针对同步脱氮除磷工艺流程复杂、基建与运行费用高等问题,从活性污泥中分离出一株兼具同步脱氮除磷能力的异养硝化细菌NP3,经16S rRNA鉴定为施氏假单胞菌(Pseudomonas stutzeri),并对其脱氮除磷特性及作用机制进行研究.结果表明,在好氧条件下,菌株NP3能够以氨氮、硝酸盐和亚硝酸盐作为单一氮源进行脱氮除磷,反应过程中间产物积累较少,氮、磷主要以同化作用去除,其生长和代谢速率NH4+-N>NO2−-N>NO3−-N.在碳源为柠檬酸钠、C/N为10、温度为30℃、pH值为7、转速为160r/min的最佳生长条件下,该菌株最大氨氮和磷酸盐去除率均接近100%.同时,反硝化和聚磷功能基因(nosZ、nirS、ppk)的成功扩增,进一步证明菌株NP3具有同步脱氮除磷能力.此外,胞外聚合物(EPS)在磷去除中起到重要作用,去除占比达到45%以上.X射线光电子能谱(XPS)分析证实EPS的表面官能团可以吸附C-PO3/P-O、PO43-/HPO42-等不同形式的磷,充当磷转移站.31P核磁共振(NMR)结果进一步表明EPS对磷的赋存形态存在较大影响,其中焦磷酸盐是EPS存在时的主要磷物种,而正磷酸盐和磷酸二酯是提取EPS后的主要磷形式.
Abstract
In order to address the issues of complex processes and high infrastructure and operational costs in simultaneous nitrogen and phosphorus removal, a heterotrophic nitrifying strain NP3 exhibiting simultaneous nitrogen and phosphorus removal capabilities was isolated from activated sludge in this study. Strain NP3 was identified as Pseudomonas stutzeri by 16S rRNA sequence analysis, and its nitrogen and phosphorus removal characteristics and mechanisms were investigated. It was showed that strain NP3 was able to utilize ammonium, nitrate, and nitrite as the sole nitrogen source for efficient nitrogen and phosphorus removal under aerobic conditions. The accumulation of intermediate products during the reaction process was minimal, and nitrogen and phosphorus were primarily removed through assimilation. The growth and metabolic rates followed the order: NH4+-N >NO2−-N >NO3−-N. Under the optimal growth conditions of sodium citrate as the carbon source, C/N=10, T=30℃, pH =7, and r=160r/min, the maximum removal rates of ammonia nitrogen and phosphate were almost 100%. Furthermore, successful amplification of denitrification and polyphosphate genes (nosZ, nirS, ppk) further confirmed the simultaneous nitrogen and phosphorus removal capability of strain NP3. X-ray Photoelectron Spectroscopy (XPS) analysis demonstrated that the functional groups on the extracellular polymeric substances (EPS) surface could adsorb different forms of phosphorus such as C-PO3/P-C, PO43-/HPO42-, acting as phosphorus transfer stations.31P nuclear magnetic resonance (NMR) results further indicated that there was a large effect of EPS on phosphorus fugitive morphology, with pyrophosphate being the main phosphorus species in the presence of EPS, whereas orthophosphate and orthophosphate diester were the major phosphorus forms after EPS extraction.
关键词
胞外聚合物 /
好氧反硝化 /
同步脱氮除磷 /
异养硝化
Key words
aerobic denitrification /
extracellular polymeric substances /
heterotrophic nitrification /
simultaneous nitrogen and phosphorus removal
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参考文献
[1] Xu N, Liao M, Liang Y, et al. Biological nitrogen removal capability and pathways analysis of a novel low C/N ratio heterotrophic nitrifying and aerobic denitrifying bacterium (Bacillus thuringiensis strain WXN-23)[J]. Environmental Research, 2021,195:110797.
[2] 刘妍,袁林江,陈希,等.一株异养硝化-好氧反硝化-积累磷的细菌Klebsiella pneumoniae A15的筛出及其特性研究[J].中国环境科学, 2024,44(4):2286-2296. Liu Y, Yuan L J, Chen X, et al. Sieving and characterization of a heterotrophic nitrifying-aerobic denitrifying-phosphorus-accumulating bacterial strain, Klebsiella pneumoniae A15[J]. China Environmental Science, 2024,44(4):2286-2296.
[3] 李艳丽,杨垒,张志昊,等.好氧反硝化细菌Burkholderia sp. ZH8的脱氮特性与生物强化作用[J].中国环境科学, 2024,44(8):4282-4291. Li Y L, Yang L, Zhang Z H, et al. Nitrogen removal characteristics and bioenhancement effects of aerobic denitrifying bacteria Burkholderia sp. ZH8[J]. China Environmental Science, 2024,44(8):4282-429.
[4] Li B, Jing F, Wu D, et al. Simultaneous removal of nitrogen and phosphorus by a novel aerobic denitrifying phosphorus-accumulating bacterium, Pseudomonas stutzeri ADP-19[J]. Bioresource Technology, 2021,321:124445.
[5] Huang M, CuiY, Huang J, et al. A novel Pseudomonas aeruginosa strain performs simultaneous heterotrophic nitrification-aerobic denitrification and aerobic phosphate removal[J]. Water Research, 2022,221:118823.
[6] Zhang M, Pan L, Su C, et al. Simultaneous aerobic removal of phosphorus and nitrogen by a novel salt-tolerant phosphate-accumulating organism and the application potential in treatment of domestic sewage and aquaculture sewage[J]. Science of the Total Environment, 2021,758:143580.
[7] Sun S, Han J, Hu M, et al. Removal of phosphorus from wastewater by Diutina rugosa BL3:Efficiency and pathway[J]. Science of the Total Environment, 2021,801:149751.
[8] Tan X, Gao W, Duan Z, et al. Synthesis of novel algal extracellular polymeric substances (EPS)-based hydrogels for the efficient removal and recovery of phosphorus from contaminated waters:Development, characterisation, and performance[J]. Journal of Environmental Chemical Engineering, 2023,11:109044.
[9] Zhang J, Wu P, Hao B, et al. Heterotrophic nitrification and aerobic denitrification by the bacterium Pseudomonas stutzeri YZN-001[J]. Bioresource Technology, 2011,102(21):9866-9869.
[10] Duan J, Fang H, Su B, et al. Characterization of a halophilic heterotrophic nitrification-aerobic denitrification bacterium and its application on treatment of saline wastewater[J]. Bioresource Technology, 2015,179:421-428.
[11] Jiang L, Wang M, Wang Y, et al. The condition optimization and mechanism of aerobic phosphorus removal by marine bacterium Shewanella sp[J]. Chemical Engineering Journal, 2018,345:611-620.
[12] Yang L, Wang X, Cui S, et al. Simultaneous removal of nitrogen and phosphorous by heterotrophic nitrification-aerobic denitrification of a metal resistant bacterium Pseudomonas putida strain NP5[J]. Bioresource Technology, 2019,285:121360.
[13] Yang L, Lu H, Wang Y, et al. Nitrogen removal characteristics and Cr (VI) tolerance mechanisms of heterotrophic nitrifying bacterium Pseudomonas putida strain LX1[J]. Journal of Water Process Engineering, 2024,64:105647.
[14] He T, Li Z, Sun Q, et al. Heterotrophic nitrification and aerobic denitrification by Pseudomonas tolaasii Y-11 without nitrite accumulation during nitrogen conversion[J]. Bioresource Technology, 2016,200:493-499.
[15] Chen S, He S, Wu C, et al. Characteristics of heterotrophic nitrification and aerobic denitrification bacterium Acinetobacter sp. T1 and its application for pig farm wastewater treatment[J]. Journal of Bioscience and Bioengineering, 2019,127(2):201-205.
[16] Cui Y, Cui Y W, Huang J L. A novel halophilic Exiguobacterium mexicanum strain removes nitrogen from saline wastewater via heterotrophic nitrification and aerobic denitrification[J]. Bioresource Technology, 2021,333:125189.
[17] Wang X, Wang W, Zhang Y, et al. Simultaneous nitrification and denitrification by a novel isolated Pseudomonas sp. JQ-H3 using polycaprolactone as carbon source[J]. Bioresource Technology, 2019, 288:121506.
[18] Li Y, Wang Y, Fu L, et al. Aerobic-heterotrophic nitrogen removal through nitrate reduction and ammonium assimilation by marine bacterium Vibrio sp. Y1-5[J]. Bioresource Technology, 2017,230:103-111.
[19] Wan C, Yang X, Lee D J, et al. Aerobic denitrification by novel isolated strain using NO2--N as nitrogen source[J]. Bioresource Technology 2011,102(15):7244-7248.
[20] Zhang Q L, Liu Y, Ai G M, et al. The characteristics of a novel heterotrophic nitrification-aerobic denitrification bacterium, Bacillus methylotrophicus strain L7[J]. Bioresource Technology, 2012,108:35-44.
[21] Zhou Y, Oehmen A, Lim M, et al. The role of nitrite and free nitrous acid (FNA) in wastewater treatment plants[J]. Water Research, 2011, 45(15):4672-4682.
[22] Chen J, Gu S, Hao H, et al. Characteristics and metabolic pathway of Alcaligenes sp. TB for simultaneous heterotrophic nitrification-aerobic denitrification[J]. Applied Microbiology and Biotechnology, 2016,100(22):9787-9794.
[23] Wei B, Luo X, Ma W, et al. Biological nitrogen removal and metabolic characteristics of a novel cold-resistant heterotrophic nitrification and aerobic denitrification Rhizobium sp. WS7[J]. Bioresource Technology, 2022,362:127756.
[24] Xia L, Li X, Fan W, et al. Heterotrophic nitrification and aerobic denitrification by a novel Acinetobacter sp. ND7 isolated from municipal activated sludge[J]. Bioresource Technology, 2020,301:122749.
[25] Wu L, Ding X, Lin Y, et al. Nitrogen removal by a novel heterotrophic nitrification and aerobic denitrification bacterium Acinetobacter calcoaceticus TY1 under low temperatures[J]. Bioresource Technology, 2022,353:127148.
[26] Fan Y, Su J, Zheng Z, et al. Denitrification performance and mechanism of a novel isolated Acinetobacter sp. FYF8 in oligotrophic ecosystem[J]. Bioresource Technology, 2021,320:124280.
[27] Carneiro Fidélis Silva L, Santiago Lima H, Antônio de Oliveira Mendes T, et al. Heterotrophic nitrifying/aerobic denitrifying bacteria:Ammonium removal under different physical-chemical conditions and molecular characterization[J]. Journal of Environmental Management, 2019,248:109294.
[28] Ma F, Sun Y, Li A. et al. Activation of accumulated nitrite reduction by immobilized Pseudomonas stutzeri T13 during aerobic denitrification[J]. Bioresource Technology, 2015,187:30-36.
[29] Rout P R, Bhunia P, Dash R R. Simultaneous removal of nitrogen and phosphorous from domestic wastewater using Bacillus cereus GS-5 strain exhibiting heterotrophic nitrification, aerobic denitrification and denitrifying phosphorous removal[J]. Bioresource Technology 2017,244:484-495.
[30] Wu C, Chen Y, Qian Z, et al. The effect of extracellular polymeric substances (EPS) of iron-oxidizing bacteria (Ochrobactrum EEELCW01) on mineral transformation and arsenic (As) fate[J]. Journal of Environmental Sciences, 2023,130:187-196.
[31] Hou R, Yang P, Qian S, et al. Understanding the mechanism of denitrifying phosphorus removal from the perspective of intracellular carbon source and extracellular polymeric substances characteristics[J]. Journal of Cleaner Production, 2022,367:133115.
[32] Yuan S J, Sun M, Sheng G P, et al. Identification of key constituents and structure of the extracellular polymeric substances excreted by Bacillus megaterium TF10 for their flocculation capacity[J]. Environmental Science& Technology, 2011,45(3):1152-1157.
[33] Zheng C, Wu Q, Hu X, et al. Adsorption behavior of heavy metal ions on a polymer-immobilized amphoteric biosorbent:Surface interaction assessment[J]. Journal of Hazardous Materials, 2021,403:123801.
[34] Yan H, Huang M, Wang J, et al. Difference in calcium ion precipitation between free and immobilized Halovibrio mesolongii HMY2[J]. Journal of Environmental Sciences, 2022,122:184-200.
[35] Li L, Pang H, He J, et al. Characterization of phosphorus species distribution in waste activated sludge after anaerobic digestion and chemical precipitation with Fe3+ and Mg2+[J]. Chemical Engineering Journal, 2019,373:1279-1285.
[36] Huang W, Cai W, Huang H, et al. Identification of inorganic and organic species of phosphorus and its bio-availability in nitrifying aerobic granular sludge[J]. Water Research, 2015,68:423-431.
[37] Jørgensen C, Jensen H S, Andersen FØ, et al. Occurrence of orthophosphate monoesters in lake sediments:significance of myo-and scyllo-inositol hexakisphosphate[J]. Journal of Environmental Monitoring, 2011,13(8):2328-2334.
[38] Wang Q, He J. Complete nitrogen removal via simultaneous nitrification and denitrification by a novel phosphate accumulating Thauera sp. strain SND5[J]. Water Research, 2020,185:116300.
[39] Long X, Tang R, Fang Z, et al. The roles of loosely-bound and tightly-bound extracellular polymer substances in enhanced biological phosphorus removal[J]. Chemosphere, 2017,189:679-688.
[40] Han J, Qiu Q, Gao M, et al. Phosphorus removal from municipal wastewater through a novel Trichosporon asahii BZ:Performance and mechanism[J]. Chemosphere, 2022,298:134329.
基金
国家自然科学基金项目(52300216,52270169);西安建筑科技大学前沿交叉领域培育专项(X20230076)
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