酚酸对连作杨树人工林土壤硝化作用的影响与机制

Nitrogen is the most essential nutrient for plants. Nitrification is a crucial step in the transformation of soil nitrogen, and it is strongly affected by the rate-limiting step known as ammonia oxidization. Both ammonia oxidizing bacteria（AOB）and ammonia oxidizing archaea（AOA）play an important role in the ammonia oxidization. However, few studies have been done on the community succession of the soil AOB and AOA influenced by the phenolic acids. Moreover, present studies have never shown how the cumulative phenolic acids affect the nitrification in the soil. According to preliminary studies, the cumulative phenolic acids in the soil were reported to have significant effects on the microorganism community and nitrogen availability in the soil of continuously cropped poplar land. The general objective of this study was to find a solution to the soil degradation problem in the continuously cropped poplar land. To achieve this objective, allelopathy was treated as a vital study object. And the triple relations of the cumulative phenolic acids, the community succession of the soil AOB and AOA, and the nitrogen availability will be studied. Based on the plot-fixed filed experiments and laboratory simulated tests, the polymerase chain reaction-denaturing gradient gelatum electrophoresis(PCR-DGGE) and stable isotope labeling (SIL) techniques will be applied to investigate the effects of cumulative phenolic acids on the community characteristics of AOB and AOA, and the nitrification in the soil of continuously cropped poplar land. With a clear explanation to the community succession of soil AOB and AOA, as well as to the mechanism about how phenolic acids affect the soil nitrification, a vital scientific basis will be provided to reveal the essence of the soil degradation in the continuously cropped poplar land, and furthermore guide the sustainable management of the continuously cropped poplar plantations.

氮素是植物最重要的营养物质，硝化作用是土壤中氮素转化的重要过程，其中氨氧化作用是整个硝化作用的限速步骤，氨氧化细菌（AOB）和氨氧化古菌（AOA）在其中发挥着重要作用。目前国内外针对酚酸作用下土壤氨氧化菌群落演变规律研究鲜有报道，现有研究无法阐明土壤酚酸累积对土壤硝化作用的影响机制。基于前期研究基础，连作杨树人工林土壤中酚酸类物质的累积对土壤微生物群落及氮素有效性有显著影响。本研究针对连作杨树人工林地力衰退问题，以化感效应为切入点，开展"酚酸物质累积-氨氧化菌群落演变-氮素有效性"三者关系研究。采用大田定位试验和实验室模拟相结合，应用PCR-DGGE技术和稳定同位素标记技术，研究酚酸累积对连作杨树人工林土壤中AOB和AOA群落结构及硝化作用的影响，阐明酚酸作用下土壤AOB和AOA群落结构的演变动态，揭示酚酸对土壤硝化作用的影响机制，为深入揭示连作杨树人工林地力衰退机理提供科学依据。

本研究以化感效应为切入点，采用田间施肥与室内培养相结合的方法，对施加15N标记的铵态氮肥后杨树人工林土壤硝化作用中氮素的迁移与转化进行定量研究，结果表明，同一林地根际土壤中铵态氮和硝态氮的变化速率及硝化活性均显著大于非根际土；与氮损有关的羟胺还原酶和亚硝酸还原酶无显著差异，而硝酸还原酶活性呈显著差异。在不同林分中，铵态氮和硝态氮的含量及其变化速率、土壤硝化活性的变化均表明连作人工林土壤中铵态氮向硝态氮转化的微生物活性下降，转化速率减小，导致转化量减少，说明杨树人工林连作对土壤硝化作用产生了抑制性，并随着时间的推移，抑制性增强。梯度酚酸作用下连作人工林土壤硝化进程研究表明：酚酸一方面通过抑制硝化微生物的活性，影响铵态氮向硝态氮的转化，抑制硝化作用的进行；另一方面促进了与反硝化作用有关的硝酸还原酶活性，使氮素的气态损失增加，土壤中氮素“库”源含量下降，使杨树可吸收利用的氮素减少，林分生产力下降。基于宏基因组测序技术筛选氮素代谢功能基因，甄别参与土壤氮循环过程中的功能微生物类群，结果表明，参与氮素循环细菌有4类11属：固氮细菌，拜叶林克氏菌属（Beijerinckia）、慢生根瘤菌（Bradyrhizobium）、根瘤菌属（Rhizobium）、弗兰克式菌属（Frankia）；硝化细菌，硝化杆菌属（Nitrobacter）、亚硝化螺菌属（Nitrosospira）；反硝化细菌，假单胞菌属（Pseudomonas）、罗尔斯通菌属（Ralstonia）、伯克氏菌属（Burkholderia）、芽孢杆菌属（Bacillus）、链霉菌属（Streptomyces）；氨化细菌，芽孢杆菌属(Bacillus)、假单胞菌属(Pseudomonas)。与Ⅰ代林相比，Ⅱ代林土壤中氮素细菌的种类没有增减，但参与氮素循环细菌总数增加4.73%，其中固氮细菌丰度增加53.44%，硝化细菌丰度没有变化，反硝化细菌丰度增加0.14%，氨化细菌丰度增加1.33%。总代谢活性降低，其中固氮细菌代谢活性增强，硝化细菌中硝化杆菌属代谢活性减弱，亚硝化螺菌属代谢活性增强，氨化细菌代谢活性减弱。总之，杨树人工林连作一代后，土壤中参与氮素循环的细菌总数增加，但代谢活性降低。