污水处理中脱氮功能微生物特性及固定化应用研究
|
摘要
随着水体富营养化的日趋严重与污水排放标准、氮素限值的日趋严格,水中氮素的去除已成为当今水污染防治的热点问题之一。生物脱氮是从废水中去除氮素污染的最为经济有效的方法,可实现真正意义的氮去除。目前为止,尽管传统生物脱氮工艺是多数废水生物脱氮处理的主要承担者,但仍存在着硝化速率低、工艺复杂、脱氮效率低等缺点。而在氮素循环过程中发现的新型脱氮微生物,可以克服传统生物脱氮工艺固有的问题,对这些微生物的研究与新型脱氮工艺开发,是对传统生物脱氮理论与实践的丰富与突破。本文从活性污泥中分离出一株异养硝化/好氧反硝化细菌A1、一株高效缺氧反硝化细菌D6,研究了两株细菌以及已有的好氧反硝化细菌AD6脱氮过程的生态影响因子。同时考察了ca2+在好氧颗粒污泥形成中的作用,确定了SBR中快速培育好氧颗粒污泥的条件。在此基础上,将分离、鉴定的3株脱氮功能菌以好氧颗粒污泥形式固定化,研究了其在开放的污水处理系统中的作用与稳定性,构建了基于微生物及环境因子的同步硝化与反硝化工艺(Simultaneous Nitrification and Denitrification, SND)。本文主要研究结果为：
从活性污泥中分离得到一株异养硝化细菌A1,通过形态学、生理生化特征并结合16S rDNA序列同源性分析,初步鉴定该菌为枯草芽孢杆菌(Bacillus subtilis).菌株A1可以自养生长。当培养液NH4+为105.58、257.23 mg/L时,菌株A1培养120h,培养液COD可达94.77、90.26mg/L,对NH4+去除率为36.32%、19.53%,对TN去除率为24.88、13.09%；当NH4+为536.21、1014.17mg/L时,菌株A1生长受到抑制,培养120h,培养液cOD分别为56.80、33.56mg/L,菌株A1对NH4+去除率仅有6.54%、2.26%,对TN去除率仅为4.07%、2.27%,基本没有脱氮效果。考察了碳源种类与数量对菌株A1脱氮特性的影响。葡萄糖、柠檬酸盐、丁二酸盐、乙酸盐4种碳源对菌株A1脱氮特性的影响没有明显差异,以乙酸盐为碳源时,最大COD、NH4+、TN去除效率分别为71.08%、60.35%、52.22%。培养液碳源为乙酸盐,菌株A1在c/N值为6、12时,对COD, TN, NH4+-去除效率较高,分别为79.35%、71.08%,59.21%、52.22%,67.07%、60.35%,培养过程中培养液DO始终大于4.01mg/L。由此,菌株A1具有异养硝化/好氧反硝化功能。
通过菌株形态观察、生理生化特征及16S rDNA序列分析,鉴定一株可进行好氧反硝化作用的细菌AD6为假单胞菌属门多萨菌(Pseudomonas mendocina)。采用摇瓶试验,研究了C/N值,培养液DO及柠檬酸盐、乙酸盐、葡萄糖3种有机碳源,NH4+、NO2-和NO3-3种氮源对好氧反硝化细菌AD6脱氮特性的影响。C/N值不仅直接影响好氧反硝化过程中碳源数量,也显著影响培养液DO变化。起始c/N值为3时,因碳源数量不够而导致AD6好氧反硝化脱氮效率仅有41.71%；当起始C/N值为23、15、8时,培养液DO先快速下降至缺氧状态然后再升至好氧状态,反应末期TN损失率分别为69.33%、70.41%、55.40%,其中好氧阶段反硝化作用引起的TN损失约为6.93%、20.09%、24.60%。培养液起始DO为7.15～8.08 mg/L, C/N值为15及以下,摇瓶(250mL三角瓶)装液量为25～100mL,摇床转速为180r/min,培养液DO先下降至缺氧状态,但可在反硝化反应进行24h后恢复至好氧状态,减少摇瓶装液量的充氧措施提高培养液DO效果有限。柠檬酸盐与乙酸盐是AD6能够高效利用的碳源,利用率可达89.64%、92.110%,而同样条件下葡萄糖的利用率仅有40.56%。乙酸盐促进了AD6的好氧反硝化功能,TN去除率、好氧反硝化脱氮效率分别比以柠檬酸盐为碳源时高13.63%、5.18%。该菌株可分别以NH4+、NO2-和N03-为起始氮源进行生长与脱氮作用,培养29～33h达到最大氮去除效率,分别为16.57%、74.84%与74.61%,AD6表现出微弱的异养硝化／好氧反硝化功能。
从活性污泥中分离筛选出一株高效缺氧反硝化细菌D6,通过形态学、生理生化特征并结合16S rDNA序列同源性分析,初步鉴定该茵为施氏假单胞菌(Pseudomonas stutzeri)。当C/N值为6、12时,菌株D6经过54h培养,COD、TN去除效率可分别达到90%、80%以上。柠檬酸盐、乙酸盐是菌株D6反硝化过程中能高效利用的碳源,当C/N值为6,培养54h后,可分别取得80.72%、74.75%的TN去除率以及90.53%、86.30%的COD去除率；同样培养条件下,菌株D6以葡萄糖为碳源,TN与COD去除率分别只有56.56%、74.64%。当C/N值为6,NO3-浓度为182.38～358.30 mg/L,菌株D6脱氮效率不受NO3-浓度影响,可达80.72～82.50%。采用摇床振荡培养与培养箱静置培养2种方式考察菌株D6的反硝化特性,发现当培养液DO大于1.0mg/L,菌株D6的反硝化特性基本被抑制。
通过运行序批式生物反应器(Sequencing Batch Reactor, SBR)与摇瓶试验相结合方法,在进水中投加不同数量Ca2+(0～200mg/L),考察了Ca在好氧颗粒污泥形成中的作用。设计了进水Ca2+含量分别为30(Ⅰ)、100 mg/L(Ⅱ)的2组SBR试验,运行20d后,两反应器中均出现了好氧颗粒污泥。运行前50d, SBR I中颗粒污泥浓度(Mixed Liquor Suspended Solid, MLSS)与污泥沉降指数(Sludge Volume Index, SVI)指标明显好于SBRⅡ;运行后期,SBRⅠ、Ⅱ中MLSS、SVI差异很小。反应器运行与摇瓶试验表明,当进水中Ca2+为0～200mg/L,进水中ca2+含量的增加对污泥Zeta电位影响很小,没有引起好氧颗粒污泥Zeta电位的明显差异,电中和在颗粒污泥形成过程中并不起重要作用。随着进水Ca2+浓度增加,颗粒污泥中的微生物数量与种类都逐渐丰富,Ca2+有助于促进污泥中微生物多样性。接种污泥及SBR中颗粒污泥的元素含量分析表明,颗粒污泥中Ca、Mg、K、Na含量均有所减少,Fe含量分别增加了4.42%、7.82%。具有良好絮凝作用的金属离子与EPS间形成的高分子生物聚合体可能是促进好氧颗粒污泥形成的主要原因。采用2套相同的模拟SBR装置,配制典型的高浓度生活污水,在上述运行条件下均形成好氧颗粒污泥。其中1套模拟SBR装置在好氧颗粒污泥形成过程中,按7天1次的频率投加了3次A1、AD6、D6的混合菌液。60d运行试验结果表明,投加功能菌的SBR中形成了具有良好脱氮效果的好氧颗粒污泥,对COD、TN、NH4+的去除效率分别达到96.53%、85.59%、99.54%,比普通好氧颗粒污泥分别高2.02%、22.16%、5.31%,反应过程中NO3-、NO2-的积累量很少,出水中COD、TN、NH4+浓度可以稳定地达到城镇污水处理厂污染物排放标准(GB18918-2002)一级A标准,具有较好的同步脱氮除碳功能。对装置运行过程中颗粒污泥PCR-DGGE图谱的动态研究分析表明,接入的异养硝化/好氧反硝化细菌A1、好氧反硝化细菌AD6、缺氧反硝化细菌D6能够固定在颗粒污泥中,在SBR反应器中具有一定的数量优势,有助于功能菌长期稳定的发挥脱氮功能。
全文研究表明,异养硝化菌株A1、好氧反硝化菌株AD6、缺氧反硝化菌株D6均具有氮脱除功能,但菌株类型不同,生态因子如有机物浓度、碳源种类、DO、氮源等对脱氮特性的影响存在明显差异。将3类脱氮功能菌组合,形成好氧颗粒污泥,能有效将功能菌固持在开放的污水处理系统中,形成具有同步脱氮除碳性能的SND工艺,稳定高效地去除污水中有机物与氮素。异养硝化细菌和好氧反硝化细菌能够在生物同步脱氮中起重要的作用,为微生物学理论解释SND现象提供了微生物学理论依据；同时,缺氧反硝化细菌D6在颗粒污泥中的固定,表明好氧颗粒污泥中存在发生SND的环境因素。研究结果为全面评价SND工艺提供了基础研究,以颗粒污泥形式固定功能菌株为新型脱氮微生物应用于污水处理实践提供了新的途径。
With the increasing severity of the eutrophication and discharge standard, the control of nitrogen pollution has become a focus problem in wastewater treatment field. The technique of biological nitrogen removal is a common and effective method in controlling nitrogen pollution at present. Because of the different requirement of the nitrifying and denitrifying bacteria for nourishment, the process of nitrogen removal is very complex and the application is limited. Recently, some new "players" in the nitrogen cycling process have been found, which provide new method for developing the application of the biological removal of nitrogen. In this experiment, a strain of bacterium with the capacity of heterotrophic nitrification and aerobic denitrification, a strain of traditional denitrifying bacterium were isolated from activated sludge. The nitrogen removal characteristics of two strains and an aerobic denitrifying bacterium were evaluated. The role of calcium in the formation of glucose-fed aerobic granular sludge in Sequencing Batch Reactor (SBR) was investigated. Meanwhile, the optimum conditions of aerobic granular sludge formation were obtained. On the basis of this analysis, nitrogen removal bacteria,for example,strains Al, AD6, and D6 were immobilized in aerobic granular sludge. The characteristics of immobilized aerobic granular sludge were studied using synthetic wastewater in two simulated sequencing batch reactors. The stability of functional bacteria, immobilized in granular sludge, was investigated in an open wastewater treatment system. SND process based on functional bacteria and microenvironment was constructed. The main results were presented as follows:
A strain of heterotrophic nitrifying bacteria, Al, isolated from actived sludge reactor was studied for its characteristics of nitrification and nitrogen removal. Based on analysis of physiological and biochemical characters and sequence analysis of the 16S rDNA, strain Al was identified as Bacillus subtilis. Nitrifying and denitrifying performances of Al at various NH4+ concentrations were investigated through flask experiment. When the concentration of NH4+ increased from 105.58 to 257.23mg/L, the COD of culture solution arrived at 94.77-90.26mg/L and NH4+,TN removal efficiency was 36.32-19.53%, and 24.88-13.09%,respectively after 120h incubation. when the concentration of NH4+ increased to 536.21-1014.17mg/L, the COD of culture arrived at 56.80-33.56mg/L and NH4+,TN removal efficiency were 6.54-2.26% and 4.07-2.27%,respectively after 120h incubation. Denitrifying performances of Bacillus subtilis Al at various carbon sources and C/N ratios were evaluated through flask experiment. The variety of carbon sources including glucose,citrate, succinate, and acetate, as near as made no difference in nitrogen removal efficiency of strain A1.After 120h culture with acetate, the COD, NH4+,TN removal efficiency by strain Al were 71.08%,60.35% and 52.22%, respectively. When the C/N ratio was 6 and 12, the COD,TN, NH4+ removal efficiency by the strain Al was 79.35-71.08%,59.21-52.22%, and 67.07-60.35%, respectively. During the incubation process, there was almost no nitrite and low nitrate accumulation, DO concentration being up 4.01mg/L. Therefore, the heterotrophic nitrifying bacteria had aerobic denitrification ability.
By checking the individual morphology, colony culture characteristics, DNA sequencing and 16S rDNA gene bank, AD6, an aerobic denitrifying bacterium was identified as Pseudomonas mendocina. Aerobic denitrifying performances of P. mendocina AD6 at various C/N ratios, dissolved oxygen (DO) concentrations, carbon sources including citrate, acetate, and glucose and nitrogen sources including ammonia, nitrate, and nitrite were evaluated through flask experiment. At an insufficient carbon concentration, namely C/N ration of 3, TN removal efficiency was 41.71% by aerobic denitrification. When the C/N ratio was increased to 8,15, and 23, the TN removal efficiency by AD6 quickly increased to 55.40%,70.41%, and 69.33%, in which 6.93%,20.09%, and 24.60% of TN losses could be contributed to the aerobic denitrification by AD6, respectively. Batch cultures were carried out in a series of 250mL conical flasks containing 100,50, and 25mL of the nitrate-supplemented basal medium with 15 of C/N ratio,7.15 mg/L-8.08 mg/L of DO and strain AD6. Flasks were shaken in a rotary shaker at 28℃and 180r/min to allow better gas exchange. DO concentration of culture medium declined rapidly to anoxia condition at the beginnings of reaction followed by an aerobic level with above 2.0 mg/L of DO after 24 h for reaction. Different carbon sources had strong influence on the aerobic denitrifying performance of strain AD6. The maximum consumption rates of citrate, acetate, and glucose by strain AD6 were 89.64%,92.11%, and 40.56%, respectively. In the medium spiked with acetate as carbon source, TN removal efficiency and N losses due to aerobic denitrification increased to 13.63% and 5.18%, respectively, with comparison to that spiked with citrate. Under the aerobic conditions, initial nitrogen sources including ammonia, nitrite, and nitrate, strain AD6 had good growth and nitrogen removal efficiency ability,16.57%,74.84%,and 74.61% of TN removal rates were obtained after 29-33 hours incubation respectively. The strain AD6 showed the weak ability of heterotrophic nitrifying when initial nitrogen sources of medium was ammonia.
A strain of denitrifying bacteria, D6, isolated from actived sludge reactor was studied for its characteristics of denitrification. The result indicated that the isolated strain D6 was most similar to Pseudomonas stutzeri based on the analysis of morphologic characteristics, physiological and biochemical properties and phylogenic analysis of 16S rDNA sequence. Denitrifying performances of P. stutzeri D6 at various C/N ratios, carbon sources, nitrate concentrations and culture method were investigated through flask experiment. When the C/N ratio was range of 6,12, after 54h culture, the COD, TN removal efficiency by D6 reached 90%,80%, respectively. The results showed that denitrification activity was influenced by different carbon source. The nitrogen removal rate by using citrate and acetate was significantly higher than that by using glucose. However denitrification efficiency by using citrate was slightly higher than by using acetate, in which the TN, COD removal efficiency was 80.72%,74.75% and 90.53%,86.30% respectively with the C/N ratio of 6,54h culture period. Under the same conditions, the TN,COD removal efficiency was only 56.56% and 74.64% with glucose. In the range of C/N ratio of 6, when the nitrate concentration increased from 182.38 to 358.30 mg/L, the nitrogen removal efficiency of AD6 varied 80.72-82.50% Under two incubate conditions of flask experiment and chemostat, denitrifying performances of D6 was evaluated. It was proved that denitrification process of D6 was inhibited when DO of culture solution was higher than 1 mg/L. Strain D6 denitrifying ability was showed only under anoxic condition.
Influence of calcium on glucose-fed aerobic granule was investigated through sequencing batch reactor (SBR) and shaking flask experiments. Granules of two SBRs spiked with 30 mg/L and 100 mg/L of Ca2+ concentration could be observed visually on day 20. In the first 50 days, the SBRⅡhad higher mixed liquor suspended solid (MLSS) and lower sludge volume index (SVI) than SBRⅠ. After 50 days, the difference of granule performance in two reactors gradually be reduced and little differences was found on day 80 or more, strain Alconcentration of influent had little influence on Zeta potential of sludge during granulation process. Microbiological observations confirmed some appreciable changes in microorganism population and diversities with the increase of Ca2+ concentration of influent. The role of Ca2+ in bioflocculation of granule can be mainly attributed to its cation bridging instead of charge neutralization. The elemental analysis of seed sludge and aerobic granule by x-ray fluorescence (XRF) showed that the content of Ca,Mg,K, and Na in granule were less than seed sludge expect for Fe, increasing 4.42%,7.82% respectively. Divalent metal ions such as Ca2+ and Fe2+ were probably constituent of biopolymer. It was possible that the binding of divalent metal ion with extracellular polymeric substances (EPS) enhanced the granulation of seed sludge. Consequently, the pollutant removal efficiency of SBRⅡwere higher than that of SBR I throughout the trial. Both NH4+-N and COD removal efficiencies reached 90%; TN and TP removal efficiency were both 65%-70% after days 70 in SBRs.With the same activated sludge inoculation and synthetic domestic sewage, granules of two simulated SBRs could be observed. One of SBRs was inoculated three strains of nitrogen removal bacteria with frequency of one time every 7 days. The results of 60 days operation revealed that the SBR inoculated nitrogen removal bacteria effluent concentration of COD,TN and NH4+-N were less than 50,15 and 5mg/L respectively, meeting the first level A criteria specified in the Discharge Standard of pollutants for municipal wastewater treatment plant (GB18918-2002). After 60 days culture, the COD, TN, N4+-N removal efficiency by the granule immobilized nitrogen removal strains were 96.53%,85.59%, and 99.54%, increasing 2.02%,22.16%, and 5.31% than common granule respectively. During the trial process, there was almost no nitrite and low nitrate accumulation in SBRs.
Microbial communities in aerobic sludge shifted obviously with granulation of aerobic sludge. DGGE analysis illustrated that aerobic granules with more abundant DGGE bands and the community structures than activated sludge in steady state. Compared to common aerobic granules, the population diversity was higher in the nitrogen-removing aerobic granules. Results demonstrated that aerobic sludge granulation may play an important role in the enrichment and retaining of nitrogen-removing microorganisms include heterotrophic nitrifying strain Al, aerobic denitrifying strain AD6, and denitrifying strain D6.
It was concluded that heterotrophic nitrifying strain Al, aerobic denitrifying strain AD6, and denitrifying strain D6 had good performance of nitrogen removal, but influencing factors such as carbon sources, DO, nitrogen sources were not the same. Three type functional bacteria were closely related to nitrogen-removing aerobic granules in SBR and showed ability of simultaneous nitrogen and carbon removal. Strains Al, AD6 play the important role in the SND, and it provides the microorganism influence factor for SND. There were strain D6 in aerobic granules, which indicated microenvironment evidence for SND. It was indicated that nitrogen-removing functional bacteria fixed with aerobic granules, as a potentially excellent immobilization technology, would play an important role in treatment of nitrogen-contaminated surface water. All these results may contribute to the establishment of new biology process to remove nitrogen from wastewater with high efficiency.
从活性污泥中分离得到一株异养硝化细菌A1,通过形态学、生理生化特征并结合16S rDNA序列同源性分析,初步鉴定该菌为枯草芽孢杆菌(Bacillus subtilis).菌株A1可以自养生长。当培养液NH4+为105.58、257.23 mg/L时,菌株A1培养120h,培养液COD可达94.77、90.26mg/L,对NH4+去除率为36.32%、19.53%,对TN去除率为24.88、13.09%；当NH4+为536.21、1014.17mg/L时,菌株A1生长受到抑制,培养120h,培养液cOD分别为56.80、33.56mg/L,菌株A1对NH4+去除率仅有6.54%、2.26%,对TN去除率仅为4.07%、2.27%,基本没有脱氮效果。考察了碳源种类与数量对菌株A1脱氮特性的影响。葡萄糖、柠檬酸盐、丁二酸盐、乙酸盐4种碳源对菌株A1脱氮特性的影响没有明显差异,以乙酸盐为碳源时,最大COD、NH4+、TN去除效率分别为71.08%、60.35%、52.22%。培养液碳源为乙酸盐,菌株A1在c/N值为6、12时,对COD, TN, NH4+-去除效率较高,分别为79.35%、71.08%,59.21%、52.22%,67.07%、60.35%,培养过程中培养液DO始终大于4.01mg/L。由此,菌株A1具有异养硝化/好氧反硝化功能。
通过菌株形态观察、生理生化特征及16S rDNA序列分析,鉴定一株可进行好氧反硝化作用的细菌AD6为假单胞菌属门多萨菌(Pseudomonas mendocina)。采用摇瓶试验,研究了C/N值,培养液DO及柠檬酸盐、乙酸盐、葡萄糖3种有机碳源,NH4+、NO2-和NO3-3种氮源对好氧反硝化细菌AD6脱氮特性的影响。C/N值不仅直接影响好氧反硝化过程中碳源数量,也显著影响培养液DO变化。起始c/N值为3时,因碳源数量不够而导致AD6好氧反硝化脱氮效率仅有41.71%；当起始C/N值为23、15、8时,培养液DO先快速下降至缺氧状态然后再升至好氧状态,反应末期TN损失率分别为69.33%、70.41%、55.40%,其中好氧阶段反硝化作用引起的TN损失约为6.93%、20.09%、24.60%。培养液起始DO为7.15～8.08 mg/L, C/N值为15及以下,摇瓶(250mL三角瓶)装液量为25～100mL,摇床转速为180r/min,培养液DO先下降至缺氧状态,但可在反硝化反应进行24h后恢复至好氧状态,减少摇瓶装液量的充氧措施提高培养液DO效果有限。柠檬酸盐与乙酸盐是AD6能够高效利用的碳源,利用率可达89.64%、92.110%,而同样条件下葡萄糖的利用率仅有40.56%。乙酸盐促进了AD6的好氧反硝化功能,TN去除率、好氧反硝化脱氮效率分别比以柠檬酸盐为碳源时高13.63%、5.18%。该菌株可分别以NH4+、NO2-和N03-为起始氮源进行生长与脱氮作用,培养29～33h达到最大氮去除效率,分别为16.57%、74.84%与74.61%,AD6表现出微弱的异养硝化／好氧反硝化功能。
从活性污泥中分离筛选出一株高效缺氧反硝化细菌D6,通过形态学、生理生化特征并结合16S rDNA序列同源性分析,初步鉴定该茵为施氏假单胞菌(Pseudomonas stutzeri)。当C/N值为6、12时,菌株D6经过54h培养,COD、TN去除效率可分别达到90%、80%以上。柠檬酸盐、乙酸盐是菌株D6反硝化过程中能高效利用的碳源,当C/N值为6,培养54h后,可分别取得80.72%、74.75%的TN去除率以及90.53%、86.30%的COD去除率；同样培养条件下,菌株D6以葡萄糖为碳源,TN与COD去除率分别只有56.56%、74.64%。当C/N值为6,NO3-浓度为182.38～358.30 mg/L,菌株D6脱氮效率不受NO3-浓度影响,可达80.72～82.50%。采用摇床振荡培养与培养箱静置培养2种方式考察菌株D6的反硝化特性,发现当培养液DO大于1.0mg/L,菌株D6的反硝化特性基本被抑制。
通过运行序批式生物反应器(Sequencing Batch Reactor, SBR)与摇瓶试验相结合方法,在进水中投加不同数量Ca2+(0～200mg/L),考察了Ca在好氧颗粒污泥形成中的作用。设计了进水Ca2+含量分别为30(Ⅰ)、100 mg/L(Ⅱ)的2组SBR试验,运行20d后,两反应器中均出现了好氧颗粒污泥。运行前50d, SBR I中颗粒污泥浓度(Mixed Liquor Suspended Solid, MLSS)与污泥沉降指数(Sludge Volume Index, SVI)指标明显好于SBRⅡ;运行后期,SBRⅠ、Ⅱ中MLSS、SVI差异很小。反应器运行与摇瓶试验表明,当进水中Ca2+为0～200mg/L,进水中ca2+含量的增加对污泥Zeta电位影响很小,没有引起好氧颗粒污泥Zeta电位的明显差异,电中和在颗粒污泥形成过程中并不起重要作用。随着进水Ca2+浓度增加,颗粒污泥中的微生物数量与种类都逐渐丰富,Ca2+有助于促进污泥中微生物多样性。接种污泥及SBR中颗粒污泥的元素含量分析表明,颗粒污泥中Ca、Mg、K、Na含量均有所减少,Fe含量分别增加了4.42%、7.82%。具有良好絮凝作用的金属离子与EPS间形成的高分子生物聚合体可能是促进好氧颗粒污泥形成的主要原因。采用2套相同的模拟SBR装置,配制典型的高浓度生活污水,在上述运行条件下均形成好氧颗粒污泥。其中1套模拟SBR装置在好氧颗粒污泥形成过程中,按7天1次的频率投加了3次A1、AD6、D6的混合菌液。60d运行试验结果表明,投加功能菌的SBR中形成了具有良好脱氮效果的好氧颗粒污泥,对COD、TN、NH4+的去除效率分别达到96.53%、85.59%、99.54%,比普通好氧颗粒污泥分别高2.02%、22.16%、5.31%,反应过程中NO3-、NO2-的积累量很少,出水中COD、TN、NH4+浓度可以稳定地达到城镇污水处理厂污染物排放标准(GB18918-2002)一级A标准,具有较好的同步脱氮除碳功能。对装置运行过程中颗粒污泥PCR-DGGE图谱的动态研究分析表明,接入的异养硝化/好氧反硝化细菌A1、好氧反硝化细菌AD6、缺氧反硝化细菌D6能够固定在颗粒污泥中,在SBR反应器中具有一定的数量优势,有助于功能菌长期稳定的发挥脱氮功能。
全文研究表明,异养硝化菌株A1、好氧反硝化菌株AD6、缺氧反硝化菌株D6均具有氮脱除功能,但菌株类型不同,生态因子如有机物浓度、碳源种类、DO、氮源等对脱氮特性的影响存在明显差异。将3类脱氮功能菌组合,形成好氧颗粒污泥,能有效将功能菌固持在开放的污水处理系统中,形成具有同步脱氮除碳性能的SND工艺,稳定高效地去除污水中有机物与氮素。异养硝化细菌和好氧反硝化细菌能够在生物同步脱氮中起重要的作用,为微生物学理论解释SND现象提供了微生物学理论依据；同时,缺氧反硝化细菌D6在颗粒污泥中的固定,表明好氧颗粒污泥中存在发生SND的环境因素。研究结果为全面评价SND工艺提供了基础研究,以颗粒污泥形式固定功能菌株为新型脱氮微生物应用于污水处理实践提供了新的途径。
With the increasing severity of the eutrophication and discharge standard, the control of nitrogen pollution has become a focus problem in wastewater treatment field. The technique of biological nitrogen removal is a common and effective method in controlling nitrogen pollution at present. Because of the different requirement of the nitrifying and denitrifying bacteria for nourishment, the process of nitrogen removal is very complex and the application is limited. Recently, some new "players" in the nitrogen cycling process have been found, which provide new method for developing the application of the biological removal of nitrogen. In this experiment, a strain of bacterium with the capacity of heterotrophic nitrification and aerobic denitrification, a strain of traditional denitrifying bacterium were isolated from activated sludge. The nitrogen removal characteristics of two strains and an aerobic denitrifying bacterium were evaluated. The role of calcium in the formation of glucose-fed aerobic granular sludge in Sequencing Batch Reactor (SBR) was investigated. Meanwhile, the optimum conditions of aerobic granular sludge formation were obtained. On the basis of this analysis, nitrogen removal bacteria,for example,strains Al, AD6, and D6 were immobilized in aerobic granular sludge. The characteristics of immobilized aerobic granular sludge were studied using synthetic wastewater in two simulated sequencing batch reactors. The stability of functional bacteria, immobilized in granular sludge, was investigated in an open wastewater treatment system. SND process based on functional bacteria and microenvironment was constructed. The main results were presented as follows:
A strain of heterotrophic nitrifying bacteria, Al, isolated from actived sludge reactor was studied for its characteristics of nitrification and nitrogen removal. Based on analysis of physiological and biochemical characters and sequence analysis of the 16S rDNA, strain Al was identified as Bacillus subtilis. Nitrifying and denitrifying performances of Al at various NH4+ concentrations were investigated through flask experiment. When the concentration of NH4+ increased from 105.58 to 257.23mg/L, the COD of culture solution arrived at 94.77-90.26mg/L and NH4+,TN removal efficiency was 36.32-19.53%, and 24.88-13.09%,respectively after 120h incubation. when the concentration of NH4+ increased to 536.21-1014.17mg/L, the COD of culture arrived at 56.80-33.56mg/L and NH4+,TN removal efficiency were 6.54-2.26% and 4.07-2.27%,respectively after 120h incubation. Denitrifying performances of Bacillus subtilis Al at various carbon sources and C/N ratios were evaluated through flask experiment. The variety of carbon sources including glucose,citrate, succinate, and acetate, as near as made no difference in nitrogen removal efficiency of strain A1.After 120h culture with acetate, the COD, NH4+,TN removal efficiency by strain Al were 71.08%,60.35% and 52.22%, respectively. When the C/N ratio was 6 and 12, the COD,TN, NH4+ removal efficiency by the strain Al was 79.35-71.08%,59.21-52.22%, and 67.07-60.35%, respectively. During the incubation process, there was almost no nitrite and low nitrate accumulation, DO concentration being up 4.01mg/L. Therefore, the heterotrophic nitrifying bacteria had aerobic denitrification ability.
By checking the individual morphology, colony culture characteristics, DNA sequencing and 16S rDNA gene bank, AD6, an aerobic denitrifying bacterium was identified as Pseudomonas mendocina. Aerobic denitrifying performances of P. mendocina AD6 at various C/N ratios, dissolved oxygen (DO) concentrations, carbon sources including citrate, acetate, and glucose and nitrogen sources including ammonia, nitrate, and nitrite were evaluated through flask experiment. At an insufficient carbon concentration, namely C/N ration of 3, TN removal efficiency was 41.71% by aerobic denitrification. When the C/N ratio was increased to 8,15, and 23, the TN removal efficiency by AD6 quickly increased to 55.40%,70.41%, and 69.33%, in which 6.93%,20.09%, and 24.60% of TN losses could be contributed to the aerobic denitrification by AD6, respectively. Batch cultures were carried out in a series of 250mL conical flasks containing 100,50, and 25mL of the nitrate-supplemented basal medium with 15 of C/N ratio,7.15 mg/L-8.08 mg/L of DO and strain AD6. Flasks were shaken in a rotary shaker at 28℃and 180r/min to allow better gas exchange. DO concentration of culture medium declined rapidly to anoxia condition at the beginnings of reaction followed by an aerobic level with above 2.0 mg/L of DO after 24 h for reaction. Different carbon sources had strong influence on the aerobic denitrifying performance of strain AD6. The maximum consumption rates of citrate, acetate, and glucose by strain AD6 were 89.64%,92.11%, and 40.56%, respectively. In the medium spiked with acetate as carbon source, TN removal efficiency and N losses due to aerobic denitrification increased to 13.63% and 5.18%, respectively, with comparison to that spiked with citrate. Under the aerobic conditions, initial nitrogen sources including ammonia, nitrite, and nitrate, strain AD6 had good growth and nitrogen removal efficiency ability,16.57%,74.84%,and 74.61% of TN removal rates were obtained after 29-33 hours incubation respectively. The strain AD6 showed the weak ability of heterotrophic nitrifying when initial nitrogen sources of medium was ammonia.
A strain of denitrifying bacteria, D6, isolated from actived sludge reactor was studied for its characteristics of denitrification. The result indicated that the isolated strain D6 was most similar to Pseudomonas stutzeri based on the analysis of morphologic characteristics, physiological and biochemical properties and phylogenic analysis of 16S rDNA sequence. Denitrifying performances of P. stutzeri D6 at various C/N ratios, carbon sources, nitrate concentrations and culture method were investigated through flask experiment. When the C/N ratio was range of 6,12, after 54h culture, the COD, TN removal efficiency by D6 reached 90%,80%, respectively. The results showed that denitrification activity was influenced by different carbon source. The nitrogen removal rate by using citrate and acetate was significantly higher than that by using glucose. However denitrification efficiency by using citrate was slightly higher than by using acetate, in which the TN, COD removal efficiency was 80.72%,74.75% and 90.53%,86.30% respectively with the C/N ratio of 6,54h culture period. Under the same conditions, the TN,COD removal efficiency was only 56.56% and 74.64% with glucose. In the range of C/N ratio of 6, when the nitrate concentration increased from 182.38 to 358.30 mg/L, the nitrogen removal efficiency of AD6 varied 80.72-82.50% Under two incubate conditions of flask experiment and chemostat, denitrifying performances of D6 was evaluated. It was proved that denitrification process of D6 was inhibited when DO of culture solution was higher than 1 mg/L. Strain D6 denitrifying ability was showed only under anoxic condition.
Influence of calcium on glucose-fed aerobic granule was investigated through sequencing batch reactor (SBR) and shaking flask experiments. Granules of two SBRs spiked with 30 mg/L and 100 mg/L of Ca2+ concentration could be observed visually on day 20. In the first 50 days, the SBRⅡhad higher mixed liquor suspended solid (MLSS) and lower sludge volume index (SVI) than SBRⅠ. After 50 days, the difference of granule performance in two reactors gradually be reduced and little differences was found on day 80 or more, strain Alconcentration of influent had little influence on Zeta potential of sludge during granulation process. Microbiological observations confirmed some appreciable changes in microorganism population and diversities with the increase of Ca2+ concentration of influent. The role of Ca2+ in bioflocculation of granule can be mainly attributed to its cation bridging instead of charge neutralization. The elemental analysis of seed sludge and aerobic granule by x-ray fluorescence (XRF) showed that the content of Ca,Mg,K, and Na in granule were less than seed sludge expect for Fe, increasing 4.42%,7.82% respectively. Divalent metal ions such as Ca2+ and Fe2+ were probably constituent of biopolymer. It was possible that the binding of divalent metal ion with extracellular polymeric substances (EPS) enhanced the granulation of seed sludge. Consequently, the pollutant removal efficiency of SBRⅡwere higher than that of SBR I throughout the trial. Both NH4+-N and COD removal efficiencies reached 90%; TN and TP removal efficiency were both 65%-70% after days 70 in SBRs.With the same activated sludge inoculation and synthetic domestic sewage, granules of two simulated SBRs could be observed. One of SBRs was inoculated three strains of nitrogen removal bacteria with frequency of one time every 7 days. The results of 60 days operation revealed that the SBR inoculated nitrogen removal bacteria effluent concentration of COD,TN and NH4+-N were less than 50,15 and 5mg/L respectively, meeting the first level A criteria specified in the Discharge Standard of pollutants for municipal wastewater treatment plant (GB18918-2002). After 60 days culture, the COD, TN, N4+-N removal efficiency by the granule immobilized nitrogen removal strains were 96.53%,85.59%, and 99.54%, increasing 2.02%,22.16%, and 5.31% than common granule respectively. During the trial process, there was almost no nitrite and low nitrate accumulation in SBRs.
Microbial communities in aerobic sludge shifted obviously with granulation of aerobic sludge. DGGE analysis illustrated that aerobic granules with more abundant DGGE bands and the community structures than activated sludge in steady state. Compared to common aerobic granules, the population diversity was higher in the nitrogen-removing aerobic granules. Results demonstrated that aerobic sludge granulation may play an important role in the enrichment and retaining of nitrogen-removing microorganisms include heterotrophic nitrifying strain Al, aerobic denitrifying strain AD6, and denitrifying strain D6.
It was concluded that heterotrophic nitrifying strain Al, aerobic denitrifying strain AD6, and denitrifying strain D6 had good performance of nitrogen removal, but influencing factors such as carbon sources, DO, nitrogen sources were not the same. Three type functional bacteria were closely related to nitrogen-removing aerobic granules in SBR and showed ability of simultaneous nitrogen and carbon removal. Strains Al, AD6 play the important role in the SND, and it provides the microorganism influence factor for SND. There were strain D6 in aerobic granules, which indicated microenvironment evidence for SND. It was indicated that nitrogen-removing functional bacteria fixed with aerobic granules, as a potentially excellent immobilization technology, would play an important role in treatment of nitrogen-contaminated surface water. All these results may contribute to the establishment of new biology process to remove nitrogen from wastewater with high efficiency.
引文
1. 胡林林,王建龙,文湘华,等.SBR中厌氧颗粒污泥向好氧颗粒污泥的转化[J].环境科学,2004,25(4)：74-77.
2. 李丛娜,吕锡武,稻森悠平.同步硝化反硝化脱氮研究.给水排水[J].给水排水,2001,27(1)：22-24.
3. 林燕,孔海南,王茸影,等.异养硝化作用的主要特点及其研究动向[J].环境科学,2008,29(11)：3291-3296.
4. 刘幽燕,何玉财,李青云,等.沸石固定化细胞降解氰化物的试验研究[J].高校化学工程学报,2005,19(4)：532-535.
5. 吕锡武,李峰,稻森悠平,等.氨氮废水处理工程中的好氧反硝化研究[J].给水排水,2000,26(4)：17-20.
6. 马放,周丹丹,王宏宇,等.一株好氧反硝化细菌生理生态特征的研究[J].哈尔滨工业大学学报,2006,38(4)：575-577.
7. 牟丽娉,黄钧,苟莎.异养硝化微生物菌剂及其好氧颗粒污泥的脱氮试验[J].应用与环境生物学报,2009,15(3)：356-360.
8. 阮文权,陈坚.同步脱氮好氧颗粒污泥的特性及其反应过程[J].中国环境科学,2003,23(4)：380-384.
9. 孙锦宜.含氮废水处理技术与应用[M].北京：化学工业出版社,2003.33.
10.汪苹,项慕飞,翟茜,等.从不同反应器筛选、鉴别好氧反硝化菌[J].环境科学研究,2007,20(4)：120-124.
11.王红娟,奚红霞,夏启斌,等.含酚废水处理技术的现状与开发前景[J].工业水处理,2002,22(6)：6-9.
12.王薇,蔡祖聪,钟文辉,等.好氧反硝化菌的研究进展[J].应用生态学报,2007,18(11)：2618-2625.
13.王旭明,从二丁,罗文龙,等.固体碳源用于异养反硝化去除地下水中的硝酸盐[J].中国科学B辑：化学,2008,38(9)：824-828.
14.叶建锋.废水生物脱氮处理新技术[M].北京：化学工业出版社,2006,10-12.
15.张立秋,张可方,张朝升,等.DO对亚硝酸型SND的影响[J].水处理技术,2008,34(8)：29-33.
16.郑平,徐向阳,胡宝兰.新型生物脱氮理论与技术[M].北京：科学出版社,2004,5-7.
17.郑兴灿,李亚新.污水除磷脱氮技术[M].北京：中国建筑工业出版社,1998,55-57.
18.朱亮,徐向阳,罗伟国,等.废水生物处理好氧污泥颗粒化进展[J].环境科学,2007,28(11)：2657-2664.
19.竺建荣,刘纯新.好氧颗粒活性污泥的培养及理化特性研究[J].环境科学,1999,20(2)：38-41.
20. Arlca M Y, Kacar Y, Gene O. Entrapment of white-rot fungus trameters versicolor in Ca-alginate beads:preparation and biosorption kinetics analysis for cadmium removal from an aqueous solution [J]. Bioresourse Technol,2001,80(2):121-129.
21. Bateman E J, Baggs E M. Contributions of nitrification and denitrification to N2O emissions from soils at different waterfilled pore space[J]. Biology and Fertility of soils,2005,41(6):379-388.
22. Bayramoglu G, Denizlia, Bektas S,et al. Entrapment of Lentinus sajor-caju into Ca-alginate gel beads for removal of Cd(II)ions from aqueous solution:preparation and biosorption kinetics analysis [J]. Micro-Chemical Journal,2002,72(1):63-76.
23. Ben J J, Heijnen J J, van Loosdrecht M C M. N-removal in a granular sludge sequencing batch airlift reactor [J].Biotechnol Bioeng,2001,75(1):82-92.
24. Brierley E D R,Wood M.Heterotrophic nitrification in an acid forest soil:isolation and characterization of a nitrifying bacterium[J].Soil Biol Biochem,2001,33:1403-1409.
25. Chen F, Xia Q, Ju L K. Aerobic denitrification of Pseudomonas aeruginosa monitored by online NAD(P) H fluorescence[J].Appl Envir Mcrobiol,2003,69(11):6715-6722.
26. Doxtader K G, Alexander M. Nitrification by heterotrophic soil microorganisms[J].Soil Sci Soc Amer Proc,1966,30:351-355.
27. Geraats S G M, Hooijmans C M, van Niel E W J. The use of a metabolically structured model in the study of growth Nitrification and denitrification by Thiosphaera pantotropha[J].Biotechnol Bioeng,1990,36:227-267.
28. Henning P,Kristie A D, Mary K F. The relative importance of autotrophic and heterotrophic nitrification in a conifer forest soil as measured by 15N tracer and pool dilution techniques[J]. Biogeochemistry,1999,44:135-150.
29. Kowalchuk G A,Stephen J R. Ammonia-oxidizing bacteria:A model for molecular microbial ecology.[J]. Annu Rev Microbiol,2001,55:485-529.
30. Kshirsagar M, Gupta A B, Gupta S K. Aerobic denitrification studies on activated sludge mixed with Thiosphaera pantotropha[J].Environ Tech,1995,16:35-43.
31. Kuai L,Verstrartr W. Autotrophic denitrification with elemental sulphur in small-scale wastewater treatment facilities[J].Environ Tech,1999,20:201-209.
32. Lin Y M, Liu Y, Tay J H. Development and characteristics of phosphorous-accumulating granules in sequencing batch reactor [J]. Appl Microbial Biotechnol,2003,62(4):25-43.
33. Liu Y, Wang Z W, Tay J H. A unified theory for upscaling aerobic granular sludge sequencing batch reactor [J]. Biotechnol Adv,2005,23(5):335-344.
34. Mchmidt I, Sliekers O, Schmid M, et al. New concepts of microbial treatment processes for the nitrogen removal in wastewater [J]. FEMS Microbiology Reviews,2003,27:481-492.
35. Metcalf, Eddy, Inc. Wastewater Engineering Treatment and Reuse.4th ed. USA:MeGraw-Hill Companies,2003
36. Muller C, Stevens R J, Laughlin R J. An N-15 tracing model to analyze N transformations in old grassland soil[J]. Soil Biology Biochemistry,2004,36(4):619-632.
37. Nishio T, Yoshikura T. Effects of organic acids on heterotrophic nitrification by Alaligenes faecalis OKKI 7[J].Biosci Biotech Biochem,1994,58:1574-1578.
38. Peng D C,Bernet N, Delgenes J P, et al. Aerobic granular sludge-a case report[J]. Water Res,1999, 33(3):890-893.
39. Robertson L A, Kuenen J G. Aerobic denitrification:A controversy revived[J]. Archives of Microbiology,1984,139:351-354.
40. Robertson L A, Van Niel E W J, Torremans R A M. Simultaneous nitrification and denitrification in aerobic chemostat cultures of Thiosphaera pantotropha[J].Appl Envir Mcrobiol,1988,54(11): 2812-2818.
41. Sen S, Demirer G N. Anaerobic treatment of real textile wastewater with a fluidized bed reactor [J]. Water Res,2003,37:1868-1878.
42. Sierra-Alvarez R,Beristain-Cardoso R,Salazar M,et al. Chemolithotrophic denitrification with elemental sulfur for groundwater treatment[J]. Water Res,2007,41:1253-1262.
43. Soares M I M, Abeliovich A. Wheat straw as substrate for water denitrification [J]. Water Res,1998, 32(12):3790-3794.
44. Stephen J R, Kowalchuk G A, Brun M V,et al. Analysis of a-subgroup proteobacteria ammonia oxidizer populations in soil by denaturing gradient gel electrophoresis analysis and hierarchical phylogentic probing [J]. Appl Environ Microbial,1998,64:2958-2965.
45. Takaya N, Catalan S M, Sakaguchi Y,et al. Aerobic denitrification bacteria that produced low levels of nitrous oxide[J].Appl Envir Mcrobiol,2003,69(6):3152-3157.
46. Yoo H,Ahn K,Lee H,et al. Nitrogen removal from synthetic wastewater by simultaneous nitrification and denitrification(SND) via nitrite in an intermittently reactor[J]. Water Res,1999, 33(1):145-154.
47. Zhao H W,Mavinic D S,Oldham W K,et al. Controlling factors for simultaneous nitrification and denitrification in a two-stage intermittent aeration process treating domestic sewage[J]. Water Res, 1999,33(4):961-970.
1. (美)奥斯伯F M.精编分子生物学试验指南.科学出版社.2005.1457-1574.
2. 北京市市政工程设计研究总院.给水排水设计手册(第5册-城镇排水)(第2版)[M].北京：中国建筑工业出版社,2004.245.
3. 东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京：科学出版社,2001.370-378.
4. 高廷耀,顾国维,周琪.水污染控制工程(第3版)[M].北京：高等教育出版社,2007.102-103.
5. 胡君利,林先贵,褚海燕,等.土壤氨氧化细菌的分离方法研究[J].土壤,2005,37(5)：569-571.
6. 周群英,高廷耀.环境工程微生物学(第二版)[M].北京：高等教育出版社,1999.122.
7. GB 11894-89,碱性过硫酸钾消解紫外分光光度法[S].
8. GB 7479-87,纳氏试剂比色法[S].
9. GB 7493-87,N-(1-萘基)-乙二胺分光光度法[S].
10.HJ/T 346-2007,紫外分光光度法[S].
11.GB 11893-89,铝酸铵分光光度法[S].
12.李阜棣,喻子牛,何绍江.农业微生物学试验技术[M].北京：中国农业出版社,1996.308.
13.土壤微生物研究会编.土壤微生物试验法[M].北京：科学出版社,2001.656.
14.沈萍,范秀容,李广武.微生物学试验(第三版)[M].北京：中国农业出版社,1998.89-87.
15.袁飞.不同农田土壤中的硝化作用及硝化细菌种群[D].南京：南京农业大学,2004.118-121.
16.张纪忠.微生物分类学[M].上海：复旦大学出版社,1990.108.
17. Edwards U, Rogall T, Blocker H,et al. Isolation and direct complete determination of entire genes. Nucleic Acids Res.1989.17:7843-7853.
18. Ferris, M J, Muyzer G, Ward D M. Denaturing gradient gel electrophoresis profiles of 16S rRNA-defined populations inhabiting a hot spring microbial mat community [J]. Appl Environ Microb,1996,62:340-346.
19. Heuer H, Krsek M, Baker P,et al. Analysis of actinomycete communities by specific amplification of genes encoding 16S rRNA and gel-electrophoretic separation in denaturing gradients[J]. Appl Environ Microb,1997,63:3233-3241.
20. Sigler W V, Miniaci C, Zeyer J. Electrophoresis time impacts the denaturing gradient gel electrophoresis-based assessment of bacterial community structure[J]. J Microbiol Methods 2004,57:17-22.
21. Teske A, Wawer C, Muyzer G,et al. Distribution of sulfate-reducing bacteria in a stratified fjord (Mariager Fjord, Denmark) as evaluated by most-probable-number counts and denaturing gradient gel electrophoresis of PCR-amplified ribosomal DNA fragments[J]. Appl Environ Microb, 1996,62:1405-1415.
22. Wilson K H, Blitchington R B, Green R C. Amplification of 16S ribosomal DNA with polymerase chain reaction. Clin Microbiol,1990.28:1942-1946.
1. 何霞,赵彬,吕剑,等.异养硝化细菌Bacillus sp.LY脱氮特性研究[J].环境科学,2007,28(6)：1404-1408.
2.胡咏梅,葛向阳,梁运祥.枯草芽孢杆菌FY99-01菌株的净水作用[J].华中农业大学学报,2006,25(4)：404-407.
3. 刘晶晶,汪苹,王欢.一株异养硝化-好氧反硝化菌的脱氮特性研究[J].环境科学研究,2008,21(3)：121-125.
4. 罗固源,杨红薇NBIAS系统中的好氧反硝化.重庆环境科学,2001,23(1)：43-46.
5. 王弘宇,马放,杨开,等.两株异养硝化细菌的氨氮去除特性[J].中国环境科学,2009,29(1)：47-52.
6. 熊伟,梁运祥,戴经元,等.枯草芽孢杆菌对斑节对虾饲养池水净化作用的初步研究[J].华中农业大学学报,2003,22(3)：247-250.
7. 杨航,黄钧,刘博.异养硝化-好氧反硝化菌Paracoccus pantotrophus ATCC 35512的研究进展[J].应用与环境生物学报,2008,14(4)：585-592.
8. 张光亚,陈美慈,韩如韩,等.一株异养硝化细菌的分离及系统发育分析[J].微生物学报,2003,43(2)：156-161.
9. Berks B C, Richardson D J, Reilly A,et al. The napEDABC gene cluster encoding the periplasmic nitrate reductase system of Thiosphaera pantotropha[J]. Biochem J,1995,309:983-992.
10. Crossman L C, Moir J W B, Enticknap J J,et al. Heterologous expression of heterotrophic nitrification genes [J]. Microbiology,1997,143(12):3775-3783.
11. Cartron M L, Roldan M D, Ferguson S J,et al. Identification of two domains and distal histidine ligands to the four haems in the bacterial c-type cytochrome NapC; the prototype connector between quinol/quinone and periplasmic oxido-reductases. [J]. Biochem J,2002,368:425-432.
12. Castignetti D, Hollocher T C. Nitrogen redox metabolism of a hetetoteophic nitrifying-denitrifying Alcaligenes sp. from soil[J]. Appl Environ Microbiol,1982,44(4):923-928.
13. Duggin J A, Voigt G K. Autotrophic and heterotrophic nitrification in response to clear-cutting northern hardwood forest.[J]. Soil Biol Biochem,1991,23(8):779-787.
14. Hooper A B, Vannelli T, Bergmann D J,et al. Enzymology of the oxidation of ammonia to nitrite by bacteria[J]. Antonie Van Leeuwenhoek,1997,71(1-2):59-67.
15. Hu T L, Kung K T. Study of heterotrophic in nitrifying bacteria from wastewater treatment systems treating acrylonitrile,butadiene and styrene resin wastewater[J]. Wat Sci Tech, 2000,42(3-4):315-322.
16. Joong K K, Kyoung J P, Kyoung S C,et al. Aerobic nitrification-denitrification by heterotrophic Bacillus strains[J]. Bioresource Technol,2005,96(17):1897-1906.
17. Kelly D P, Euzeby J P, Enticknap J J,et al. Redefining Paracoccus denitrificans and Paracoccus pantotrophus and the case for a reassessment of the strains held by international culture collections [J]. Int J Syst Evol Microbiol,2006,56(10):2495-2500.
18. Pedersen H, Dunkin K A, Firestone M K. The relative importance of autotrophic and heterotrophic nitrification in a conifer forest soil as measured by N-15 tracer and pool dilution techniques[J]. Biogeochemistry,1999,44(2):135-150.
19. Richardson D J, Ferguson S J. The influence of carbon substrate on the activity Thiosphaera pantotropha of the periplasmic nitrate reductase in aerobically grown[J]. Arch Microbiol,1992,157 (6):535-537.
20. Schimel J P, Firestone M K. Identification of heteroteophic nitrification in a sierran forest soil[J]. Appl Environ Microbiol,1984,48(4):802-806.
21. Stouthamer A H, De Boer A P N, Van der Oost J,et al. Emerging principles of inorganic nitrogen metabolism in Paracoccus denitrificans and related bacteria[J]. Antonie Van Leeuwenhoek, 1997,71(1-2):33-41.
22. Wehrfritz J M, Reilly A, Spiro S,et al. Purification of hydroxylamine oxidase from Thiosphaera pantotropha. Identification of electron acceptors that couple heterotropbic nitrification to aerobic denitrification[J]. FEBS Lett,1993,335(2):246-250.
23. Witzel K P, Overbeck H J. Heteroteophic nitrification by Arthtobacter sp.(strain 9006) as influenced by different cultural conditions growth state and acetate metabolism[J]. Arch Microbiol,1979,122:137-143.
1. 孔庆鑫,李君文,王新为,等.一种新的好氧反硝化细菌筛选方法的建立及新菌株的发现[J].应用与环境生物学报,2005,11(2)：222-225.
2. 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,2006,26(11)：3718-3724.
3. 马放,周丹丹,王弘宇,等.一株好氧反硝化细菌生理生态特征的研究[J].哈尔滨工业大学学报,2006,38(4)：575-577.
4. 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,2007,28(7)：1548-1552.
5. 许保玖,龙腾锐.当代给水与废水处理原理(第2版)[M].北京：高等教育出版社,2000.350.
6. 郑平,徐向阳,胡宝兰.新型生物脱氮理论与技术[M].北京：科学出版社,2004.10-11.
7. 周立祥,黄峰源,王世梅.好氧反硝化菌株的分离及其在土壤氮素转化过程中的作用[J].土壤学报,2006,43(3)：430-435.
8. 朱晓宇.两株高效好氧反硝化细菌的分离鉴定及其好氧反硝化特性研究[D].南京：南京农业大学,2008.38-40.
9. Arts P A M, Robertson L A, Kuenen J G. Nitrification and denitrification by Thiosphaea pantotropha in aerobic chemostat cultures[J]. FEMS Microbiol Ecol,1995,18(4):305-315.
10. Gupta A B, Gupta S K. Simultaneous carbon and nitrogen removal from high strength domestic wastewater in an aerobic RBC biofilm[J].Water Res,2001,35(7):1714-1722.
11. Holman J B, Wareham D G. COD,ammonia and dissolved oxygen time profiles in the simultaneous nitrification/denitrification process[J]. Biochem Eng J,2005,22(2):125-133.
12. Hwang S, Hanaki K. Effects of oxygen concentration and moisture content of refuse on nitrification,denitrification and nitrous oxide production[J]. Bioresourse Technol, 2000,71(2):159-165.
13. Joo H S, Hirai M, Shoda M. Characteristids of ammonium removal by hererotrophic nitrification-aerobic denitrification by Alcaligenes faecalis no.4 and L1[J]. J Biosci Bioeng, 2005,100(2):184-73191.
14. Joo H S, Hirai M, Shoda M. Improvement in ammonium removal efficiency in wastewater treatment by mixed culture of Alcaligenes faecalis no.4 and L1[J]. J Biosci Bioeng, 2007,103(1):66-73.
15. Katarzyna B, Wojnowska-Baryta I. Carbon source in aerobic denitrification[J]. Biochem Eng J, 2007,36(2):116-122.
16. Kim J K, Park K. J, Cho K S,et al. Aerobic nitrification-denitrification by heterotrophic Bacillus strains[J]. Bioresource Technol,2005,96(17):1897-1906.
17. Kim M, Jeong S, Yoon S J,et al. Aerobic denitrification of Pseudomonas putida AD-21 at different C/N rations[J]. J Biosci Bioeng,2008,106(5):498-502.
18. Oguz M T, Robinson K G,Layton A C,et al. Volatile fatty acid impacts on nitrite oxidation and carbon dioxide fixation in activated sludge.[J]. Water Res,2006,40(4):665-674.
19. Patureau D, Bernet N, Delgenes J P,et al. Effect of dissolved oxygen and carbon-nitrogen loads on denitrification by an aerobic consortium[J]. Appl Microbiol Biot,2000,54(4):535-542.
20. Patureau D, Bernet N, Moletta R. Study of the denitrifying enzymatic system of Comamonas sp. Strain SGLY2 under various aeration conditions with a particular view on nitrate and nitrite reductases.[J]. Curr Microbiol,1996,32:25-32.
21. Robertson L A, Kuenen J G. Combined heterotrophic nitrification and aerobic denitrification in Thiosphaera pantotropha and other bacteria [J]. Antonie Van Leeuwenhoek,1990,57(5):139-152.
22. Sears H J,Sawers G,Berks B C. Control of periplasmic nitrate reductase gene expression (napEDABC) from Paracoccus pantotrophus in response to oxygen and carbon substrates. [J]. Micro,2000,146(11):2977-2985.
23. Su J J, Liu B Y, Liu C Y. Comparison of aerobic denitrification under high oxygen atmosphere by Thiosphaera pantotropha ATCC 35512 and Pseudomonas stutzeri SU2 newly isolated from the activated sludge of a piggery wastewater treatment system[J]. J Appl Microbiol, 2001,90(3):457-462.
24. Thomas K L,Lloyd D,Boddy L. Effects of oxygen, pH and nitrate concentration on denitrification by Pseudomonas species.[J]. FEMS Microbiol Lett,1994,118(1-2):181-186.
25. van Niel E W J, Braber K J, Robertson L A,et al. Heterotrophic nitrification and aerobic denitrification in Alcaligenes faecalis strain TUD[J]. Anton Leeuw Int J G,1992,62:231-237.
1. 黄慎勇,付中志,赵忠富,等.氮的排放标准与设计[J].西南给排水,2006,28(3)：7-9.
2. 唐建国,林洁梅.城镇污水处理厂的氮负荷分析[J].给水排水,2005,31(10)：31-35.
3. 王琳,李季,康文力,等.河流沉积物中反硝化细菌的分离及脱氮除磷研究[J].环境科学,2009,30(1)：91-95.
4. 王淑莹,孙洪伟,杨庆,等.传统生物脱氮反硝化过程的生化机理及动力学[J].应用与环境生物学报,2008,14(5)：732-736.
5. 叶姜瑜,罗固源,吉芳英.污水生物处理功能微生物的多样性[J].重庆大学学报：自然科学版,2005,28(10)：119-123.
6. 尹艳娥,沈新强,晁敏,等.反硝化技术对模拟养殖池塘修复的研究[J].华中农业大学学报,2009,28(8)：1727-1732.
7. 曾庆武,梁运祥,葛向阳.反硝化细菌的分离筛选及其反硝化特性的初步研究[J].农业环境科学学报,2008,27(5)：616-620.
8. 张小玲,梁运祥.一株反硝化细菌的分离筛选及其反硝化特性的研究[J].淡水渔业,2006,36(5)：28-32.
9. Aslan S, Cakici H. Biological denitrification of drinking water in a slow sand filter[J]. Journal of Hazardous Materials,2007,148:253-258.
10. Bark P S, Dold P L. General model for biological nutrient removal in activated sludge system: Model presentation[J]. Water Environ Res,1997,68(5):969-984.
11. Fernandez N, Alvarez R S, Field J A,et al. Microbial community dynamics in a chemolithotrophic denitrification reactor inoculatedwith methanogenic granular sludge[J]. Chemosphere,2008,70:462-468.
12. Gomeza M A,Gonzalez-L6pez J,Hontoria-Garcia E. Influence of carbon source on nitrate removal of contaminated groundwater in a denitrifying submerged filter[J]. Hazard Mater,2000, B80:69-80.
13. Jong O K, John U, Rick W Y. Diversity of oxygen and N-oxide regulation of nitrite reductases in denitrifying bacteria [J]. FEMS Microbiol Lett,1997,156 (1):55-60
14. Kenneth J, Forsh A Y, Emily Y H,et al. Rapid nitrate loss and denitrification in a temperate river floodplain[J]. Biogeo chemistry,2005,75:43-64.
15. Kirstein K, Bock E. Close genetic relation and characterization of the periplasmic reductase from Thiosphaerchia coli nitrate reductase [J]. Arch Microbial,1993,160:447-453.
16. Loosdrecht M C, Jetten S M. Microbilogical conversion in nitrogen removal[J]. Water Sci Ttechnol.1998,38:1-8.
17. Mahood T, Ali R, Malik K A,et al. Seasonal pattern of denitrification under an irrigated wheat-maize cropping system fertilized with urea and farmyard manure in different combinations[J]. Biol Fertil Soils,2005,42(1):1-9.
18. Modina O, Fukushib K, Yamamotoc K. Denitrification with methane as external carbon source[J]. Water Res,2007,41 (12):2726-2738.
19. Ovez B. Batch biological denitrification usingArundo donax,Glycyrrhiza glabra, andGracilaria verrucosaas carbon source[J].Process Biochemistry,2006,41:1289-1295.
20. Qiu X Y, Hurt R A, Wu L Y,et al. Detection and quantification of copper-denitrifying bacteria by quantitative competitive PCR [J]. J Microbiol Methods,2004,59(2):199-210.
21. Rijn J V, Tal Y, Schreier H J. Denitrification in recirculating systems:Theory and applications[J].Aquacul Eng,2006,34:364-376.
22. Rocca C D, Belgiorno V, Meric S. Heterotrophic/autotrophic denitrification (HAD) of drinking water:prospective use for permeable reactive barrier[J]. Desalination,2007,210:194-204.
23. Sharma B,Ahlert R C.Nitrification and nitrogen removal [J]. Water Res,1977,11(10):897-925.
24. Wang C C, Lee C M. Isolation of the ε-caprolactam denitrifying bacteria from wastewater treatment system manufactured with acrylonitrile-butadiene-styrene resin[J]. J Hazardous Mat, 2007,145:136-141.
25. Ye R W,Avrill B A,Tiedje J M. Denitrification:Production and consumption of nitric oxide[J]. Appl EnvironMicrobial,1994,60 (4):1053-1058.
26. Zumft W G, Kroneck P M H. Respiratory transformation of nitrous oxide(N20)to dinitrogen by bacteria and arcbaea [J]. Adv Microb Physiol,2006,52:107-227.
1. 管运涛,吴晓磊,钱易,等.生物钙法好氧污泥颗粒化条件研究[J].给水排水,1996,22(2)：27-29.
2. 江滢.有关Zeta电位的若干问题[J].中国造纸,1976,(2)：54-55.
3. 李浩,袁林江.好氧SBR反应器中污泥颗粒化过程的成核研究[J].中国给水排水,2008,24(13)：42-45.
4. 李久义,吴晓清,陈福泰,等.Fe(Ⅲ)对活性污泥絮体结构和生物絮凝的影响[J].环境科学学报,2003,23(5)：582-587.
5. 刘丽,任婷婷,徐得潜,等.高强度好氧颗粒污泥的培养及特性研究[J].中国环境科学,2008,28(4)：360-364.
6. 刘倩倩,李小明,杨麒,等.Mg2+对SBR中好氧颗粒污泥培养的影响研究[J].中国给水排水,2008,24(17)：31-35.
7. 王建龙,张子健,吴伟伟.好氧颗粒污泥的研究进展[J].环境科学学报,2009,29(3)：449-473.
8. 王建龙.生物固定化技术与水污染控制[M].北京：科学出版社,2002.122-125.
9. 吴立波,王建龙,黄霞,等.自固定化技术强化高效菌种活性的保持[J].环境科学,2000,21(1)：32-35.
10.张丽丽,陈效,陈建孟,等.胞外多聚物在好氧颗粒污泥形成中的作用机制[J].环境科学,2007,28(4)：795-799.
11. Adav S S, Lee D J, Show K Y,et al. Aerobic granular sludge:recent advances [J].Biotechnol Adv, 2008,26(5):411-423.
12. Apostolos V, Elli M B, Sofia M. Influence of ferrous iron on the granularity of a UASB reactor[J].Chem Eng J,2009,146(1):49-56.
13. Batstone D J, Landelli J, Saunders A,et al. The influence of calcium on granular sludge in a full-scale UASB treating paper mill wastewater [J].Water Sci Technol,2002,45(10):187-193.
14. Jackson C R, Roden E E, Church ill P F. Denaturing gradient gel electropho resis can fail to separate 16S rDNA fragment w ith multip le base differences[J]. Microbiol Today,2000,1 (2): 49-51.
15. Jiang H L, Tay J H, Liu Y,et al. Ca2+ augumentation for enhancement of aerobically grown microbial granules in sludge blanket recators[J]. Biotechnol Lett,2003,25(2):95-99.
16. Laspidou C S, Rittmann B E. A unified theory for extracellular polymeric substances, soluble microbial products, and active and inert biomass [J]. Water Res,2002,36(11):2711-2720.
17. Li X M, Liu Q Q, Yang Q,et al. Enhanced aerobic sludge granulation in sequencing batch reactor by
18. Mg2+ augmentation [J].Bioresource Technol,2009,100(1):64-67.
19. Muyzer G, Waal E C de, Uitterlinden A G. Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol,1993,59:695-700.
20. Muyzer G, Smalla K. Application of denaturing gradient gel electrophoresis (DGGE) and temperature gradient gel electrophoresis (TGGE) in microbial ecology[J]. Antonie Van Leeuwenhoek,1998,73:127-41.
21. Muyzer G. DGGE/TGGE a method for identifying genes from natural ecosystems [J]. Current Microbiol,1999,2:317-322.
22. Pavoni J L, Tenny M W, Echelberger Jr W F. Bacterial extracellular polymers and biological flocculation [J]. Water Pollut Control Fed,1972,44(3):414-431.
23. Qin L, Tay J H, Liu Y. Selection pressure is a driving force of aerobic granulation in sequencing batch reactor [J]. Process Biochem,2004,39(5):79-84.
24. Ren T T, Liu L, Sheng G P,et al. Calcium spatial distribution in aerobic granules and its effects on granule structure, strength and bioactivity[J].Water Res,2008,42(3):3343-3352.
25. Rose R K. The role of calcium in oral streptococcal aggregation and the implications for biofilm formation and rentation[J].Biochim Biophys Acta,2000,1475(1):76-82.
26. Schmidt J E, Ahring B K. Granular sludge formation in upflow anaerobic sludge blanket(UASB) reactors[J]. Biotechnol Bioeng,1996,49(3):229-246.
27. Sigler W V, Miniaci C, Zeyer J. Electrophoresis time impacts the denaturing gradient gel electrophoresis-based assessment of bacterial community structure[J]. J Microbiol Methods 2004,57:17-22.
28. Wang Z W, Li Y, Liu Y. Mechanism of calcium accumulation in acetate-fed aerobic granule [J].Appl Microbiol Biot,2007,74(2):467-473.
29. Yu H Q, Fang H H H, Tay J H. Effects of Fe2+ on sludge granulation in upflow anaerobic sludge blanket reactors[J]. Water Sci Technol,2000,41(12):199-205.
30. Yu H Q, Fang H H H, Tay J H. The roles of calcium in sludge granulation during UASB reactor start-up[J].Water Res,2001,35(4):1052-1060.
31. Yu H Q, Tay J H, Fang H H H. Enhanced sludge granulation in upflow anaerobic sludge blanket(UASB) reactors by aluminum chloride[J].Chemosphere,2001,44(1):31-3.
2. 李丛娜,吕锡武,稻森悠平.同步硝化反硝化脱氮研究.给水排水[J].给水排水,2001,27(1)：22-24.
3. 林燕,孔海南,王茸影,等.异养硝化作用的主要特点及其研究动向[J].环境科学,2008,29(11)：3291-3296.
4. 刘幽燕,何玉财,李青云,等.沸石固定化细胞降解氰化物的试验研究[J].高校化学工程学报,2005,19(4)：532-535.
5. 吕锡武,李峰,稻森悠平,等.氨氮废水处理工程中的好氧反硝化研究[J].给水排水,2000,26(4)：17-20.
6. 马放,周丹丹,王宏宇,等.一株好氧反硝化细菌生理生态特征的研究[J].哈尔滨工业大学学报,2006,38(4)：575-577.
7. 牟丽娉,黄钧,苟莎.异养硝化微生物菌剂及其好氧颗粒污泥的脱氮试验[J].应用与环境生物学报,2009,15(3)：356-360.
8. 阮文权,陈坚.同步脱氮好氧颗粒污泥的特性及其反应过程[J].中国环境科学,2003,23(4)：380-384.
9. 孙锦宜.含氮废水处理技术与应用[M].北京：化学工业出版社,2003.33.
10.汪苹,项慕飞,翟茜,等.从不同反应器筛选、鉴别好氧反硝化菌[J].环境科学研究,2007,20(4)：120-124.
11.王红娟,奚红霞,夏启斌,等.含酚废水处理技术的现状与开发前景[J].工业水处理,2002,22(6)：6-9.
12.王薇,蔡祖聪,钟文辉,等.好氧反硝化菌的研究进展[J].应用生态学报,2007,18(11)：2618-2625.
13.王旭明,从二丁,罗文龙,等.固体碳源用于异养反硝化去除地下水中的硝酸盐[J].中国科学B辑：化学,2008,38(9)：824-828.
14.叶建锋.废水生物脱氮处理新技术[M].北京：化学工业出版社,2006,10-12.
15.张立秋,张可方,张朝升,等.DO对亚硝酸型SND的影响[J].水处理技术,2008,34(8)：29-33.
16.郑平,徐向阳,胡宝兰.新型生物脱氮理论与技术[M].北京：科学出版社,2004,5-7.
17.郑兴灿,李亚新.污水除磷脱氮技术[M].北京：中国建筑工业出版社,1998,55-57.
18.朱亮,徐向阳,罗伟国,等.废水生物处理好氧污泥颗粒化进展[J].环境科学,2007,28(11)：2657-2664.
19.竺建荣,刘纯新.好氧颗粒活性污泥的培养及理化特性研究[J].环境科学,1999,20(2)：38-41.
20. Arlca M Y, Kacar Y, Gene O. Entrapment of white-rot fungus trameters versicolor in Ca-alginate beads:preparation and biosorption kinetics analysis for cadmium removal from an aqueous solution [J]. Bioresourse Technol,2001,80(2):121-129.
21. Bateman E J, Baggs E M. Contributions of nitrification and denitrification to N2O emissions from soils at different waterfilled pore space[J]. Biology and Fertility of soils,2005,41(6):379-388.
22. Bayramoglu G, Denizlia, Bektas S,et al. Entrapment of Lentinus sajor-caju into Ca-alginate gel beads for removal of Cd(II)ions from aqueous solution:preparation and biosorption kinetics analysis [J]. Micro-Chemical Journal,2002,72(1):63-76.
23. Ben J J, Heijnen J J, van Loosdrecht M C M. N-removal in a granular sludge sequencing batch airlift reactor [J].Biotechnol Bioeng,2001,75(1):82-92.
24. Brierley E D R,Wood M.Heterotrophic nitrification in an acid forest soil:isolation and characterization of a nitrifying bacterium[J].Soil Biol Biochem,2001,33:1403-1409.
25. Chen F, Xia Q, Ju L K. Aerobic denitrification of Pseudomonas aeruginosa monitored by online NAD(P) H fluorescence[J].Appl Envir Mcrobiol,2003,69(11):6715-6722.
26. Doxtader K G, Alexander M. Nitrification by heterotrophic soil microorganisms[J].Soil Sci Soc Amer Proc,1966,30:351-355.
27. Geraats S G M, Hooijmans C M, van Niel E W J. The use of a metabolically structured model in the study of growth Nitrification and denitrification by Thiosphaera pantotropha[J].Biotechnol Bioeng,1990,36:227-267.
28. Henning P,Kristie A D, Mary K F. The relative importance of autotrophic and heterotrophic nitrification in a conifer forest soil as measured by 15N tracer and pool dilution techniques[J]. Biogeochemistry,1999,44:135-150.
29. Kowalchuk G A,Stephen J R. Ammonia-oxidizing bacteria:A model for molecular microbial ecology.[J]. Annu Rev Microbiol,2001,55:485-529.
30. Kshirsagar M, Gupta A B, Gupta S K. Aerobic denitrification studies on activated sludge mixed with Thiosphaera pantotropha[J].Environ Tech,1995,16:35-43.
31. Kuai L,Verstrartr W. Autotrophic denitrification with elemental sulphur in small-scale wastewater treatment facilities[J].Environ Tech,1999,20:201-209.
32. Lin Y M, Liu Y, Tay J H. Development and characteristics of phosphorous-accumulating granules in sequencing batch reactor [J]. Appl Microbial Biotechnol,2003,62(4):25-43.
33. Liu Y, Wang Z W, Tay J H. A unified theory for upscaling aerobic granular sludge sequencing batch reactor [J]. Biotechnol Adv,2005,23(5):335-344.
34. Mchmidt I, Sliekers O, Schmid M, et al. New concepts of microbial treatment processes for the nitrogen removal in wastewater [J]. FEMS Microbiology Reviews,2003,27:481-492.
35. Metcalf, Eddy, Inc. Wastewater Engineering Treatment and Reuse.4th ed. USA:MeGraw-Hill Companies,2003
36. Muller C, Stevens R J, Laughlin R J. An N-15 tracing model to analyze N transformations in old grassland soil[J]. Soil Biology Biochemistry,2004,36(4):619-632.
37. Nishio T, Yoshikura T. Effects of organic acids on heterotrophic nitrification by Alaligenes faecalis OKKI 7[J].Biosci Biotech Biochem,1994,58:1574-1578.
38. Peng D C,Bernet N, Delgenes J P, et al. Aerobic granular sludge-a case report[J]. Water Res,1999, 33(3):890-893.
39. Robertson L A, Kuenen J G. Aerobic denitrification:A controversy revived[J]. Archives of Microbiology,1984,139:351-354.
40. Robertson L A, Van Niel E W J, Torremans R A M. Simultaneous nitrification and denitrification in aerobic chemostat cultures of Thiosphaera pantotropha[J].Appl Envir Mcrobiol,1988,54(11): 2812-2818.
41. Sen S, Demirer G N. Anaerobic treatment of real textile wastewater with a fluidized bed reactor [J]. Water Res,2003,37:1868-1878.
42. Sierra-Alvarez R,Beristain-Cardoso R,Salazar M,et al. Chemolithotrophic denitrification with elemental sulfur for groundwater treatment[J]. Water Res,2007,41:1253-1262.
43. Soares M I M, Abeliovich A. Wheat straw as substrate for water denitrification [J]. Water Res,1998, 32(12):3790-3794.
44. Stephen J R, Kowalchuk G A, Brun M V,et al. Analysis of a-subgroup proteobacteria ammonia oxidizer populations in soil by denaturing gradient gel electrophoresis analysis and hierarchical phylogentic probing [J]. Appl Environ Microbial,1998,64:2958-2965.
45. Takaya N, Catalan S M, Sakaguchi Y,et al. Aerobic denitrification bacteria that produced low levels of nitrous oxide[J].Appl Envir Mcrobiol,2003,69(6):3152-3157.
46. Yoo H,Ahn K,Lee H,et al. Nitrogen removal from synthetic wastewater by simultaneous nitrification and denitrification(SND) via nitrite in an intermittently reactor[J]. Water Res,1999, 33(1):145-154.
47. Zhao H W,Mavinic D S,Oldham W K,et al. Controlling factors for simultaneous nitrification and denitrification in a two-stage intermittent aeration process treating domestic sewage[J]. Water Res, 1999,33(4):961-970.
1. (美)奥斯伯F M.精编分子生物学试验指南.科学出版社.2005.1457-1574.
2. 北京市市政工程设计研究总院.给水排水设计手册(第5册-城镇排水)(第2版)[M].北京：中国建筑工业出版社,2004.245.
3. 东秀珠,蔡妙英.常见细菌系统鉴定手册[M].北京：科学出版社,2001.370-378.
4. 高廷耀,顾国维,周琪.水污染控制工程(第3版)[M].北京：高等教育出版社,2007.102-103.
5. 胡君利,林先贵,褚海燕,等.土壤氨氧化细菌的分离方法研究[J].土壤,2005,37(5)：569-571.
6. 周群英,高廷耀.环境工程微生物学(第二版)[M].北京：高等教育出版社,1999.122.
7. GB 11894-89,碱性过硫酸钾消解紫外分光光度法[S].
8. GB 7479-87,纳氏试剂比色法[S].
9. GB 7493-87,N-(1-萘基)-乙二胺分光光度法[S].
10.HJ/T 346-2007,紫外分光光度法[S].
11.GB 11893-89,铝酸铵分光光度法[S].
12.李阜棣,喻子牛,何绍江.农业微生物学试验技术[M].北京：中国农业出版社,1996.308.
13.土壤微生物研究会编.土壤微生物试验法[M].北京：科学出版社,2001.656.
14.沈萍,范秀容,李广武.微生物学试验(第三版)[M].北京：中国农业出版社,1998.89-87.
15.袁飞.不同农田土壤中的硝化作用及硝化细菌种群[D].南京：南京农业大学,2004.118-121.
16.张纪忠.微生物分类学[M].上海：复旦大学出版社,1990.108.
17. Edwards U, Rogall T, Blocker H,et al. Isolation and direct complete determination of entire genes. Nucleic Acids Res.1989.17:7843-7853.
18. Ferris, M J, Muyzer G, Ward D M. Denaturing gradient gel electrophoresis profiles of 16S rRNA-defined populations inhabiting a hot spring microbial mat community [J]. Appl Environ Microb,1996,62:340-346.
19. Heuer H, Krsek M, Baker P,et al. Analysis of actinomycete communities by specific amplification of genes encoding 16S rRNA and gel-electrophoretic separation in denaturing gradients[J]. Appl Environ Microb,1997,63:3233-3241.
20. Sigler W V, Miniaci C, Zeyer J. Electrophoresis time impacts the denaturing gradient gel electrophoresis-based assessment of bacterial community structure[J]. J Microbiol Methods 2004,57:17-22.
21. Teske A, Wawer C, Muyzer G,et al. Distribution of sulfate-reducing bacteria in a stratified fjord (Mariager Fjord, Denmark) as evaluated by most-probable-number counts and denaturing gradient gel electrophoresis of PCR-amplified ribosomal DNA fragments[J]. Appl Environ Microb, 1996,62:1405-1415.
22. Wilson K H, Blitchington R B, Green R C. Amplification of 16S ribosomal DNA with polymerase chain reaction. Clin Microbiol,1990.28:1942-1946.
1. 何霞,赵彬,吕剑,等.异养硝化细菌Bacillus sp.LY脱氮特性研究[J].环境科学,2007,28(6)：1404-1408.
2.胡咏梅,葛向阳,梁运祥.枯草芽孢杆菌FY99-01菌株的净水作用[J].华中农业大学学报,2006,25(4)：404-407.
3. 刘晶晶,汪苹,王欢.一株异养硝化-好氧反硝化菌的脱氮特性研究[J].环境科学研究,2008,21(3)：121-125.
4. 罗固源,杨红薇NBIAS系统中的好氧反硝化.重庆环境科学,2001,23(1)：43-46.
5. 王弘宇,马放,杨开,等.两株异养硝化细菌的氨氮去除特性[J].中国环境科学,2009,29(1)：47-52.
6. 熊伟,梁运祥,戴经元,等.枯草芽孢杆菌对斑节对虾饲养池水净化作用的初步研究[J].华中农业大学学报,2003,22(3)：247-250.
7. 杨航,黄钧,刘博.异养硝化-好氧反硝化菌Paracoccus pantotrophus ATCC 35512的研究进展[J].应用与环境生物学报,2008,14(4)：585-592.
8. 张光亚,陈美慈,韩如韩,等.一株异养硝化细菌的分离及系统发育分析[J].微生物学报,2003,43(2)：156-161.
9. Berks B C, Richardson D J, Reilly A,et al. The napEDABC gene cluster encoding the periplasmic nitrate reductase system of Thiosphaera pantotropha[J]. Biochem J,1995,309:983-992.
10. Crossman L C, Moir J W B, Enticknap J J,et al. Heterologous expression of heterotrophic nitrification genes [J]. Microbiology,1997,143(12):3775-3783.
11. Cartron M L, Roldan M D, Ferguson S J,et al. Identification of two domains and distal histidine ligands to the four haems in the bacterial c-type cytochrome NapC; the prototype connector between quinol/quinone and periplasmic oxido-reductases. [J]. Biochem J,2002,368:425-432.
12. Castignetti D, Hollocher T C. Nitrogen redox metabolism of a hetetoteophic nitrifying-denitrifying Alcaligenes sp. from soil[J]. Appl Environ Microbiol,1982,44(4):923-928.
13. Duggin J A, Voigt G K. Autotrophic and heterotrophic nitrification in response to clear-cutting northern hardwood forest.[J]. Soil Biol Biochem,1991,23(8):779-787.
14. Hooper A B, Vannelli T, Bergmann D J,et al. Enzymology of the oxidation of ammonia to nitrite by bacteria[J]. Antonie Van Leeuwenhoek,1997,71(1-2):59-67.
15. Hu T L, Kung K T. Study of heterotrophic in nitrifying bacteria from wastewater treatment systems treating acrylonitrile,butadiene and styrene resin wastewater[J]. Wat Sci Tech, 2000,42(3-4):315-322.
16. Joong K K, Kyoung J P, Kyoung S C,et al. Aerobic nitrification-denitrification by heterotrophic Bacillus strains[J]. Bioresource Technol,2005,96(17):1897-1906.
17. Kelly D P, Euzeby J P, Enticknap J J,et al. Redefining Paracoccus denitrificans and Paracoccus pantotrophus and the case for a reassessment of the strains held by international culture collections [J]. Int J Syst Evol Microbiol,2006,56(10):2495-2500.
18. Pedersen H, Dunkin K A, Firestone M K. The relative importance of autotrophic and heterotrophic nitrification in a conifer forest soil as measured by N-15 tracer and pool dilution techniques[J]. Biogeochemistry,1999,44(2):135-150.
19. Richardson D J, Ferguson S J. The influence of carbon substrate on the activity Thiosphaera pantotropha of the periplasmic nitrate reductase in aerobically grown[J]. Arch Microbiol,1992,157 (6):535-537.
20. Schimel J P, Firestone M K. Identification of heteroteophic nitrification in a sierran forest soil[J]. Appl Environ Microbiol,1984,48(4):802-806.
21. Stouthamer A H, De Boer A P N, Van der Oost J,et al. Emerging principles of inorganic nitrogen metabolism in Paracoccus denitrificans and related bacteria[J]. Antonie Van Leeuwenhoek, 1997,71(1-2):33-41.
22. Wehrfritz J M, Reilly A, Spiro S,et al. Purification of hydroxylamine oxidase from Thiosphaera pantotropha. Identification of electron acceptors that couple heterotropbic nitrification to aerobic denitrification[J]. FEBS Lett,1993,335(2):246-250.
23. Witzel K P, Overbeck H J. Heteroteophic nitrification by Arthtobacter sp.(strain 9006) as influenced by different cultural conditions growth state and acetate metabolism[J]. Arch Microbiol,1979,122:137-143.
1. 孔庆鑫,李君文,王新为,等.一种新的好氧反硝化细菌筛选方法的建立及新菌株的发现[J].应用与环境生物学报,2005,11(2)：222-225.
2. 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,2006,26(11)：3718-3724.
3. 马放,周丹丹,王弘宇,等.一株好氧反硝化细菌生理生态特征的研究[J].哈尔滨工业大学学报,2006,38(4)：575-577.
4. 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,2007,28(7)：1548-1552.
5. 许保玖,龙腾锐.当代给水与废水处理原理(第2版)[M].北京：高等教育出版社,2000.350.
6. 郑平,徐向阳,胡宝兰.新型生物脱氮理论与技术[M].北京：科学出版社,2004.10-11.
7. 周立祥,黄峰源,王世梅.好氧反硝化菌株的分离及其在土壤氮素转化过程中的作用[J].土壤学报,2006,43(3)：430-435.
8. 朱晓宇.两株高效好氧反硝化细菌的分离鉴定及其好氧反硝化特性研究[D].南京：南京农业大学,2008.38-40.
9. Arts P A M, Robertson L A, Kuenen J G. Nitrification and denitrification by Thiosphaea pantotropha in aerobic chemostat cultures[J]. FEMS Microbiol Ecol,1995,18(4):305-315.
10. Gupta A B, Gupta S K. Simultaneous carbon and nitrogen removal from high strength domestic wastewater in an aerobic RBC biofilm[J].Water Res,2001,35(7):1714-1722.
11. Holman J B, Wareham D G. COD,ammonia and dissolved oxygen time profiles in the simultaneous nitrification/denitrification process[J]. Biochem Eng J,2005,22(2):125-133.
12. Hwang S, Hanaki K. Effects of oxygen concentration and moisture content of refuse on nitrification,denitrification and nitrous oxide production[J]. Bioresourse Technol, 2000,71(2):159-165.
13. Joo H S, Hirai M, Shoda M. Characteristids of ammonium removal by hererotrophic nitrification-aerobic denitrification by Alcaligenes faecalis no.4 and L1[J]. J Biosci Bioeng, 2005,100(2):184-73191.
14. Joo H S, Hirai M, Shoda M. Improvement in ammonium removal efficiency in wastewater treatment by mixed culture of Alcaligenes faecalis no.4 and L1[J]. J Biosci Bioeng, 2007,103(1):66-73.
15. Katarzyna B, Wojnowska-Baryta I. Carbon source in aerobic denitrification[J]. Biochem Eng J, 2007,36(2):116-122.
16. Kim J K, Park K. J, Cho K S,et al. Aerobic nitrification-denitrification by heterotrophic Bacillus strains[J]. Bioresource Technol,2005,96(17):1897-1906.
17. Kim M, Jeong S, Yoon S J,et al. Aerobic denitrification of Pseudomonas putida AD-21 at different C/N rations[J]. J Biosci Bioeng,2008,106(5):498-502.
18. Oguz M T, Robinson K G,Layton A C,et al. Volatile fatty acid impacts on nitrite oxidation and carbon dioxide fixation in activated sludge.[J]. Water Res,2006,40(4):665-674.
19. Patureau D, Bernet N, Delgenes J P,et al. Effect of dissolved oxygen and carbon-nitrogen loads on denitrification by an aerobic consortium[J]. Appl Microbiol Biot,2000,54(4):535-542.
20. Patureau D, Bernet N, Moletta R. Study of the denitrifying enzymatic system of Comamonas sp. Strain SGLY2 under various aeration conditions with a particular view on nitrate and nitrite reductases.[J]. Curr Microbiol,1996,32:25-32.
21. Robertson L A, Kuenen J G. Combined heterotrophic nitrification and aerobic denitrification in Thiosphaera pantotropha and other bacteria [J]. Antonie Van Leeuwenhoek,1990,57(5):139-152.
22. Sears H J,Sawers G,Berks B C. Control of periplasmic nitrate reductase gene expression (napEDABC) from Paracoccus pantotrophus in response to oxygen and carbon substrates. [J]. Micro,2000,146(11):2977-2985.
23. Su J J, Liu B Y, Liu C Y. Comparison of aerobic denitrification under high oxygen atmosphere by Thiosphaera pantotropha ATCC 35512 and Pseudomonas stutzeri SU2 newly isolated from the activated sludge of a piggery wastewater treatment system[J]. J Appl Microbiol, 2001,90(3):457-462.
24. Thomas K L,Lloyd D,Boddy L. Effects of oxygen, pH and nitrate concentration on denitrification by Pseudomonas species.[J]. FEMS Microbiol Lett,1994,118(1-2):181-186.
25. van Niel E W J, Braber K J, Robertson L A,et al. Heterotrophic nitrification and aerobic denitrification in Alcaligenes faecalis strain TUD[J]. Anton Leeuw Int J G,1992,62:231-237.
1. 黄慎勇,付中志,赵忠富,等.氮的排放标准与设计[J].西南给排水,2006,28(3)：7-9.
2. 唐建国,林洁梅.城镇污水处理厂的氮负荷分析[J].给水排水,2005,31(10)：31-35.
3. 王琳,李季,康文力,等.河流沉积物中反硝化细菌的分离及脱氮除磷研究[J].环境科学,2009,30(1)：91-95.
4. 王淑莹,孙洪伟,杨庆,等.传统生物脱氮反硝化过程的生化机理及动力学[J].应用与环境生物学报,2008,14(5)：732-736.
5. 叶姜瑜,罗固源,吉芳英.污水生物处理功能微生物的多样性[J].重庆大学学报：自然科学版,2005,28(10)：119-123.
6. 尹艳娥,沈新强,晁敏,等.反硝化技术对模拟养殖池塘修复的研究[J].华中农业大学学报,2009,28(8)：1727-1732.
7. 曾庆武,梁运祥,葛向阳.反硝化细菌的分离筛选及其反硝化特性的初步研究[J].农业环境科学学报,2008,27(5)：616-620.
8. 张小玲,梁运祥.一株反硝化细菌的分离筛选及其反硝化特性的研究[J].淡水渔业,2006,36(5)：28-32.
9. Aslan S, Cakici H. Biological denitrification of drinking water in a slow sand filter[J]. Journal of Hazardous Materials,2007,148:253-258.
10. Bark P S, Dold P L. General model for biological nutrient removal in activated sludge system: Model presentation[J]. Water Environ Res,1997,68(5):969-984.
11. Fernandez N, Alvarez R S, Field J A,et al. Microbial community dynamics in a chemolithotrophic denitrification reactor inoculatedwith methanogenic granular sludge[J]. Chemosphere,2008,70:462-468.
12. Gomeza M A,Gonzalez-L6pez J,Hontoria-Garcia E. Influence of carbon source on nitrate removal of contaminated groundwater in a denitrifying submerged filter[J]. Hazard Mater,2000, B80:69-80.
13. Jong O K, John U, Rick W Y. Diversity of oxygen and N-oxide regulation of nitrite reductases in denitrifying bacteria [J]. FEMS Microbiol Lett,1997,156 (1):55-60
14. Kenneth J, Forsh A Y, Emily Y H,et al. Rapid nitrate loss and denitrification in a temperate river floodplain[J]. Biogeo chemistry,2005,75:43-64.
15. Kirstein K, Bock E. Close genetic relation and characterization of the periplasmic reductase from Thiosphaerchia coli nitrate reductase [J]. Arch Microbial,1993,160:447-453.
16. Loosdrecht M C, Jetten S M. Microbilogical conversion in nitrogen removal[J]. Water Sci Ttechnol.1998,38:1-8.
17. Mahood T, Ali R, Malik K A,et al. Seasonal pattern of denitrification under an irrigated wheat-maize cropping system fertilized with urea and farmyard manure in different combinations[J]. Biol Fertil Soils,2005,42(1):1-9.
18. Modina O, Fukushib K, Yamamotoc K. Denitrification with methane as external carbon source[J]. Water Res,2007,41 (12):2726-2738.
19. Ovez B. Batch biological denitrification usingArundo donax,Glycyrrhiza glabra, andGracilaria verrucosaas carbon source[J].Process Biochemistry,2006,41:1289-1295.
20. Qiu X Y, Hurt R A, Wu L Y,et al. Detection and quantification of copper-denitrifying bacteria by quantitative competitive PCR [J]. J Microbiol Methods,2004,59(2):199-210.
21. Rijn J V, Tal Y, Schreier H J. Denitrification in recirculating systems:Theory and applications[J].Aquacul Eng,2006,34:364-376.
22. Rocca C D, Belgiorno V, Meric S. Heterotrophic/autotrophic denitrification (HAD) of drinking water:prospective use for permeable reactive barrier[J]. Desalination,2007,210:194-204.
23. Sharma B,Ahlert R C.Nitrification and nitrogen removal [J]. Water Res,1977,11(10):897-925.
24. Wang C C, Lee C M. Isolation of the ε-caprolactam denitrifying bacteria from wastewater treatment system manufactured with acrylonitrile-butadiene-styrene resin[J]. J Hazardous Mat, 2007,145:136-141.
25. Ye R W,Avrill B A,Tiedje J M. Denitrification:Production and consumption of nitric oxide[J]. Appl EnvironMicrobial,1994,60 (4):1053-1058.
26. Zumft W G, Kroneck P M H. Respiratory transformation of nitrous oxide(N20)to dinitrogen by bacteria and arcbaea [J]. Adv Microb Physiol,2006,52:107-227.
1. 管运涛,吴晓磊,钱易,等.生物钙法好氧污泥颗粒化条件研究[J].给水排水,1996,22(2)：27-29.
2. 江滢.有关Zeta电位的若干问题[J].中国造纸,1976,(2)：54-55.
3. 李浩,袁林江.好氧SBR反应器中污泥颗粒化过程的成核研究[J].中国给水排水,2008,24(13)：42-45.
4. 李久义,吴晓清,陈福泰,等.Fe(Ⅲ)对活性污泥絮体结构和生物絮凝的影响[J].环境科学学报,2003,23(5)：582-587.
5. 刘丽,任婷婷,徐得潜,等.高强度好氧颗粒污泥的培养及特性研究[J].中国环境科学,2008,28(4)：360-364.
6. 刘倩倩,李小明,杨麒,等.Mg2+对SBR中好氧颗粒污泥培养的影响研究[J].中国给水排水,2008,24(17)：31-35.
7. 王建龙,张子健,吴伟伟.好氧颗粒污泥的研究进展[J].环境科学学报,2009,29(3)：449-473.
8. 王建龙.生物固定化技术与水污染控制[M].北京：科学出版社,2002.122-125.
9. 吴立波,王建龙,黄霞,等.自固定化技术强化高效菌种活性的保持[J].环境科学,2000,21(1)：32-35.
10.张丽丽,陈效,陈建孟,等.胞外多聚物在好氧颗粒污泥形成中的作用机制[J].环境科学,2007,28(4)：795-799.
11. Adav S S, Lee D J, Show K Y,et al. Aerobic granular sludge:recent advances [J].Biotechnol Adv, 2008,26(5):411-423.
12. Apostolos V, Elli M B, Sofia M. Influence of ferrous iron on the granularity of a UASB reactor[J].Chem Eng J,2009,146(1):49-56.
13. Batstone D J, Landelli J, Saunders A,et al. The influence of calcium on granular sludge in a full-scale UASB treating paper mill wastewater [J].Water Sci Technol,2002,45(10):187-193.
14. Jackson C R, Roden E E, Church ill P F. Denaturing gradient gel electropho resis can fail to separate 16S rDNA fragment w ith multip le base differences[J]. Microbiol Today,2000,1 (2): 49-51.
15. Jiang H L, Tay J H, Liu Y,et al. Ca2+ augumentation for enhancement of aerobically grown microbial granules in sludge blanket recators[J]. Biotechnol Lett,2003,25(2):95-99.
16. Laspidou C S, Rittmann B E. A unified theory for extracellular polymeric substances, soluble microbial products, and active and inert biomass [J]. Water Res,2002,36(11):2711-2720.
17. Li X M, Liu Q Q, Yang Q,et al. Enhanced aerobic sludge granulation in sequencing batch reactor by
18. Mg2+ augmentation [J].Bioresource Technol,2009,100(1):64-67.
19. Muyzer G, Waal E C de, Uitterlinden A G. Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol,1993,59:695-700.
20. Muyzer G, Smalla K. Application of denaturing gradient gel electrophoresis (DGGE) and temperature gradient gel electrophoresis (TGGE) in microbial ecology[J]. Antonie Van Leeuwenhoek,1998,73:127-41.
21. Muyzer G. DGGE/TGGE a method for identifying genes from natural ecosystems [J]. Current Microbiol,1999,2:317-322.
22. Pavoni J L, Tenny M W, Echelberger Jr W F. Bacterial extracellular polymers and biological flocculation [J]. Water Pollut Control Fed,1972,44(3):414-431.
23. Qin L, Tay J H, Liu Y. Selection pressure is a driving force of aerobic granulation in sequencing batch reactor [J]. Process Biochem,2004,39(5):79-84.
24. Ren T T, Liu L, Sheng G P,et al. Calcium spatial distribution in aerobic granules and its effects on granule structure, strength and bioactivity[J].Water Res,2008,42(3):3343-3352.
25. Rose R K. The role of calcium in oral streptococcal aggregation and the implications for biofilm formation and rentation[J].Biochim Biophys Acta,2000,1475(1):76-82.
26. Schmidt J E, Ahring B K. Granular sludge formation in upflow anaerobic sludge blanket(UASB) reactors[J]. Biotechnol Bioeng,1996,49(3):229-246.
27. Sigler W V, Miniaci C, Zeyer J. Electrophoresis time impacts the denaturing gradient gel electrophoresis-based assessment of bacterial community structure[J]. J Microbiol Methods 2004,57:17-22.
28. Wang Z W, Li Y, Liu Y. Mechanism of calcium accumulation in acetate-fed aerobic granule [J].Appl Microbiol Biot,2007,74(2):467-473.
29. Yu H Q, Fang H H H, Tay J H. Effects of Fe2+ on sludge granulation in upflow anaerobic sludge blanket reactors[J]. Water Sci Technol,2000,41(12):199-205.
30. Yu H Q, Fang H H H, Tay J H. The roles of calcium in sludge granulation during UASB reactor start-up[J].Water Res,2001,35(4):1052-1060.
31. Yu H Q, Tay J H, Fang H H H. Enhanced sludge granulation in upflow anaerobic sludge blanket(UASB) reactors by aluminum chloride[J].Chemosphere,2001,44(1):31-3.