| Ephedra sinica Stapf(E.sinica)is widely used in the treatment of diseases due to its rich and diverse pharmacological effects.The roots and stems of E.sinica have the same origin but different effects,and the quality of E.sinica is different in different years.In recent years,it has been discovered that endophytes are the important factor affecting plant quality.They co-exist with host plants and exhibit dynamic changes in different years.During the process of co-evolution and growth with plants,they potentially affect the physiological and metabolic processes of host plants,thereby affecting the quality of host plants.In this study,the composition of endophytic bacteria population,dynamic succession characteristics and changes in metabolite formation in E.sinica in different ages(8 years,13 years,18 years,23 years,28 years)were studied.The results are as follows:1.Different DNA barcodes(ITS2,trn H-psb A,rbc L,rpo C1)were used for species identification and phylogenetic analysis of E.sinica in different years.The results showed that the four kinds of DNA barcodes could identify the species as E.sinica.Phylogenetic analysis showed that rpo C1 has the finest distinction on different years of E.sinica,providing ideas for subsequent researches on changes of endophytic bacteria and metabolites in different years.2.Using nuclear magnetic resonance technology to analyze the changes of metabolites in the roots and stems of E.sinica in different years,combined with T-test and multivariate statistical analysis,it was found that there are 38 different marker metabolites in the roots and stems,and there were 16 unique compounds in the roots,including two alkaloids,two flavonoids,seven tannins,two organic acids,one volatile oil,amino acid and sugar.Also,there were 11 unique compounds in the stems,including six alkaloids,two flavonoids,one urea,organic acid and sugar.In addition,there were 11 compounds in common in roots and stems,including eight amino acids,one volatile oil and two sugars.These compounds change dynamically during the cultivation of E.sinica,and they were attached to 14 metabolic pathways.3.The metagenomics method was used to analyze the structural distribution characteristics of endophytes in the roots and stems of E.sinica in different years.It was found that the distribution of endophytes in E.sinica in different years were different,and there were different dominant endophytes in each medicinal part of each age.The abundance of endophytes in the roots and stems of E.sinica in different ages was also different.The abundance of endophytes in the roots of different ages was 1112,1582,968,840,966,and the abundance of stems was 15,16,29,22,230.In addition,there were significant positive and negative correlations among endophytes,in which the interactive complex network relationship was formed.4.The dynamic changes of endophytes in the top 50 of E.sinica roots and stems in different years show that there were no significant changes in E.sinica stems;but significant and irregular changes in the abundance of endophytes in roots were existed at class levels(2),order levels(8),family levels(12),and genus levels(12),respectively.Correlation analysis between dominant bacteria and differential compounds in E.sinica roots of different years shows that 40 dominant bacteria in roots are mostly positively correlated with 16 differential marker metabolites,while 14 dominant bacteria in stems are mostly negatively correlated with the formation of 11 differential metabolites.In this paper,the relationship between the ecological succession of endophytes and metabolite accumulation of E.sinica in different years was studied,and the effect of endophytes on E.sinica in different years was systematically revealed.The results will provide a scientific basis in evaluating and monitoring the quality of E.sinica in the process of cultivation,and provide the scientific references for the use of species interactions to improve the quality of E.sinica. |
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