The Change In Microflora And Screening Of Key Microorganisms In Wild R. Glutinosa Rhizosphere Under Consecutive Monoculture

Consecutive monoculture problem is a common issue in the cultivation of medicinal plants.There is also serious soil disease in the production of Rehmannia glutinosa,and previous experiments found that both cultivated and wild R.glutinosa represented typical consecutive monoculture problems,such as abnormal plant growth,unable enlargement of root tubers,severe diseases and insect pests,etc.In this study,wild R.glutinosa were artificially transplanted into field and consecutively monocultured.The rhizospheric soils sampled from the four different treatments,including the newly planted,the two-year monocultured,the wild R.glutinosa and the control,were used to study the shifts of the soil microflora by T-RFLP analysis.It showed that the bacterial community structure was significantly different among the wild R.glutinosa rhizospheric soils with the field soils monocultured for different years.The bacteria belong to Proteobacteria and Firmicutes were predominent in the wild and newly planted R.glutinosa soils.Furthermore,some beneficial biocontrol bacteria(such as Bacillus,Pseudomonas,etc)gathered in the rhizosphere of the wild and newly planted R.glutinosa,however a large number of pathogenic bacteria bred in the rhizosphere of the two-year monocultured R.glutinosa,such as Clostridium,Flexibacter polymorphus,Clostridium ghoni,etc.which might go anainst the colonization of beneficial bacteria and cellulose degradation bacteria.Based on this,the screening and quatification of antagonistic bacteria were performed in this study.Two effective antagonistic bacteria,Bacillus cereus and Pseudomonas TL,were screened and identified in this work.It was indicated that these two strains mentioned above had obvious antagonistic effects on R.glutinosa pathogen(Fusarium oxysporum).Meanwhile,the pathogenicity of F.oxysporum and the antagonistic effects of antagonistic bacteria were evaluated by using tissue culture seedlings of R.glutinosa.The results showed that the tissue culture seedlings of R.glutinosa treated by F.oxysporum soon appeared disease symptom,withered and died finally.However,in experiment I using Bacillus cereus to inhibit F.oxysporum,the disease symptom of tissue culture seedlings came slowly,but the seedlings withered and ultimately died.In experiment ? using Pseudomonas TL to inhibit F.oxysporum,the transplanted tissue culture seedling of R.glutinosa grew well during the whole experiment period.For further study,qRT-PCR techinque was applied to detect the contents of F.oxysporum and antagonistic bacteria.It displayed that the content of F.oxysporum was the lowest in experiment ?(treated by Pseudomonas TL),then in.experiment ?(treated by Bacillus cereus),the highest in control(without antagonistic bacteria).While the contents of antagonistic bacteria in experiment ? and ? were higher than that in control,which were consistent with the results of field trial.Namely,the content of F.oxysporum in the two-year monoculture soil was higher than those in the newly planted and wild R.glutinosa soils.However,the content of antagonistic bacteria in two-year monoculture soil was lower than those in the newly planted and wild R.glutinosa soils.In conclusion,the pathogenic microbes bred seriously in the rhizospheric soil of wild R.glutinosa under the monoculture regime,and yet the growth of beneficial bacteria was restrained,which led to the serious soil-born disease and abnormal growth of R.glutinosa.However,exogenously applied antagonistic bacteria could prevent R.glutinosa from the infection of pathogenic F.oxysporum to some degree,implying the biocontrol effects of these two antagonistic bacteria on R.glutinosa root rot pathogen.