Study On The Dynamics Of Rhizosphere Microbial Community In Different Development Stages Of The Chinese Medicinal Herb "Beishashen"

“Beishashen” is the swollen root of Glehnia Littoralis Fr.Schmidt ex Miq.in the genus Glehnia of the Apiaceae family.G.littoralis is a traditional Chinese herb with very important medicinal value and a long history of cultivation.Currently,it’s wild resources are in an endangered state,and the G.littoralis herbs on the market mainly come from artificial cultivation,but the long-term cultivation in fixed areas has led to a decrease in the yield and quality of the herbs.Rhizosphere microorganisms play an important role in influencing plant quality and productivity through their interaction with the plant root system.Understanding the community composition,structural function,and dynamic changes of rhizosphere microorganisms during the cultivation of G.littoralis is important for using them to improve plant resistance,disease resistance,and yield and quality,and may provide an idea to solve the bottleneck problem of declining yield and quality of G.littoralis.In this study,we selected rhizosphere soils of G.littoralis at different developmental stages(seedling stage,vigorous growth stage and harvest stage)in two main cultivation areas(Laiyang,Shandong and Chifeng,Inner Mongolia)as the research objects,and systematically investigated the rhizosphere bacterial and environmental factors of G.littoralis by examining the physicochemical properties of rhizosphere soils of G.littoralis,combined with the high-throughput sequencing analysis of rhizosphere microorganisms and the symbiotic network analysis of microorganisms and environmental factors.The community composition,diversity,structure,function of rhizosphere microorganisms and correlation between rhizosphere microorganisms and environmental factors at different developmental stages were systematically investigated.The main findings were as follows:(1)All soil samples were sequenced to obtain abundant bacterial operational taxonomic units(OTUs),1885 OTUs belonging to 25 phyla and 479 genera were obtained from Laiyang in Shandong,and 1913 bacterial OTUs belonging to 25 phyla and 470 genera were obtained from Chifeng in Inner Mongolia.The main dominant phyla were Proteobacteria,Acidobacteriota and Actinobacteriota.The relative abundance of Proteobacteria increased significantly from the seedling stage to the vigorous growth stage,but remained basically stable from the vigorous growth stage to the harvest stage.For the analysis of fungal OTUs in the two regions,we learned that there were 903 OTUs belonging to 10 phyla and 312 genera in Laiyang,Shandong Province,and 1093 OTUs belonging to 11 phyla and 320 genera in Chifeng,Inner Mongolia,all of which indicated abundant OTUs.Ascomycota,Basidiomycota,Peridiomycota were the dominant phylum.Ascomycota is an important pathogenic phylum in the fungal kingdom,and Ascomycota was significantly elevated during the vigorous stage with the development of “Bei Sha Shen” in both regions,which may indicate that the invasion of pathogenic fungi occurred during the vigorous growth stage.(2)The microbial diversity of the rhizosphere exhibited changes with the growth and development of G.littoralis.Bacterial diversity in Laiyang,Shandong showed a gradual decline,whereas bacterial diversity in Chifeng,Inner Mongolia showed a decline followed by a rise,and fungal diversity at both locations showed a decline followed by an increase.It was concluded that the diversity of the different microbial communities in the two regions decreased during the vigorous growth stage,and was generally accompanied by a decline in the richness index that was similar to previous studies indicating that secretion of G.littoralis at the vigorous growth stage may have exhibited an effect of selection on the microbial community,and soil type shifted from a bacterialdominant to a fungal-dominant state.(3)Through annotation analysis and comparison of soil rhizosphere bacterial sequences,significant differences were observed between seedling stage and other developmental stages through functional difference analysis and principal component analysis.The relative abundance of "environmental adaptation" functions at harvest stage was significantly higher than that at seedling stage,and this function is mainly related to "plant-pathogen interactions",suggesting that bacterial communities may be involved in disease resistance at harvest stage.For fungal communities,functional differentiation also occurs in early and late developmental stages.The abundance of saprophytic,symbiotic and pathogenic saprophytic symbiotic compound fungi increased gradually with the growth and development of G.littoralis.These fungi may speed up the decomposition of soil organic matter and convert insoluble minerals into nutrients available to G.littoralis.(4)The correlation analysis between rhizosphere microorganisms and soil physicochemical properties showed that eight key environmental factors(p H,organic carbon,available phosphorus,sucrase,nitrate nitrogen,ammonium nitrogen,available potassium and urease)may drive the change of microflora at different developmental stages and provide a list of bacteria and fungi at different developmental stages.In addition,symbiotic network analysis also shows that there is a complex network relationship between rhizosphere microorganisms and soil physicochemical factors.Understanding the influence of various environmental factors on rhizosphere microbial community can help us to construct beneficial microbial community by regulating environmental factors,promote the healthy growth of plants and improve the yield of medicinal materials.(5)Based on the root microbiome data of the two regions,biomarkers were identified by random forest method.47 bacterial biomarkers and 22 fungal biomarkers were selected in Laiyang of Shandong Province.Twenty-one bacterial biomarkers and 34 fungal biomarkers were selected from Chifeng in Inner MongoliaCombined with the results of the microbial community composition,diversity analysis and community function,it can be inferred that G.littoralis exhibited some selectivity towards the microbial community at the growth and development stages.Understanding the enrichment preference of microorganisms during the growth and development of G.littoralis will be beneficial for improving the quality and yield of G.littoralis and provide data-based support for the field management and optimal cultivation of Chinese medicinal materials.