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Article(id=1234106392538305264, tenantId=1146029695717560320, journalId=1234093305789726721, issueId=1234106384963400440, articleNumber=null, orderNo=null, doi=null, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1730995200000, receivedDateStr=2024-11-08, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1772163492569, onlineDateStr=2026-02-27, pubDate=1750348800000, pubDateStr=2025-06-20, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1772163492569, onlineIssueDateStr=2026-02-27, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1772163492569, creator=13701087609, updateTime=1772163492569, updator=13701087609, issue=Issue{id=1234106384963400440, tenantId=1146029695717560320, journalId=1234093305789726721, year='2025', volume='45', issue='6', pageStart='2961', pageEnd='3552', issueExtLink='null', onlineDate='null', pubDate='null', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=1, specialIssue=null, createTime=1772163490763, creator=13701087609, updateTime=1772163969484, updator=13701087609, preIssue=null, nextIssue=null, ext={EN=IssueExt(id=1234108392948682946, tenantId=1146029695717560320, journalId=1234093305789726721, issueId=1234106384963400440, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1234108392948682947, tenantId=1146029695717560320, journalId=1234093305789726721, issueId=1234106384963400440, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null}, startPage=3037, endPage=3045, ext={EN=ArticleExt(id=1234106393293280033, articleId=1234106392538305264, tenantId=1146029695717560320, journalId=1234093305789726721, language=EN, title=Effects of co-existence of quinoline/microplastics on anammox biofilm reactor, columnId=1234106386360103680, journalTitle=China Environmental Science, columnName=Water Pollution Control, runingTitle=null, highlight=null, articleAbstract=

An anaerobic sequencing batch biofilm reactor was used to explore the combined effect mechanism of anammox under the co-existence conditions of quinoline (50~200mg/L) and microplastics (PET-MPs) (20~100mg/L). With the increase of the concentrations of quinoline and PET-MPs, the performance of Anammox first decreases and then gradually recovers, and recovery time of reversible inhibition was positively correlated with the concentration of combined pollutants. The specific anammox activity (SAA) decreased from 22.8mg N/(g VSS·h) in stage C1 to 16.2mg N/(g VSS·h) in stage C3, while the corresponding reactive oxygen species (ROS) production increased by 55.7%, indicating that the inhibition of Anammox was enhanced under combined pollution. Extracellular polymer (EPS) analysis revealed that an increase in the concentrations of quinoline and PET-MPs would lead to a rapid decrease in the EPS content of the biofilm from 75.3mg/g VSS to 39.2mg/g VSS. The significant reduction in protein (PN) secretion, which in turn led to a significant decrease in PN/PS, indicates a decline in the structural stability of the Anammox biofilm. High-throughput sequencing revealed that the concentration of quinoline /PET-MPs increased, while the microbial community diversity and richness indices decreased. The relative abundance of Candidatus_Brocadia decreased from 1.73% to 1.24%, while the relative abundance of Denitratisoma changed little. However, the relative abundance of anaerobic heterocyclic degrading bacteria increased significantly.

, correspAuthors=Xin ZHOU, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, authorCompany=null, fund=null, authors=null, authorsList=Yu-han XIE, Xin ZHOU, Bing-xin NIU), CN=ArticleExt(id=1234106398146089090, articleId=1234106392538305264, tenantId=1146029695717560320, journalId=1234093305789726721, language=CN, title=喹啉/微塑料共存对Anammox生物膜反应器的影响, columnId=1234106386565624579, journalTitle=中国环境科学, columnName=水污染与控制, runingTitle=null, highlight=null, articleAbstract=

本文采用厌氧序批式生物膜反应器探究了喹啉(50~200mg/L)和微塑料(PET-MPs)(20~100mg/L)共存条件下对厌氧氨氧化(Anammox)的复合影响机制.结果表明，随着喹啉和PET-MPs浓度增高，Anammox性能先下降，再逐步回升，且可逆性抑制恢复时间与复合污染物浓度呈正相关.厌氧氨氧化活性(SAA)从C1阶段的22.8mg N/(g VSS·h)下降至C3阶段的16.2mg N/(g VSS·h)，而相应活性氧(ROS)产生量提高55.7%，表明复合污染下对Anammox抑制程度增强.胞外聚合物(EPS)分析发现喹啉和PET-MPs浓度增加会导致生物膜EPS含量从75.3mg/g VSS快速下降至39.2mg/g VSS，蛋白质(PN)分泌量减少显著进而导致PN/PS显著下降表明Anammox生物膜结构稳定性下降.高通量测序发现喹啉/PET-MPs浓度升高，微生物群落多样性和丰富度指数均下降，Candidatus_Brocadia相对丰度从1.73%下降至1.24%，Denitratisoma相对丰度变化不大，而厌氧杂环降解菌的相对丰度则显著增加.

, correspAuthors=周鑫, authorNote=null, correspAuthorsNote=
* 责任作者，教授，
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谢宇涵(2000-)，男，江西宜春人，太原理工大学环境工程硕士研究生.发表论文2篇..

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谢宇涵(2000-)，男，江西宜春人，太原理工大学环境工程硕士研究生.发表论文2篇..

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谢宇涵(2000-)，男，江西宜春人，太原理工大学环境工程硕士研究生.发表论文2篇..

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Operating conditions

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运行阶段运行天数(d)喹啉(mg/L)PET(mg/L)
C11~255020
C226~5510050
C356~95200100
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运行条件

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运行阶段运行天数(d)喹啉(mg/L)PET(mg/L)
C11~255020
C226~5510050
C356~95200100
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Nitrogen removal performance at different stages

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阶段运行天数(d)NH4+-N去除(%)NO2--N去除(%)TN去除(%)
C11~2569.7~82.484.7~90.377.2~85.5
C226~5557.2~81.781.5~91.466.7~86.4
C356~9554.0~82.573.6~92.959.2~85.0
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各阶段脱氮性能

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阶段运行天数(d)NH4+-N去除(%)NO2--N去除(%)TN去除(%)
C11~2569.7~82.484.7~90.377.2~85.5
C226~5557.2~81.781.5~91.466.7~86.4
C356~9554.0~82.573.6~92.959.2~85.0
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Microbial community diversity and richness index

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样品OTUs多样性指数丰富度指数
ShannonSimpsonChaoAce
C128625.960.01428622862
C224725.5080.01624722472
C38974.6690.037897897
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微生物群落多样性和丰富度指数

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样品OTUs多样性指数丰富度指数
ShannonSimpsonChaoAce
C128625.960.01428622862
C224725.5080.01624722472
C38974.6690.037897897
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喹啉/微塑料共存对Anammox生物膜反应器的影响
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谢宇涵 , 周鑫 * , 牛冰心
中国环境科学 | 水污染与控制 2025,45(6): 3037-3045
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中国环境科学 | 水污染与控制 2025, 45(6): 3037-3045
喹啉/微塑料共存对Anammox生物膜反应器的影响
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谢宇涵 , 周鑫* , 牛冰心
作者信息
  • 太原理工大学环境与生态学院，山西 晋中 030600

    • 谢宇涵(2000-)，男，江西宜春人，太原理工大学环境工程硕士研究生.发表论文2篇..

通讯作者:

* 责任作者，教授，
Effects of co-existence of quinoline/microplastics on anammox biofilm reactor
Yu-han XIE , Xin ZHOU* , Bing-xin NIU
Affiliations
  • College of Environment and Ecology, Taiyuan University of Technology, Jinzhong 030600, China
出版时间: 2025-06-20
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摘要
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本文采用厌氧序批式生物膜反应器探究了喹啉(50~200mg/L)和微塑料(PET-MPs)(20~100mg/L)共存条件下对厌氧氨氧化(Anammox)的复合影响机制.结果表明，随着喹啉和PET-MPs浓度增高，Anammox性能先下降，再逐步回升，且可逆性抑制恢复时间与复合污染物浓度呈正相关.厌氧氨氧化活性(SAA)从C1阶段的22.8mg N/(g VSS·h)下降至C3阶段的16.2mg N/(g VSS·h)，而相应活性氧(ROS)产生量提高55.7%，表明复合污染下对Anammox抑制程度增强.胞外聚合物(EPS)分析发现喹啉和PET-MPs浓度增加会导致生物膜EPS含量从75.3mg/g VSS快速下降至39.2mg/g VSS，蛋白质(PN)分泌量减少显著进而导致PN/PS显著下降表明Anammox生物膜结构稳定性下降.高通量测序发现喹啉/PET-MPs浓度升高，微生物群落多样性和丰富度指数均下降，Candidatus_Brocadia相对丰度从1.73%下降至1.24%，Denitratisoma相对丰度变化不大，而厌氧杂环降解菌的相对丰度则显著增加.

喹啉  /  PET-MPs  /  厌氧氨氧化  /  微生物群落  /  影响机制

An anaerobic sequencing batch biofilm reactor was used to explore the combined effect mechanism of anammox under the co-existence conditions of quinoline (50~200mg/L) and microplastics (PET-MPs) (20~100mg/L). With the increase of the concentrations of quinoline and PET-MPs, the performance of Anammox first decreases and then gradually recovers, and recovery time of reversible inhibition was positively correlated with the concentration of combined pollutants. The specific anammox activity (SAA) decreased from 22.8mg N/(g VSS·h) in stage C1 to 16.2mg N/(g VSS·h) in stage C3, while the corresponding reactive oxygen species (ROS) production increased by 55.7%, indicating that the inhibition of Anammox was enhanced under combined pollution. Extracellular polymer (EPS) analysis revealed that an increase in the concentrations of quinoline and PET-MPs would lead to a rapid decrease in the EPS content of the biofilm from 75.3mg/g VSS to 39.2mg/g VSS. The significant reduction in protein (PN) secretion, which in turn led to a significant decrease in PN/PS, indicates a decline in the structural stability of the Anammox biofilm. High-throughput sequencing revealed that the concentration of quinoline /PET-MPs increased, while the microbial community diversity and richness indices decreased. The relative abundance of Candidatus_Brocadia decreased from 1.73% to 1.24%, while the relative abundance of Denitratisoma changed little. However, the relative abundance of anaerobic heterocyclic degrading bacteria increased significantly.

quinoline  /  PET-microplastics  /  anammox  /  microbial community  /  influencing mechanism
谢宇涵, 周鑫, 牛冰心. 喹啉/微塑料共存对Anammox生物膜反应器的影响. 中国环境科学, 2025 , 45 (6) : 3037 -3045 .
Yu-han XIE, Xin ZHOU, Bing-xin NIU. Effects of co-existence of quinoline/microplastics on anammox biofilm reactor[J]. China Environmental Science, 2025 , 45 (6) : 3037 -3045 .
正文
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喹啉(C9H7N)是一种典型的含氮杂环化合物(NHCs).喹啉及其衍生物由于其具有的杂环结构，在水环境中溶解性高，容易迁移和积累.此外，喹啉还具有较高的遗传毒性和致突变性，已被美国环保署列为优先污染物，对自然生态乃至人类健康具有潜在的巨大危害[1].微塑料(MPs)一般指直径小于5mm的塑料颗粒物，由于其尺寸小，比表面积大等特点，MPs作为其他持久性有毒有机物的载体，能造成水体的复合污染[2]，是近年来备受关注的新污染物之一.
厌氧氨氧化(Anammox)是一种新型自养生物脱氮技术，与传统的硝化/反硝化生物脱氮工艺相比，脱氮效率可高达89%，同时降低60%以上曝气能耗，节省100%有机碳源，减少90%污泥产量，几乎不产生N2O等温室气体[3]，因而在污水脱氮节能低碳处理潜力巨大.
在一些印染行业排放的废水中，喹啉浓度可高达200mg/L以上[4]，且同时含有高浓度MPs(900个/L以上)[5].由于MPs通过沉淀和静电吸附很容易在Anammox处理系统中保留[6]，进而导致喹啉和MPs共存下的复合污染[7].这对于Anammox在该类废水实际工程应用提出了很大的挑战.目前喹啉和MPs单独作用下对Anammox影响已有报道[8].Chen等[9]发现Anammox菌群短期条件下，对喹啉的半抑制浓度(IC50)为13.07mg/L.Hong等[10]发现短期投加1.0g/L的PET使Anammox活性降低了16.2%.这表明单一污染物在特定浓度下对Anammox菌群活性具有显著抑制作用，且抑制程度与污染物的种类和浓度密切相关.然而，目前对于喹啉与MPs共同胁迫下Anammox的脱氮性能及其微生物群落的影响机制尚缺乏明确的认识.
聚对苯二甲酸乙二醇酯(PET)是污水处理系统中被检出的最常见的MP类型之一[11].因此，本研究探究喹啉与PET共同胁迫下Anammox的脱氮性能及微生物群落特征，旨在了解含杂环芳烃PET-MPs对于废水Anammox脱氮的影响机制，并为Anammox在该类废水工程应用提供参考.
1　材料和方法
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1.1　实验装置
采用有效容积为6L的厌氧序批式生物膜反应器，如图1所示.反应器由有机玻璃制成，内部填充50%的聚氨酯海绵填料.反应器密闭，整体用黑布覆盖避免光照产氧.接种污泥取自实验室膨胀颗粒污泥床反应器(EGSB)培养良好的生物量12g/L的Anammox颗粒污泥.反应器温度控制在(30±1)℃，底部装有推进器，保证搅拌良好.反应器采用间歇进出水，换水比为1/2，换水周期为1d.
1.2　运行条件
喹啉(分析纯)，购自天津市大茂化学试剂厂；PET，白色颗粒状，粒径约为300µm，密度为1.37~1.39g/cm3，购自东莞市樟木头晴天塑胶原料经营部.进水采用人工配水，添加100mg/L NH4Cl(分析纯)和130mg/L NaNO2(分析纯)，以N/P比为5:1的浓度比添加KH2PO4(分析纯)以保证厌氧微生物营养需求.进水pH值为(7.5±0.2).通入高纯氮气将DO浓度控制至0.2mg/L以下.待反应器成功启动后，NH4+-N和TN去除率均大于90%，投加不同浓度喹啉和PET-MPs，进入运行阶段，运行条件如表1所示，共运行95d.
1.3　水质分析方法
水样在测定前经过0.45μm的滤膜过滤.NH4+-N采用哈希快速测定法(HACH DR 1900，美国)分析，NO2--N采用N-(1-萘基)-乙二胺分光光度法(GB 7493-87)，NO3--N采用紫外分光光度法(GB/T 5750.5-2023).DO浓度使用DO仪(WTW Multi510IDS，WTW，德国)进行测定，pH值用pH计(FE-20，梅特勒，中国)进行测定.喹啉浓度采用紫外分光光度法，以蒸馏水为参比，在λ=270nm处进行吸光度测定[12].
1.4　Anammox活性测定
第25d，第55d，第95d进行Anammox活性(SAA)测定.通过工作体积为250mL的血清瓶进行批次测定.首先将瓶内填充适量的生物膜填料，再注入人工配水至刻度线处.人工配水中以1:1.32的物质的量比添加氨氮和亚硝态氮，以N/P为5:1的比例加入磷酸二氢钾，添加碳酸氢钠将pH值控制在(7.6±0.2).进水中通入高纯氮气20min，使反应器内的DO浓度低于0.2mg/L.之后将血清瓶密封并放置于恒温振荡箱中运行，温度设置为30℃，转速设为60r/min.以一定的时间间隔抽取水样，经过0.45μm滤膜过滤后测定出水水样氨氮和亚硝态氮的含量.根据式(1)计算SAA[13].
式中：和、和分别为进水和出水氨氮和亚硝酸盐的浓度，mg/L;T为反应时间，h；MLVSS为混合液中的挥发性悬浮固体，g/L.
1.5　活性氧水平测试
第25d，第55d，第95d进行生物膜样品中的活性氧(ROS)含量测定.使用荧光探针2,7-二氯荧光素二乙酸盐(DCFH-DA)进入细胞，DCFH-DA可以被细胞内的酯酶水解，形成荧光物质DCFH，通过测量荧光强度可以得出ROS的含量.每份样品测试3次.
1.6　EPS测定
第25d，第55d，第95d，分别从反应器中随机取出一块填料，采用超声法使填料上的生物膜脱落，将脱落的生物膜全部倒入离心管中.加入去离子水至30mL，在3600r/min的转速下离心5min，加入去离子水对细胞膜沉淀清洗3次.保留细胞膜沉淀用来提取胞外聚合物(EPS)，随后在污泥沉淀物中加入磷酸缓冲溶液，将其体积补充至30mL.在2000g离心力下离心15min，之后通过0.45μm的滤膜过滤上清液，得到扩散层EPS(S-EPS).将剩余的污泥沉淀物继续用磷酸缓冲溶液补充至30mL，并在5000g离心力的条件下离心15min，过滤上清液后得到松散层EPS(LB-EPS).继续加入磷酸缓冲溶液至30mL，将离心管放入70℃的水浴锅加热30min，取出后在5000g离心力的条件下离心15min，将上清液过滤后得到紧密层EPS(TB-EPS)[14].蛋白质(PN)采用考马斯亮兰比色法测定，多糖(PS)使用蒽酮-硫酸比色法测定.
1.7　EPS表征
EPS采用三维荧光光谱(3D-EEM)进行分析，使用荧光分光光度计(CARYEclipse，VARIAN，美国)进行扫描测定，以去离子水作为空白样.扫描参数为：激发光波长(Ex)为200~550nm，步长2nm，发射光波长(Em)为200~550nm，步长为5nm;扫描速度为12000nm/min，狭缝宽度为10nm，扫描间隔为2nm.以上数据结果采用Origin 2021进行处理与分析.
1.8　高通量测序
第25d，第55d，第95d取生物样品，采用美国OMEGA公司提供的M5635-0型E.Z.N.ATM Mag-Bind Soil DNA Kit试剂盒对DNA进行提取，引物为V3-V4区域的通用引物341F(5'-CCTACGGGNGGCWGCAG-3')和805R (5'-GACTACHVGGGTATCTAATCC-3').采用上海生物工程股份有限公司Illumina高通量测序[10](登陆号：PRJNA1095704).
2　结果与讨论
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2.1　喹啉去除影响
由图2可见，在1~25d投加的喹啉浓度为50mg/L且PET-MPs浓度为20mg/L，随着反应的进行，反应器内出水喹啉浓度逐渐下降，喹啉去除率从72.2%逐渐上升至87.1%.此时，反应器内的微生物处于对杂环芳烃适应期，低浓度喹啉能够作为NOx-N反硝化的电子供体，而被降解利用[15].第26d时，添加的喹啉浓度上升至100mg/L且PET-MPs浓度上升至50mg/L，出水喹啉浓度达31.7mg/L，喹啉去除率迅速下降至68.3%.反应进行到第55d时，喹啉去除率再次恢复至86.5%.这表明，在较低浓度喹啉对于微生物毒性抑制为可逆的;第56d时，喹啉的添加浓度继续上升至200mg/L且PET-MPs浓度上升至100mg/L时，在高浓度有毒有机物和PET-MPs的冲击下，喹啉去除率仅为60.5%，且经过40d后，喹啉去除率仅部分恢复至80.3%.分析认为，在高浓度喹啉和PET-MPs联合冲击条件下，导致对异养微生物产生叠加毒性抑制，恢复时间延长，抑制难以完全恢复.
2.2　脱氮影响
由图3、表2可见，在C1初始阶段，反应器内脱氮性能迅速恶化，Anammox过程受到抑制，NH4+-N、NO2--N去除率快速下降至(69.7%±1.4%)、(84.7%±2.5%)而此时出水NO3--N浓度较低.有研究报道短期喹啉暴露对Anammox的IC50仅为13~31mg/L[9,16].这表明，低浓度的喹啉即可对Anammox微生物产生较强的毒性抑制作用.当进行到21d时，反应器脱氮性能恢复，TN去除率提高至80.0%以上.25d时，Anammox性能彻底恢复，此时NH4+-N和NO2--N去除率分别可达到(82.4%±2.1%)和(90.3%±0.4%)的去除率，TN去除率可达到(85.5%±0.3%).这些说明，低浓度喹啉对Anammox微生物产生的影响是可逆性抑制，且Anammox恢复时间较短.
第C2阶段，随着喹啉和PET-MPs浓度分别升高到100和50mg/L，Anammox明显受到杂环芳烃和PET-MPs毒性抑制，NH4+-N和NO2--N去除率很快下降至(57.2%±1.3%)和(81.5%±0.5%)，TN去除率仅为(66.7%±0.2%).这可能归因于含氮杂环的加入使得异养菌迅速增殖，而AnAOB的活性受到竞争性抑制，在与兼性反硝化菌竞争NO2--N过程处于不利位置.经过11d的驯化后，NH4+-N和NO2--N的去除率分别达到70.0%和85.0%左右，TN去除率恢复至70.0%以上，Anammox性能开始恢复.反应进行到第55d时，NH4+-N和NO2--N的去除率恢复至(81.7%±1.7%)和(91.4%±1.8%)，此时TN去除率达到(86.4%±1.7%).在低浓度杂环芳烃存在下，有机物的存在不会对Anammox产生长期抑制，相反还促进了反硝化菌群的增殖，增强了对系统NO2--N和NO3--N去除.因此，在杂环芳烃存在下，通过同步Anammox反硝化保证了系统较理想的脱氮效果[17].C3阶段，喹啉浓度和PET-MPs浓度继续升高到200和100mg/L时，此时出水NH4+-N和NO2--N浓度迅速升高，分别为46.3mg/L和33.9mg/L，NH4+-N和NO2--N去除率仅为(54.0%±0.4%)和(73.6%±1.4%).出水中残留较高浓度NO3--N，从C1阶段(3.2±1.6) mg/L到C3阶段(8.4±4.0) mg/L.尽管PET-MPs单独存在下，100mg/ L对Anammox不足以产生抑制作用.然而，当高浓度的喹啉同时存在下，PET-MPs由于极强的吸附能力，可作为高毒有机物喹啉的载体输送至微生物细胞内[18]，进而加剧了对Anammox及反硝化菌负面影响.
经过长达40d运行后，反应器对NH4+-N和NO2--N去除率分别恢复至(82.5%±0.7%)和(92.9%±2.1%)，TN去除率恢复至(85.0%±0.8%).随着抑制性底物浓度的大幅度增加，Anammox脱氮性能的恢复时间也相应延长.对比前人研究，喹啉降至5mg/L不足以恢复颗粒污泥系统厌氧氨氧化性能[9]，本研究Anammox生物膜系统可以耐受更高浓度的喹啉.
2.3　SAA和ROS分析
如图4(a)，C1阶段，SAA处于最高，为22.8mg N/(g VSS·h)，随着喹啉和PET-MPs浓度的逐步上升，反应器的Anammox性能开始下降，导致C2阶段SAA下降为21.2mg N/(g VSS·h)，而C3阶段SAA值继续降至16.2mg N/(g VSS·h)，其活性仅为C1阶段的71%.Chen等[13]也发现随着喹啉从0提高到100mg/L，Anammox颗粒污泥SAA呈急速下降.图4b所示，C1阶段生物膜ROS的水平为(1418.1±57.3)pg/mL，在100mg/L喹啉和50mg/L PET-MPs的添加量下，细胞内ROS的产生量轻微上升至(1451.1±31.6)pg/mL;而随着喹啉和PET-MPs浓度继续提高至200和100mg/L，细胞内ROS的产生量显著增加至(2205.7±46.9) pg/mL.C2~C3阶段ROS值突然增高，可能原因是PET-MPs增加了细胞的通透性，导致喹啉更容易进入细胞.当AnAOB受到高浓度喹啉和PET-MPs的复合胁迫时，微生物会产生大量的ROS诱发氧化应激，过度高水平的氧化应激会破坏细胞的DNA链，并抑制酶的活性，从而降低AnAOB的生存能力[19].
2.4　EPS分析
由图5(a)可见，C1阶段总EPS含量最高为94.0mg/g VSS.而C2和C3阶段总EPS含量分别下降了12.0%和40.3%，由于喹啉浓度和PET-MPs浓度的增加，微生物受到的毒性抑制叠加，导致EPS分泌大量减少[20].这与胡璇等[21]发现聚酯纤维MPs的持续暴露抑制EPS的分泌结果相似. EPS组分方面，在每个阶段，PN的含量都要远远高于PS，表明蛋白质类物质的存在对生物膜的形成、结构和维持起着重要作用.本研究中，随着喹啉浓度和PET-MPs浓度的增加，EPS中PS含量呈现先增大后减少的趋势，分析认为：在喹啉和PET-MPs复合污染下，微生物通过释放大量PS来缓解二者对它的影响，但随着复合胁迫浓度持续增加，微生物分泌EPS不足以抵抗复合污染冲击，导致EPS含量从75.3mg/g VSS快速下降至39.2mg/g VSS.
PN/PS可评价EPS稳定性，PN/PS越低，表明生物膜凝聚力以及结构的紧密性和稳定性越差[22].图5(a)显示，随着喹啉和PET-MPs浓度提高，PN/PS的比值从4.0逐渐下降到2.4、2.3，表明污染物浓度提升导致Anammox生物膜结构稳定性下降，进而造成脱氮性能恶化.
图5(b)为各阶段下生物膜各层EPS含量的变化情况.本研究中TB-EPS作为EPS最主要的成分，TB-EPS可将细胞连接成簇，促进微生物的粘附和聚集，TB-EPS的浓度越高，生物膜结构越紧凑，有利于其在脱氮过程中的持续活性和稳定性[23].TB-EPS含量从C1阶段41.1mg/g VSS下降至C3阶段21.5mg/g VSS.LB-EPS含量的下降与喹啉/PET-MPs联合冲击有关.LB-EPS的产生可能与细胞外氧化还原活动有关，有利于微生物耐受和难降解有机物利用[24].各阶段LB-EPS含量分别为31.4、25.9和19.8mg/g VSS，表明微生物对于难降解喹啉耐受性能和利用能力降低.而S-EPS的含量呈现先增加后降低，分析认为：C2阶段增加，意味着细胞膜释放出更多的营养物质和胞外酶，可以被微生物利用.而C3阶段，由于高浓度喹啉和PET-MPs的联合胁迫，微生物正常代谢过程受到阻碍.
2.5　三维荧光光谱分析
图6显示了各阶段下S-EPS、LB-EPS和TB-EPS的3D-EEM.共检测到5个荧光峰，F1峰(Ex/Em=278~285nm/309~341nm)和F2峰(Ex/Em=275~279nm/327~349nm)代表可溶性微生物代谢产物，F3峰(Ex/Em=219~229nm/330~388nm)、F4峰(Ex/Em=215~219nm/287~294nm)和F5峰(Ex/Em=307~317nm/390~414nm)分别属于色氨酸类蛋白、酪氨酸类蛋白[25]以及腐殖酸类物质的荧光响应区.
在C1和C2阶段，在生物膜的SB-EPS中观察到F1峰、F2峰和F3峰，表明SB-EPS存在溶解性微生物代谢产物和色氨酸类蛋白.而在C3阶段生物膜SB-EPS的F2峰消失，出现了新的峰F4峰和F5峰，代表酪氨酸类蛋白和腐殖酸类物质的产生.C1~C3阶段生物膜的LB-EPS中都观察到F1峰、F2峰、F3峰和F4峰.而在C3阶段生物膜SB-EPS的F2峰消失，出现了新的峰F5峰.C1和C2阶段在生物膜的TB-EPS中观察到F2峰和F3峰，表明存在溶解性微生物代谢产物和色氨酸类蛋白.而在C3阶段发现TB-EPS比C1和C2阶段具有更弱的荧光强度，而S-EPS出现了F3峰和F5峰荧光强度增加，表明EPS分泌了更多的色氨酸和腐殖酸类物质.这说明在喹啉和PET-MPs的共存下，生物膜EPS不同层通过分泌不同类别蛋白类物质达到适应和抵御毒性胁迫环境的目的[26].
2.6　微生物群落分析
表3显示，随着喹啉和PET-MPs浓度增加，操作分类单元(OTUs)数量呈显著降低.Shannon指数不断减小而Simpson指数不断升高，这意味着毒性胁迫之下，群落种类多样性不断减少，系统中一些不能适应环境变化的细菌被淘汰.Chao指数和Ace指数也呈现不断下降趋势，表明微生物物种丰富度降低.可见，喹啉和PET-MPs不同浓度添加对微生物群落的多样性和丰富度具有显著影响.
由图7(a)可见，所有样品中的优势菌门为Proteobacteria(22.70%~27.49%)、Chloroflexi(15.68%~18.73%)、Bacteroidota(13.67%~18.95%)、Planctomycetota(8.52%~10.58%)、NB1-j(4.67%~12.73%)和Acidobacteriota(3.15%~5.44%).其中，Anammox功能菌属于Planctomycetota，其相对丰度的下降暗示了反应器中的Anammox反应受到了高浓度喹啉和PET-MPs的抑制.Proteobacteria中的微生物可以利用有机碳源进行反硝化[28].各阶段Proteobacteria的相对丰度分别为27.49%、22.70%和23.27%，张之钰等[29]与Xiao等[30]发现单独投加PET-MPs或者投加低浓度喹啉导致Proteobacteria菌门的相对丰度提高，而本文发现Proteobacteria菌门的相对丰度降低，这表明喹啉和PET-MPs存在着复合胁迫作用，对Proteobacteria产生毒性叠加抑制.而随着喹啉浓度和PET-MPs浓度的不断增加，Chloroflexi的相对丰度则从15.83%增加到18.73%.Chloroflexi通常会在Anammox反应器存在，可以降解有机物，对生物膜的形成与稳定发挥着重要的作用[27].
由图7(b)可见，Candidatus_Brocadia是本研究中唯一检测到的AnAOB.Candidatus_Brocadia相对丰度从1.73%下降至1.24%，与SAA变化趋势一致，其中，对比上个工况，C2~C3阶段中，ROS值提高51.9%，以及SAA快速降低23.6%，表明在高浓度喹啉和PET-MPs协同作用下加大了对Anammox菌属的不利影响.此外，Denitratisoma是相对丰度最高的菌属，是一种可以利用NO3--N为电子受体的异养反硝化菌[31]，其相对丰度受喹啉和PET浓度影响不大.表明该菌属能够耐受高浓度的复合胁迫并在系统同步Anammox反硝化脱氮中发挥至关重要的作用.而Limnobacter、Desulforhabdus、Rivibacter、Pseudomonas等菌属的相对丰度均呈现先变化不大再到急剧下降的趋势，这些菌属均与反硝化过程有关[32].而Anaerolinea具有厌氧降解喹啉功能[33]，其相对丰度则显著增加.因此，进水底物中喹啉和PET-MPs浓度将会直接决定微生物群落中各功能菌的数量占比和优势水平，进而影响系统脱氮除碳性能.
3　结论
收起
3.1　Anammox生物膜系统对喹啉和PET-MPs的耐受浓度分别为100和50mg/L.当这两种污染物的浓度分别升至200和100mg/L时，Anammox脱氮效率将难以恢复至正常水平.
3.2　随着喹啉和PET-MPs浓度增高，SAA从22.8mg N/(g VSS⋅h)下降至16.2mg N/(g VSS⋅h)，而相应ROS产量提高了55.7%.
3.3　随着喹啉和PET-MPs浓度增高，生物膜EPS含量从75.3mg/g VSS快速下降至39.2mg/g VSS，PN/PS从4.0显著下降到2.3，降低了生物膜稳定性.
3.4　随着喹啉和PET-MPs浓度增高，微生物群落的多样性和丰富度均出现下降，Candidatus_Brocadia相对丰度从1.73%下降至1.24%.
基金
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  • 国家自然科学基金项目(21607111)
  • 山西省专利转化计划资助项目(202401011)
  • 山西省基础研究计划项目(202403021221049)
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  • 接收时间：2024-11-08
  • 首发时间：2026-02-27
  • 出版时间：2025-06-20
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  • 收稿日期：2024-11-08
基金
国家自然科学基金项目(21607111)
山西省专利转化计划资助项目(202401011)
山西省基础研究计划项目(202403021221049)
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    太原理工大学环境与生态学院，山西 晋中 030600

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鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
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