研究.山东畜牧兽医,2000(1):6-7.
[22]Meseguer VF,Ortu?o JF,Aguilar MI,Pinzón-Bedoya ML,Lloréns M,Sáez J,Pérez-Marín AB.Biosorption of cadmium(II)from aqueous solutions by natural and modified non-living leaves of Posidonia oceanica.Environmental Science and Pollution Research,2016,23(23):24032-24046.
[23]Zheng HS,Guo WQ,Li S,Wu QL,Yin RL,Feng XC,Du JS,Ren NQ,Chang JS.Biosorption of cadmium by a lipid extraction residue of lipid-rich microalgae.RSC Advances,2016,6(24):20051-20057.
[24]Jin Y,Qu JJ,Li Y,Gu HD,Yan LL,Sun XB.Isolation,identification and Pb(II)biosorption characterization of a lead-resistant strain.Acta Scientiae Circumstantiae,2013,33(8):2248-2255.(in Chinese)金羽,曲娟娟,李影,顾海东,闫立龙,孙兴滨.一株耐铅细菌的分离鉴定及其吸附特性研究.环境科学学报,2013,33(8):2248-2255.
[25]Chen MB,Guo JH,Yao Q,Duan JM,Feng GD,Cai ZP,Zhu HH.Isolation and identification,and biological characteristics of cadmium-tolerant bacteria from the Dabao Mine.Microbiology China,2012,39(12):1720-1733.(in Chinese)陈美标,郭建华,姚青,段锦梅,冯广达,蔡卓平,朱红惠.大宝山矿区耐Cd2+细菌的分离鉴定及其生物学特性.微生物学通报,2012,39(12):1720-1733.
[26]Geng YY,Wang XM,Wang HQ,Tong XC.Screening and identification of cadmium-tolerant bacteria in polluted soil and characteristics.Journal of Northeast Agricultural University,2010,41(11):59-65.(in Chinese)耿印印,王旭梅,王红旗,佟秀春.污染土壤中耐镉菌株的筛选、鉴定及吸附试验研究.东北农业大学学报,2010,41(11):59-65.
[27]Liu Y,Dang Z,Liu Y,Yi XY,Guo CL,Lu GN,Zhou XQ.Isolation and identification of a sulfate-reducing bacteria DSRBa and its characterization.Journal of Agro-Environment Science,2011,30(1):176-182.(in Chinese)刘艳,党志,刘云,易筱筠,郭楚玲,卢桂宁,周兴求.一株硫酸盐还原菌DSRBa的分离鉴定及特性分析.农业环境科学学报,2011,30(1):176-182.
[28]Huang F,Dang Z,Guo CL,Lu GN,Gu RR,Liu HJ,Zhang H.Biosorption of Cd(II)by live and dead cells of Bacillus cereus RC-1 isolated from cadmium-contaminated soil.Colloids&Surfaces B:Biointerfaces,2013,107:11-18.
[29]周赓.耐镉放线菌的筛选及特性研究.湖南农业大学硕士毕业论文,2016.
[30]Sun YL,Li X,Wang J,Wang JF,Li WJ,Wang YB.Screening of a Cd2+-resistant bacterium and its biosorption.Chinese Journal of Environmental Engineering,2014,8(3):1235-1242.(in Chinese)孙永亮,李欣,王洁,王菊芳,李文建,王弋博.一株抗Cd2+菌株的筛选及其吸附性能.环境工程学报,2014,8(3):1235-1242.
[31]Hokkanen S,Bhatnagar A,Sillanp??M.A review on modification methods to cellulose-based adsorbents to improve adsorption capacity.Water Research,2016,91:156-173.
[32]Goswami S,Syiem MB,Pakshirajan K.Cadmium removal by Anabaena doliolum Ind1 isolated from a coal mining area in Meghalaya,India:associated structural and physiological alterations.Environmental Engineering Research,2015,20(1):41-50.
[2]Dong JQ,Zhang HZ,Wang JN,Zhang TZ.Modeling of cadmium speciation in Longjiang River during an emergent environmental incident.China Environmental Science,2015,35(10):3046-3052.(in Chinese)董璟琦,张红振,王金南,张天柱.龙江河突发环境事件河流镉污染化学形态模拟.中国环境科学,2015,35(10):3046-3052.
[3]Zhang XJ,Lin PF,Chen C.Emergent water supply in environmental accident of cadmium,hallium and arsenic in Xiannvhu Lake in Xinyu City.China Water&Wastewater,2017,33(9):1-10.(in Chinese)张晓健,林朋飞,陈超.江西新余仙女湖镉铊砷突发环境事件应急供水.中国给水排水,2017,33(9):1-10.
[4]Tran TA,Popova LP.Functions and toxicity of cadmium in plants:recent advances and future prospects.Turkish Journal of Botany,2013,37(1):1-13.
[5]Godt J,Scheidig F,Grosse-Siestrup C,Esche V,Brandenburg P,Reich A,Groneberg DA.The toxicity of cadmium and resulting hazards for human health.Journal of Occupational Medicine and Toxicology,2006,1:22.
[6]Cao Z,Luo SL,Zeng GM,Xiao X,Wan Y,Su F.Removal of Cd2+by an endophytic bacteria SDE06 obtained from Solanum Nigrum L..Microbiology China,2009,36(3):328-333.(in Chinese)曹喆,罗胜联,曾光明,肖潇,万勇,苏峰.一株龙葵内生细菌SDE06去除Cd2+的实验.微生物学通报,2009,36(3):328-333.
[7]Fu FL,Wang Q.Removal of heavy metal ions from wastewaters:a review.Journal of Environmental Management,2011,92(3):407-418.
[8]Ye JJ,Xiao HL,Xiao BL,Xu WS,Gao LX,Lin G.Bioremediation of heavy metal contaminated aqueous solution by using red algae Porphyra leucosticta.Water Science&Technology,2015,72(9):1662-1666.
[9]Xiao CW,Luo XY,Tian Y,Lu XY.Research progress of bioremediation of heavy metal cadmium pollution.Chemistry&Bioengineering,2013,30(8):1-4.(in Chinese)肖春文,罗秀云,田云,卢向阳.重金属镉污染生物修复的研究进展.化学与生物工程,2013,30(8):1-4.
[10]Lin XY,Mou RX,Cao ZY,Zhu ZW,Chen MX.Isolation and cadmium adsorption mechanisms of cadmium-resistant bacteria strains.Journal of Agro-Environment Science,2015,34(9):1700-1706.(in Chinese)林晓燕,牟仁祥,曹赵云,朱智伟,陈铭学.耐镉细菌菌株的分离及其吸附镉机理研究.农业环境科学学报,2015,34(9):1700-1706.
[11]Wang JL,Chen C.Research advances in heavy metal removal by biosorption.Acta Scientiae Circumstantiae,2010,30(4):673-701.(in Chinese)王建龙,陈灿.生物吸附法去除重金属离子的研究进展.环境科学学报,2010,30(4):673-701.
[12]Xiao W,Zhang SY,Zhao Q,Wang YX,Lai YH,Li ZY,Cui XL.Diversity and heavy-metal tolerance of bacteria isolated from Gejiu tin mining area of Yunnan.Acta Microbiologica Sinica,2013,53(11):1158-1165.(in Chinese)肖炜,张仕颖,赵琴,王永霞,赖泳红,李治滢,崔晓龙.云南个旧锡矿区可培养细菌多样性及其重金属抗性.微生物学报,2013,53(11):1158-1165.
[13]Igwe JC,Abia AA.A bioseparation process for removing heavy metals from waste water using biosorbents.African Journal of Biotechnology,2006,5(12):1167-1179.
[14]Loukidou MX,Karapantsios TD,Zouboulis AI,Matis KA.Diffusion kinetic study of cadmium(II)biosorption by Aeromonas caviae.Journal of Chemical Technology&Biotechnology,2004,79(7):711-719.
[15]Ziagova M,Dimitriadis G,Aslanidou D,Papaioannou X,Litopoulou E,Liakopoulou-Kyriakides M.Comparative study of Cd(II)and Cr(VI)biosorption on Staphylococcus xylosus and Pseudomonas sp.in single and binary mixtures.Bioresource Technology,2007,98(15):2859-2865.
[16]Flouty R,Estephane G.Bioaccumulation and biosorption of copper and lead by a unicellular algae Chlamydomonas reinhardtii in single and binary metal systems:a comparative study.Journal of Environmental Management,2012,111:106-114.
[17]KadukováJ,Vir?íkováE.Comparison of differences between copper bioaccumulation and biosorption.Environment International,2005,31(2):227-232.
[18]黄飞.蜡状芽孢杆菌对水体中镉的吸附特性与机理研究.华南理工大学博士学位论文,2013.
[19]Li HF,Lin YB,Guan WM,Chang JL,Xu L,Guo JK,Wei GH.Biosorption of Zn(II)by live and dead cells of Streptomyces ciscaucasicus strain CCNWHX 72-14.Journal of Hazardous Materials,2010,179(1/3):151-159.
[20]Limcharoensuk T,Sooksawat N,Sumarnrote A,Awutpet T,Kruatrachue M,Pokethitiyook P,Auesukaree C.Bioaccumulation and biosorption of Cd2+and Zn2+by bacteria isolated from a zinc mine in Thailand.Ecotoxicology and Environmental Safety,2015,122:322-330.
[21]Tang KX,Niu ZX,Sun CX,Pang X.Research on growth characteristics of Bacillus cereus.Shandong Journal of Animal Science and Veterinary Medicine,2000(1):6-7.(in Chinese)唐珂心,牛钟相,孙存孝,庞昕.蜡样芽胞杆菌生长特性的研究.山东畜牧兽医,2000(1):6-7.
[22]Meseguer VF,Ortu?o JF,Aguilar MI,Pinzón-Bedoya ML,Lloréns M,Sáez J,Pérez-Marín AB.Biosorption of cadmium(II)from aqueous solutions by natural and modified non-living leaves of Posidonia oceanica.Environmental Science and Pollution Research,2016,23(23):24032-24046.
[23]Zheng HS,Guo WQ,Li S,Wu QL,Yin RL,Feng XC,Du JS,Ren NQ,Chang JS.Biosorption of cadmium by a lipid extraction residue of lipid-rich microalgae.RSC Advances,2016,6(24):20051-20057.
[24]Jin Y,Qu JJ,Li Y,Gu HD,Yan LL,Sun XB.Isolation,identification and Pb(II)biosorption characterization of a lead-resistant strain.Acta Scientiae Circumstantiae,2013,33(8):2248-2255.(in Chinese)金羽,曲娟娟,李影,顾海东,闫立龙,孙兴滨.一株耐铅细菌的分离鉴定及其吸附特性研究.环境科学学报,2013,33(8):2248-2255.
[25]Chen MB,Guo JH,Yao Q,Duan JM,Feng GD,Cai ZP,Zhu HH.Isolation and identification,and biological characteristics of cadmium-tolerant bacteria from the Dabao Mine.Microbiology China,2012,39(12):1720-1733.(in Chinese)陈美标,郭建华,姚青,段锦梅,冯广达,蔡卓平,朱红惠.大宝山矿区耐Cd2+细菌的分离鉴定及其生物学特性.微生物学通报,2012,39(12):1720-1733.
[26]Geng YY,Wang XM,Wang HQ,Tong XC.Screening and identification of cadmium-tolerant bacteria in polluted soil and characteristics.Journal of Northeast Agricultural University,2010,41(11):59-65.(in Chinese)耿印印,王旭梅,王红旗,佟秀春.污染土壤中耐镉菌株的筛选、鉴定及吸附试验研究.东北农业大学学报,2010,41(11):59-65.
[27]Liu Y,Dang Z,Liu Y,Yi XY,Guo CL,Lu GN,Zhou XQ.Isolation and identification of a sulfate-reducing bacteria DSRBa and its characterization.Journal of Agro-Environment Science,2011,30(1):176-182.(in Chinese)刘艳,党志,刘云,易筱筠,郭楚玲,卢桂宁,周兴求.一株硫酸盐还原菌DSRBa的分离鉴定及特性分析.农业环境科学学报,2011,30(1):176-182.
[28]Huang F,Dang Z,Guo CL,Lu GN,Gu RR,Liu HJ,Zhang H.Biosorption of Cd(II)by live and dead cells of Bacillus cereus RC-1 isolated from cadmium-contaminated soil.Colloids&Surfaces B:Biointerfaces,2013,107:11-18.
[29]周赓.耐镉放线菌的筛选及特性研究.湖南农业大学硕士毕业论文,2016.
[30]Sun YL,Li X,Wang J,Wang JF,Li WJ,Wang YB.Screening of a Cd2+-resistant bacterium and its biosorption.Chinese Journal of Environmental Engineering,2014,8(3):1235-1242.(in Chinese)孙永亮,李欣,王洁,王菊芳,李文建,王弋博.一株抗Cd2+菌株的筛选及其吸附性能.环境工程学报,2014,8(3):1235-1242.
[31]Hokkanen S,Bhatnagar A,Sillanp??M.A review on modification methods to cellulose-based adsorbents to improve adsorption capacity.Water Research,2016,91:156-173.
[32]Goswami S,Syiem MB,Pakshirajan K.Cadmium removal by Anabaena doliolum Ind1 isolated from a coal mining area in Meghalaya,India:associated structural and physiological alterations.Environmental Engineering Research,2015,20(1):41-50.
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