Application Of ZnO NPs On The Effect Of Legumes On Nitrogen Fixation And Soil Microbial Community

While nanoparticles are widely used in industrial and agricultural production,they will undoubtedly bring certain risks to the ecological environment.Plants are a basic part of the ecosystem,and studying the potential effects of ZnO NPs on plants is an important aspect of its environmental risk assessment.In this study,germination experiments and pot experiments were used to explore the effects of nano zinc oxide(ZnO NPs)at different concentrations(10?50?100?200?400?800 mg L^-1)on the seed germination,seedling growth and zinc content of edible and feed legumes peas,mung beans,alfalfa,and white clover,and the effects of different ZnO NPs treatments(ZnO NPs:10?50?250?750 mg kg^-1;Zn SO₄:50 mg kg^-1)on the growth,potential nitrogen fixation of alfalfa and soil microorganisms.The main findings are as follows:(1)There were no significant difference in the germination rates of the four legumes with ZnO NP concentrations(10,50,100,200,400,800 mg L^-1)compared with the control.The biomasses of pea and mung bean seedlings increased first and then decreased with the increase in ZnO NP concentrations,while alfalfa and white clover tended to decrease.Pea,mung bean,alfalfa,and white clover seedlings showed different sensitivities to ZnO NPs;the dose-response thresholds with root length were 200,100,50,and 50 mg L^-1,respectively.The four seedlings root length decreased with the increase of ZnO NPs concentration when exceeding these thresholds.The root growth inhibition rates with the800 mg L^-1 ZnO NP treatment for pea,mung bean,alfalfa,and white clover were 68%,75%,83%,and 85%,respectively,indicating phytotoxicity.The malondialdehyde(MDA)content in the seedlings was also determined.The stress effect of high concentration(100?800 mg L^-1)ZnO NPs was stronger for pea and white clove seedlings than on mung bean and alfalfa.Zinc contents in pea,mung bean,alfalfa,and white clover seedlings increased from 9.17 mg kg^-1,12.04 mg kg^-1,8.98 mg kg^-1,and 17.84 mg kg^-1 with the control,to 83.96 mg kg^-1,82.96 mg kg^-1,212.48 mg kg^-1,and 263.21 mg kg^-1with the highest ZnO NPs treatments,respectively.In summary,the sensitivity of different types of legumes to ZnO NPs differed in the order of white clover,alfalfa,mung bean,and pea from high to low.(2)In pot experiments,S50 treatment was more beneficial to the accumulation of alfalfa plant biomass,root length and plant height elongation than N50 treatment.High-concentration ZnO NPs(250?750 mg kg^-1)treatments all reduced the biomass,root length and plant height of alfalfa but the inhibitory effect is not significant.Consistent with the change in biomass,S50 treatment significantly increased the chlorophyll content of alfalfa plant leaves,While the chlorophyll content of plant leaves treated with N250 and N750 was significantly lower than control.The zinc content of alfalfa roots,stems leaves and soil available zinc content all increased with the increase of ZnO NPs concentration,and there was no significant difference in the zinc content between N50 and S50 treatments.the transfer efficiency from root to stem is lower than that from stem to leaf under two zinc species treatments.N750 treatment significantly reduced the number of alfalfa nodules,and there were ineffective nitrogen fixation areas in the nodules.and ZnO NPs treatment would break and bend the nodule cell wall,and reduce the number of bacteria in the root nodule cells,thereby reducing the nitrogen fixation efficiency of the plant.(3)Observing the distribution of nanoparticles in the nodule tissue,N50 treatment,there are more nanoparticles distributed in the nodule cells,and the aggregate particle size is small;N750 treatment,there are less nanoparticles distributed in the nodule cells,but the aggregate size is larger.Similar to the distribution of nanoparticles in nodules,in the root tissue of alfalfa,N50 and N750 treatments have visible nanoparticles,but the phenomenon is not observed in nodules treated with S50.(4)N750 treatment significantly reduced the abundance and diversity of soil bacterial communities for alfalfa growth.N750 treatment increased the relative abundance of Proteobacteria by 39%,but it reduces the relative abundance of the phylum Chloroflexi,which is 57%less than the control.The composition of the bacterial community sample treated with N750 was quite different from that of the control treatment,which changed the bacterial community structure.Changes in soil bacterial community structure are mainly driven by soil available zinc content(DTPA-Zn).Under the same treatment,the abundance of soil fungi grown by alfalfa was lower than that of bacteria,N50 treatment reduced the abundance of soil fungi,N750 treatment increased the abundance of soil fungi.N50treatment reduced the relative abundance of the Basidiomycota.Compared with the control,it was reduced by 61%,N750 and S50 treatments changed the composition of the fungal community,Soil physical and chemical properties p H,SOM,TN,NH₄⁺-N,NO₃^—N and DTPA-Zn cannot explain changes in soil fungal community structure,and changes in fungal community structure may be affected by other factors.In summary,the sensitivity of different types of legumes to ZnO NPs differed in the order of white clover,alfalfa,mung bean,and pea from high to low.The rules of ZnO NPs entering plant tissues were closely related to the aggregation of particles.ZnO NPs reduces the nitrogen fixation efficiency of alfalfa by reducing the number of bacteria in nodules,and changed the soil microbial community structure.