徐翩翩,孟佳,汪聪,李建政
(城市水资源与水环境国家重点实验室(哈尔滨工业大学),哈尔滨 150090)
摘要:
为解决低碳氮比有机废水生物脱氮的难题,经济高效的厌氧氨氧化（ANAMMOX）技术得到广泛研究,但ANAMMOX菌群对环境要求苛刻且富集困难,系统启动缓慢,限制了其工程应用.前期研究中,构建了用于处理高氨氮低碳氮比养猪废水的升流式限氧生物膜反应器（UOLBR）,经180 d的启动运行实现了以ANAMMOX为主导的脱氮功能.将UOLBR闲置2个月,以COD与TN比为0.6～1.0的干清粪养猪废水为进水,在水力停留时间10 h、25 ℃和出水回流比25∶1的条件下再次启动,考察其处理效果和ANAMMOX功能的恢复情况.结果表明,经过溶解氧分别为2.5～3.0和0.2～0.5 mg/L两个阶段,共计53 d的连续运行,UOLBR的处理效能得以恢复并达到稳定状态,对COD、NH+4-N、TN的平均去除率分别为64.0%,96.5%和91.7%,出水质量浓度分别仅为1,7.0和16.4 mg/L左右.微生物群落结构和物料平衡分析表明,UOLBR系统恢复了以ANAMMOX为主要脱氮途径的特征.该研究可为低碳氮比有机废水ANAMMOX处理系统的启动和运行管理提供指导.
关键词: 限氧 废水 低COD与TN比 生物脱氮 生物膜
DOI：10.11918/j.issn.03676234.201801014
分类号:X703.1
文献标识码:A
基金项目:中国博士后科学基金(2017M611376)
Restart-up of an oxygen limitation wastewater treatment process for the recovery of ANAMMOX
XU Pianpian,MENG Jia,WANG Cong,LI Jianzheng
(State Key Laboratory of Urban Water Resource and Environment (Harbin Institute of Technology), Harbin 150090, China)
Abstract:
Anaerobic ammonium oxidation (ANAMMOX) is known as the most effective process for nitrogen removal from organic wastewater with low ratio of carbon to nitrogen. But ANAMMOX population is difficult to enrich and sensitive to environment change, which results in a long start-up process. Thus, the tardy start-up of ANAMMOX process is a major issue for engineering application. In preliminary research, an upflow oxygen limitation biofilm reactor (UOLBR) was constructed to treat piggery wastewater with high ammonium (NH+4-N) and low C/N ratio. After the 180-day start-up process, an excellent synchronous removal of chemical oxygen demand (COD) and total nitrogen (TN) was achieved with ANAMMOX as the dominant nitrogen removal pathway. Idled for 2 months, the UOLBR was restarted at 25 ℃ with a hydraulic retention time of 10 h and an effluent reflux ratio of 25∶1. Fed with manure-free piggery wastewater with the COD/TN ratio of 0.6-1.0, performance of the UOLBR during the restart-up process was evaluated. Operation process of the reactor was divided into 2 stages with a dissolved oxygen of 2.5-3.0 and 0.2-0.5 mg/L, respectively. The results showed that the UOLBR could recover from the idle state and a new steady state was reached within 53 days. Within the steady phase, the removal rates of COD, NH+4-N, and TN in the reactor averaged 63.96%, 96.5% and 91.7%, with residues of about 1,7.0 and 16.4 mg/L in the effluent, respectively. Analysis of microbial community along with mass balance indicated that the recovered UOLBR was again characterized by ANAMMOX as the dominant approach for nitrogen removal from the wastewater. This work would be helpful for the start-up and management of ANAMMOX processes.
Key words: oxygen limitation wastewater low COD/TN biological nitrogen removal biofilm
徐翩翩, 孟佳, 汪聪, 李建政. 限氧废水处理系统的再启动与ANAMMOX功能恢复[J]. 哈尔滨工业大学学报, 2019, 51(2): 32-38. DOI: 10.11918/j.issn.0367-6234.201801014.
XU Pianpian, MENG Jia, WANG Cong, LI Jianzheng. Restart-up of an oxygen limitation wastewater treatment process for the recovery of ANAMMOX[J]. Journal of Harbin Institute of Technology, 2019, 51(2): 32-38. DOI: 10.11918/j.issn.0367-6234.201801014.
基金项目 中国博士后科学基金(2017M611376) 作者简介 徐翩翩(1993—)，女，硕士研究生;
李建政(1965—)，男，教授，博士生导师 通信作者 李建政，ljz6677@163.com 文章历史 收稿日期: 2018-01-08
Contents -->Abstract Full text Figures/Tables PDF
限氧废水处理系统的再启动与ANAMMOX功能恢复
徐翩翩, 孟佳, 汪聪, 李建政
城市水资源与水环境国家重点实验室(哈尔滨工业大学)，哈尔滨 150090
收稿日期: 2018-01-08
基金项目: 中国博士后科学基金(2017M611376)
作者简介: 徐翩翩(1993—)，女，硕士研究生;
李建政(1965—)，男，教授，博士生导师
通信作者: 李建政，ljz6677@163.com
摘要: 为解决低碳氮比有机废水生物脱氮的难题，经济高效的厌氧氨氧化(ANAMMOX)技术得到广泛研究，但ANAMMOX菌群对环境要求苛刻且富集困难，系统启动缓慢，限制了其工程应用.前期研究中，构建了用于处理高氨氮低碳氮比养猪废水的升流式限氧生物膜反应器(UOLBR)，经180 d的启动运行实现了以ANAMMOX为主导的脱氮功能.将UOLBR闲置2个月，以COD与TN比为0.6~1.0的干清粪养猪废水为进水，在水力停留时间10 h、25 ℃和出水回流比25: 1的条件下再次启动，考察其处理效果和ANAMMOX功能的恢复情况.结果表明，经过溶解氧分别为2.5~3.0和0.2~0.5 mg/L两个阶段，共计53 d的连续运行，UOLBR的处理效能得以恢复并达到稳定状态，对COD、NH4+-N、TN的平均去除率分别为64.0%, 96.5%和91.7%，出水质量浓度分别仅为61，7.0和16.4 mg/L左右.微生物群落结构和物料平衡分析表明，UOLBR系统恢复了以ANAMMOX为主要脱氮途径的特征.该研究可为低碳氮比有机废水ANAMMOX处理系统的启动和运行管理提供指导.
关键词: 限氧 废水 低COD与TN比 生物脱氮 生物膜
Restart-up of an oxygen limitation wastewater treatment process for the recovery of ANAMMOX
XU Pianpian, MENG Jia, WANG Cong, LI Jianzheng
State Key Laboratory of Urban Water Resource and Environment (Harbin Institute of Technology), Harbin 150090, China
Abstract: Anaerobic ammonium oxidation (ANAMMOX) is known as the most effective process for nitrogen removal from organic wastewater with low ratio of carbon to nitrogen. But ANAMMOX population is difficult to enrich and sensitive to environment change, which results in a long start-up process. Thus, the tardy start-up of ANAMMOX process is a major issue for engineering application. In preliminary research, an upflow oxygen limitation biofilm reactor (UOLBR) was constructed to treat piggery wastewater with high ammonium (NH4+-N) and low C/N ratio. After the 180-day start-up process, an excellent synchronous removal of chemical oxygen demand (COD) and total nitrogen (TN) was achieved with ANAMMOX as the dominant nitrogen removal pathway. Idled for 2 months, the UOLBR was restarted at 25 ℃ with a hydraulic retention time of 10 h and an effluent reflux ratio of 25: 1. Fed with manure-free piggery wastewater with the COD/TN ratio of 0.6-1.0, performance of the UOLBR during the restart-up process was evaluated. Operation process of the reactor was divided into 2 stages with a dissolved oxygen of 2.5-3.0 and 0.2-0.5 mg/L, respectively. The results showed that the UOLBR could recover from the idle state and a new steady state was reached within 53 days. Within the steady phase, the removal rates of COD, NH4+-N, and TN in the reactor averaged 63.96%, 96.5% and 91.7%, with residues of about 61, 7.0 and 16.4 mg/L in the effluent, respectively. Analysis of microbial community along with mass balance indicated that the recovered UOLBR was again characterized by ANAMMOX as the dominant approach for nitrogen removal from the wastewater. This work would be helpful for the start-up and management of ANAMMOX processes.
Keywords: oxygen limitation wastewater low COD/TN biological nitrogen removal biofilm
畜禽养殖废水和许多工业有机废水都具有高氨氮和低碳氮比的特点，城镇生活污水的碳氮比也呈现出下降趋势，给传统的生物脱氮处理技术带来了严峻挑战[1-3].在传统生物脱氮处理工艺基础上，增加铵(NH4+)的物化去除设施，或外加碳源以调升废水的碳氮比，是目前广为采用的以达到良好脱氮效果的措施，但也显著提高了废水处理成本[2, 4].厌氧氨氧化(anaerobic ammonium oxidation，ANAMMOX)以亚硝酸盐(NO2-)和NH4+分别作为电子受体和电子供体发生反应，生成N2和少量硝酸盐(NO3-)，被认为是最为经济高效的生物脱氮途径，适宜于高氨氮低碳氮比废水的脱氮处理[5].然而，ANAMMOX菌群的增殖缓慢，在成分复杂的废水或污水生物处理系统中，其生长代谢极易受到增殖较快的化能异养微生物的抑制，如何实现有效富集并使之与其他功能菌群达到代谢上的平衡，是ANAMMOX技术工程应用的关键[6-9].
研究表明，在溶解氧(DO)为0.3~1.0 mg/L的微氧(亦称限氧)条件下，好氧、厌氧和兼性细菌可共存于同一污泥相中，在单一反应器中即可实现碳氮的同步去除，具有工程投资省、剩余污泥少、耗能低等优点[10-12].针对干清粪养猪废水高氨氮低碳氮比的特点，前期设计并启动运行了升流式限氧生物膜反应器(upflow oxygen limitation biofilm reactor，UOLBR)，并成功富集了ANAMMOX功能菌群，在水力停留时间(HRT)8 h、27 ℃和DO为0.40 mg/L的条件下，对化学需氧量(COD)、铵态氮(NH4+-N)和总氮(TN)的去除负荷分别平均高达0.60，0.94和0.91 kg/(m3 · d)，实现了碳氮的高效同步去除，但启动进程缓慢，达180 d以上[13].在实际应用中，污水和废水生物处理系统还会因季节性生产、年度维护及设备故障等原因而中断运行数天乃至数月.重新启动运行时，系统往往需要一定的恢复期以达到良好的处理效能[14-15].因此，实现生物处理系统闲置后的快速恢复对实际工程应用具有重要意义.本文在前期启动运行并实现以ANAMMOX为主导的脱氮途径后，将UOLBR闲置2个月，在HRT 10 h、25 ℃和出水回流比25: 1的条件下再次启动，考察其处理效果和ANAMMOX功能的恢复情况，并对系统的微生物群落结构和脱氮途径进行分析，以期为限氧生物处理系统的启动和运行管理提供指导.
1 实验 1.1 实验装置图 1为UOLBR养猪废水处理系统的装置示意.其中，UOLBR由有机玻璃制成，反应区域容积4.9 L，反应柱高0.5 m，内径0.1 m，顶部设有固-液-气三相分离器，底部为锥形，与进水管和出水回流管连接.在反应器内的中上部，装填有规格为Φ16×10 mm的PVC填料，填料床高200 mm，自然堆积孔隙率在95%左右.UOLBR进水由蠕动泵泵入，顶部出水排入容积为10 L蓄水箱.蓄水箱一分为二，其一用于缓存排放出水，另一部分用于存储出水并曝气，由蠕动泵回流以维持UOLBR内的限氧环境.UOLBR配有在线溶解氧监测仪对回流水的曝气量进行控制，将反应系统的DO维持在设定值.UOLBR外壁缠有电热丝，由温控仪将反应器内的温度控制在(25±1)℃.
Fig. 1
图 1 UOLBR系统装置示意 Fig. 1 Schematic diagram of the UOLBR system 1.2 实验用水实验用水为取自哈尔滨市某养猪场的干清粪养猪废水，其COD、NH4+-N和TN质量浓度分别为217~1 410，104.3~471.6和104.9~472.5 mg/L，COD与TN比在0.7~4.3内波动，是典型的高氨氮低碳氮比有机废水.基于前期研究，利用序批式反应器(sequencing batch reactor，SBR)对养猪废水进行预处理，通过COD的去除，将UOLBR进水的COD与TN比控制在0.6~1.0内[13, 16-17].在重新启动运行过程中，UOLBR进水的COD、NH4+-N和TN质量浓度分别为114~263，171.4~281.7和171.6~281.1 mg/L.
1.3 UOLBR前期和重新启动的控制参数UOLBR在前期已成功启动并连续运行[13].在HRT 10 h、25 ℃、出水回流比25: 1，以及进水COD、NH4+-N、TN和pH分别为(193±35)，(251.2±31.1)，(251.9±31.0) mg/L和(8.2±0.2)的条件下，UOLBR在稳定运行状态下对COD、NH4+-N和TN的平均去除率分别为62.3%, 95.3%和88.3%，此时反应器内悬浮污泥的混合液悬浮固体(MLSS)和挥发性悬浮固体(MLVSS)总量分别为5.18和3.60 g/L，生物膜的分别为8.47和2.81 g/L.在满水状态下，反应器停止运行2个月，之后重新启动并连续运行，HRT、温度和出水回流比等控制参数与停运前相同.依据系统内的DO水平，UOLBR的重新启动运行过程分为两个阶段：前30 d为第1阶段，将DO控制在2.5~3.0 mg/L的较高水平，以富集硝化菌群；自第31天以后的运行为第2阶段，将DO调控为0.2~0.5 mg/L，使系统处于限氧状态并持续运行.UOLBR在这两个阶段的控制参数和水质如表 1所示.
表 1
表 1 UOLBR重新启动过程的阶段和控制参数 Tab. 1 Stages and control parameters within the restart-up process of the UOLBR 阶段 控制参数 进水 DO/(mg·L-1) 水力停留时间/h 回流比 COD/(mg·L-1) ρ(NH4+-N)/(mg·L-1) ρ(TN)/(mg·L-1) COD与TN比(平均值)
1 2.68±0.11 10 25:1 160±30 236.5±17.4 236.9±17.3 0.67
2 0.42±0.12 10 25:1 178±38 218.5±34.1 219.6±33.9 0.81
表 1 UOLBR重新启动过程的阶段和控制参数 Tab. 1 Stages and control parameters within the restart-up process of the UOLBR
1.4 分析方法在UOLBR的启动运行过程中，每天定时采集进水和出水样品进行水质分析.COD、NH4+、NO2-和NO3-分别采用重铬酸钾法、纳式试剂光度法、N-(1-萘基)-乙二胺光度法和紫外分光光度法[18].pH采用pH计(Switzerland Mettler Toledo，DELTA 320)检测，DO采用溶解氧在线检测仪(台湾衡欣，AZ8403)测定.水样中的TN以NH4+-N、NO2--N及NO3--N之和计.
1.5 生物量与微生物群落结构分析在UOLBR重新启动运行的第68天，从填料床随机采集填料3个，利用涡漩振荡器(Kylin-bell，VORTEX-5)将填料上的生物膜剥离，烘干；从反应器侧面最下端取样口(图 1)采集泥水混合液100 mL，烘干.将以上生物膜和悬浮污泥恒重，并计算系统内的MLSS和MLVSS[18].
借助于细菌16S rRNA基因的高通量测序，分别对悬浮污泥和生物膜污泥进行微生物群落结构分析.其中，样品的总DNA利用Power soil DNA试剂盒(美国，MOBIO)提取，细菌16S rRNA基因的V3~ V4区扩增引物为341F(5′-CCTACGGGAGGCAGCAG-3′)和805R(5′-GACTACHVGGGTATCTAATCC-3′)，PCR产物的高通量测序仪器为Illumina Miseq PE300(美国，Illumina)[16].依据高通量测序提供的信息，进行微生物群落多样性和构成分析[19].
2 结果与讨论 2.1 COD的去除如图 2所示，UOLBR系统在重新启动运行的第1天，其COD去除率为59.2%.随着运行的延续，系统对COD的去除率逐渐上升，并在第1阶段的最后10 d(第19~第30天)维持在62.1%左右.自第31天进入第2运行阶段后，尽管系统中的DO由第1阶段的2.68 mg/L降低到0.42 mg/L左右，但UOLBR对COD的去除率并未受到显著影响.在运行的最后16 d(第53~第68天)，UOLBR的COD去除率平均为64.0%，出水COD平均只有61 mg/L.经过68 d的恢复运行，UOLBR系统悬浮污泥的MLSS和MLVSS分别为6.38和4.09 g/L，生物膜的分别为9.27和2.95 g/L，表征污泥活性的MLVSS与MLSS比分别为0.64和0.32，与反应器在2个月前停运时的0.70和0.33相近.以上结果表明，尽管UOLBR闲置长达2个月之久，但其中的微生物仍然保持着良好的活性，对COD的去除能力可得到迅速恢复.
Fig. 2
图 2 UOLBR在重新启动过程的COD去除 Fig. 2 COD removal in the UOLBR during the restart-up process 2.2 NH4+-N和TN的去除干清粪养猪废水是典型的高氨氮低碳氮比有机废水，有效脱氮是其处理的核心和难点[13, 16-17].由于干清粪养猪废水的TN主要由NH4+-N贡献(表 1)，欲使UOLBR达到良好的脱氮效能，首先必须使氨氧化菌群(AOB)得到富集，将NH4+-N氧化为NO2--N.为此，在UOLBR重新启动运行的第1阶段，将系统内的DO控制在了2.68 mg/L这一较高的水平(表 1).如图 3(a)所示，UOLBR在重新启动运行的第1天，由于反应器内混合液对进水的稀释作用，其NH4+-N去除率高达97.1%，但随着运行的延续，系统表现出NH4+-N去除率持续下降，至第1阶段在第30天结束时，只有57.9%.尽管NH4+-N去除率不断下降，出水中的NO3--N呈现出持续增加趋势(图 3(b))，在第30天达到了52.2 mg/L，表明包括AOB和亚硝酸盐氧化菌群(NOB)在内的硝化菌群得到了富集和活性恢复.由于较高的DO水平会严重抑制包括反硝化和ANAMMOX等脱氮功能菌群的活性[20]，在DO为2.68 mg/L左右的第1运行阶段，UOLBR并未呈现出显著的脱氮效应，所呈现出的TN去除率逐步下降的趋势，主要也是由反应器内原有混合液对进水的稀释作用引起的(图 3 (c)).
Fig. 3
图 3 NH4+-N、NOx--N和TN在UOLBR重新启动过程中的变化 Fig. 3 NH4+-N, NOx--N, and TN in the UOLBR during the restart-up process 自第31天将UOLBR内的DO下调到0.42 mg/L左右后(表 1)，系统对NH4+-N的去除虽有明显波动，但仍表现为继续下降趋势，直到第39天达到最低值30.9%(图 3a).同一时期，TN去除率则表现出了恢复趋势，但在第39天也只有30.4%(图 3(c)).在第31~第39天的运行中，由于DO的显著降低，硝化作用受到了显著影响，出水中的NO3--N迅速下降，同时出现了NO2--N的短期积累(图 3(b)).第39天以后，UOLBR的NH4+-N和TN去除率均呈现出了快速增加趋势，并在最后的16 d(第53~第68天)保持了相对稳定，分别平均高达96.5%和91.7%，出水平均质量浓度分别只有7.0和16.4 mg/L，说明系统的生物脱氮功能得到了有效恢复.值得注意的是，在最后16 d的稳定运行期，UOLBR系统中出现了一定量的NO3--N积累，出水质量浓度平均为8.2 mg/L，而NO2--N质量浓度则保持在1.0 mg/L左右的低水平.这一结果暗示，ANAMMOX在系统生物脱氮功能中发挥了重要作用[5, 10].
2.3 UOLBR系统的微生物群落结构分析为了解UOLBR恢复后的脱氮机制，在最后的稳定运行期分别采集悬浮污泥和生物膜，借助于细菌16S rRNA基因的高通量测序技术，对其微生物群落结构进行了分析.
如表 2所示，从UOLBR悬浮污泥和生物膜样品中分别获得47 446和47 922条基因序列，以及1 725和1 916个操作分类单元(operational taxonomic units，OTUs)，2个样品的基因文库覆盖率均达到了99%，说明测序结果能够反映样本的真实情况.Alpha多样性可以反映微生物群落的丰度和多样性，其中Chao1和ACE指数表征的是群落丰度，Shannon-Wiener和Simpson指数表征的是群落多样性[21].比较分析发现(表 2)，在UOLBR系统中，悬浮污泥与生物膜微生物群落的物种丰富度都很高，但生物膜中的物种丰度和多样性均高于悬浮污泥，说明填料的布设更利于复杂微生物群落的构建，具有提升系统生物量和污染物降解途径的显著功效.
表 2
表 2 UOLBR悬浮污泥和生物膜的Alpha多样性分析 Tab. 2 Alpha diversity of the sampled suspended sludge and biofilm in the UOLBR 样品 原始序列 筛选序列 OUTs Chao1指数 ACE指数 Shannon指数 Simpson指数 样品覆盖率/% 悬浮污泥 49 235 47 446 1 725 2 117.45 2 084.2 5.22 0.02 99
生物膜 59 652 47 922 1 916 2 291.66 2 257.3 5.55 0.01 99
表 2 UOLBR悬浮污泥和生物膜的Alpha多样性分析 Tab. 2 Alpha diversity of the sampled suspended sludge and biofilm in the UOLBR
基于分类到属的OTUs分析发现，功能恢复后的UOLBR系统，其悬浮污泥中存在大量与生物脱氮相关的功能菌群，如硝化菌群、反硝化菌群和ANAMMOX菌群等(图 4(a)).其中，硝化菌群相对丰度为0.41%(包括0.29%的AOB和0.12%的NOB)，AOB主要有Nitrosomonas[22]，Prosthecobacter[23]和Sphingomonas[24]，NOB主要有Nitrobacter[25]和Nitrospira[26]；反硝化菌群的相对丰度为14.87%，主要包括Limnobacter[27]、Azospira[28]、Methyloversatilis[29]、Azohydromonas[30]、Lautropia[31]和Longilinea[32]，相对丰度分别为4.06%，2.66%，1.48%，1.13%，1.09%和1.05%；ANAMMOX菌群的相对丰度为0.78%，主要是Candidatus brocadia、C.kuenenia和C.anammoxoglobus[33]，相对丰度分别为0.58%，0.20%和0.002%.
Fig. 4
图 4 UOLBR悬浮污泥和生物膜的菌属相对丰度 Fig. 4 Relative abundance of genus in suspended sludge and biofilm of the UOLBR 如图 4(b)所示，硝化菌群，包括Nitrosomonas、Prosthecobacter和Sphingomonas在内的AOB，以及包括Nitrobacter和Nitrospira在内的NOB，同样大量存在于UOLBR的生物膜中，但其总体相对丰度均比悬浮污泥中的高，分别达到了0.56%和0.18%.生物膜中的反硝化菌群结构与悬浮污泥的有显著差异，优势菌属主要是Azospira[28]、Thermogutta[34]、Thauera[28]、Comamonas[35]、Limnobacter[27]、Rhodobacter[28]和Methyloversatilis[29]，其相对丰度分别为3.07%，2.76%，1.83%，1.21%，1.18%，1.11%和1.1%，总体丰度达到了18.41%，显著高于悬浮污泥的14.87%.与悬浮污泥相比，生物膜中的ANAMMOX菌属相对丰度较低，为0.46%，主要包括C.kuenenia(0.28%)和C.brocadia(0.18%).
比较发现[36]，在停止运行的UOLBR中，其AOB、NOB、反硝化菌群和ANAMMOX菌群的相对丰度范围分别为0.25%~4.10%，0.10%~0.34%，11.43%~19.17%和0.08%~0.93%.而在重新启动并达到稳定运行状态后，UOLBR中的AOB、NOB、反硝化细菌和ANAMMOX的相对丰度均达到或超过了停止运行前的水平.
以上结果表明，悬浮污泥和生物膜的共存，不仅可以使AOB、NOB、硝酸盐还原菌群、亚硝酸盐还原菌群和ANAMMOX菌群等共栖于UOLBR限氧生物处理系统中，而且增加了生物脱氮功能菌群的多样性，为全程硝化反硝化脱氮、短程硝化反硝化脱氮和ANAMMOX脱氮奠定了生物学基础.
2.4 UOLBR系统的脱氮途径分析除了细胞合成外，废水生物脱氮途径有三，即全程硝化反硝化、短程硝化反硝化和ANAMMOX[19, 20].图 4所示的微生物群落结构表明，在UOLBR系统中，共栖着AOB、NOB、硝酸盐还原菌、亚硝酸盐还原菌和ANAMMOX菌等生物脱氮功能菌群，全程硝化反硝化、短程硝化反硝化和ANAMMOX均有发生的可能.如式(1)和式(2)所示，NO3--N和NO2--N的反硝化脱氮均需有机碳源作为电子供体，而ANAMMOX的脱氮过程(式(3))无需有机碳源[5, 37].
$5\text{C+2}{{\text{H}}_{\text{2}}}\text{O+4NO}_{3}^{-}\to 2{{\text{N}}_{2}}+4\text{O}{{\text{H}}^{\text{-}}}+5\text{C}{{\text{O}}_{2}}, $ (1)
$3\text{C+2}{{\text{H}}_{\text{2}}}\text{O+4NO}_{2}^{-}\to 2{{\text{N}}_{2}}+4\text{O}{{\text{H}}^{\text{-}}}+3\text{C}{{\text{O}}_{2}}, $ (2)
$\begin{align} &\text{NH}_{4}^{+}+1.32\text{NO}_{2}^{-}+0.066\text{HCO}_{3}^{-}+0.13{{\text{H}}^{+}}\to \\ &1.02{{\text{N}}_{2}}+0.26\text{NO}_{3}^{-}+0.066\text{C}{{\text{H}}_{\text{2}}}{{\text{O}}_{\text{0}\text{.15}}}+2.03{{\text{H}}_{2}}\text{O}\text{.} \\ \end{align}$ (3)
由式(1)、(2)可知，通过反硝化作用，将NO3--N还原为N2所需的COD去除与TN去除比为2.86，还原NO2--N所需要的COD去除与TN去除比也要达到1.71.如图 2和图 3(c)所示，UOLBR在第53~第68天的相对稳定运行阶段，其COD和TN去除率分别平均为64.0%和91.7%，COD去除与TN去除比平均为0.61，且没有明显的NO2--N积累(图 3(b)).假设系统去除的COD全部被用于NO3--N或NO2--N的还原，全程硝化反硝化和短程硝化反硝化对系统TN去除率的最大贡献分别也只有21.33%和35.67%.因此，在处于稳定运行时期的UOLBR系统中，至少有64.33%的TN是由ANAMMOX途径贡献的.可见，恢复后的UOLBR系统保持了生物脱氮途径的多样性，而ANAMMOX仍是其最主要的生物脱氮途径[13].
3 结论1) 闲置2个月之久的UOLBR，可直接重新启动，并在53 d内达到稳定运行.在HRT 10 h、25 ℃和出水回流比25: 1的条件下，其COD、NH4+-N和TN去除率分别维持在64.0%，96.5%和91.7%的较高水平.
2) 生物脱氮功能菌群在UOLBR悬浮污泥和生物膜中的分布存在显著差异.其中，生物膜富集了更多的硝化菌群和反硝化菌群，而悬浮污泥中的ANAMMOX更为丰富.
3) 功能恢复后的UOLBR系统，保留了全程硝化反硝化、短程硝化反硝化以及ANAMMOX等多种脱氮功能菌群及脱氮途径共存的特征，其中ANAMMOX依然是系统的主要脱氮途径.
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