Study On Response Mechanism Of Rhizosphere Soil Microenvironment And Leaf Nitrogen Metabolism Of Alfalfa To Nitrogen And Phosphorus Addition In Alkaline Soil

Soil salinization is one of the main obstacles to the sustainable development of agriculture.The northern part of Ningxia is located in arid and semi-arid region,which is an area with a more serious degree of salinization.According to literature studies,the alkaline soil is more harmful to plants than the saline soil.Alfalfa has the advantages of strong saline-alkali resistance,high yield,high quality and wide adaptability,and is an important high-quality forage for comprehensive improvement and rational utilization of saline-alkali land.It is of great significance to solve the shortage of land resources,improve ecological environment and alleviate food security.At present,the yield and protein content of alfalfa are lower in China,especially in northwest China,compared with developed countries and regions at home and abroad,due to the lack of basic research on alfalfa planting and back-ward production and management technology.Fertilization is the main measure to improve the yield and protein content of alfalfa,especially in saline-alkali land with poor fertility.According to literatures at home and abroad,the research on how nitrogen(N)and phosphorus(P)addition affect alfalfa yield and protein in alkaline soil is not systematic enough,and the mechanism is not very clear.Therefore,N and P addition experiments were set in 0 6 and 12kg/667m2 of N,and in 0 9 and 18kg/667m2 of P on moderate alkaline soil from 2019 to 2021 in Yinbei,Ningxia,the growth and development of alfalfa,rhizosphere soil microenvironment and nitrogen cycling microorganisms,key products and enzyme activities of leaf nitrogen metabolism were studied,so as to provide theoretical basis and technical guidance for scientific fertilization of alfalfa with high yield and high quality in alkaline soil.(1)Medium nitrogen(ZN)and medium phosphorus(ZP)promoted non-rhizosphere soil nutrient accumulation and alfalfa’s nutrient absorption,accumulation,utilization,growth and development.N addition increased carbon(C)and N content,decreased P content,were 14.081.27 and 1.25g/kg under ZN treatment in 20cm soil layer every year on average,respectively,improving N/P and C/P,reducing C/N in non-rhizosphere soil,P addition increased C and P content,decreasing N content,improving C/N,reducing C/P and N/P in non-rhizosphere soil,moderate N and P addition improved non-rhizosphere soil fertility,prevented nutrient restriction and waste,and promoted nutrient absorption,utilization,accumulation and growth of alfalfa in alkaline soil,but excessive addition had opposite effects.(2)The N and P addition promoted leaf nitrogen metabolism and protein formation of alfalfa.Proper N and P addition increased leaf N,P,nitrate nitrogen(NO3--N),ammonium nitrogen(NH4+-N),protein and chlorophyll content,but decreased leaf chlorophyll a/b,Proper N and P addition increased enzyme activities of leaf nitrate reductase(NR),glutamine synthase(GS),glutamate synthase(GOGAT)and glutamic-pyruvate transaminase(GOT)and glutamic-oxalacetic transaminase(GPT),but decreased amino acid content and protease activity.In general,Proper N and P addition increased the key products and enzymes of leaf nitrogen metabolism of alfalfa in alkaline soil,and promoted formation and accumulation of protein.(3)Different N and P addition had significant effects on soil chemical property in the rhizosphere soil of alfalfa(P<0.05).GN treatment significantly increased organic carbon(OC),total nitrogen(TN),ammonium nitrogen(AN)and nitrate nitrogen(XN)contents in rhizosphere soil,but significantly decreased the s total phosphorus(TP),available phosphorus(AP)and total salt(TS)contents(P<0.05),and decreased pH,GP treatment significantly increased the OM OC TP and AP contents in rhizosphere soil,significantly decreased TN XN AN and TS contents(P<0.05),and decreased the pH.N addition increased C/N C/P and N/P,increased by 23.08 51.93 and 40.24%under GN treatment every year on average,respectively,P addition increased C/N C/P,and decreased N/P,and GN and GP treatments reached significant levels(P<0.05).There were significant positive correlation between OM OC TN and AN contents of rhizosphere soil in N and P addition after correlation analysis,XN was positively correlated with TN and AN contents,TP was positively correlated with AP content,TP and AP were positively correlated with OC(OM)contents,but negatively correlated with TN,XN and AN contents,OC(OM)was negatively correlated with pH,PH was positively correlated with TS content(P<0.05).So Proper N and P addition reduced the pH and TS content of rhizosphere soil of alfalfa in alkaline soi,and improved the comprehensive fertility level.(4)N and P addition significantly changed the activities of nitrogen-related enzymes and bacterial diversity in rhizosphere soil(P<0.05).① ZN and ZP treatments significantly increased the activities of nitrogen-related enzymes in rhizosphere soil(P<0.05),GN treatment decreased enzyme activities of NR,nitrite reductase(NiR),GS,GOGAT and glutamate dehydrogenase(GDH),but increased enzyme activity of urease(UE),GP treatment significantly increased enzyme activity of NR,NiR and GS(P<0.05),increased by 40.01 26.52 and 30.42%every year on average,respectively,but decreased enzyme activities of GOGAT,GDH and UE.Correlation analysis showed that enzyme activity of NR was positively correlated with enzyme activities of GS and GDH in rhizosphere soil,and enzyme activity of GS was significantly positively correlated with enzyme activity of GOGAT(P<0.05).② ZN and ZP treatments significantly increased the Sobs and Chao1 indexes of bacteria(P<0.05),and had a great influence on the relative abundance of phylum and genus levels.Correlation analysis showed that dominant phylum of bacteria was positively affected by AN,XN and TN content in N addition,dominant phylum of bacteria was positively affected by pH、AP、TP and TS content in P addition,and dominant phylum of bacteria both was positively affected by enzyme activities of NiR and GOGAT in N and P addition.In general,proper N and P addition improved the activities of nitrogen-related enzymes,changing bacterial diversity and community structure,and promoted the uptake and utilization of nutrients by roots.(5)Different N and P additions had different effects on the diversity and community structure of nitrifying bacteria in rhizosphere soil.① GN treatment significantly increased the Shannon index of ammonia oxidizing archaea(AOA),ZP treatment significantly increased Sobs,Shannon and Chaol indexes(P<0.05),were 65.67,2.66 and 71.25,respectively,GN and ZP treatments had a significant and sensitive effect on the community structure of AOA at phylum and genus levels,dominant phylum of AOA was positively affected by pH,AN,XN,AP,TP content in N addition(P<0.05).② In general,GN and ZP treatments significantly increased the Alpha diversity of ammonia oxidizing bacteria(AOB),and P addition had a greater effect on AOB than N addition,The effects of P addition on dominant phylum of AOB were greater than that of N addition,and ZN and GP treatments had greater effects on the dominant genus of AOB.dominant phylum of AOB was positively affected by XN content in N addition,while dominant phylum of AOB was significantly affected by pH and AP content in P addition(P<0.05).the N and P addition increased nitrification of rhizosphere soil and promoted the nitrification capacity of nitrogen.(6)N and P addition significantly affected the Alpha diversity and community structure of nitrite oxidizing bacteria(NOB).ZN treatment significantly increased Sobs and Chao1 indexes of NOB species(P<0.05),were 109.67 and 137.32,respectively,GP treatment significantly decreased Chao1 index(P<0.05).N and P addition had little effect on the relative abundance of dominant Nitrospirae and Nitrospira.dominant phylum of NOB was positively affected by TN,AN and XN content in N addition,dominant phylum of was positively affected by AP content and pH in P addition(P<0.05).In general,the N and P addition increased nitrite oxidation and XN content in rhizosphere soil.(7)N and P addition changed diversity and community structure of nitrogen-fixing bacteria in rhizosphere soil to a certain extent.ZN and ZP treatments both significantly increased Sobs and Chaol indexes(P<0.05),GN and GP treatments were the opposite,and GN treatment reached significant level(P<0.05),N and P addition had little effect on the relative abundance of nitrogen-fixing bacteria at phylum,but had a great effect on the genus,the dominant phylum is Proteobacteria,the dominant and influential genus are Proteobacteria,unclassified_p_Proteobacteria,Skermanella,unclassified_c_Deltaproteobacteria and Azoarcus,domina nt phylum of nitrogen-fixing bacteria was positively affected by TN content in N addition,dominant phylum of nitrogen-fixing bacteria was positively affected by AP TP XN content and pH in P addition(P<0.05).In general,the N and P addition significantly affected the diversity and microbial community structure of nitrogen-fixing bacteria in rhizosphere soil,and promoted the fixation of nitrogen.In conclusion,the 6kg/667m2 of N and and 9kg/667m2 of P improved the microenvironment of alfalfa’s rhizosphere soil that Chemical properties,enzyme activity and bacterial make in alkaline soil,optimize the diversity and community structure of AOA,AOB,NOB and nitrogen-fixing bacteria,improve the leaf key enzymes and products of nitrogen metabolism of alfalfa,enhance the nitrogen cycle efficiency,and improve the protein and yield of alfalfa,providing theoretical basis and technical guidance for scientific fertilization for production of high yield and high quality of alfalfa in moderately alkaline soil.