生物黑炭调控苹果根区土壤硝酸盐代谢的生物学机制

The excessive accumulation of nitrate is the primary cause of N2O emission, nitrogen loss, nutrient imbalances and other problem in soil, the effect apple roots on soil is profounder than annual crop. This project will investigate black carbon and metabolism of nitrate in apple root-zone soil and then carry on the experiments of biochar addition and nitrate carrying capacity under the system of orchard, potted plant，root chamber underground and pure soil separately. The effect of biochar on apple roots, production of nitric oxide，hydroxylamine reductase, and the activity and gene expression of nitric oxide reductase in soil, ect, will be explored for resolving the relationship of biochar → apple root → nitrate metabolism in soil. At the same time, microbial activity characterized by TTC-ETS and microbial biomass nitrogen and carbon in soil will be determined and the microbial community structure and functional diversity represented by the denitrifying bacteria, the bacteria capable of dissimilatory nitrate reduction to ammonium (DNRA), ammonia-oxidizing bacteria (AOB) and archaea (AOA) will be analyzed for detecting the relationship of biochar → microorganism → nitrate metabolism in soil. With that, the mechanism of nitrate metabolism regulated by biochar through roots and microorganism will be revealed in apple root-zone soil. This will provide the theoretical basis for the reduction of N2O emission and nitrogen waste and provide the theoretical guidance for the creation of "environment-friendly" technology of soil management in orchard.

硝酸盐积累是土壤N2O排放、氮素流失和养分失衡的根源；苹果根系对土壤影响比一年生作物深刻。本项目在调查苹果根区土壤黑炭与硝酸盐代谢的基础上，通过土施生物黑炭和硝酸盐承载试验，在果园、盆栽、根窖和纯土壤体系下，探讨苹果根系发育、土壤和根系一氧化氮（NO）释放、土壤羟胺还原酶（HyR）和一氧化氮还原酶（NoR）活性及其基因对黑炭的反应，解析"生物黑炭→苹果根系→土壤硝酸盐代谢"的关系；同时检测黑炭处理后土壤微生物TTC-ETS活性和微生物量氮和碳，分析反硝化细菌、硝酸盐异化还原为铵（DNRA）的细菌、氨氧化细菌（AOB）和氨氧化古菌（AOA）所表征的微生物群落结构及其功能多样性，摸清"生物黑炭→土壤微生物→硝酸盐代谢"的关系，进而揭示生物黑炭通过根系和微生物调控根区土壤硝酸盐代谢的机制，为减少果园土壤N2O排放和氮素损耗提供理论依据，为研发"环保型"果园土壤管理技术提供理论指导。

硝酸盐代谢是影响土壤氮素流失、氮氧化物排放和养分失衡的重要因素；生物黑炭由秸秆、枝条等生物质材料在缺氧条件下高温炭化而成，施入土壤显著影响土壤和根系硝酸盐代谢。.本项目调查了苹果根区土壤黑炭及硝酸盐代谢，分析了生物炭对根系与土壤硝酸盐代谢以及对土壤硝酸盐承载能力的影响，研究了炭化苹果枝对土壤硝化-反硝化作用、硝酸盐代谢酶、一氧化氮（NO）、一氧化二氮（N2O）、细菌和真菌多样性、氨氧化细菌（AOA）、氨氧化古菌（AOB）和反硝化细菌，以及对苹果根系硝酸盐代谢及其相关基因的调节，探讨了“生物炭--苹果根系---微生物---硝酸盐代谢”的关系，进行了根系转录组分析，研究了发酵果木屑和地面覆盖对土壤有机碳、微生物及根系硝酸盐代谢的影响。.主要结果表明，根区土壤有机碳和硝酸盐代谢明显受生物炭影响，1.0%-2%（w/w）的炭化苹果枝施入土壤明显提高土壤对硝酸盐的承载力，并可通过调节硝酸盐代谢而减少土壤氮氧化物释放，还可通过促进根系对硝酸盐的还原而提高苹果砧木的氮素吸收利用率，降低硝酸盐达到最大生理效应时的用量。施入1%的炭化苹果枝明显影响根区土壤细菌和真菌多样性，提高根区土壤细菌丰富度，并可通过降低土壤反硝化细菌、AOA和AOB基因的拷贝数而降低土壤NO和N2O浓度。土壤施入1%的炭化苹果枝可引起根系中核糖体、氧化磷酸化、碳代谢、氨基酸生物合成等代谢通路中大量基因的差异表达，使硝酸盐-亚硝酸盐共运蛋白家族基因以及硝酸还原酶调控基因表达上调。此外，平邑甜茶MhNCED基因可通过ABA促进硝酸盐还原进而增强根系内源NO生成。.本研究揭示了生物炭调控根系和根区土壤硝酸盐代谢的机理，为解决硝酸盐积累所引发的土壤氮素流失、氮氧化物排放和养分失衡等问题提了供理论依据，也为果园废弃枝条的资源化利用提高了新途径。.项目执行期间发表16篇论文，授权5项专利，培养6名研究生，完成计划内容和指标。