Study On The Separation And Identification Of Antimicrobial Active Substances From Streptomyces Sp.CX3 And Its Induced Resistance

Blueberry canker is a fungal disease caused by Botryosphaeria dothidea,resulting in a large yield reduction of blueberry fruit.In this study,B.dothidea was used as the target bacteria,and the antagonistic strain Streptomyces sp.CX3,which was independently screened by our laboratory,was used as the research carrier.Field experiments were carried out to evaluate its control effect,and the whole genome sequencing of Streptomyces sp.CX3 was performed to analyze the function.Then,preliminary isolation of antimicrobial active substances from Streptomyces sp.CX3 was also conducted,and the changes in the substance content and enzyme activity of phenylpropane metabolism and salicylic acid were studied to explore whether the fermentation broth can regulate the host plant to resist the invasion of blueberry canker bacteria.The research results are as follows:1.Whole genome sequencing of Streptomyces sp.CX3 based on the Illum ina platform showed that the genome was 8516989 bp after assembly,with 7504 coding genes,and GC content of 72.73%.The annotation results in COG and KEGG database showed that genes related to biosynthesis of antibiotics and biosynthesis of secondary metabolites were-largely enriched.The anti SMASH was used to predict the gene clusters of secondary metabolites in Streptomyces sp.CX3 genome,and the results indicated that 32 gene clusters was found,which were involved in the synthesis of bacteriocins,terpenes,butyrolactones,lanthiopeptides and siderophores.Among them,the compounds Dutomycin,Ficellomycin,Kirromycin,Lactonamycin and Nocathiacin all have antibacterial effects,and ECO-02301 has strong antifungal activity.2.Due to the diversity of the secondary metabolite gene clusters in Streptomyces sp.CX3,the physio chemical properties and preliminary separation and identification of the active substances in the Streptomyces sp.CX3 fermentation broth were investigated.The results showed that Streptomyces sp.CX3 fermentation broth had strong UV stability,good thermal stability,alkali resistance but not acid resistance.The results of paper chromatography showed that at least two active components were present in Streptomyces sp.CX3 fermentation broth,one of which might be water-soluble type I antibiotic.The results of Betina solvent system experiment showed that the active ingredient was insoluble in ethyl acetate and benzene,and had moderate polarity.The results of paper electrophoresis experiments showed that one of the active components was neutral and the other was weak acid.3.The artificial inoculation method was used to carry out the field control experiment of the fermentation broth of Streptomyces sp.CX3.The results showed that the fermentation broth has a good protective effect,reaching 53.7%,which is significantly better than the two commonly used chemical pesticides tested.In view of this,we determined whether Streptomyces sp.CX3 fermentation broth can induce host disease resistance.The results showed that Streptomyces sp.CX3 fermentation broth significantly increased the activities of PAL and CHI in blueberry leaves,and significantly increased the content of antibacterial substances such as total phenols,flavonoids,lignin,proanthocyanidins,salicylic acid,etc.,activate the phenylpropane metabolic pathway and the salicylic acid pathway in the host,thereby improving the host's disease resistance.In conclusion,the fermentation broth of Streptomyces sp.CX3 are relatively stable,and has a good biocontrol effect on blueberry canker in field.Meanwhile,the fermentation broth can stimulate the phenylpropane metabolic pathway and the salicylic acid pathway to induce the disease resistance of blueberry plants indicating the promising development value and application prospects of Streptomyces sp.CX3.