一株碳酸盐矿化菌的分离鉴定及其对Cu的固定作用
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摘要
有机肥的大量使用使得菜田土壤重金属含量呈快速的累积趋势,对蔬菜安全生产构成了严重的威胁.本文从长期施加鸡粪猪粪等农家肥的大棚蔬菜根际土壤中筛选出了一株能降解尿素释放碳酸根,进而通过生物矿化作用固结重金属的细菌,系统发育分析和生理生化特征表明该菌株为氧化木糖无色杆菌,将其命名为LAX2.菌株LAX2可在含尿素的培养基中快速生长,所产脲酶的活力可达140 U·m L~(-1),发酵液的pH可达9.06.菌株LAX2可耐受浓度高达115 mg·L~(-1)的铜离子.X-射线衍射分析(XRD)、傅里叶红外光谱分析(FTIR)、扫描电子显微镜(SEM)和能谱(EDS)分析表明菌株LAX2可通过生物矿化作用形成微球形的Cu_2(OH)_2CO_3,其直径可达2μm.菌体细胞吸附、生物矿化和化学沉淀对溶液中Cu~(2+)的去除率分别为93%、85%和72%.菌株LAX2对土壤中有效态Cu的固定作用呈现适应-快速-慢速3个阶段,培养5、10和30 d后土壤有效态Cu的含量分别下降了52.3%、73.7%和87.4%.与菌体细胞吸附和化学沉淀相比,生物矿化作用固定的Cu对短期内的淹水和反复冻融具有很强的抗性作用.以上结果表明菌株LAX2可通过生物成矿作用形成性质较稳定的碳酸铜矿物,在土壤铜污染修复方面具有重要的应用价值.
Heavy metal content in vegetable soil shows obvious accumulation trend due to the large use of organic fertilizer,which poses a serious threat to the safety of vegetable production. A bacterium strain,carring the ability to degrade urea and precipitate heavy metals through bio-mineralization,was isolated from the vegetable rhizosphere soil with long-term application of farm manure,such as chicken and pig manure. According to the results of phylogenetic analysis,physiological and biochemical characterization,this strain was named as Achromobacter xylosoxidans LAX2. Strain LAX2 grew rapidly in urea-containing medium,producing urease with the highest activity of 140 U·m L~(-1)under pH 9.06 in fermentation broth. Strain LAX2 is copper resisted with the highest tolerance concentration of 115 mg·L~(-1). Results of X-ray diffraction( XRD),Fourier transform infrared spectroscopy( FTIR),scanning electron microscopy( SEM) and energy dispersive spectroscopy( EDS) analysis show that the bio-mineralized product was Cu2( OH)2CO3and micro-spherical shape with diameter of 2 μm. The removal rate of Cu2+by bacterial cell adsorption,biomineralization and chemical precipitation were93%,85% and 72% respectively. The immobilization of available Cu in soil by strain LAX2 presented three phases,such as adaption,fast and slow rate phases. The content of available Cu in soil decreased by 52. 3%,73. 7% and 87. 4%,respectively,after 5,10 and 30 days of bacrterium cultivation.Immobilized Cu shows strong resistance to short-term flooding and repeated freezing and thawing,compared with bioadsorption and chemical precipitation.Results of this study indicate that strain LAX2 could form stable carbonate minerals by biomineralization and have application potential in Cu-polluted soil remediation.
Heavy metal content in vegetable soil shows obvious accumulation trend due to the large use of organic fertilizer,which poses a serious threat to the safety of vegetable production. A bacterium strain,carring the ability to degrade urea and precipitate heavy metals through bio-mineralization,was isolated from the vegetable rhizosphere soil with long-term application of farm manure,such as chicken and pig manure. According to the results of phylogenetic analysis,physiological and biochemical characterization,this strain was named as Achromobacter xylosoxidans LAX2. Strain LAX2 grew rapidly in urea-containing medium,producing urease with the highest activity of 140 U·m L~(-1)under pH 9.06 in fermentation broth. Strain LAX2 is copper resisted with the highest tolerance concentration of 115 mg·L~(-1). Results of X-ray diffraction( XRD),Fourier transform infrared spectroscopy( FTIR),scanning electron microscopy( SEM) and energy dispersive spectroscopy( EDS) analysis show that the bio-mineralized product was Cu2( OH)2CO3and micro-spherical shape with diameter of 2 μm. The removal rate of Cu2+by bacterial cell adsorption,biomineralization and chemical precipitation were93%,85% and 72% respectively. The immobilization of available Cu in soil by strain LAX2 presented three phases,such as adaption,fast and slow rate phases. The content of available Cu in soil decreased by 52. 3%,73. 7% and 87. 4%,respectively,after 5,10 and 30 days of bacrterium cultivation.Immobilized Cu shows strong resistance to short-term flooding and repeated freezing and thawing,compared with bioadsorption and chemical precipitation.Results of this study indicate that strain LAX2 could form stable carbonate minerals by biomineralization and have application potential in Cu-polluted soil remediation.
引文
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郭莹,秦玉莹,鞠天琛,等.2016.一株穗花狐尾藻内生菌的分离鉴定及其溶磷特征研究[J].环境科学学报,36(12):4352-4360
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Hadi P,Barford J,Mc Kay G.2013.Toxic heavy metal capture using a novel electronic waste-based material-mechanism,modeling and comparison[J].Environmental Science&Technology,47(15):8248-8255
Ivan S,Egon M.2003.Homogeneous precipitation by enzymecatalyzed reactions-strontium and barium carbonates[J].Chemistry of Materials,15(6):1322-1326
Jin H P,Bolan N,Megharaj M,et al.2011.Isolation of phosphate solubilizing bacteria and their potential for lead immobilization in soil[J].Hazardous Materials,185:829-836
李辉,连宾,龚国洪,等.2011.碳酸钙颗粒的细菌诱导形成[J].高校地质学报,17(1):112-117
Li L,Qian C X,Cheng L,et al.2010.A laboratory investingation of microbe-inducing Cd CO3precipitate treatment in Cd2+contaminated soil[J].Journal of Soils and Sediments,10:248-254
刘凤芝,马锦秋.2010.土壤监测分析实用手册[M].北京:化学工业出版社
陆兆文,钱春香,许燕波,等.2013.不同污染条件下微生物矿化固结Zn2+的作用及机理[J].东南大学学报,43(2):365-370
钱春香,许燕波,胡黎明,等.2011.一种微生物固结污染体系中Cu2+的研究[J].环境科学与技术,34(12H):33-36
钱春香,王瑞兴,詹其伟.2015.微生物矿化的工程应用基础[M].北京:科学出版社
Shin M N,Shim J,You Y,et al.2012.Characterization of lead resistant endophytic bacillus sp.MN3-4 and its potential for promoting lead accumulation in metal hyperaccmulator Alnus firma[J].Hazardous Materials,200(199):314-320
Tamura K,Stecher G,Peterson D,et al.2013.MEAG6:molecular evolutionary genetics analysis version 6.0[J].Molecular Biology Evolution,30(12):2725-2729
Van R S,Vanbroekhoven K,Dejonghe W,et al.2006.Immobilization of heavy metals in the saturated zone by sorption and in situ bioprecipitation processes[J].Hydrometallurgy,83(1):195-203
王明明,钱春香.2013.磷酸盐矿化菌矿化重金属离子Zn2+的研究[J].功能材料,3(44):393-395
王瑞兴,钱春香,吴淼,等.2007.微生物矿化固结土壤中重金属研究[J].功能材料,9(38):1523-1527
徐晶,姚武.2013.微生物非脲解作用诱导碳酸钙沉积研究[J].同济大学学报(自然科学版),41(10):1542-1546
许燕波,钱春香,陆兆文.2013.柠檬酸粗化铜离子矿化产物颗粒粒径的研究[J].材料导报B,27(1):92-95
许燕波,钱春香,陆兆文,等.2012.微生物矿化修复铅离子污染的研究[J].化工时刊,26(6):14-17
许燕波,钱春香,陆兆文,等.2013.微生物矿化修复重金属污染土壤[J].环境工程学报,7(7):2763-2768
周维芝,李伟伟,张玉忠,等.2009.深海适冷菌Pseudoalteromonas sp.SM9913胞外多糖对Pb2+和Cu2+的吸附性能研究[J].环境科学,30(1):200-205
赵越,姚俊,王天齐,等.2016.碳酸盐矿化菌的筛选与其吸附矿化Cd2+的特性[J].中国环境科学,36(12):3800-3806
Chai L,Huang S,Yang Z,et al.2009.Cr(Ⅵ)remediation by indigenous bacteria in soils contaminated by chromium-containing slag[J].Journal of Hazardous Materials,167(1/3):516-522
陈庆丽,景澄茗,付韵鑫,等.2014.寒区水体中溶藻铜绿假单胞菌的分离和性质研究[J].环境科学学报,35(3):692-698
Chen Y X,Wang Y P,Lin Q,et al.2005.Effect of copper-tolerant rhizosphere bacteria on mobility of copper in soil and copper accumulation by Elsholtzia splendens[J].Environment International,31(6):861-866
成亮,钱春香,王瑞兴,等.2008.碳酸盐矿化菌株A固结土壤Cd2+的矿化过程[J].硅酸盐学报,36(S1):215-221
董明,宋卫锋,程亚杰.2016.苯胺黑药高效降解菌(Bacillus vallismortis)胞外聚合物去除重金属的研究[J].环境科学学报,36(12):4367-4375
东秀珠,蔡妙英,等.2001.常见细菌系统鉴定手册[M].北京:科学出版社
Francesca P,Paola Z.2000.Heavy metal coprecipitation with hydrozincite[Zn5(CO3)2(OH)6]from mine waters caused by photosynthetic microorganisms[J].Applied and Environmental Microbiology,66(11):5092-5098
Fujita Y,Redden G D,Ingram J C,et al.2004.Strontium incorporation into calcite generated bybacterial ureolysis[J].Geochimicaet Cosmochimica Acta,68(15):3261-3270
郭莹,秦玉莹,鞠天琛,等.2016.一株穗花狐尾藻内生菌的分离鉴定及其溶磷特征研究[J].环境科学学报,36(12):4352-4360
Gwenzi W,Musarurwa T,Nyamugafata P,et al.2014.Adsorptiom of Zn2+and Ni2+in a binary aqueous solution by biosorbents derived from sawdust and water hyacinth(Eichhorniacrassipes)[J].Water Science&Technology,70(8):1419-1427
Hadi P,Barford J,Mc Kay G.2013.Toxic heavy metal capture using a novel electronic waste-based material-mechanism,modeling and comparison[J].Environmental Science&Technology,47(15):8248-8255
Ivan S,Egon M.2003.Homogeneous precipitation by enzymecatalyzed reactions-strontium and barium carbonates[J].Chemistry of Materials,15(6):1322-1326
Jin H P,Bolan N,Megharaj M,et al.2011.Isolation of phosphate solubilizing bacteria and their potential for lead immobilization in soil[J].Hazardous Materials,185:829-836
李辉,连宾,龚国洪,等.2011.碳酸钙颗粒的细菌诱导形成[J].高校地质学报,17(1):112-117
Li L,Qian C X,Cheng L,et al.2010.A laboratory investingation of microbe-inducing Cd CO3precipitate treatment in Cd2+contaminated soil[J].Journal of Soils and Sediments,10:248-254
刘凤芝,马锦秋.2010.土壤监测分析实用手册[M].北京:化学工业出版社
陆兆文,钱春香,许燕波,等.2013.不同污染条件下微生物矿化固结Zn2+的作用及机理[J].东南大学学报,43(2):365-370
钱春香,许燕波,胡黎明,等.2011.一种微生物固结污染体系中Cu2+的研究[J].环境科学与技术,34(12H):33-36
钱春香,王瑞兴,詹其伟.2015.微生物矿化的工程应用基础[M].北京:科学出版社
Shin M N,Shim J,You Y,et al.2012.Characterization of lead resistant endophytic bacillus sp.MN3-4 and its potential for promoting lead accumulation in metal hyperaccmulator Alnus firma[J].Hazardous Materials,200(199):314-320
Tamura K,Stecher G,Peterson D,et al.2013.MEAG6:molecular evolutionary genetics analysis version 6.0[J].Molecular Biology Evolution,30(12):2725-2729
Van R S,Vanbroekhoven K,Dejonghe W,et al.2006.Immobilization of heavy metals in the saturated zone by sorption and in situ bioprecipitation processes[J].Hydrometallurgy,83(1):195-203
王明明,钱春香.2013.磷酸盐矿化菌矿化重金属离子Zn2+的研究[J].功能材料,3(44):393-395
王瑞兴,钱春香,吴淼,等.2007.微生物矿化固结土壤中重金属研究[J].功能材料,9(38):1523-1527
徐晶,姚武.2013.微生物非脲解作用诱导碳酸钙沉积研究[J].同济大学学报(自然科学版),41(10):1542-1546
许燕波,钱春香,陆兆文.2013.柠檬酸粗化铜离子矿化产物颗粒粒径的研究[J].材料导报B,27(1):92-95
许燕波,钱春香,陆兆文,等.2012.微生物矿化修复铅离子污染的研究[J].化工时刊,26(6):14-17
许燕波,钱春香,陆兆文,等.2013.微生物矿化修复重金属污染土壤[J].环境工程学报,7(7):2763-2768
周维芝,李伟伟,张玉忠,等.2009.深海适冷菌Pseudoalteromonas sp.SM9913胞外多糖对Pb2+和Cu2+的吸附性能研究[J].环境科学,30(1):200-205
赵越,姚俊,王天齐,等.2016.碳酸盐矿化菌的筛选与其吸附矿化Cd2+的特性[J].中国环境科学,36(12):3800-3806