我国南方酸性土壤完全氨氧化微生物多样性及其功能研究

Nitrification, the microbial oxidation of ammonia to nitrate via nitrite, was conventionally assumed as a two-step process in which ammonia oxidation was thought to be catalyzed by ammonia-oxidizing bacteria (AOB) and archaea (AOA), as well as nitrite oxidation by nitrite-oxidizing bacreria (NOB). Recently, the long-held assumption of labour division between the two functional groups, however, was challenged by the unexpected discovery of complete ammonia oxidizers (Comammox) that are capable of converting ammonia to nitrate in a single organism. Nitrification in acidic soils was also the focus of the practice of agricultural production. It will be of significant importance to explore the nitrification of acidic soils and its microbial driving mechanism. In this proposed project, we aimed to study the microbially medicated nitrification mechanism in acidic soils of sourthern China. Traditional and cutting-edge molecular methods ( real-time PCR and Illumina Miseq sequencing) in combination with stable isotope probing techniques were employed to decipher the contribution of Comammox、AOB and AOA to nitrification in acidic soils; the distribution and ecological characteristics of Comammox in large scale acidic soils; the effects of long-term fertilization on the abundance, community composition of Comammox in acidic soils. Overrall, the implementation of this project will be very important to understand the microbial mechanism of nitrification and ecological characteristics of Comammox in widely-distributed acidic soils.

硝化过程是氨被微生物经亚硝酸盐氧化为硝酸盐的过程，被认为主要由氨氧化微生物（AOB和AOA）和亚硝酸盐氧化细菌（NOB）主导完成。近来，完全氨氧化微生物（Comammox)能够经过一步反应完成氨到硝酸盐的转化，这一发现有力地挑战了长久以来的理论认识。酸性土壤硝化过程是农业生产实践的热点问题，探究酸性土壤硝化作用及其微生物驱动机制对农业生产具有重要意义。本项目拟对我国南方酸性土壤中参与硝化过程的关键微生物进行深入研究，运用实时荧光定量PCR技术、Illumina Miseq测序技术和稳定同位素探针技术深入全面的研究Comammox、AOB和AOA对酸性土壤硝化过程的相对贡献，Comammox在大尺度范围内的分布特征及环境影响因素；长期施肥管理对酸性农田土壤Comammox的丰度、群落组成的影响。研究结果有望揭示酸性土壤中Comammox参与硝化作用的机理及其生态学特征。

本项目对我国典型农业土壤参与硝化过程的关键微生物进行研究。主要研究内容有：（1）探究热带森林向香蕉园转变后参与硝化过程的关键微生物；（2）探索不同施肥措施对酸性土壤硝化微生物群落的影响；（3）探索不同施肥措施对黑土硝化微生物群落的影响；（4）探索施肥方式与硝化抑制剂对土壤中硝化微生物群落的影响。主要结果如下：.1、对自然林地和连作2年的健康蕉园土壤来说，DNA-SIP标记试验发现，单独添加FOC（尖孢镰刀菌）会限制AOB被标记，但是添加尿素后能AOB被成功标记。对连作12年的发病香蕉土壤来说，AOB已经占据主导地位，高氮条件下反而会抑制AOA。结果表明林地-蕉园长期转换使得由AOA主导的氨氧化作用转变为AOB。.2、通过对长期施肥监测的红壤进行研究发现，有机肥与化肥混施可以显著提高土壤典型硝化微生物（AOA、AOB、NOB）和完全氨氧化物（Comammox）丰度和物种多样性，促进土壤硝化作用，增强氮循环过程。而单施含氮化肥会导致其丰度降低，物种多样性减少；长期施肥情况下，pH值和OM含量是影响硝化微生物丰度、群落结构和功能的主要驱动因子。.3、长期施肥处理显著提高了土壤的硝化活性，含氮化肥施加显著降低了AOA的丰度，增加了AOB和Comammox的丰度。然而，有机肥与化肥配施显著提高了AOA、AOB和Comammox的丰度。长期施肥改变了Comammox的群落结构。土壤pH值和NH4+-N是决定Comammox群落关键因素。.4、分次施肥可以提高红壤净硝化速率，而黑土一次性施肥处理中净硝化速率更高，DMPP抑制土壤硝化过程。红壤中施加氮肥增加了氨氧化菌和亚硝酸盐氧化菌丰度，而DMPP则抑制AOB的生长。黑土中施加氮肥增加了硝化微生物的丰度，DMPP则显著抑制了AOB和Comammox的生长。两种类型土壤中，硝化微生物群落对硝化抑制剂和施肥的响应不同。