黄河三角洲滨海湿地铁循环微生物多样性和电活性

	黄河三角洲滨海湿地铁循环微生物多样性和电活性
	张洪霞 1,2
学位类型	博士
导师	刘芳华
	2017-05-18
学位授予单位	中国科学院大学
学位授予地点	北京
关键词	滨海湿地河口铁循环菌多样性电活性
其他摘要	铁是滨海湿地环境中重要的变价金属元素，铁还原菌和铁氧化菌介导的铁的氧化还原过程与碳、氮等元素的循环相耦合，在生物地球化学循环过程中起到重要的驱动作用。阐明滨海湿地沉积物中铁还原菌和铁氧化菌群落结构的变化和分布特征，及其主要环境驱动因素，对于认识河海交互作用区域铁还原菌和铁氧化菌在生物地球化学循环过程中作用具有重要意义。黄河三角洲作为典型的滨海湿地，是世界上陆-河-海交互作用最活跃的区域之一。本研究以黄河三角洲滨海潮滩湿地、河口及近海沉积物为研究对象，通过对原位土壤/沉积物及其富集培养样品进行基于16S rRNA和rRNA高通量测序以及对原位样品环境因子的测定，解析了滨海区域不同空间和生态演替尺度下细菌群落组成的变化规律，及其主要环境影响因素；分析了河海交互作用区域铁还原菌和铁氧化菌群落结构组成和变化特征；揭示了潮滩湿地不同盐度植物带土壤铁还原菌和铁氧化菌群落结构差异；分离得到具有铁还原能力和电化学活性的团聚体，发现电子穿梭体蒽醌-2,6-二磺酸(AQDS)和活性炭能够明显促进团聚体的胞外电子传递；证实了Geobacteraceae和Methanosarcina mazei的共生体存在于滨海河流沉积物富集体系中。研究中得到的主要结论如下： 1、黄河三角洲滨海潮滩湿地、河口及近海不同生态系统沉积物细菌群落组成存在明显差异。在陆地-河流-海洋交互作用的典型滨海区域，细菌群落多样性和空间分布特征受环境因素和生境的影响。河流沉积物中具有最高的细菌丰度和多样性，主要优势菌为在氢、氮、铁循环中起重要作用的Hydrogenophaga、Nitrospira、Pseudomonas和Thiobacillus；潮滩湿地优势细菌主要为耐盐菌Planctomyces、Marinobacter、Halomonas、Salinivibrio和Salinibacter；而Lutimonas、Desulfococcus、Photobacterium、Propionigenium和Vibrio相对丰度在近海沉积物中较高。盐度、pH和SO₄^2-是显著影响滨海区域细菌群落结构变化的环境因素。 2、黄河三角洲滨海潮滩湿地铁还原菌和铁氧化菌相对丰度随盐度梯度带的变化呈现不同的变化趋势和分布特征。铁还原菌相对丰度和铁还原能力与土壤盐度呈负相关；反之，铁氧化菌相对丰度与土壤盐度呈正相关。Geobacter、Pseudomonas、Thiobacillus、Bacillus、Anaeromyxobacter和Gallionella、Thermomonas分别为黄河三角洲滨海潮滩湿地较低盐度植物带（白茅-芦苇、芦苇）原位土壤中占主要优势的铁还原菌和铁氧化菌；Bacillus、Desulfovibrio、Clostridum和Marinobacter分别为高盐度植物带（碱蓬、柽柳）原位土壤中占优势的铁还原菌和铁氧化菌。土壤盐度是显著影响滨海湿地铁还原菌和铁氧化菌群落结构的重要环境变量。滨海潮滩湿地低盐度土壤样品具有较高的铁还原活性。 3、黄河口及近海不同生态系统沉积物中铁还原菌和铁氧化菌丰度及多样性表现出空间异质性。在河海交互作用区域沉积物中，铁还原菌和铁氧化菌多样性和分布特征显著受环境因子和生境类型的影响。河口沉积物中铁还原菌和铁氧化菌相对丰度和多样性相对高于近海沉积物样品。Pseudomonas、Thiobacillus、Geobacter、Rhodoferax、Clostridium和Azospira、Gallionella、Rhodobacter、Thermomonas分别为河口沉积物中占主要优势的铁还原菌和铁氧化菌；而近海沉积物中占主要优势的铁还原菌和铁氧化菌分别为Vibrio、Shewanella、Thiobacillus 和Marinobacter。对富集培养样品分析发现，海洋沉积物样品的铁还原菌相对丰度明显高于河流沉积物样品，海洋沉积物样品具有一定的铁还原活性，但河口沉积物样品的铁还原活性相对高于海洋沉积物样品。说明原位海洋沉积物铁还原菌相对丰度尽管较低，但可能添加铁氧化物能够促进海洋沉积物中某些种类铁还原菌的生长，从而促进其铁还原活性。在河口及近海区域铁循环菌和已知氮循环菌（属水平）呈现出相同的规律性分布特征，在河海交互作用区域（BHB02和YR3）沉积物中铁循环菌和已知氮循环菌相对丰度和多样性都最高，并且铁循环菌和氮循环菌相对丰度可能呈正相关。此外，在黄河口及近海区域，沉积物TN、NH₄⁺-N和NO₃^--N是显著影响铁还原菌和铁氧化菌群落结构的环境变量。 4、在滨海湿地环境中分离到一种由细菌与细菌（Desulfovibrio和Pseudomonas）组成并具有铁还原能力的团聚体C1。在不同类型有机底物（乙酸和乙醇）条件下Desulfovibrio和Pseudomonas相对丰度发生变化；且在乙醇为底物时，团聚体铁还原能力受盐度影响较小。电子穿梭体AQDS和活性炭能够明显促进团聚体C1与Fe(III)氧化物之间的电子传递。团聚体在有透析袋包裹电极的微生物燃料电池中具有较高的电活性，说明其可能通过电子穿梭体进行胞外电子传递。 5、通过对滨海河流界河沉积物样品富集培养，并基于16S rRNA/rRNA 的分子生物学方法，证实了Geobacteraceae和Methanosarcina mazei的共生体存在于界河沉积物富集体系中，并且在铁还原富集体系中具有铁还原活性和产甲烷能力。微生物种间常以团聚体的形式进行直接种间电子传递，本研究以乙醇为底物时经过3代转接培养，M. mazei和Geobacteraceae形成紧密结合的团聚体，这表明两者可能能够进行直接种间电子传递。界河沉积物富集样品中主要优势细菌和古菌分别为Geobacteraceae、Bacilli和Methanosarcinaceae。 ; Iron is one of the most abundant variant valence metal elements. Iron-reducing bacteria (IRB) and iron-oxidizing bacteria (IOB) play pivotal biogeochemical roles in the coupling of iron and carbon or nitrogen redox cycles. In the coastal wetland environments, pattern in abundance, diversity of IRB and IOB can provide evidence of their contribution to biogeochemical cycles in assessing their community structural distinctions associated with spatial variations of nutrients in different habitats. As a typical coastal wetland, the Yellow River Delta is one of the most active regions of wetland-river-sea interaction in the world. In this study, spatial patterns and community structures of IRB and IOB in soil/sediment samples from the Yellow River estuary through intertidal wetland to coastal marine environments were investigated. Deep sequencing of bacterial 16S rRNA gene and rRNA in field and enrichment samples revealed significant shifts in diversity and distribution of bacterial community, iron-reducing and -oxidizing bacterial communities in sediment samples from the Yellow River estuary to Bohai Sea, and in wetland soils with different salinity gradient. The effects of biogeochemical variables along the gradient on the abundance and biogeographic patterns of bacteria, putative IRB and IOB in sediments were investigated. Aggregates with iron reduction and electrical activity were obtained. AQDS and granular activated carbon can obviously promote extracellular electron transfer for the aggregate. The co-occurrence of Geobacteraceae and Methanosarcina mazei and their activities in an iron(III)-reducing microbial community from a coastal gold mining river has been found. The main results were summarized as follows: Differences in bacterial community composition indicate that community diversity and spatial distribution of bacteria in sediments from the typical coastal area with land-river-sea interaction are mainly determined by biogeochemistry and habitats. Yellow River sediment had the highest diversity and was dominated by hydrogen-, nitrogen- and iron-cycling bacteria, such as Hydrogenophaga, Nitrospira, Pseudomonas, and Thiobacillus. The intertidal wetland had a haloduric community with diverse function (such as Planctomyces, Marinobacter, Halomonas, Salinivibrio and Salinibacter). The Bohai Sea sediment had higher relative abundances of Lutimonas, Desulfococcus, Photobacterium, Propionigenium, and Vibrio. Salinity, pH and SO₄^2- concentration were key parameters, which contributed to variations in the bacterial communities. Relative abundance of IRB and IOB in samples from intertidal wetland with different salinity gradients showed distinct responses to salinity, revealing considerable spatial heterogeneity even within this relatively small site.. The diversity of iron-reducing bacterial populations was negatively correlated with soil salinity, while the diversity of iron-oxidizing bacterial populations was positively correlated with soil salinity. Iron-reducing bacteria Geobacter, Pseudomonas, Thiobacillus, Bacillus and Anaeromyxobacter, together with iron-oxidizing bacteria Gallionella and Thermomonas species were predominanted in soil samples with low salinity (Blendimings and Phragmites), respectively, whereas Bacillus, Desulfovibrio, Clostridum and Marinobacter species were the dominant iron-reducing and -oxidizing bacteria in soil samples with high salinity (Suaeda and Tamarix), respectively. Soil samples with low salinity have higher activity of iron(III) reduction. Change in relative abundance and spatial distribution of iron-reducing and -oxidizing bacterial communities in sediment samples from the Yellow River estuary to Bohai Sea, revealed substantial spatial heterogeneity between two ecosystems. Community diversity and spatial distribution of the IRB and IOB in sediments from the typical coastal area with river-sea interaction are mainly determined by biogeochemical factors and habitat type. Relative abundance and diversity of IRB and IOB in sediments from the Yellow River estuary were higher than those from the Bohai Sea. We found that Pseudomonas, Thiobacillus, Geobacter, Rhodoferax, Clostridium and Azospira, Gallionella, Rhodobacter, Thermomonas were the most abundance genera of IRB and IOB in sediments from the Yellow River estuary. And the dominant genera in sediments from the Bohai Sea were Vibrio, Shewanella, Thiobacillus and Marinobacter. The opposite trend was observed in enrichment cultures compared to in situ samples, indicating that relative abundance of IRB in marine sediments can be promoted in the presence of amorphous iron oxides in enrichment culture. However, sediment samples from river have higher activity of Fe(III) reduction than those from marine. Shifts in diversity of putative iron-reducing, iron-oxidizing and nitrogen-cycling bacteria discovered a novel distribution pattern in Yellow River estuary and adjacent Bohai Sea. There are the higher relative abundances and diversities of putative iron-reducing, iron-oxidizing and nitrogen-cycling bacteria at coastal transitional sites. Relative abundance of well-known nitrogen-cycling bacteria was positively correlated with relative abundance of iron-cycling bacteria. Additionally, concentrations of TN, NH₄⁺-N and NO₃^--N contributed to variations of iron-reducing and -oxidizing bacterial communities. Aggregate C1 with iron reduction activities formed by Desulfovibrio and Pseudomonas was isolated from coastal wetland. Differences in relative abundances of Desulfovibrio and Pseudomonas in aggregate C1 were observed when offered with different types of organic substrates. The effect of salinity on Fe (III) reduction of aggregate C1 was little in cultures with ethanol as the only electron donor. AQDS and granular activated carbon can obviously promote electron transfer between aggregate C1 and Fe (III) oxide. Aggregate produced higher electrical density in microbial fuel cell with dialysis bag, revealing it may use electron shuttle for extracellular electron transfer. The co-occurrence of Geobacteraceae and Methanosarcina mazei and their activities in an iron(III)-reducing microbial community from a coastal gold mining river, Jiehe River, has been found and confirmed by culture independent molecular techniques based on 16S rRNA and rRNA, respectively. Aggregates, as close physical contact necessary for direct interspecies electron transfer (DIET), have been found in the culture with ethanol as electron donor after three transfers, and indicated the possibility of DIET between them. The enrichment cultures actively contained the bacteria belong to Geobacteraceae and Bacilli, exclusively dominated by the archaea belong to Methanosarcinaceae.
文献类型	学位论文
条目标识符	http://ir.yic.ac.cn/handle/133337/22445
专题	中国科学院烟台海岸带研究所知识产出_学位论文
作者单位	1.中国科学院烟台海岸带研究所 2.中国科学院大学
第一作者单位	中国科学院烟台海岸带研究所
推荐引用方式 GB/T 7714	张洪霞. 黄河三角洲滨海湿地铁循环微生物多样性和电活性[D]. 北京. 中国科学院大学,2017.

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