The Effect And Mechanism Of Biocontrol Strain GZ-34 Against Ralstonia Solanacearum

Ralstonia solanacearum infected vascular bundle of the host plants and resulted in the occurrence of bacterial wilt,which caused the death of the plant,commonly known as "plant blast".It is an important plant disease worldwide in tropical and subtropical regions.Ralstonia solanacearum can invade several hundreds plants species of 54 families.So far,there is no effective agricultural measures,physical measures or chemical measures to prevent bacterial wilt.Therefore,biological prevention and control has attracted more and more attention.This study is divided into three parts: 1?EGFP tagging Ralstonia solanacearum GMI1000 with PBBR1-2.2?Screening and identification of antagonistic bacteria against Ralstonia solanacearum and greenhouse inoculation experiment.3?The extraction,isolation,identification and optimization of fermentation conditions of antagonistic bacteria GZ-34,and the inhibition of antagonistic bacteria GZ-34 against smut fungus and rice blast fungus.The main results are as follows:1?The Ralstonia solanacearum strain was chromosomally tagged with the marker gene eGFP with PBBR1-2 by electroporation.2?30 samples were collected from Guangdong,Henan,Hunan,and other 30 sites.Using soil diluted suspension separation method,coated LB plate,then pick a single colony in R.solanacearum plate through confrontation screened 384 strains of growth have obvious antagonistic effect,then the screened strains were secondary screened,and 56 strains with stable antagonistic effect were obtained.3?The screened strains of antagonistic bacteria were identified,and the result was: GZ-27 is a single strain of Acinetobacter soli,the preservation number is CCTCC NO:M 2016351.GZ-33 is a single strain of Pantoea coffeiphila,the preservation number is CCTCC NO:M 2016352.GZ-34 is a single strain of Escherichia coli,the preservation number is CCTCC NO:M 2016353.GZ-39 is a single strain of Dickeya dadantii,the preservation number is CCTCC NO:M 2016354.The above strains are preserved in China Center for Type Culture Collection,Wuhan University in Wuhan,China.4 ? Greenhouse experimental study on tomato seedling inoculated with strain GZ-27,33,34,39,the root canal inoculation method,the above 4 antagonistic bacteria have good control effect.5?The antimicrobial substances of antagonistic strain GZ-34 were extracted,isolated,purified and identified.The crude extract was obtained by fermentation and extracted and its antibacterial activity was tested.The results showed that the ethyl acetate extract had high antibacterial activity against Ralstonia solanacearum,and the ethyl extract inhibited the growth and sexual compatibility of Sporisorium scitamineum.Then the isolation and identification of antibacterial substances,purified by gel column chromatography,separation and purification of HPLC,identification by ESI-MS and NMR,identification of the antibacterial substance has two kinds,cyclo-((S)-pro-(R)(-Ile)and cyclo-(S)-pro-(S)-phe).The cyclo-(S)-pro-(S)-phe)also inhibited sporulation of Magnaporthe grisea.6?The minimal inhibitory concentration(MIC)of antibacterial substance against Ralstonia solanacearum.7?We continued to study whether the two compounds affect the expression levels of virulence contributors in R.solanacearum.Such as extracellular polysaccharides,motility,T3 SS,TTE,T4 SS pili,chemotaxis and the global virulence regulator PhcA.The experiments were tested in the presence of compounds at 100 ?M.Because the concentration of antibacterial material does not affect the growth of Ralstonia solanacearum.