Infection And Transcriptome Analysis Of Zoysia Japonica Response To Rhizoctonia Solani

Brown patch on Turfgrass was a fungal disease caused by Rhizoctonia solani Kuhn. Spread of the disease in speed showed a trend of increasing, and gradually became a worldwide epidemic disease. The infection relationship of Z. japonica and R. solani was established by the artificial embedded methods. In this research, several technical means were used, such as histological staining transparent technology, paraffin sections technology, transmission electron microscope technology, defense enzymes detection technology and the transcriptome sequencing technology. The objective of this research was to study the interaction process of R.solani and Z.japonica and screen some differentially expressed genes related to host plant resistance. Molecular interactions mechanism of Z japonica and R. solani were discussed at the resistance gene expression level. The main results we obtained were as follows:(1) The hyphae attach to the root surface and not invade to the root organization after the inoculations. After the 24 hours of the inoculations, hyphae formed embryonic infection cushions and penetrated to the root tissue through intercellular spaces. Root cells and leaf cells still kept its complete structure and leaves had no visible symptoms at this time. At the time of 36 hours after the inoculations, the complete infection cushions had formed. The large amount of hyphae invaded and extended inward the root cells. At this moment, root cells and leaf cells also kept its intact structure and visible symptoms occurred in leaves. After the 48 hours of the inoculations, the large number of hyphae infected along the root surface widely and invaded the cortical cells, and gradually extended inward the vascular system. Root cells also keep its complete structure, but more and more serious disease symptoms developed from the bottom and leaf cells rupture. After the 96 hours of the inoculations, the whole roots surface were almost covered by hyphae which invaded the cortical cells and gradually extended inward all the tissues except xylem vessels, but its morphological structure were still intact. The whole blade which were covered with infection structure wilting. Leaf cells structure were in degradation. They were suggested that the lesions of Z. japonica grass has gradient and also indicated that it had no direct contact with the direct infection of R. solani. At the same time, the interaction relationship between R. solani and Z. japonica and disease symptoms provided evidence for the selection of time point for the transcriptome sequencing. We should selected the root samples after 12-48 h of the inoculations to obtain differentially expressed genes profile related to resistance from Z. japonica by transcriptome sequencing.(2) (3-1,3-glucanase, Chitinase, PAL and LOX enzyme activity in the root showed the trend of rising first and downward after while these enzymes activity in the leaves showed a trend of falling first and rising after. It is noteworthy that the enzyme activity in the root was mostly higher than that of activity in leaves, suggested that roots played an important role in resistance to pathogen infection.(3) GO analysis of the transcriptome data showed that the 12 h and 24 h after infection by R. solani are the key time point for Z. japonica to resist fungal invasion.(4) 12 up-regulated differentially expressed genes which their main function was to build structure obstacle against pathogen invasion were screened. Up-regulated unigenes of "Zenith" root fungal resistance were involved in ligin biosynthesis, phytoalexin synthesis, oxidative burst, wax biosynthesis. Cysteine protease protease inhibitors which its function was to inhibit protease in the pathogen showed increased expression steadily, it’s proved that disease symptoms of the zoysia grass was irrelevant to the direct infection of R. solani; Also 2 genes which were leucine rich repeat receptor protein kinase and subtilisin-like protease may act as recognition receptors of pathogen perception and plant signal peptide in the host plant, repectively. While two down-regulated unigenes encoding leucine-rich repeat receptor protein kinase and subtilisin-like protease indicated repression of "Zenith" pathogen-associated molecular patterns perceiver which activated the expression of plant specific defense pathway. Their down regulated expression speculated that R.solani may manipulate defence pathways of host by secreting effector to favor its invasion.