Effect Of Verticillium Dahliae Infection On Physiological Parameters And Gene Expression In Eggplant Seedlings(Solanum Melongena L.)

Eggplant(Solanum melongena L.)originates in the tropical regions of India and Southeast Asia.Eggplant verticillium wilt is one of the three major diseases of eggplant.The pathogen that causes eggplant verticillium wilt in China is mainly Verticillium dahliae.In this study,expansion of Verticillium dahliae in plant tissues were monitored across stages of infection using light and electron microscopy.The Verticillium dahliae marked with GFP fusion was also established.In addition,the effects of Verticillium dahliae infection on the growth,physiological parameters of eggplant were determined.A transcriptome analysis of differentially expressed genes during the Verticillium dahliae infection were performered.The mainresults of this study are as follows:1.The invation progress of Verticillium dahliae in plant tissuesThere was very few mycelia in the roots 3 days post inoculation by Verticillium dahliae,very few appeared.5 days post inoculation,conidia were formed with hyphae,but no pathogen was observed in the stems and leaves.7 days post inoculation,a large number of conidia and mycelia were observed in the roots and stems of plant tissues,and lots of conidia were also present in the leaves.At this time,the edge of the leaves of plants showed slight wilting.10 to 15 days post inoculation,the number of mycelia and conidia showed changes in roots,stems and leaves,and the most obvious character was the significant decrease in mycelia and conidia in roots.The plants exhibited obvious symptoms of Verticillium wilt with wilting and yellow-edged leaves.2.Labling Verticillium dahlia using GFPThe plasmid pSilent-1 and pEGFP were digested and ligated to obtain the GFP fungal expression vector..The transformants of Verticillium dahlia were obtain by the protoplast transformation and identified by microscopic examination,growth rate,GFP gene sequence amplified and pathogenicity.The results showed that GFP was transferred into the genome of Verticillium dahliae,and there were no significant differences in pathogenicity and growth between the transformants and the wild strains.So the GFP-labled Verticillium dahliae strains can be used for the study of the infection process.3.Effects of Verticillium dahliae infection on growth and physiological parametes of plantsOur results showed that growth of eggplant plants was inhibited by Verticillium dahlia infection,with the inhibition rate of stems by 7.81%and that of height by 17.32%,but it has no significant effect on the root-top ratio.Chlorophyll content in leaves was slightly affected at early stage of Verticillium dahlia infection,but it decreased at later stage.During the infection of Verticillium dahliae,the content of soluble sugar of leaves increased,and the content of soluble protein increased at early stage and decreased later.The content of malondialdehyde of leaves showed a similar tendency as soluble protein.The activities of SOD,PPO and POD enzymes all increased in leaves.Accumulated callose in leaves increased at early stage and decreased later.We also observed the distribution of superoxide anion and hydrogen peroxide differed in the leaf tissues,and the content of hydrogen peroxide increased,while the superoxide anion decreased as the disease became severe.4.Transcriptome analysis of differentially expressed genesAfter inoculation with Verticillium dahliae,significant difference in the number of differentially expressed genes(DEGs)was observed.There were 1529,2287 and 9085 DEGs 1,7 and 13 days post inoculation,respectively.GO analysis of DEGs indicated they were mainly found in groups of metabolic processes,cellular processes,single-organism process,response to stimulus,cell parts,membranes,binding,catalytic activity,etc..Analysis of DEGs by KEGG pathway revealed that the DEGs are mainly involved in starch and sucrose metabolism,stilbenoid,diarylheptanoid and gingerol biosynthesis,glutathione metabolism,plant pathogen interaction,phenylalanine metabolism,flavonoid biosynthesis,terpenoid backbone biosynthesis,sesquiterpene and triterpenoid biosynthesis.Among them,the most important DEGs involved in plant-pathogen interaction pathway are transcription factors such as WRKY25 and WRKY29,the FRK1 and PR1 protein genes,and genes involved in hypersensitivity,such as RIN4,RPM1 and RPS2,and all these genes were obviously upregulated.