反硝化作用对生态沟渠氮削减的贡献及机理研究

Eutrophication of water bodies has become an urgent problem in China. Nitrogen released from agriculture is a significant pollutant source, as fertilizer inputs to crops are generally higher than the amount of nitrogen required to maximize plant productivity. Ecological ditch (Eco-ditch) is considered as the best management to tackle diffuse pollution from agriculture and has gained more and more attentions. It is well known that denitrification contributes mostly for the nitrogen removal of contaminated water, which is mainly conducted by denitrifying bacteria. Microbial community structure is always related with the ecological function, the study of denitrifers in the eco-ditches seems indispensable for better understanding of the N pollution-purification of eco-ditches. Firstly, we intend to detect the dynamic emissions of N2 and N2O from the eco-ditch, through which to determine the contribution of denitrification to eco-ditch N removal; Then, we employed the molecular technologies , such as q-PCR and Illumina Miseq, to reveal the temporal and spatial variations of the denitrifying bacterial abundance, community and diversity; Finally, 15N-DNA-SIP is used to confirm the active denitrifers in the eco-ditches. Meanwhile, we measure the environmental parameters which influence the denitrification rate. Through this study, we will have a more general understanding of the ecological functions of eco-ditches in N pollutant purification and help to provide theoretical guidance for the intensification of biological denitrification .

我国水体富营养化问题严峻，其中农业源氮是重要的污染源。利用农田排水沟渠构建生态拦截型沟渠用于削减、去除农田径流和渗漏损失的氮磷已成为国内外学者开展农业面源污染防控与治理研究的热点。生物反硝化脱氮是水体氮去除的重要途径，主要由反硝化细菌参与完成。反硝化细菌的群落组成与其生态功能密切相关，对生态沟渠系统中的反硝化细菌进行研究十分必要。本项目拟首先通过对生态沟渠系统释放的氮气和氧化亚氮进行动态测定，明确反硝化作用对生态沟渠氮削减的贡献；然后利用q-PCR和Illumina Miseq测序技术对反硝化细菌的群落组成、丰度和多样性进行研究，揭示生态沟渠反硝化细菌的时空变化特征；最后利用15N-DNA-SIP技术确认生态沟渠系统中的活性反硝化菌群。本工作的开展及其结果将有助于提高对沟渠系统净化农田排水氮功能和反硝化微生物的认识，可以为强化生态沟渠反硝化脱氮提供理论依据和技术支持。

生物反硝化脱氮是水体氮去除的重要途径，主要由反硝化细菌参与完成，其群落组成和物种丰度与氮净化功能密切相关。本文以沟底种植常绿苦草的三种类型的生态沟渠—水泥光面（CED）、常规土面（SED）和植草砖面（GED）为主要研究对象，同时以未植苦草的普通水泥排水沟为对照，利用定量PCR和Illumina高通量测序技术对生态沟渠系统的细菌和反硝化细菌的物种丰度和群落组成进行分析，揭示不同类型生态沟渠系统细菌和反硝化细菌分布特征，并进一步探究其与污水氮净化的关系。.本研究发现（1）三种生态沟渠对不同形态无机氮的去除效果存在差异，SED对NH4+去除效果较差，CED对NO3-、NO2-去除效果不佳，GED对三种无机氮的去除效果均较好。对总无机氮（TN）的去除效果为：GED>SED>CED；（2）沟渠类型对苦草叶面附着细菌和浮游细菌群落组成有显著影响（P<0.05），而对沉积物细菌群落组成影响不显著（P>0.05），变形菌门和拟杆菌门在生态沟渠系统中占据优势地位；（3）沟渠反硝化细菌丰度依次为：水体>苦草叶面>沉积物，并且沟渠底部种植苦草更有利于增加水体反硝化细菌的丰度。浮游细菌优势反硝化菌属主要为Limnohabitans、Fluviicola和黄杆菌属；苦草叶面主要优势反硝化菌属依次为红细菌属、嗜氢菌属、气单胞菌属和黄杆菌属；沉积物中占优势的反硝化菌属依次为硝化螺菌、Ignavibacterium和地杆菌属；（4）RDA分析结果表明，GED和SED沟渠nirS型反硝化菌群落组成更相似，DO和EC显著影响其群落组成；不同沟渠nirK型反硝化菌群落组成分异不明显，水温和NH4+对其群落有显著影响。生态沟渠苦草叶面nirK型反硝化菌α多样性低于nirS型反硝化菌，对应不同沟渠多样性分别为SED >CED>GED和SED>GED>CED。因此，nirS型反硝化菌可能与硝氮去除密切相关。陶厄氏菌属与DO显著负相关，其在SED和GED中丰度大于CED，表明CED高DO不利于该菌生长从而抑制其反硝化除氮。