Asymbiotic Nitrogen Fixation And Influencing Factors Of Soil And Plant Leaf In Alpine Grassland

Asymbiotic nitrogen fixation is an important input form of exogenous nitrogen in natural ecosystems,especially in those lacking symbiotic nitrogen fixation.Grassland on the Tibet Plateau lacks symbiotic nitrogen fixation mechanism due to the environmental conditions of arcticalpine and hypoxia.However,alpine grassland is rich in nitrogen nutrient.So,does asymbiotic nitrogen fixation mechanism make an important contribution?At the same time,what factors affect asymbiotic nitrogen fixation of main alpine grassland?The answer to this question is an important content for further understanding the mechanism of nitrogen transformation and cycling ecology,which can provide an important scientific basis for nitrogen management.In order to answer this scientific question,this study focused on alpine meadow（AM）,swamp meadow（SM）and alpine steppe（AS）in Qilian County,Gonghe County and Maqin County of Qinghai Province,respectively.The asymbiotic nitrogen fixation rates of soil,plant leaves,mosses and cyanobacteria in different alpine grassland communities were measured by acetylene reduction method and ¹⁵N₂ feeding method.At the same time,soil physical and chemical factors,plant community characteristics,soil asymbiotic nitrogen-fixing microbial community characteristics,soil related metal elements were tested.The effects of environmental factors on soil asymbiotic nitrogen fixation microbial community were analyzed,and the key influencing factors of soil and phyllospheric asymbiotic nitrogen fixation rates were identified.The annual asymbiotic nitrogen fixation rates of soil and leaves in three alpine grassland communities were estimated.The main results are as follows:（1）There were significant differences in compositions of soil asymbiotic nitrogen-fixing bacteria among different sample sites（p=0.001）.The species richness index and diversity index of swamp meadow were higher than other two alpine grassland communities.g＿＿unclassified＿k＿＿norank＿d＿＿Bacteria,g＿＿unclassified＿p＿＿Proteoba-cteria,and Skermanblla were dominant asymbiotic itrogen-fixing bacteria in alpine grassland.Among them,the nif H gene of g＿＿unclassified＿k＿＿norank＿d＿＿Bacteria and Frankia had high sequence similarity.Theβdiversity of soil asymbiotic nitrogen-fixing microbial community decreased significantly with the increase of soil p H and increased significantly with the increase of soil P:MO ratio and elevation.（2）Both soil and phyllospheric asymbiotic nitrogen fixation rates were the highest in swamp meadow.Among all the plots,swamp meadow in Qilian County（QL-SM）had the highest rates of soil and phyllospheric asymbiotic nitrogen fixation which were2.016×10^-7 mmol N₂·g^-1·d^-1 and 4.568×10^-7 mmol N₂·g^-1·d^-1,respectively.In different alpine grassland plant communities,the asymbiotic nitrogen fixation rates of soil and leaves were the highest in swamp meadow,followed by alpine meadow and alpine steppe.（3）The asymbiotic nitrogen fixation rates of leaves in alpine grassland ecosystem were 2.3 to 12.2 times as fast as soil,and the key factors affecting asymbiotic nitrogen fixation were different between two substrates.Soil asymbiotic nitrogen fixation rate was significantly inhibited by soil nitrate nitrogen,while stimulated by asymbiotic nitrogen-fixing bacteria and soil N:MO ratio.The latitude variation had a significant positive effect on phyllospheric asymbiotic nitrogen fixation rate.（4）The amount of nitrogen imported into these alpine grasslands by asymbiotic nitrogen fixation is estimated to be 0.1 T g per year on the Tibet Plateau.The annual asymbiotic nitrogen fixation rates of soil in different alpine grassland communities were35.5 to 54.5 times that of leaves in the same area.The annual total asymbiotic nitrogen fixation rate of swamp meadow was the highest(1.547 kg N·ha^-1·yr^-1),which was more than two times of that of alpine meadow(0.696 kg N·ha^-1·yr^-1)or alpine steppe(0.683kg N·ha^-1·yr^-1).（5）Mosses and cyanobacteria had greater potential for asymbiotic nitrogen fixation in alpine grassland.The asymbiotic nitrogen fixation rates of mosses and cyanobacteria in alpine grassland ecosystems were 7.4 to 99.0 times higher than those of soils in the corresponding plots.The asymbiotic nitrogen fixation potential of cyanobacteria(nitrogen fixation rate:64.806×10^-7 mmol N₂·g^-1·d^-1)was higher than mosses(1.655×10^-7 mmol N₂·g^-1·d^-1).In conclusion,asymbiotic nitrogen fixation contributed significantly to nitrogen in alpine grassland.The composition of soil asymbiotic nitrogen-fixing microbial community in swamp meadow was the most complex and the annual asymbiotic nitrogen fixation rate was the largest.Soil p H had extremely significant negative effects onβdiversity of soil asymbiotic nitrogen-fixing microbial community,while elevation and P:MO ratio had extremely significant positive effects.Asymbiotic nitrogen fixation of soil in alpine grassland was dominated by soil nitrate,PD-whole-tree index（pedigree diversity）and soil N:MO ratio,while phyllospheric asymbiotic nitrogen fixation was dominated by latitude.Therefore,it is suggested that further research can be carried out to explore efficient asymbiotic nitrogen-fixing microorganisms and excavate beneficial asymbiotic nitrogen-fixing microbial resources.It will provide efficient nitrogen for restoration of degraded alpine grassland and promote nitrogen management of alpine grassland ecosystem.