Effects Of Traffic Volume On Root Exudate And Rhizosphere Microbes Of Ligularia Virgaurea In Alpine Meadow

As human activity increases,road traffic is becoming an important driver of natural environmental change,particularly on plant communities and soils on the roadside.Plant root exudates and rhizosphere microorganisms are sensitive indicators to the impact of road traffic.Currently,the alpine meadow ecosystem is also severely affected by road traffic on the Qinghai-Tibetan Plateau.To clarify this impact,we collected rhizosphere and non-rhizosphere soil samples from 11 experimental sites on the roadside with high（>150 vehicles/hour）,medium（50-150 vehicles/hour）and low（<50 vehicles/hour）traffic volumes in Maqin County,Golog Tibetan Autonomous Prefecture,Qinghai Province.We selected Ligularia virgaurea,a typical poisonous herb in alpine meadows,as the study target.For the soil samples analyzing,7 soil physicochemical properties and 12 metal elements were determined.Soil bacterial community structure was determined by high-throughput sequencing.The components and relative contents of Ligularia virgaurea root exudates were determined by Gaschromatography-mass spectrometry（GC-MS）.The effects of road traffic on Ligularia virgaurea root exudates and soil microorganisms were analyzed by correlation analysis and construction of the bacterial molecular ecological network model.The main results of the study are as following points:（1）Traffic volume significantly affected the structure of the plant communities on the roadside.The dominant species of the grassland on the roadside under different traffic volume disturbances were Ligularia virgaurea、Knorringia sibirica、Potentilla.However,the effect of traffic volume on the plant speciesα-diversity was not significantly.（2）Traffic volume significantly affected soil metal elements content（p<0.05）.The highest contents of Ni(30.2 mg.Kg^-1),Cu(20.8 mg.Kg^-1)and Co(11.1 mg.Kg^-1)were found in the high-traffic volume sample sites.Pb(45.7 mg.Kg^-1)and Cd(3.5mg.Kg^-1)contents were significantly higher in the non-rhizosphere soils than in the rhizosphere soils.In contrast,As(18.6 mg.Kg^-1)and Cr(51.7 mg.Kg^-1)contents were significantly lower（p<0.05）in the non-rhizosphere soils than in the rhizosphere soils.（3）Traffic volume significantly influenced the components of root exudates as well as their relative content（p<0.05）.The root exudates of Ligularia virgaurea consisted of lipids and lipid-like molecules,Benzenoids,and Organic cyanides compounds.Compared to the medium and low traffic volume sample sites,Ligularia virgaurea’s organic acids in high traffic volume sample sites were 1.2-5.0 times higher.Meanwhile,Citramalic acid was 15 times higher,and the relative content of pentadecanoic acid,vanillic acid,stearic acid and palmitic acid was also significantly higher in high traffic volume sample sites too.These root exudate’s relative content were significantly and positively correlated with soil total carbon（TC）,total nitrogen（TN）,total phosphorus（TP）,total organic carbon（TOC）,N:P and C:P（p<0.05）.（4）Traffic volume significantly influenced the composition and diversity of the soil bacterial community.The dominant phylums were Proteobacteria（38.27%）,Acidobacteria（14.62%）and Planctomycetes（11.08%）.The bacterial community composition of the high-traffic volume sample sites was significantly with the medium and low-traffic volume sample sites（p<0.05）.The bacterialα-diversity was lowest in the high-traffic volume sample sites（p<0.05）,while theβ-diversity tended to decrease and then increase when increasing road traffic volume.（5）Bacterial richness andβ-diversity were significantly higher in the non-rhizosphere soil than in the rhizosphere soil.Bacterial communities of non-rhizosphere soil were positively correlated with TC,TN,TP,TOC,C:N,N:P,C:P,NO₃-N,NH₄-N,pH,bulk density and water content（p<0.05）,and not significantly correlated with metal elements.In contrast,the rhizosphere soil bacterial community was negatively correlated with soil metal elements（p<0.05）.The structural stability of the bacterial community tended to decrease with increasing road traffic volume,and the dominant bacterial phylum changed from Actinobacteria to Bacteroidetes.（6）The results of the molecular ecological network analysis showed that the number of nodes of the dominant soil bacterial community increased and then decreased with increasing road traffic volume.The number of nodes were lowest in the high-traffic sample（121）and highest in the medium-traffic sample（147）.In the dominant bacterial community network,the degree of association between species nodes gradually increased.In contrast,in the differential bacterial community network,the number of species nodes decreased with increasing road traffic volume,and the degree of association between species nodes increased and then decreased.The dominant phylums in the important nodes of the network were Acidobacteria,Proteobacteria,Bacteroidetes and Actinobacteria.（7）Pb was an important indicator affecting key nodes of soil bacterial community,key nodes bacterial genera such as Edaphobacter,Sphingomonas,Phyllobacterium,Nocardioides,Microvirga,RB41,Rhizorhapis,MND1 were significantly negatively correlated with Pb（p<0.05）.Among the root exudates compounds,small molecule organic acids are important components involved in the interaction between soil bacterial communities and soil properties.Organic acids such as Citramalic acid,4-hydroxybutyric acid,3,4-hydroxybutyric acid,Hydroxypropionic acid and glyceric acid were significantly negatively correlated with key nodes of soil bacterial community（p<0.05）.In summary,traffic volume did not affect plant species diversity on the roadside in this study,but traffic volume could significantly affect soil heavy metal elements content and bacterial community diversity.Traffic volume effects were related to the relative content of Ligularia virgaurea’s root exudates,organic acids.The root exudates relative content of Ligularia virgaurea,such as organic acids was closely related to road traffic volume.Rhizosphere and non-rhizosphere soil microorganisms were sensitive to changes in traffic volume.There was a negative correlation between rhizosphere soil microorganisms of Ligularia virgaurea and soil metal elements.Thus,root exudates and the rhizosphere soil bacterial community are key to the stability of the physiological and ecological functions of Ligularia virgaurea in the condition of metal accumulation caused by traffic.