Resistance Evaluation And Mechanism Of Different Fall Dormancy Alfalfa (Medicago Sativa L.) On Stemphyllium Leaf Spot Disease

The Stemphylium leaf spot of alfalfa which development serious in 3 years alfalfa field in Wanzhuang town of Hebei province was investigated and the pathogen was separated and identified.And based on it,evaluated the disease resistance to Stemphylium botryosum of 50 alfalfa cultivars,and analyzed the connection between stoma and fall dormancy with disease resistance of germplasm resources.And 8 representative cultivars which represent the different resistance to disease grade were selected to research the physiological and biochemical characteristic in the disease resistance process.The main results of research are as followings:1.The disease investigation of alfalfa in Langfang area showed that Stemphylium leaf spot disease in alfalfa was the new report in this area,and the disease development widely, the main factor to disease development was the rainfall.Through separating and identifying and pathogenicity test of disease pathogen,adopted the Booth classification standard and system,the pathogeny of Stemphylium leaf spot of alfalfa was identifyied as the Stemphylium botryosum Wilson.,which belonged to the sub-phylum of deuteromycotina,class of moniliales,family of trappoid and genus of stemphylium.The species of the pathogen is Stemphylium botryosum Wilson2.The microstructure and the biological character of the Stemphylium botryosum Wilson was studied unprecedentedly in alfalfa in China.The results showed that the pathogen growth more rapidely in the V8 culture medium,and it got to generate spores from the fourth days,the conidiophore has diversiforms of clavated,column and ellipse.the size of the conidium was 18～25um×22～30um.In the initial stages,the conidium had 1 traverse septum,with the maturation of the conidiurn,the traverse septum increased to 1～4,and 0～3 longitudinal septum comed forth.The conidiophore was singly inserted on the top of the peduncle,and spore trace can be observed in the base of the conidium,the conidiophore rarely had branchs.In the axenic water with the culture temperature of 25℃,the pathogen germinated after 4h,and reached the apex after 14h.The optimal culture humidity,temperature and PH were above 95%,about 25℃and PH5～8 respectively.3.Several effect factors of the pathogen inoculation were studied in two mode of inoculation,individual plant inoculation and single leaf inoculation.results showed that in former mode the optimal inoculation condition was:at the culture temperature of 25℃,by the method of daubing,the concentration of the conidial suspension was 5.0×10~5-1.0×10~6 /ml,and the optimal investigation stage of disease degree was 10-15 days after the inoculation.The method of single leaf inoculation could also be used in the for Stemphylium leaf spot disease resistant evaluation,In this mode,the dipping method was the optimal inoculation ways,and the mature degree of the leaf had impact on the inoculation effct.mature leaf was the optimal material.On the basis of the study,the inoculation technic system for the Stemphylium leaf spot disease evaluation was established.4.methods of nature infection in field,individual plant inoculation and single leaf inoculation were compared for the evaluation effect.The results showed that the outcome of the 3 evaluation methods were correlative remarkably,and method of the individual plant inoculation can evaluate more variation in the 50 entries.So this method was adopted in the germplasm evaluation for the Stemphylium leaf spot disease.Results showed that there were 3 entries of resistant,17 entries of moderate resistant and 23 entries of moderate sensitive in the 50 materials,there was no high resistant material observed in the 50 entries.On the basis of the evaluation,8 entries of different resistant and sensitive degree were choosed fot the study of disease resistance mechanism.5.Correlation analysis results showed that stoma density had the remarkably negative correlation with the Stemphylium leaf spot disease resistant degree.Stoma density can be treated as the index of the Stemphylium leaf spot disease evaluation.Fall dormancy rate had the remarkably positive with the disease resistant degree.It indicated that with the fall dormancy rate increased,the material may more sensitive to this disease.6.Studing of the change of chiorophy content and the activity of POD,PAL,PPO in different disease resistant degree material after the infection.The physiological mechanism of the disease resistance had been analysised.which showed that after the disease infection, the chlorophy descend in all the materials,and the resistant material had less descend extent than the sensitive material.7 After the disease infection,the activity of POD,PAL and PPO were increased remarkably,the range of activity of POD and PALincreasing in the resisitant material were remarkably larger than which in the sensitive material.The increasing range of PPO activity has no prominent correlation with resistance degree.