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题名: 典型水稻土中胞外呼吸菌多样性及其对碳氮元素转化的影响
作者: 王宁
学位类别: 博士
答辩日期: 2015-05
授予单位: 中国科学院研究生院
授予地点: 北京
导师: 朱永官
关键词: 典型水稻土,碳氮,胞外呼吸,阳极菌,耦合效应,Typical paddy soil, Carbon and nitrogen, Extracellular respiration, anode respiring bacteria, Coupling.
其他题名: Distribution of extracellular-respiring microbes and their functions in carbon and nitrogen cycles in Chinese paddy soils
学位专业: 环境科学
中文摘要:     水稻土是我国主要的土地利用方式,是保障我国粮食安全的重要自然资源。和旱地土壤系统不同,水稻土经历频繁的干湿交替,使其物理化学和生物性质发生深刻的变化,特别适合于厌氧或微好氧微生物的生长。由于其独特的生物地球化学过程,水稻土常被当作模式土壤生态系统,用来研究温室气体排放的微生物学机制。其中,胞外呼吸菌在水稻土生物地球化学循环中发挥了不可替代的作用,但是目前国际上尚没有对这一类功能菌群结构和功能开展系统性的研究。过去尽管有不少关于胞外呼吸菌对金属,如铀的还原及其生物修复的研究,但是大部分集中在亚表层环境。而对于水稻土这类典型的地表环境中胞外呼吸菌的组成、铁的微生物还原及其与生物地球化学过程的互相作用机制仍缺乏系统的认识。
    本论文以我国典型水稻土作为研究出发点,初步研究了不同类型水稻土壤中碳氮铁生物地球化学循环及微生物群落多样性分布特征,借助微生物燃料电池(Microbial Fuel Cells, MFCs)装置的构建,逐步深入研究了不同典型水稻土壤中胞外呼吸菌的多样性及其主要控制因子。为了进一步了解土壤性质对阳极菌的影响,选择多种类型的水稻土详细探究了不同种类的有机酸和尿素施用对于阳极菌群落分布的影响,探讨了阳极菌与碳氮转化的相关关系。具体研究结果:
1)五种不同类型的水稻土中,淹水条件下,尿素施用对水稻土中碳氮铁循环的影响存在着显著的差异。尿素施用明显促进了嘉兴(JX)、鹰潭(YT)、湖南常德红壤(CR)以及雷州(LZ)土壤中甲烷(CH4)释放速率,尤其在淹水初期。然而对于湖南常德冲击土(CA)中CH4释放速率并无显著影响;在淹水后期,尿素施用明显促进了水稻土JX、YT和CA中一氧化氮(N2O)释放速率。在淹水期间,水稻土JX、YT、CR、LZ中溶解性Fe2+先升高后又逐渐降低,仅有水稻土CA中其含量是逐渐升高的。而对于水稻土JX和CA,尿素施用明显抑制了溶解性Fe2+ 含量,但对于其他三种水稻土并无显著影响。
2)五种不同类型的水稻土中,淹水使细菌群落组成发生明显迁移,而尿素处理对细菌群落组成影响并不大,但明显改变了一些细菌种群的相对丰度。不同类型的水稻土中,受尿素施用影响的微生物种群亦不同。孔隙水中二价铁(Fe2+)、硝酸根(NO3–)、氨根(NH4+)和溶解性有机碳(DOC)是影响土壤细菌群落的重要因子,对土壤细菌群落组成的变异贡献量为33.4%。因此尿素施用到土壤中后可能主要通过改变土壤性质,影响了一些细菌群种群的相对丰度。
3)通过构建土壤微生物燃料电池,研究发现,与对照(没有MFC运行的土壤)相比,五种类型的水稻土中,MFC运行明显改变了阳极菌群落结构。对于低产电量的水稻土LZ、JX 和 YT,阳极上Betaproteobacteria纲和Burkholderiales科明显被富集。而对于高产电量的水稻土CA 和 CR,阳极上Deltaproteobacteria纲和Geobacter属明显被富集。另外,对于所有类型的水稻土(除LZ外), 阳极上Bacilli纲和Bacillus属被明显富集。而雷州土壤中阳极菌Gammaproteobacteria 被明显富集。对于所有类型的水稻土,MFC运行并没有导致阳极上Clostridia纲和Clostridium属富集,然而Clostridia在水稻土YT、CA和CR上阳极菌群落组成上仍占有相当大的比例。
    环境因子与阳极菌群落组成的冗余分析显示,在所有的环境因子中,孔隙水NH4+、DOC和Fe2+是五种类型的水稻土中阳极菌群落组成差异性的主要贡献因子。这些因子解释了阳极菌群落变异量的55.1%。四种主要的阳极种群(Burkholderiales、Bacillus、Clostridium和Geobacter) 与孔隙水NH4+、DOC和Fe2+相关分析显示,Burkholderiales与DOC、NH4+呈明显的负相关;Clostridium、Geobacter与DOC、NH4+呈明显的正相关;而Bacillus仅与DOC呈明显的正相关。不过孔隙水Fe2+与这四种微生物种群并无显著相关性。这些发现表明土壤性质是影响阳极菌群落组成的重要因素。
4)基于微生物燃料电池装置,探究了两种类型的水稻土壤中尿素施用对阳极菌多样性的影响。研究发现对于有机碳含量较高的水稻土中,尿素施用对其阳极上细菌群落结构影响并不显著,而对于有机碳含量较低的水稻土中,尿素施用可以改变其阳极上细菌群落结构。其影响机制可能是厌氧下阳极上富集的Bacillu和Clostridium及未富集的Geobacter很可能参与了土壤微生物燃料电池中氨的相关转化。
5)选取九种有机碳种类和含量有差异的水稻土[湖南桃源(HNTY)、贵州(GZ)、四川(SC)、湖北(HB)、江西鹰潭(JXYT)、浙江红壤(ZJHR)、安徽(AH)、哈尔滨(HEB)、广东(GD)],构建微生物燃料电池。对于环境因子与阳极菌群落组成的冗余分析显示,在所有的环境因子中,土壤pH和孔隙水DOC解释了变异量的50.83%,结果表明pH和DOC是阳极微生物群落组成的重要影响因子。不同类型的水稻土阳极微生物呈现出较为丰富的多样性,优势种群较多。这可能是由于小分子有机酸作为有机碳的重要组成部分,能被许多阳极菌所利用。然而根据主成分分析结果显示,不同类型的水稻土中阳极微生物群落仍有显著的差异,这可能与土壤中有机酸的种类和含量的差异有关,其中丙酸和乳酸的作用较为明显。其影响机制可能是厌氧下阳极上富集的阳极菌直接或间接的参与了土壤微生物燃料电池中碳的相关转化,但碳与阳极菌之间的相互作用机制仍不够清晰,仍需进一步的研究。
本论文初步研究了典型水稻土中元素循环及微生物群落变化,并基于微生物燃料电池深入研究了不同类型的水稻土中微生物胞外呼吸菌组成,分布以及对土壤中碳氮的生物地球化学循环的影响,可为胞外电子传递菌的群落特征及微生物介导下的氧化还原反应的研究提供科学依据。
英文摘要:     Paddy field is a typical agricultural ecosystem and plays an important role in food supply and ecosystem sustainability. When it is flooded, the redox potential below overlying water immediately decreases and becomes negative. Paddy soil is a good model of studing soil microbial ecology due to its unqiue conditions. Numerous studies have focused on paddy soils to investigate the microbial processes of greenhouse gases emission. The finding of the microorganisms with the extracellular respiration functionality provides a new perspective for understanding the evolution of microbial respiration and element biogeochemical cycling in paddy soils. Previous studies have shown the effect of the extracellular respiration microbes on heavy metal remediation in subsurface environment. However, more studies would be needed to focuse on the distribution of extracellular-respiring microbes and their functions in coupled biogeochemistry of carbon and nitrogen for paddy soils.
    In this study, serveal typical paddy soils with various physicochemical properties were collected from China. First, the biogeochemistry of carbon, nitrogen and iron and the bacterial community structure were investigated. Then, we further investigated the distribution of anode-respiring bacteria and the influence of soil characteristics on anode bacterial community composition by constructed paddy soil microbiual fuel cells (MFCs). To futher elucidate the role of soil properties in anode microbial community composition, serveal typical paddy soils were selected to investigate the influence of the organic acid and urea addition on anode microbial community. Results showed that:
1) Urea amendment had obvious different effects on soil carbon and nitrogen cycling in various paddy soils during the flooding incubation. Urea amendment significantly promoted CH4 production for JX, YT, CR and LZ, especially in the intial period of flooding. After 8 days, urea amendment significantly promoted N2O production for JX, YT, and CA
2) Flooding incubation without urea addition (control treatment) significantly changed bacterial community composition of 5 soils, but had no obvious effects on soil bacterial community composition shift. However, urea amendment significantly increased the relative abundances of some genera, which are related to the C and N cycling. The effects of urea amendment on specific genera varied among soils. Porewater Fe2+, NO3–, NH4+, and DOC that significantly correlated with the bacterial community composition were selected in RDA analysis. These factors together explained 33.4% of the soil bacterial community composition variation. Different soil characteristics might explain why effects of urea amendment on specific genera varied among soils.
3) Compared to control treatments (without MFC treatment), MFC running significantly altered bacterial community composition at anodes in each soil. At class level, MFC running significantly enriched Betaproteobacteria for LZ, JX, and YT soils by a factor of 4–30 times, which showed low Pd. In the soils with high Pd (CA and CR), Deltaproteobacteria was enriched 4–20 times at anodes. In addition, Bacilli was also significantly enriched at anodes for all soils except for LZ. In contrast, Gammaproteobacteria was only enriched in the soil LZ. For all soils, MFC running did not lead to an enrichment of Clostridia. Although not enriched, Clostridia shared a great portion of microbial communities at anodes for YT, CA, and CR. The relative abundance of order Burkholderiales which dominates     Betaproteobacteria was significantly enriched by MFC running in soils LZ, JX, and YT, while genus Bacillus which dominates Bacilli was enriched in all soils except for LZ. Geobacter which are the dominant genus of Deltaproteobacteria was significantly enriched with MFC running in soils CA and CR. However, in all soils with MFC running, the relative abundance of Clostridium (belongs to Clostridia) was not increased.
    Redundancy analysis (RDA) showed that porewater NH4+, DOC, and Fe2+ were major factors that attributed to the variation of microbial community composition among all possible influence factors (porewater DOC, NH4+, Fe2+, soil TC and TN, overlying water pH and EC, and Eah). In total, these factors could explain 55.1% of the soil microbial community composition variation. The correlations between relative abundance of the 4 main bacterial taxons (Burkholderiales, Bacillus, Clostridium, and Geobacter) and porewater NH4+, DOC, and Fe2+ were developed, showing that Burkholderiales was significantly negatively correlated with DOC and NH4+. The relative abundance of Clostridium and Geobacter were significantly positively correlated with NH4+ and DOC, and that of Bacillus was only positively correlated with DOC. Significant correlations with Fe2+ were not found for all the 4 bacterial taxons.
5) Two types of paddy soils were collected to investigate the influence of N (urea) on ERB community composition Urea amendment did not change anode bacterial community structure in soils with high DOC, suggesting that DOC rather than NH4+ was the major factor controlling bacterial community. However, when DOC was relatively lower, added NH4+ significantly altered the bacterial community structure. These findings imply that soil properties shape the soil bacterial communities and provide new insights into the microbial–mediated nitrogen cycling in paddy soils. Results suggest that Bacillus, Clostridium and Geobacter might be involved in NH4+ transformation, while the mechanism behind the interactions between NH4+ and ERB require further studies.
4) Nine paddy soils were collected from China with various organic carbon and constructed paddy soil MFCs. Redundancy analysis (RDA) showed that soil pH and DOC were major factors that attributed to the variation of microbial community composition among all possible influence factors (porewater DOC, NH4+, NO3-, SO42- and pH). The dominant population of anode microbial community composition with more species in various paddy soils might due to the small organic organic acids which could be used by mang iron reducing bacteria. The microbial community compositions at anodes varied among the 9 soils with MFC running according to Adonis analysis (p < 0.05). Soil organic acid, especially Latate and propionic, might play an intial role in the variation of microbial community composition among soils. These findings imply that anode microbes might be involved in carbon transformation, while the mechanism behind the interactions between carbon and extracellular respiring bacteria (ERB) require further studies.
    In this study, the biogeochemistry of carbon, nitrogen and iron and the bacterial community structure were investigated. Then, we further investigated the distribution of anode-respiring bacteria and the influence of soil characteristics on anode bacterial community composition by constructed paddy soil microbiual fuel cells (MFCs). MFC experiments highlight the importance of soil properties in shaping bacterial communities at MFC anode and point to a need of future studies to study how both quantity and style of DOC and NH4+ influence MFC bacterial communities, especially AEB composition. The mechanism between NH4+ and AEB has yet to be fully understood and require further studies. Our study provides new insights into the microbial–mediated carbon and nitrogen cycling in paddy soils.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/34437
Appears in Collections:中澳联合土壤环境研究室_学位论文

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Recommended Citation:
王宁. 典型水稻土中胞外呼吸菌多样性及其对碳氮元素转化的影响[D]. 北京. 中国科学院研究生院. 2015.
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