耐镉甲基营养芽胞杆菌吸附条件的优化及其抗逆性
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摘要
【目的】对所筛选的1株耐镉甲基营养芽胞杆菌NTGB29进行了环境抗逆性的研究,及影响菌株吸附镉离子效率的条件优化。【方法】以发酵液活菌数为指标,研究其对不同NaCl浓度、酸碱度、镉离子浓度的耐受情况;进一步通过单因素实验和响应面法优化影响菌株镉离子吸附效率的发酵条件;以有效镉离子含量为指标,验证菌株在镉污染土壤中的吸附效果。【结果】结果表明,菌株NTGB29对NaCl浓度、酸碱度、Cd~(2+)浓度的最大耐受值分别为10%、pH11.0、50mg/L;菌株在发酵液初始Cd~(2+)浓度10 mg/L、起始pH 6.4、培养温度37°C、NaCl浓度4.2%、装液量50 mL/250 mL、培养时间24 h时,对Cd~(2+)的吸附率达到79.70%;菌株能有效降低镉污染土壤中的有效镉离子含量,吸附率为29.65%。【结论】菌株NTGB29在较高浓度Cd~(2+)浓度、NaCl浓度及强碱环境条件下仍然能够生长,具有良好的环境抗逆性及Cd~(2+)吸附能力,在镉污染土壤调理剂及微生物功能菌剂的研制方面能够提供有价值的菌种资源。
[Objective] To assess the anti-stress characteristics and to optimize the conditions affecting its cadmium ions adsorption efficiency of a cadmium-tolerant Bacillus methylotrophicus strain NTGB29. [Methods] The anti-stress characteristics were studied by plate colony-counting method. In addition, we used single factor experiment and response surface methodology to optimize the fermentation conditions of cadmium ions adsorption of the strain. Moreover, the available cadmium ions contents were used to verify the cadmium ions adsorption ability of the strain in the cadmium contaminated soils. [Results] The maximum tolerance of NTGB29 to NaCl, pH and concentration of Cd~(2+) was 10%, 11.0 and 50 mg/L, respectively. The optimum cadmium adsorption conditions were as follows: initial pH 6.4, culture temperature 37 °C, NaCl concentration 4.2%, liquid volume 50 mL in the250 mL flask, and culture time 24 h. Under the conditions, the cadmium ions adsorption percentage reached to79.70%. In addition, the available cadmium ions of the cadmium contaminated soils was decreased significantly by adding strain NTGB29, and the adsorption percentage was 29.65%. [Conclusion] Strain NTGB29 has good environmental resistance and Cd~(2+) adsorption capacity, and it could be a kind of valuable strain source to develop the heavy metal contaminated soil conditioners and microbial functional agents.
[Objective] To assess the anti-stress characteristics and to optimize the conditions affecting its cadmium ions adsorption efficiency of a cadmium-tolerant Bacillus methylotrophicus strain NTGB29. [Methods] The anti-stress characteristics were studied by plate colony-counting method. In addition, we used single factor experiment and response surface methodology to optimize the fermentation conditions of cadmium ions adsorption of the strain. Moreover, the available cadmium ions contents were used to verify the cadmium ions adsorption ability of the strain in the cadmium contaminated soils. [Results] The maximum tolerance of NTGB29 to NaCl, pH and concentration of Cd~(2+) was 10%, 11.0 and 50 mg/L, respectively. The optimum cadmium adsorption conditions were as follows: initial pH 6.4, culture temperature 37 °C, NaCl concentration 4.2%, liquid volume 50 mL in the250 mL flask, and culture time 24 h. Under the conditions, the cadmium ions adsorption percentage reached to79.70%. In addition, the available cadmium ions of the cadmium contaminated soils was decreased significantly by adding strain NTGB29, and the adsorption percentage was 29.65%. [Conclusion] Strain NTGB29 has good environmental resistance and Cd~(2+) adsorption capacity, and it could be a kind of valuable strain source to develop the heavy metal contaminated soil conditioners and microbial functional agents.
引文
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[14]Sharma P,Sharma N,Pathania S,Handa S.Purification and characterization of lipase by Bacillus methylotrophicus PS3under submerged fermentation and its application in detergent industry.Journal of Genetic Engineering and Biotechnology,2017,15(2):369-377.
[15]Frikha-Gargouri O,Ben Abdallah D,Ghorbel I,Charfeddine I,Jlaiel L,Triki MA,Tounsi S.Lipopeptides from a novel Bacillus methylotrophicus 39b strain suppress Agrobacterium crown gall tumours on tomato plants.Pest Management Science,2016,73(3):568-574.
[16]Jemil N,Ayed H B,Manresa A,Nasri M,Hmidet N.Antioxidant properties,antimicrobial and anti-adhesive activities of DCS1 lipopeptides from Bacillus methylotrophicus DCS1.BMC Microbiology,2017,17:144.
[17]Rajendiran T,Gopakumaran N,Bhuyan U,Kumari S,Ramesh KV.Optimization and immobilization of lipase from Bacillus subterraneus TNUS15:comparison between response surface methodology and artificial neural network.Der Pharma Chemica,2017,9(12):83-91.
[18]Wang XB,Li XR,Mao CQ,Jiang NP,Guo TL.The characteristics of cadmium adsorption by Cd-resisting Kluyveromyces marxianus YS-K1.Mycosystema,2013,32(5):868-875.(in Chinese)王小波,李学如,茆灿泉,江南屏,郭泰林.耐镉马克思克鲁维酵母重金属镉吸附特性的研究.菌物学报,2013,32(5):868-875.
[19]Liu M,Liu FZ,Liu BF.Determination of available lead and cadmium in soil.Journal of Agro-Environment Science,2007,26(S1):300-302.(in Chinese)刘铭,刘凤枝,刘保锋.土壤中有效态铅和镉的测定.农业环境科学学报,2007,26(S1):300-302.
[20]Fu J,Xie XH,Qian L,Liu JS.Cadmium tolerance and bio-accumulation mechanisms of Ralstonia pickettii strain DX-T3-01.Chinese Journal of Applied&Environmental Biology,2011,17(5):717-721.(in Chinese)付瑾,谢学辉,钱林,柳建设.皮氏罗尔斯通氏菌株DX-T3-01的耐镉性能及镉富集机理.应用与环境生物学报,2011,17(5):717-721.
[21]Li Y,Pang HD,He LY,Wang Q,Sheng XF.Cd immobilization and reduced tissue Cd accumulation of rice(Oryza sativa wuyun-23)in the presence of heavy metal-resistant bacteria.Ecotoxicology and Environment Safety,2017,138:56-63.
[22]Manikandan M,Gopal J,Kumaran RS,Kannan V,Chun S.Purification and characterization of a highly active chromate reductase from endophytic Bacillus sp.DGV19 of Albizzia lebbeck(L.)Benth.actively involved in phytoremediation of tannery effluent-contaminated sites.Preparative Biochemistry Biotechnology,2016,46(2):192-199.
[23]Xia JJ,Sheng XF,Jiang CY.Screening of cadmium-resistant strains and their effects on cadmium activation.Chinese Journal of Ecology,2005,24(11):1357-1360.(in Chinese)夏娟娟,盛下放,江春玉.重金属镉抗性菌株的筛选及其对镉活化作用的研究.生态学杂志,2005,24(11):1357-1360.
[24]Xia BB,Zhong CB,Wei DZ,Cheng J,Wang XB.Biosorption of Zn2+,Cd2+by Bacillus mucilaginosus.Biotechnology,2008,18(3):80-84.(in Chinese)夏彬彬,仲崇斌,魏德州,程静,王秀兵.胶质芽孢杆菌对Zn2+、Cd2+的生物吸附.生物技术,2008,18(3):80-84.
[25]Shen QY,Cao ZQ,Zhu YF,Shi WL.Isolation of a Cd-resistant bacterium and optimization of its bio-accumulation condition.Soils,2016,48(3):615-620.(in Chinese)沈秋悦,曹志强,朱月芳,施维林.一株耐镉细菌的分离鉴定及其吸附条件的优化.土壤,2016,48(3):615-620.
[26]Yan CM,He Z,Ge ZB,Sheng XF,He LY.Comparative study on adsorption characteristics of lead and cadmium by two heavy metal resistant bacteria.Acta Scientiae Circumstantiae,2018,38(9):3597-3604.(in Chinese)燕传明,贺卓,葛占标,盛下放,何琳燕.两株重金属抗性细菌对铅镉吸附特性的比较研究.环境科学学报,2018,38(9):3597-3604.
[27]Sun J,Li H,Liu LG,Zhan PP,Zhang CD,Su XR.Bioaccumulation capacity and mechanisms of cadmium by Bacillus licheniformis.Journal of Nuclear Agricultural Science,2013,27(11):1644-1651.(in Chinese)孙静,李晔,刘联国,詹萍萍,张春丹,苏秀榕.地衣芽孢杆菌富集镉的特性及机理研究.核农学报,2013,27(11):1644-1651.
[28]Li H,Zhang GF,Shi L,Gong CJ.Isolation and identification of cadmium resistance Bacillus cereus SY and its culture conditions optimization.Hubei Agricultural Sciences,2010,49(6):1353-1355.(in Chinese)李辉,张国芳,石璐,巩常军.耐镉蜡状芽孢杆菌SY的筛选鉴定及其培养条件优化.湖北农业科学,2010,49(6):1353-1355.
[29]Xiao HQ,Li YZ.Research progress in the optimization of microbial culture medium.Liquor-making Science&Technology,2010,(1):90-94.(in Chinese)肖怀秋,李玉珍.微生物培养基优化方法研究进展.酿酒科技,2010,(1):90-94.
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[31]Kim SY,Jin MR,Chung CH,Yun YS,Jahng KY,Yu KY.Biosorption of cationic basic dye and cadmium by the novel biosorbent Bacillus catenulatus JB-022 strain.Journal of Bioscience and Bioengineering,2015,119(4):433-439.
[32]Kumar A,Cameotra SS,Gupta S.Screening and characterization of potential cadmium biosorbent Alcaligenes strain from industrial effluent.Journal of Basic Microbiology,2012,52(2):160-166.
[2]Yang QL,Wu ZZ,Chen JL,Liu XG,Wang WH,Liu YW.Research status of phytoremediation of heavy metals contaminated soil and prospects of water and fertilizer regulating technology.Ecology and Environmental Sciences,2015,24(6):1075-1084.(in Chinese)杨启良,武振中,陈金陵,刘小刚,王卫华,刘艳伟.植物修复重金属污染土壤的研究现状及其水肥调控技术展望.生态环境学报,2015,24(6):1075-1084.
[3]Sochor J,Zitka O,Hynek D,Jilkova D,Krejcova L,Trnkova L,Adam V,Hubalek J,KynickyJ,Vrba R,Kizek R.Bio-sensing of cadmium(Ⅱ)ions using Staphylococcus aureus.Sensors,2011,11(11):10638-10663.
[4]El-Habit OH,Moneim AEA.Testing the genotoxicity,cytotoxicity,and oxidative stress of cadmium and nickel and their additive effect in male mice.Biological Trace Element Research,2014,159(1/3):364-372.
[5]Li JH,Li WX,Yin H,Zhang B,Zhu W.Effect of cadmium on TET enzymes and DNA methylation changes in human embryonic kidney cell.Chinese Journal of Preventive Medicine,2015,49(9):822-827.(in Chinese)李金慧,李文学,殷花,张波,朱伟.镉暴露对人胚肾细胞TET酶表达及DNA甲基化水平的影响.中华预防医学杂志,2015,49(9):822-827.
[6]Skowerski M,Jasik K,Konecki J.Effects of interaction between cadmium and selenium on heart metabolism in mice:the study of RNA,protein,ANP synthesis activities and ultrastructure in mouse heart.Medical Science Monitor:International Medical Journal of Experimental&Clinical Research,2000,6(2):258-265.
[7]Wu YM.Patent bibliometric analysis on the remediation techniques of soil heavy metal pollution.Ecology and Environmental Sciences,2013,22(5):901-904.(in Chinese)吴耀楣.中国土壤重金属污染修复技术的专利文献计量分析.生态环境学报,2013,22(5):901-904.
[8]Zhang DX,Wen SL,Huang WB,Cai ZP,Li SH,Qian W,Yuan LH,Yang WJ.Effects of electrode material on the electrokinetic remediation of cadmium-contaminated soil.Guangdong Chemical Industry,2017,44(7):205-207.(in Chinese)张大鑫,温思玲,黄伟斌,蔡宗平,黎森辉,钱伟,苑丽红,杨炜杰.电极材料对电动修复重金属镉污染土壤效果的影响.广东化工,2017,44(7):205-207.
[9]Gao Y,Cheng Q,Zhang MJ,Zhu ZY,Hu TT,Yang Y.Research advance on remediation technology of cadmium contaminated soil.Biotechnology Bulletin,2017,33(10):103-110.(in Chinese)高宇,程潜,张梦君,朱振宇,胡婷婷,杨宇.镉污染土壤修复技术研究.生物技术通报,2017,33(10):103-110.
[10]Ni ZY,Xie GX,Zhang MK.Research progress on remediation technology of cadmium-contaminated agricultural soils.Anhui Agricultural Science Bulletin,2017,23(6):115-120.(in Chinese)倪中应,谢国雄,章明奎.镉污染农田土壤修复技术研究进展.安徽农学通报,2017,23(6):115-120.
[11]Mala JGS,Sujatha D,Rose C.Inducible chromate reductase exhibiting extracellular activity in Bacillus methylotrophicus for chromium bioremediation.Microbiological Research,2015,170:235-241.
[12]Chen JL,Liu XW,Zhang YJ,Fang XH.Identification of four tolerance bacteria and the preliminary study for pollution remediation of heavy metals.Ecology and Environmental Sciences,2014,23(7):1199-1204.(in Chinese)陈佳亮,刘晓文,张雅静,方晓航.耐性细菌的分离鉴定及重金属污染修复初步研究.生态环境学报,2014,23(7):1199-1204.
[13]Li T,Wang J,Wei XL,Zhao HY,Zhao ZX,Liu HB.Identification and characterization of a Bacillus methylotrophicus strain with high flocculating activity.RSCAdvances,2015,5(111):91766-91775.
[14]Sharma P,Sharma N,Pathania S,Handa S.Purification and characterization of lipase by Bacillus methylotrophicus PS3under submerged fermentation and its application in detergent industry.Journal of Genetic Engineering and Biotechnology,2017,15(2):369-377.
[15]Frikha-Gargouri O,Ben Abdallah D,Ghorbel I,Charfeddine I,Jlaiel L,Triki MA,Tounsi S.Lipopeptides from a novel Bacillus methylotrophicus 39b strain suppress Agrobacterium crown gall tumours on tomato plants.Pest Management Science,2016,73(3):568-574.
[16]Jemil N,Ayed H B,Manresa A,Nasri M,Hmidet N.Antioxidant properties,antimicrobial and anti-adhesive activities of DCS1 lipopeptides from Bacillus methylotrophicus DCS1.BMC Microbiology,2017,17:144.
[17]Rajendiran T,Gopakumaran N,Bhuyan U,Kumari S,Ramesh KV.Optimization and immobilization of lipase from Bacillus subterraneus TNUS15:comparison between response surface methodology and artificial neural network.Der Pharma Chemica,2017,9(12):83-91.
[18]Wang XB,Li XR,Mao CQ,Jiang NP,Guo TL.The characteristics of cadmium adsorption by Cd-resisting Kluyveromyces marxianus YS-K1.Mycosystema,2013,32(5):868-875.(in Chinese)王小波,李学如,茆灿泉,江南屏,郭泰林.耐镉马克思克鲁维酵母重金属镉吸附特性的研究.菌物学报,2013,32(5):868-875.
[19]Liu M,Liu FZ,Liu BF.Determination of available lead and cadmium in soil.Journal of Agro-Environment Science,2007,26(S1):300-302.(in Chinese)刘铭,刘凤枝,刘保锋.土壤中有效态铅和镉的测定.农业环境科学学报,2007,26(S1):300-302.
[20]Fu J,Xie XH,Qian L,Liu JS.Cadmium tolerance and bio-accumulation mechanisms of Ralstonia pickettii strain DX-T3-01.Chinese Journal of Applied&Environmental Biology,2011,17(5):717-721.(in Chinese)付瑾,谢学辉,钱林,柳建设.皮氏罗尔斯通氏菌株DX-T3-01的耐镉性能及镉富集机理.应用与环境生物学报,2011,17(5):717-721.
[21]Li Y,Pang HD,He LY,Wang Q,Sheng XF.Cd immobilization and reduced tissue Cd accumulation of rice(Oryza sativa wuyun-23)in the presence of heavy metal-resistant bacteria.Ecotoxicology and Environment Safety,2017,138:56-63.
[22]Manikandan M,Gopal J,Kumaran RS,Kannan V,Chun S.Purification and characterization of a highly active chromate reductase from endophytic Bacillus sp.DGV19 of Albizzia lebbeck(L.)Benth.actively involved in phytoremediation of tannery effluent-contaminated sites.Preparative Biochemistry Biotechnology,2016,46(2):192-199.
[23]Xia JJ,Sheng XF,Jiang CY.Screening of cadmium-resistant strains and their effects on cadmium activation.Chinese Journal of Ecology,2005,24(11):1357-1360.(in Chinese)夏娟娟,盛下放,江春玉.重金属镉抗性菌株的筛选及其对镉活化作用的研究.生态学杂志,2005,24(11):1357-1360.
[24]Xia BB,Zhong CB,Wei DZ,Cheng J,Wang XB.Biosorption of Zn2+,Cd2+by Bacillus mucilaginosus.Biotechnology,2008,18(3):80-84.(in Chinese)夏彬彬,仲崇斌,魏德州,程静,王秀兵.胶质芽孢杆菌对Zn2+、Cd2+的生物吸附.生物技术,2008,18(3):80-84.
[25]Shen QY,Cao ZQ,Zhu YF,Shi WL.Isolation of a Cd-resistant bacterium and optimization of its bio-accumulation condition.Soils,2016,48(3):615-620.(in Chinese)沈秋悦,曹志强,朱月芳,施维林.一株耐镉细菌的分离鉴定及其吸附条件的优化.土壤,2016,48(3):615-620.
[26]Yan CM,He Z,Ge ZB,Sheng XF,He LY.Comparative study on adsorption characteristics of lead and cadmium by two heavy metal resistant bacteria.Acta Scientiae Circumstantiae,2018,38(9):3597-3604.(in Chinese)燕传明,贺卓,葛占标,盛下放,何琳燕.两株重金属抗性细菌对铅镉吸附特性的比较研究.环境科学学报,2018,38(9):3597-3604.
[27]Sun J,Li H,Liu LG,Zhan PP,Zhang CD,Su XR.Bioaccumulation capacity and mechanisms of cadmium by Bacillus licheniformis.Journal of Nuclear Agricultural Science,2013,27(11):1644-1651.(in Chinese)孙静,李晔,刘联国,詹萍萍,张春丹,苏秀榕.地衣芽孢杆菌富集镉的特性及机理研究.核农学报,2013,27(11):1644-1651.
[28]Li H,Zhang GF,Shi L,Gong CJ.Isolation and identification of cadmium resistance Bacillus cereus SY and its culture conditions optimization.Hubei Agricultural Sciences,2010,49(6):1353-1355.(in Chinese)李辉,张国芳,石璐,巩常军.耐镉蜡状芽孢杆菌SY的筛选鉴定及其培养条件优化.湖北农业科学,2010,49(6):1353-1355.
[29]Xiao HQ,Li YZ.Research progress in the optimization of microbial culture medium.Liquor-making Science&Technology,2010,(1):90-94.(in Chinese)肖怀秋,李玉珍.微生物培养基优化方法研究进展.酿酒科技,2010,(1):90-94.
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