Collecting For Identification Of The Pathogen Of Mulberry Wilt Disease In South China And A Preliminary Study On The Mechanism Of Infection

China is the birthplace of the silk industry, wich has more than5,000years ofhistory. South China has always been one of the major sericulture production base inChina, the area of mulberry field is more than6,000hectares, accounting for1/3ofthe total area in the country. In south china mulberry bacterial wilt is a devastatingdisease and more serious harm. The disease caused great economic losses and aserious impact on sericulture production. Predecessors had carried out a series ofstudies on the incidence rules and control methods of bacterial wilt of mulberry, someprogress has been made.But understanding of mulberry bacterial wilt pathogens andpathogenic mechanism is still relatively small. The comprehensive and systematicunderstanding of the mulberry bacterial wilt is sill lacking.In the study, the pathogen of mulberry bacterial wilt in southern China werecollected, separated and identified by molecular identification. The biovarclassification, the inoculation conditions and Pathogenicity Differentiation arecompleted, in order to in-depth understand the pathogenesis of mulberry bacterial wilt.The results provide theoretical and technical support for mulberry breeding thatresistance to bacterial wilt, and preventing R. solanacearum dissemination andspreading. The main results of the study are as follows:(1) Through the investigation of important bacterial wilt epidemic areas in SouthChina, there were two kinds of symptoms. In the early stages,the main difference isthat, one symptom is the green withered leaves, another is the brown wither leaves.The both of symptoms are basically similar during the later stage and the entiredisease process. In the early onset,the3rd&4th leaf under the terminal bud firstwithered,and light yellow or brown stripes appear in the xylem of roots.In the middlestage, the most of the leaves wilt and the xylem of roots appears a large number ofblack and brown stripes. The late of the onset, All of the leaves wither curly, andgradually fall. Also,the roots’ skin began to rot and the whole xylem become darkbrown.(2) A total of370pathogens were isolated from the samples of117mulberrybacterial wilt, collected in South China. By16S rDNA-PCR sequencing, there are223strains belonging to R. solanacearum,44the Pseudomonas,72the Enterobacteriaceae,31other bacteria. This results shows that the pathogen is Ralstonia solanacearum inSouth China. (3) The form of3types of bacteria is essentially similar. In TTC solid mediumthe colony performance red, mostly round, smooth and shiny, wet mucoid,and thegram stain was negative. Under the SEM, the Ralstonia solanacearum andPseudomonas are short rod. Ralstonia solanacearum is about1.1×0.4μM, andPseudomonas is about1.69×0.4μ M. The main differences among the three types ofbacteria are the growth rate and colony color. The best culture time of Ralstoniasolanacearum is48h, the Enterobacteriaceae and Pseudomonas is24h. The Coloniesof the Ralstonia bacteria and Pseudomonas are dark red In TTC solid medium.At thelate growth stage Ralstonia solanacearum colonies’ surface will appear a layer ofwhite film, and E. coli colonies usually pink.(4)By the biovar classification of110stains Ralstonia solanacearum, there are89strains of biochemical type I,4strains of type Ⅲ,17strains of type V.The resultshows that the Ralstonia solanacearum from mulberry in South China is abiochemical type I.(5) By orthogonal experiment, and explore the infection’ s conditions ofmulberry seedlings. The results showed that the best conditions of the hydroponicinjury root inoculation method: seedling age of30days, a temperature of30℃,inoculum109cfu/mL. Through water culture incoulation method,the pathogenicity of30eucalypt R.solanacearum isolates from South China were identified. Virulence ofdifferent strains are quite different12-50-1,11-41and11-42, the3strain virulence isstronger and stable.(6) by SEM,the process of Ralstonia solanacearum infecting mulberry showedthat: First, a small amount of R. solanacearum colonize in vascular bundles of themulberry root.120h after inoculation, the results show that the large number ofbacteria present in the vascular catheter and cell gap, plug the catheter and cause theplants showed symptoms of wilt.