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题名: 湿地系统氨氧化微生物的分布及对氨氮去除的贡献
作者: 苏瑜1
学位类别: 硕士
答辩日期: 2017-05
授予单位: 中国科学院大学
授予地点: 北京
导师: 王为东
关键词: 芦苇湿地,氨氮,氨氧化微生物,氨氧化速率,群落生态学 ; reed wetland, ammonium nitrogen, ammonia-oxidizing microorganisms,ammonium oxidation rate, community ecology
其他题名: Distribution of AOA and AOB, and Their Contributions to Ammonium Oxidation in Wetland Ecosystems
学位专业: 环境工程
中文摘要: 湿地生态系统是生物地球化学循环的活性区域,对流域中的氮素等营养物质 具有显著的拦截、过滤和净化作用。尤其在湿地水 -土边界处,频繁的水位波动 形成“厌氧-好氧”交替的环境,水和土壤之间进行着频繁的水文、物质和能量 交换,使其具有特殊的“边界”和“梯度”效应。湿地生态系统非均一的环境为 微生物的繁殖提供了充足的养分,在此处氨氧化活性和氮损失速率都较高,成为 微生物反应的活性“热区”。 在湿地系统中,微生物对氮素的去除约占 90%,氨氧化过程作为无机氮转化 为氮气的第一步,对生态系统中氮素去除起着重要作用。氨氧化过程由氨氧化细 菌(AOB)和氨氧化古菌(AOA)两种生理特性不同的微生物共同催化,并且 AOA和 AOB广泛存在于各种环境中。但是目前 AOA和 AOB对氨氮去除的相 对贡献尚不明确,因此本文利用双氰胺和 1-辛炔双抑制剂的方法测得 AOA和 AOB分别的活性,剖析 AOA和 AOB在不同湿地生态系统中的作用。本文利用 包括定量 PCR、克隆文库、454高通量测序技术在内的分子生物学手段与氨氧化 微生物活性测定相结合,在湿地生态系统中不同功能区、植物床 -沟壕系统的结 构单元以及全国尺度芦苇湿地研究氨氧化微生物的分布、多样性以及对氨氮去除 的贡献,旨在阐述在湿地生态系统中微生物对氨氮去除的机理,为人工湿地的构 建和运行提供理论指导,为岸边带修复提供技术支撑。主要研究结果如下: (1)对石臼漾构筑根孔湿地运行的第三到五年(2011年-2013年)的研究 发现 AOA amoA功能基因的丰度随着湿地的演替以一个数量级趋势降低,在不 同功能区 AOA丰度变化规律不同:在预处理区和深度净化区, AOA丰度在 2011-2012年基本保持不变,在 2013年降低;在植物床-沟壕系统中,AOA丰度 随着湿地演替逐渐下降。AOA多样性随着湿地的演替,在时间上基本保持不变, 而在空间上有略微波动趋势。AOA的群落结构随着湿地的演替发生了水平迁移, 并且 AOA群落结构的生态位分离首先发生在植物床-沟壕系统,虽然在湿地沉积 物中 AOA以 Nitrososphaera属为主导,但随着湿地运行,Nitrosopumilus属和 Nitrososphaera sister属有增加的趋势。综合反映湿地水文、水质和相应功能的运 行时间是驱动 AOA丰度和群落结构生态位分离的关键因素。总之,在石臼漾湿 地运行的第三到五年,AOA丰度在时间上不断降低,空间上趋于一致;AOA多 样性在时间上略微降低,空间上呈波动趋势;AOA的群落结构从相对均一状态 向多样的群落结构演替,并且在空间上发生了明显的水平转移。 (2)基于氨氧化反应在湿地边界处的“热区”效应以及水陆交错带的生态 效应,提出对植物床-沟壕系统进行形态优化改进,并在贯泾港构筑根孔湿地中得以验证。研究结果表明,在植物床-沟壕系统中,氨氧化反应的活性区域发生 在小沟中心、植物床边界处;开沟后,氨氧化速率在植物床 -沟壕系统中由相对 均一的分布变为高异质性分布,开沟增强了氨氧化速率(从 1.07±0.30到 1.22±1.27 mg N kg-1 d-1),氨氧化速率在两个活性区域得到明显增强(在小沟中心增强 2.13 倍,在植物床边界增强 1.76倍)。AOA和 AOB在植物床-沟壕系统中占据不同的 生态位,AOA在植物床中主导氨氧化过程,AOB在小沟中主导氨氧化过程;开 沟后 AOB的活性得到显著增强,并且 AOB活性的变化是氨氧化速率提高的驱 动因子;AOB的活性变化与丰度没有显著相关关系,而开沟后 Nitrosomonas属 的富集是 AOB活性增强的主要因素。氨氧化微生物的系统发育树表明,开沟后 AOB由 Nitrosospira主导变为 Nitrosospira和 Nitrosomonas共同在植物床-沟壕 系统中占主导;开沟也改变了 AOA的群落结构。 (3)对全国尺度 21个湿地生态系统表层土壤/沉积物样品中氨氧化微生物 的分布和活性研究结果表明,AOA是全国尺度湿地生态系统氨氧化反应的主要 驱动者。在湿地生态系统中,AOA amoA功能基因丰度平均比 AOB高 1-2个数 量级(平均值分别为 9.85×10 7 copies g-1 和 4.01×10 6 copies g-1);并且 AOA丰 度变化影响着湿地生态系统中氨氧化微生物丰度的变化, pH、有机质和总氮含 量也是影响氨氧化微生物分布的主要因子。在湿地生态系统中,AOA的多样性 高于 AOB。AOB群落结构在大部分湿地中以 Nitrosospira属为主导,在碱性较 高的湿地中以 Nitrosomonas属占主导;AOA群落结构大部分以 Nitrososphaera 属占主导,7个湿地中以 Nitrosopumilus属为主导。AOA和 AOB群落结构的聚 类受到地理位置的影响。全国尺度湿地生态系统的氨氧化微生物活性为 0.08-4.60 mg kg-1 d-1,具有很高的异质性;总氨氧化速率与 AOA和 AOB活性均呈现显著性正相关关系,说明 AOA和 AOB两种氨氧化微生物共同发挥作用,减轻湿地生态系统中氮负荷;但是在湿地生态系统中 AOA主导氨氧化过程,氨氧化速率 与 AOA活性相关性更强也说明 AOA是湿地生态系统中氨氧化反应的驱动因子。
英文摘要: Wetland ecosystems are active zones for biogeochemical cycle, which can intercept, filter, and remove the nutrients such as nitrogen (N) from the watershed. In particular, the soil-water interface in the wetlands has special “gradient effect” and “boundary effect” because of the intensive hydrological, material and energy exchange as well as the alternation of anoxic and aerobic conditions. The nonuniform conditions in the wetlands provide adequate nutrition for microbes, and wetland has long been considered as hotspots for microorganism activity with high rates of ammonium oxidation and nitrogen loss. Microorganism activity account for about 90% for nitrogen removal in wetland ecosystems, and ammonium oxidation process as the first step to convert inorganic N to dinitrogen plays an important role in nitrogen removal. The ammonium oxidation process was catalyzed jointly by ammonia oxidizing bacteria (AOB) and ammonia oxidizing archaea (AOA), and AOB and AOA were reported to exist in various ecosystems. Since the relative contributions of AOB and AOA for ammonium oxidation were uncertain, in this study, Dicyandiamide (DCD) and 1-octyne inhabitation methods were exploited to quantify the AOB activity and AOA activity. We also evaluated the contribution of AOB and AOA to ammonium oxidation in various wetland ecosystems. This paper studied the distribution, diversity and contributions of ammonia oxidizing organisms in different function zones of wetlands, plant-bed/ditch system and the nation-scale reed wetlands, using qPCR, clone library and 454 pyrosequencing, combined with AOB activity and AOA activity. The overall objective of the study is to explore ammonium removal mechanism by microorganism, thus providing theoretical guidance for the construction and operation of constructed wetlands and technical support for the restoration of riparian zones. The results are summarized as follows: (1) The study of AOA behavior in Shijiuyang constructed wetland (SJY-CW) from the third year (2011) to the fifth year (2013) showed that AOA abundance ended up with one order of magnitude lower being spatially consistent with the succession of wetlands, in the pre- and post-treatment ponds, it remained invariant in 2011–2012 and decreased in 2013, while the abundance in the plant-bed/ditch system decreased gradually from 2011 to 2013. AOA diversity showed little temporal differentiation with slight decrease trends for community diversity in 2011–2013. Hierarchical clustering and redundancy analysis verified the horizontal shifts of AOA communities, and the shifts occurred preferentially in the central plant-bed/ditch system. AOA community was dominated by Nitrososphaera cluster accompanying with Nitrosopumilus cluster and Nitrososphaera sister cluster increasing with the succession of wetland. Operational duration of the wetland which comprehensively reflected the wetland hydrology, water quality, and associated functions became a key factor driving the AOA abundance and community shift in SJY-CW sediments. To summarize, with the three-year through five-year succession of SJY-CW, the sedimentary AOA tended to be characterized by temporally decreased and spatially consistent abundance, while spatially variable and temporally decreased richness and diversity; regarding AOA community structure, temporally from local species and relatively uniform distribution to mixing with allochthonous species and highly diversified distribution, while spatially occurring significant horizontal shifts of AOA amoA genes. (2) Based on the fact of ammonium oxidation hotspots at the soil-water interface and ecological effect of riparian zones, we proposed to optimize the plant-bed/ditch system, and the research was carried in Guanjinggang constructed wetland (GJG-CW). The results indicated that the ditch center and plant-bed fringe were two active zones for ammonium oxidation, and the ammonium oxidation rate in the plant-bed/ditch system changed from relative uniform to highly heterogeneity after ditching. Ditching in the original plant-bed can improve ammonium oxidation activity (from 1.07±0.30 to 1.22±1.27 mg N kg-1 d -1 ), especially in both active zones, and it increased by 2.13 and 1.76 times (ditch center and plant-bed fringe) respectively. AOA and AOB had different ecological niches in the plant-bed/ditch system; AOB activity was dominant in ditch and AOA dominated in plant-bed. After ditching, AOB activity increased greatly, and the stimulated AOB-activity was thought to be responsible for the enhanced ammonium oxidation rate in the improved plant-bed/ditch system. While we didn’t find significant correlation between AOB activity and its abundance, and the enrichment of Nitrosomonas was more contributed to high AOB-activity and ammonium oxidation activity. Phylogenetic analysis of ammonia oxidizers amoA gene indicated a shift of AOB community from dominant genera Nitrosospira in original plant-bed/ditch system to dominant genera Nitrosospira and Nitrosomonas in improved plant-bed/ditch system. Ditching in the original plant-bed also changed the AOA community structure. (3) In addition, this paper studied the distribution and activity of ammonia-oxidizing microorganisms in 21 typical wetland ecosystems in China. The results indicated that AOA was the main driver for the ammonium oxidation process. In the wetland ecosystems, AOA amoA gene abundance was higher than AOB (1.68×10 5 -7.86×10 8 copies g -1 and 1.00×10 5 -3.00×10 7 copies g-1 dry soil respectively). What’s more, the AOA abundance drove the change of ammonia oxidizing organism abundance in the wetland ecosystems, and the pH, organic matter and TN also influenced the ammonia oxidizing organism abundance. The AOA diversity was higher than AOB diversity. AOB community structure was dominated by Nitrosospira in most wetlands, and in the wetlands with higher alkalinity, it was dominated by Nitrosomonas. AOA composition was dominated by Nitrososphaera in 14 wetlands, and others was dominated by Nitrosopumilus. Based on the Bray-Curtis similarity matrix, the distribution of AOB and AOA community structure was influenced by geographical factors. The ammonium oxidation rate in wetland ecosystems was 0.08-4.60 mg kg -1 d -1 harboring high heterogeneity, and ammonium oxidation rate was positively correlated with AOA activity and AOB activity. It indicated that AOA and AOB played important roles in ammonium removal jointly; AOA was dominated in ammonium oxidation and the higher correlation between AOA activity and ammonium oxidation rate also indicated AOA was the main contributor for ammonium oxidation in wetland ecosystems.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/38694
Appears in Collections:环境水质学国家重点实验室_学位论文

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作者单位: 1.中国科学院生态环境研究中心

Recommended Citation:
苏瑜. 湿地系统氨氧化微生物的分布及对氨氮去除的贡献[D]. 北京. 中国科学院大学. 2017.
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