Isolation And Identification Of Endophytic Bacteria Of Panax Ginseng And Evaluation Of Antibacterial Effect On Ginseng Rhizoctonia Solani And Pythium Debaryanum

In this paper,the endophytic bacterium Bacillus velezensis LXS-N2,which has inhibitory effect on the ginseng pathogenic fungus Rhizoctonia solani and the ginseng pathogenic fungus Pythium debaryanum,was isolated from the 15-year-old ginseng.The physicochemical characteristics of Bacillus velezensis LXS-N2 were studied and the culture conditions were optimized,and the functional characteristics,colonization ability and antibacterial mechanism of Bacillus velezensis LXS-N2 were investigated.In order to provide theoretical reference for the development of germplasm resources of ginseng endophytic bacteria,the commercialization of Bacillus velezensis LXS-N2 and the control of two kinds of ginseng diseases.The main study results are presented as follows:1、 In Fusong County,Jilin Province,twelve 15-year-old Panax ginseng were collected,endophytic bacteria that have inhibitory effects on the pathogen of Rhizoctonia solani and Pythium debaryanum are isolated and screened,obtain strains LXS-N2.Combining morphological observation,physiological and biochemical reaction and molecular biological methods,the endophytic bacteria LXS-N2 was identified as Bacillus velezensis.2、 The fermentation medium components and fermentation conditions of Bacillus velezensis LXS-N2 were optimized by single factor test,and the proportion of the medium components was determined by orthogonal test.The results showed that the influence level of each factor was inorganic salt>carbon source>nitrogen source,and the optimal ratio should be yeast extract powder 9g/L,peptone 9g/L,and magnesium sulfate 6g/L.Under this medium,the p H,temperature,inoculation amount and rotation speed of the fermentation medium of strain LXS-N2 were optimized.The optimal fermentation conditions were p H 6,temperature 30 ℃,and inoculation amount 5%,the rotation rate is 180r/min.The biomass of the optimized strain LXS-N2 fermentation broth was improved,the bacteriostatic rate to Rhizoctonia solani of ginseng was increased by about 25.56%,and the bacteriostatic rate to Pythium debaryanum of ginseng was increased by about 20.79%.3、 The functional characteristics and colonization ability of Bacillus velezensis LXS-N2 were preliminarily explored.The results showed that,(1)the strain LXS-N2 has a good inhibitory effect on 8 kinds of ginseng pathogenic bacteria,such as A:Pythium debaryanum,Rhizoctonia solani,Cylindrocarpon destructans,Colletotrichun panacicola,Botrytis cinerea,Alternaria panax Whetz,Fusarium solani,Fusarium oxysporum.The overall inhibitory rate is between 50% and 80%,with antifungal broad-spectrum;(2)The strain LXS-N2 can produce protease and cellulase,and has the ability of dissolving potassium,dissolving phosphorus,producing IAA.(3)After antibiotic labeling,the colonization rate of strain LXS-N2 in the root of ginseng was detected.The results showed that as a dominant group of bacteria symbiotic with plants,Bacillus velezensis LXS-N2 could colonize in ginseng very well,and the colonization rule in the root soil of ginseng showed a down-up-down-maintaining stability;the rule of colonization in the root of Panax ginseng showed ascending-descending-maintaining stability.4、 The bacteriostasis mechanism and control effect of Bacillus velezensis LXS-N2 were further discussed.The results showed that,(1)The fermentation liquid of strain LXS-N2 can inhibit the mycelial growth of Rhizoctonia solani and Pythium debaryanum,which cause the mycelial branches of Rhizoctonia solani to increase,thin and break;the hyphae of Pythium debaryanum were uneven in thickness,the nodules were swollen,and the cell wall became thinner.(2)The bacteriostatic rate of strain LXS-N2 was the highest after cultivation for 22 h.The degree of lipid peroxidation in the mycelium of two pathogenic bacteria increased significantly with the increase of LXS-N2 culture time.At 22 h,the content of MDA(malondialdehyde)reached the maximum value,and then decreased,and the bacteriostatic rate also decreased.(3)The bacteriostatic rate of the fermentation broth,sterile filtrate and mycelium of strain LXS-N2 was tested,and it was found that bacteriostatic rate of the mycelium and the fermentation broth was good.After the cell fragmentation of the mycelium,it was found that it still had good bacteriostatic activity against Rhizoctonia solani and Pythium debaryanum.(4)In Ex vitro experiment of ginseng root,the fermentation broth of strain LXS-N2 showed good control effect.