不同环境因子和碳氮源对短小芽孢杆菌BP-171无机氮降解特性的影响
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摘要
本文研究了不同环境因子和碳氮源对分离自海水养殖环境的一株短小芽孢杆菌(Bacillus pumilus,编号BP-171)无机氮降解特性的影响。环境因子设置了C∶N(质量比)、盐度和pH三种,碳源设为乙酸钠、丁二酸钠、柠檬酸钠、蔗糖、乳糖和葡萄糖,氮源设置为氨态氮、亚硝酸态氮、硝酸态氮及三者混合氮源(1∶1∶1)。研究表明,在温度28℃下,菌株BP-171在碳源为葡萄糖、C/N=18、盐度20~40和pH=7.0~8.0条件下对降解液中无机氮有较好的去除作用。在上述脱氮条件下,进一步研究3种单一氮源和1种混合氮源对菌株BP-171无机氮降解特性的影响。研究表明,当氮源为单一的氨态氮、亚硝酸态氮、硝酸态氮时,5天后培养液中氨态氮、亚硝酸态氮、硝酸态氮的去除率最高可分别达74.49%、53.61%和83.39%,总氮则分别降低了2.91%、23.89%和30.98%;在以三者混合作为氮源时,降解液中氨态氮、亚硝酸态氮、硝酸态氮的最高去除率分别为97.89%、78.73%和46.57%,总氮则降低了15.14%。不同条件下培养液中不同形态氮含量的变化情况表明,菌株BP-171对无机氮的转化和去除包括菌体细胞的同化作用、异养硝化作用和好氧反硝化作用等生物学过程,不同条件下具体机制有所差异。
In this study,the effects of environmental factors and different nitrogen source on the inorganic nitrogen removing characteristics of Bacillus pumilus BP-171 were described.The environmental factors were set as C∶N(mass ratio),salinity and pH while carbon sources were set as sodium acetate,sodium succinate,sodiumcitric acid,sucrose,lactose and glucose,and nitrogen sourceswere set as ammonia nitrogen,nitrite nitrogen,nitrate nitrogen,and mixed nitrogen(1∶1∶1 of the former three).The results showed that the optimal experimental conditions for strain BP-171 to have the best inorganic nitrogen removal performance were glucose as the carbon source,the C∶N ratio 18,the salinity 20~40 and pH=7.0~8.0.Under the optimal nitrogen removal performance conditions,the sole nitrogen source and the mixed nitrogen source were used as basic solution to study the effects of different nitrogen source on the inorganic nitrogen removing characteristics of strain BP-171.In the experiment of the sole nitrogen,the highest removal rate of NH+4-N,NO-2-N and NO-3 N reached 74.49%,53.61%and83.39%,respectively,and the total nitrogen were reduced by 2.91%,23.89%and 30.98%,respectively.In the experiment of the mixed nitrogen,the highest removal rate of NH+4-N,NO-2-N and NO-3 N reached 97.89%,78.73% and 46.57%,respectively,and the total nitrogen was reduced by15.14%.Overall,strain BP-171 can use different nitrogen sources in assimilation,heterotrophic nitrification and aerobic denitrification,but the specific mechanism under different conditions may be different.
In this study,the effects of environmental factors and different nitrogen source on the inorganic nitrogen removing characteristics of Bacillus pumilus BP-171 were described.The environmental factors were set as C∶N(mass ratio),salinity and pH while carbon sources were set as sodium acetate,sodium succinate,sodiumcitric acid,sucrose,lactose and glucose,and nitrogen sourceswere set as ammonia nitrogen,nitrite nitrogen,nitrate nitrogen,and mixed nitrogen(1∶1∶1 of the former three).The results showed that the optimal experimental conditions for strain BP-171 to have the best inorganic nitrogen removal performance were glucose as the carbon source,the C∶N ratio 18,the salinity 20~40 and pH=7.0~8.0.Under the optimal nitrogen removal performance conditions,the sole nitrogen source and the mixed nitrogen source were used as basic solution to study the effects of different nitrogen source on the inorganic nitrogen removing characteristics of strain BP-171.In the experiment of the sole nitrogen,the highest removal rate of NH+4-N,NO-2-N and NO-3 N reached 74.49%,53.61%and83.39%,respectively,and the total nitrogen were reduced by 2.91%,23.89%and 30.98%,respectively.In the experiment of the mixed nitrogen,the highest removal rate of NH+4-N,NO-2-N and NO-3 N reached 97.89%,78.73% and 46.57%,respectively,and the total nitrogen was reduced by15.14%.Overall,strain BP-171 can use different nitrogen sources in assimilation,heterotrophic nitrification and aerobic denitrification,but the specific mechanism under different conditions may be different.
引文
[1]吴建江,王兆阳,许培雅.一株高效异养硝化菌的分离、鉴定及其氨氮去除特性[J].中国环境科学,2013,33(7):1309-1315.Wu J J,Wang Z Y,Xu P Y.Isolation and identification of an efficiently heterotrophic nitrifier,and its removal characterization of ammonia nitrogen[J].China Environmental Science,2013,33(7):1309-1315.
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[5]孙雪梅,李秋芬,张艳,等.一株海水异养硝化-好氧反硝化菌系统发育及脱氮特性[J].微生物学报,2012,52(6):687-695.Sun X M,Li Q F,Zhang Y,et al.Phylogenetic analysis and nitrogen removal characteristicsof a heterotrophic nitrifying-aerobic denitrifying bacteriastrain from marine environment[J].Acta Microbiologica Sinica,2012,52(6):687-695.
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[7]王红卫,赵新鲜,张晶.紫外分光光度计测定水中硝酸盐氮方法的探讨[J].水利技术监督,2007(4):4-5.Wang H W,Zhao X X,Zhang J.Determine of the nitrate nitrogen in water by ultraviolet speetrophotometry[J].Water Conservancy and Technical Supervision,2007(4):4-5.
[8]薛爱群,贾锋,齐顺章.细菌总蛋白含量测定方法的改进[J].微生物学通报,1994(1):58-59.Xue A Q,J,Q.Improvement of determination method for baeterial total proteins[J].Microbiology China,1994(1):58-59.
[9]李秋芬,有小娟,张艳,等.象山港中部养殖区细菌群落结构的特征及其在生境修复过程中的变化[J].中国水产科学,2013,20(6):1234-1246.Li Q F,You X J,Zhang Y,et al.Changes in the characteristics of the bacterial community structureduring habitat restoration of aquaculture sites in Xiangshan Bay[J].Journalof Fishery Sciences ofChina,2013,20(6):1234-1246.
[10]李卫芬,傅罗琴,邓斌,等.1株好氧反硝化菌的分离鉴定及反硝化特性研究[J].环境科学,2011,32(8):2403-2408.Li W F,Fu L Q,Deng B,et al.Identification and denitrification characteristics of an aerobic denitrifier[J].Environmental Science,2011,32(8):2403-2408.
[11]姜磊,徐成斌,马溪平,等.1株好氧反硝化菌的分离鉴定和反硝化特性研究[J].环境科学与技术,2013,36(3):12-15.Jiang L,Xu C B,Ma X P,et al.Identification and denitrification characteristics of an aerobic denitirfier[J].Environmental Science&Technology,2013,36(3):12-15.
[12]陈玲,白洁,赵阳国,等.分离于河口区芦苇湿地1株好氧反硝化菌的鉴定及其反硝化特性[J].微生物学报,2016,56(8):1314-1325.Chen L,Bai J,Zhao Y G,etal.Identification and denitrification characteristics of an aerobicdenitrifier in estuary phragmites wetland[J].Acta Microbiologica Sinica,2016,56(8):1314-1325.
[13]李兵,林炜铁.1株好氧反硝化芽孢杆菌的脱氮特性研究[J].水生态学杂志,2009,30(3):48-52.Li B,Lin W T.Study on aerobic denitrification characteristics of bacillus strain D5[J].Journal of Hydroecology,2009,30(3):48-52.
[14]安健,宋增福,杨先乐,等.好氧反硝化细菌YX-6特性及鉴定分析[J].中国水产科学,2010,17(3):561-569.An J,Song Z F,Yang X L,et al.Characteristics of aerobic denitrifying strain YX-6and identification[J].Journal of Fishery Sciences of China,2010,17(3):561-569.
[15]鲜思淑,陈茂霞,熊蓉,等.异养硝化-好氧反硝化影响因素研究进展[J].水处理技术,2016,42(1):1-6.Xian S S,Chen M X,Xiong R,et al.Research advances of heterotrophic nitrification-aerobic denitrification under different factors[J].Technology of Water Treatment,2016(1):1-6.
[16]席星林,潘玉洁.基于环境因子的异养硝化菌Providenciarettgeristrain YL脱氮性能研究[J].环境工程,2014,32(S1):144-147Xi X L,Pan Y J.The denitrification performance study of heterotrophic nitrifying bacteria Providencia rettgeri strain YL based on environmental factors[J].Environmental Engineering,2014,32(S1):144-147.
[17]宋宇杰,李屹,刘玉香,等.碳源和氮源对异养硝化好氧反硝化菌株Y1脱氮性能的影响[J].环境科学学报,2013,33(9):2491-2497.Song YJ,Li Y,Liu YX,et al.Effect of carbon and nitrogen sources on nitrogen removal by a heterotrophicnitrification-aerobic denitrificationstrain Y1[J].Acta Scientiae Circumstantiae,2013,33(9):2491-2497.
[18]陈月,郭亮,孙美.好氧颗粒污泥对污泥碳源的反硝化出水处理效果[J].中国海洋大学学报(自然科学版),2017,47(9):104-109.Chen Y,Guo L,Sun M.Study on effect of aerobic granules treatment on denitrification effluent used sludge hy-drolysate as carbon sources[J].Periodical of Ocean University of China,2017,47(9):104-109.
[19] A B Gupta.Thiosphaerapantotropha:A sulphur bacterium capable of simultaneous heterotrophic nitrification and aerobic denitrification[J].Enzyme and Microbial Technology,1997,21(8):589-595.
[20]王弘宇,马放,苏俊峰,等.好氧反硝化菌株的鉴定及其反硝化特性研究[J].环境科学,2007(7):1548-1552.Wang H Y,Ma F,Su J F,et al.Identification and characterization of a bacterial strain C3capable of aerobic denitrification[J].Environmental Science,2007(7):1548-1552.
[21]廖绍安,郑桂丽,王安利,等.养虾池好氧反硝化细菌新菌株的分离鉴定及特征[J].生态学报,2006(11):3718-3724.Liao S A,Zheng G L,Wang A L,et al.Isolation and characterization of a novel aerobic denitrifier from shrimp pond[J].Acta Ecologica Sinica,2006(11):3718-3724.
[22]张彦灼,李军,陈光辉,等.NaCl对好氧颗粒污泥短程硝化反硝化的影响[J].环境科学研究,2015,28(5):823-830.Zhang YZ,Li J,Chen GH,et al.Effects of NaCl concentration on partial nitrification and denitrification by aerobic granular sludge[J].Research of Environmental Sciences,2015,28(5):823-830.
[23]孙庆花,于德爽,张培玉,等.1株海洋异养硝化-好氧反硝化菌的分离鉴定及其脱氮特性[J].环境科学,2016,37(2):647-654.Sun Q H,Yu D S,Zhang P Y,et al.Identification and nitrogen removal characteristics of a heterotrophic nitrification-aerobic denitrification strainIsolated from marine environment[J].Environmental Science,2016,37(2):647-654.
[24] Qing Ling Zhang,YingLiu,Guo Min Ai,et al.The characteristics of a novel heterotrophic nitrification–aerobic denitrification bacterium,Bacillus methylotrophicus strain L7[J].Bioresource Technology,2012,108:35-44.
[25] Soumesh Kumar Padhi,Nikhil Kumar Maiti.Molecular insight into the dynamic central metabolic pathways of Achromobacter xylosoxidans CF-S36 during heterotrophic nitrogen removal processes[J].Journal of Bioscience and Bioengineering,2017,123(1):46-55.
[26] Hai-Yan Zheng,YingLiu,Xi-Yan Gao,et al.Characterization of a marine origin aerobic nitrifying-denitrifying bacterium[J].Journal of Bioscience and Bioengineering,2012,114(1):33-37.
[27]梁贤,任勇翔,杨垒,等.异养硝化-好氧反硝化菌YL的脱氮特性[J].环境科学,2015,36(5):1749-1756.Liang X,Ren Y X,Yang L,et al.Characteristics of nitrogen removal by a heterotrophic nitrification-aerobic denitrification bacterium YL[J].Environmental Science,2015,36(5):1749-1756.
[28] Richardson D J,Wehrfritz J M,Keech A.The diversity of redox proteins involved in bacterial heterotrophic nitrification and aerobic denitrification[J].Biochemical Society Transactions,1998,26(3):401-408.
[29] Stewart V.Nitrate respiration in relation to facultative metabolism in enterobacteria[J].Microbiological Reviews,1988,52(2):190-232.
[30]闫法军,田相利,董双林,等.一株低温降解菌的分离鉴定及其降解特性研究[J].中国海洋大学学报(自然科学版),2011,41(3):24-30.Yan F J,Tian X L,Dong S L,et al.Isolation,Identification and characterization of a low-temperature degrading bacterium[J].Periodical of Ocean University of China,2011,41(3):24-30.
[2]曾庆梅,司文攻,李志强,等.一株高效异养硝化菌的选育、鉴定及其硝化条件[J].微生物学报,2010,50(6):803-810.Zeng Q M,Si W G,Li Z Q,et al.Isolation,identification and characterization of a high efficient heterotrophic nitrifier[J].Acta Microbiologica Sinica,2010,50(6):803-810.
[3] Ahn Y H.Sustainable nitrogen elimination biotechnologies:A review[J].Process Biochemistrry,2006,41(8):1709-1721.
[4]何霞,赵彬,吕剑,等.异养硝化细菌Bacillus sp.LY脱氮性能研究[J].环境科学,2007,28(6):1404-1408.He X,Zhao B,He Y L,et al.Nitrogen removal by Bacillus sp.LY with heterotrophic nitrification ability[J].Environmental Science,2007,28(6):1404-1408.
[5]孙雪梅,李秋芬,张艳,等.一株海水异养硝化-好氧反硝化菌系统发育及脱氮特性[J].微生物学报,2012,52(6):687-695.Sun X M,Li Q F,Zhang Y,et al.Phylogenetic analysis and nitrogen removal characteristicsof a heterotrophic nitrifying-aerobic denitrifying bacteriastrain from marine environment[J].Acta Microbiologica Sinica,2012,52(6):687-695.
[6]雷衍之.养殖水环境化学实验[M].北京:中国农业出版社,2006:31-35.Lei Y Z.The Chemical Experiment of Water in Aquaculture Environment[M].Beijing:China Agriculture Press,2006:31-35.
[7]王红卫,赵新鲜,张晶.紫外分光光度计测定水中硝酸盐氮方法的探讨[J].水利技术监督,2007(4):4-5.Wang H W,Zhao X X,Zhang J.Determine of the nitrate nitrogen in water by ultraviolet speetrophotometry[J].Water Conservancy and Technical Supervision,2007(4):4-5.
[8]薛爱群,贾锋,齐顺章.细菌总蛋白含量测定方法的改进[J].微生物学通报,1994(1):58-59.Xue A Q,J,Q.Improvement of determination method for baeterial total proteins[J].Microbiology China,1994(1):58-59.
[9]李秋芬,有小娟,张艳,等.象山港中部养殖区细菌群落结构的特征及其在生境修复过程中的变化[J].中国水产科学,2013,20(6):1234-1246.Li Q F,You X J,Zhang Y,et al.Changes in the characteristics of the bacterial community structureduring habitat restoration of aquaculture sites in Xiangshan Bay[J].Journalof Fishery Sciences ofChina,2013,20(6):1234-1246.
[10]李卫芬,傅罗琴,邓斌,等.1株好氧反硝化菌的分离鉴定及反硝化特性研究[J].环境科学,2011,32(8):2403-2408.Li W F,Fu L Q,Deng B,et al.Identification and denitrification characteristics of an aerobic denitrifier[J].Environmental Science,2011,32(8):2403-2408.
[11]姜磊,徐成斌,马溪平,等.1株好氧反硝化菌的分离鉴定和反硝化特性研究[J].环境科学与技术,2013,36(3):12-15.Jiang L,Xu C B,Ma X P,et al.Identification and denitrification characteristics of an aerobic denitirfier[J].Environmental Science&Technology,2013,36(3):12-15.
[12]陈玲,白洁,赵阳国,等.分离于河口区芦苇湿地1株好氧反硝化菌的鉴定及其反硝化特性[J].微生物学报,2016,56(8):1314-1325.Chen L,Bai J,Zhao Y G,etal.Identification and denitrification characteristics of an aerobicdenitrifier in estuary phragmites wetland[J].Acta Microbiologica Sinica,2016,56(8):1314-1325.
[13]李兵,林炜铁.1株好氧反硝化芽孢杆菌的脱氮特性研究[J].水生态学杂志,2009,30(3):48-52.Li B,Lin W T.Study on aerobic denitrification characteristics of bacillus strain D5[J].Journal of Hydroecology,2009,30(3):48-52.
[14]安健,宋增福,杨先乐,等.好氧反硝化细菌YX-6特性及鉴定分析[J].中国水产科学,2010,17(3):561-569.An J,Song Z F,Yang X L,et al.Characteristics of aerobic denitrifying strain YX-6and identification[J].Journal of Fishery Sciences of China,2010,17(3):561-569.
[15]鲜思淑,陈茂霞,熊蓉,等.异养硝化-好氧反硝化影响因素研究进展[J].水处理技术,2016,42(1):1-6.Xian S S,Chen M X,Xiong R,et al.Research advances of heterotrophic nitrification-aerobic denitrification under different factors[J].Technology of Water Treatment,2016(1):1-6.
[16]席星林,潘玉洁.基于环境因子的异养硝化菌Providenciarettgeristrain YL脱氮性能研究[J].环境工程,2014,32(S1):144-147Xi X L,Pan Y J.The denitrification performance study of heterotrophic nitrifying bacteria Providencia rettgeri strain YL based on environmental factors[J].Environmental Engineering,2014,32(S1):144-147.
[17]宋宇杰,李屹,刘玉香,等.碳源和氮源对异养硝化好氧反硝化菌株Y1脱氮性能的影响[J].环境科学学报,2013,33(9):2491-2497.Song YJ,Li Y,Liu YX,et al.Effect of carbon and nitrogen sources on nitrogen removal by a heterotrophicnitrification-aerobic denitrificationstrain Y1[J].Acta Scientiae Circumstantiae,2013,33(9):2491-2497.
[18]陈月,郭亮,孙美.好氧颗粒污泥对污泥碳源的反硝化出水处理效果[J].中国海洋大学学报(自然科学版),2017,47(9):104-109.Chen Y,Guo L,Sun M.Study on effect of aerobic granules treatment on denitrification effluent used sludge hy-drolysate as carbon sources[J].Periodical of Ocean University of China,2017,47(9):104-109.
[19] A B Gupta.Thiosphaerapantotropha:A sulphur bacterium capable of simultaneous heterotrophic nitrification and aerobic denitrification[J].Enzyme and Microbial Technology,1997,21(8):589-595.
[20]王弘宇,马放,苏俊峰,等.好氧反硝化菌株的鉴定及其反硝化特性研究[J].环境科学,2007(7):1548-1552.Wang H Y,Ma F,Su J F,et al.Identification and characterization of a bacterial strain C3capable of aerobic denitrification[J].Environmental Science,2007(7):1548-1552.
[21]廖绍安,郑桂丽,王安利,等.养虾池好氧反硝化细菌新菌株的分离鉴定及特征[J].生态学报,2006(11):3718-3724.Liao S A,Zheng G L,Wang A L,et al.Isolation and characterization of a novel aerobic denitrifier from shrimp pond[J].Acta Ecologica Sinica,2006(11):3718-3724.
[22]张彦灼,李军,陈光辉,等.NaCl对好氧颗粒污泥短程硝化反硝化的影响[J].环境科学研究,2015,28(5):823-830.Zhang YZ,Li J,Chen GH,et al.Effects of NaCl concentration on partial nitrification and denitrification by aerobic granular sludge[J].Research of Environmental Sciences,2015,28(5):823-830.
[23]孙庆花,于德爽,张培玉,等.1株海洋异养硝化-好氧反硝化菌的分离鉴定及其脱氮特性[J].环境科学,2016,37(2):647-654.Sun Q H,Yu D S,Zhang P Y,et al.Identification and nitrogen removal characteristics of a heterotrophic nitrification-aerobic denitrification strainIsolated from marine environment[J].Environmental Science,2016,37(2):647-654.
[24] Qing Ling Zhang,YingLiu,Guo Min Ai,et al.The characteristics of a novel heterotrophic nitrification–aerobic denitrification bacterium,Bacillus methylotrophicus strain L7[J].Bioresource Technology,2012,108:35-44.
[25] Soumesh Kumar Padhi,Nikhil Kumar Maiti.Molecular insight into the dynamic central metabolic pathways of Achromobacter xylosoxidans CF-S36 during heterotrophic nitrogen removal processes[J].Journal of Bioscience and Bioengineering,2017,123(1):46-55.
[26] Hai-Yan Zheng,YingLiu,Xi-Yan Gao,et al.Characterization of a marine origin aerobic nitrifying-denitrifying bacterium[J].Journal of Bioscience and Bioengineering,2012,114(1):33-37.
[27]梁贤,任勇翔,杨垒,等.异养硝化-好氧反硝化菌YL的脱氮特性[J].环境科学,2015,36(5):1749-1756.Liang X,Ren Y X,Yang L,et al.Characteristics of nitrogen removal by a heterotrophic nitrification-aerobic denitrification bacterium YL[J].Environmental Science,2015,36(5):1749-1756.
[28] Richardson D J,Wehrfritz J M,Keech A.The diversity of redox proteins involved in bacterial heterotrophic nitrification and aerobic denitrification[J].Biochemical Society Transactions,1998,26(3):401-408.
[29] Stewart V.Nitrate respiration in relation to facultative metabolism in enterobacteria[J].Microbiological Reviews,1988,52(2):190-232.
[30]闫法军,田相利,董双林,等.一株低温降解菌的分离鉴定及其降解特性研究[J].中国海洋大学学报(自然科学版),2011,41(3):24-30.Yan F J,Tian X L,Dong S L,et al.Isolation,Identification and characterization of a low-temperature degrading bacterium[J].Periodical of Ocean University of China,2011,41(3):24-30.