Study On The Cherry Resistance Mechanism Of Stem Rot Disease And The Function Of The RxLR Effector PnAvh241

To cultivate large numbers of cherry seedlings as quickly as possible,Many seedling companies and fruit farmers use the method of sowing in the spring,grafting in the summer,and growing seedlings in the fall,This led to the death of cherry seedlings grafted in the rainy summer months and even the failure of entire nurseries.It is called cherry stem rot because it causes the leaves to die and the stems to rot.The pathogen of stem rot of cherry is Phytophthora nicotianae,belong to the Oomycota,Once occurred,the rapid expansion,destructive,difficult to prevent and control.The strain P1639 was isolated from stem rot of cherry,We confirmed the difference of disease resistance of different varieties of cherry through inoculation test,All 11 sweet cherry varieties tested were susceptible to disease,and all 6 rootstock varieties were resistant to disease;In order to determine the resistance mechanism of cherry to stem rot,we measured the resistance differences of leaves of different varieties,young branches and lignified branches of cherry,and analyzed the differences of leaf thickness,epidermal cell thickness and stomatal size between cherry varieties with different resistance and measured the activities changes of SOD,POD and CAT before and after inoculate the pathogenic bacteria.The results showed that,branches and leaves of resistant varieties were more resistant than susceptible varieties,leaves and new branches of susceptible varieties are susceptible to disease,and lignified branches are resistant to disease;there was no significant correlation between leaf thickness,epidermal cell thickness and stomatal size and disease resistance;the activity of the three enzymes in the resistant varieties increased significantly before and after the disease,while the activity of the three enzymes in the susceptible varieties showed no significant change or increased trend,which was significantly lower than that in the resistant varieties.Therefore,the lignification of branches plays an important role in resisting stem rot of cherries.SOD,POD and CAT of resistant varieties play important roles in pathogen resistance during tobacco epidemic.A large number of studies have shown that Rx LR effector molecules play a very important role in the process of phytophthora infection.We have screened an Rx LR effector molecule PnAvh241 through bioinformatics analysis,The gene sequence of the effector molecule is 582 bp,and the protein has 193 amino acids.We predicted the first 23 amino acids as signal peptide sequences in the Signal P prediction website,We performed functional analysis on the protein sequence and found that it has a conserved Rx LR d EER motif,in addition to two W motifs and two Y motifs.By analyzing the expression of PnAvh241 in the life cycle of phytophthora nicotianae,We determined that PnAvh241 was up-regulated at the and down-regulated at the initial stage of infection.The relative expression level of spore germination was decreased at 0.5h after infection.With the increase of infection time,the expression increased gradually.When the infection lasted for 24 h,the expression level reached the level close to that of the mycelium stage.We had transient expression the PnAvh241-GFP on the tobacco leaf and determined that PnAvh241 protein is located on the cell membrane of plant cells.In the process of phytophthora nicotianae interacts with its host,and the protein can cause allergic necrosis of plant cells.Based on the above results,we preliminarily determined that the effector molecule PnAvh241 can be recognized by the protein on the cell membrane after entering plant cells,which can cause allergic necrosis of plant cells,the phytophthora nicotianae can avoid being recognized by host cells by down-regulating the expression of this gene in the early stage of infection.The biological and chemical fungicides with better control effect against phytophthora nicotianae were screened by indoor toxicity test,which provided scientific theoretical basis for the control of cherry stem rot in the field.The toxicity test results showed that,biological fungicides such as eugenol,ethylicin and osthole had better inhibition on phytophthora nicotianae,their EC₅₀ were 0.0153 mg/L,1.2925 mg/L and10.6309 mg/L;The chemical fungicides such as Pyraclostrobin,Alkyl Metalaxyl,Cyanazole-enomorpholine,Metalaxyl Cyanazole,cyazofamid,Cyanogen frost and Chlorothalonil,Dimethomorph,Epoxiconazole enestroburin,Thiram had better control effect on phytophthora nicotianae,their EC₅₀ were 0.1539 mg/L,0.426 mg/L,0.5829 mg/L,0.8224 mg/L,0.831 mg/L,1.1033 mg/L,6.0517 mg/L,7.7237 mg/L,14.4061 mg/L.