Elevated CO₂ Alleviates The Effects Of Lead On The Root Transcriptome And Rhizosphere Bacterial Communities Of Trifolium Repens L.

Various environmental problems caused by elevated atmospheric CO₂concentration and lead pollution in soil have become great challenges faced for contemporary society,affectings the plant ecosystems.When soil is contaminated with lead,its ecological processes and environmental risks will inevitably be affected by the increased atmospheric CO₂concentration.Currently,there are large uncertainties and even some conflicting phenomena regarding the impacts of elevated atmospheric CO₂concentration on heavy metal uptake by plants.This is not only related to the type of heavy metals,but also to the physiological response characteristics of plants and the critical rhizospheric processes regulated by microbes.T.repens.is a heavy metal lead tolerant plant that can adapt widely to a variety of stress situations and is a good plant resource for heavy metal soil remediation.In this study,T.repens was used as the test plant,and the toxic effects of different degrees of lead pollution on T.repens and the effects of heavy metal enrichment characteristics were systematically investigated,and the gene regulation mechanism and rhizosphere bacterial community changes of T.repens under stress environment were further analyzed by transcriptomics and16S r RNA amplicon.In order to provide scientific support for the application of CO₂gas fertilizer in enhnced phytoremediation technology,it will also serve as a reference for a comprehensive assessment of soil pollution risk.The main results are as follows:（1）The effects of high concentration CO₂on root length,plant height,biomass,root morphology,photosynthetic parameters,ROS level and lead enrichment of T.repens were investigated by pot experiment and hydroponic experiment.The results showed that under different Pb stresses（CK,PL,PM）,high concentration of CO₂（800 ppm）could significantly increase the fresh weight and plant height of T.repens In hydroponic system,under different Pb stresses（CK,PL,PM）,high concentration of CO₂（800 ppm）can significantly increase the root length,plant height,biomass and lead content of T.repens,promote the lateral root growth and photosynthesis of T.repens,and reduce the oxidative damage of plants.Compared with conventional CO₂（460 ppm）,the root length and plant height of T.repens with high CO₂concentration increased by 22.57%～32.33%and 22.21%～27.93%respectively.Biomass（aboveground and underground）increased by 216.53%～382.26%,164.71%～465.57%,lead content（aboveground and underground）increased by11.07%～34.62%,43.9%～53.53%,and net photosynthetic rate increased by 24.65%～63.53%.The detection and analysis of ROS levels in leaves and roots showed that the accumulation levels of H₂O₂and superoxide anion O₂^-in T.repens under lead stress were decreased under high concentration of CO₂.The above results suggest that a high concentration of CO₂can alleviate the damage of Pb stress to T.repens to some extent,and may improve the stress resistance and enrichment ability of T.repens.（2）The molecular response mechanism of root system of T.repens under stress environment was studied by differential gene expression analysis,GO enrichment analysis and KEGG enrichment analysis.The results showed that,compared with lead stress alone,the DEGs induced by combined high concentration CO₂and Pb stress were less,and it mainly involved in the expression of related genes such as translation,peptide biosynthesis and peptide metabolism by GO analysis.In KEGG enrichment analysis,compared with single high concentration CO₂,lead stress under high concentration CO₂treatment can induce more genes related to ribosome and phenylpropanoid biosynthesis.The results of RNA-seq were verified by q RT-PCR analysis of the screened genes,and it was found that high concentration CO₂treatment may affect the metabolic signal transduction and secondary metabolite synthesis of T.repens under lead stress.（3）16S rRNA amplicon was used to compare the changes of rhizosphere bacterial community structure of T.repens in lead contaminated soil under different CO₂concentrations.The results showed that under all Pb stress concentrations,elevated concentration of CO₂decreased the p H of soil and increased the activities of urease,sucrase and acid phosphatase,but had no significant effect on theα-diversity of soil bacterial community.Pb stress concentration is the biggest source affecting the variation of bacterial community structure,followed by different CO₂concentrations.The results of Welch’s t-test difference analysis of different treatments at the genus level showed that Pb stress significantly increased the relative abundance of Pseudoxanthomonas in the rhizosphere soil of T.repens.The relative abundance of Bosea and unclassified＿Xanthomonadales increased significantly under the treatment of high concentration of CO₂.FAPROTAX analysis showed that T.repens rhizosphere microbes were mainly characterized by chemotaxis and heterotrophy,with the relative abundance of iron respiration and methane nutrition increased under high concentration of CO₂.