Identification Of Pathogen Species Causing Apple Alternaria Blotch And Their Fungicides Resistance Analysis

Alternaria leaf blotch of apple is an important disease in apple production,which occurred widely in the world.It mainly affects apple leaves and fruit,and causes massive defoliation in sever condition,and finally affects tree strength and flower bud formation,results in a decline in fruit quality.It is also one of the most resistant pathogens to produce the high resistance to the fungicides.But,there is still a lack of systematic research on the pathogen species and fungicides resistance level in China.In this study,based on the collection and isolation of Alternaria species in the main production areas,we carried out research on the clarify the species of Alternaria spp.associated with Alternaria leaf blotch of apple in China,the determination of pathogen susceptibility to fungicides and the detection of resistance technology,which provided a methodological and theoretical basis for the efficient management of Alternaria leaf blotch of apple.The main results were as follows:(1)The pathogens of Alternaria species associated with Alternaria leaf blotch of apple from some production areas in China were clarified.A total of 210 samples of diseased samples were randomly collected from six major apple producing areas in Liaoning,Shandong,Shanxi,Henan,Gansu and Anhui,and 289 Alternaria species isolates were obtained by isolation,colony morphology,conidial morphology and pathogenicity determination of the pathogenic fungi.Four different species of Alternaria were identified,namely A.alternata,A.longipes,A.arborescen and A.gaisen,among of them,the dominant species are A.alternata with the ratio of 53.19 %.(2)Detected the resistance of Alternaria mali to the main fundicides in six apple producing areas in China.The results showed that The resistant frequency of test strains to tebuconazole,polyoxin,pyraclostrobin and cyprodinil was 6.50%,90.60%,73.14% and 4.70% respectively in six apple producing areas,and the presence of resistant strains to iprodione was not monitored.The frequency of resistance to tebuconazole and pyraclostrobin varied widely among different regions,with the highest frequency of 38.89% in Gansu and less than 10.00% in other regions;the frequency of resistance to pyraclostrobin was higher in Gansu,Henan and Anhui,at 83.3%,78.00% and 74.70% respectively,while the frequency of resistance to pyraclostrobin was higher in Liaoning.The resistance frequency of strains in Liaoning was lower,only 9.00%.The tested strains in all regions showed high resistance to polyoxin and low resistance frequency to iprodione.(3)A dual fluorescence assay method for the detection of Fungicide-resistant strains was developed.Three different pathogenic fungi were used as objects,multiple stains were screened and staining conditions were optimized.To evaluate the feasibility of the method for detecting active spores of fungal spore metabolism,samples with pre-defined percentages of dead spores were detected by the dual fluorescence assay.In addition,comparing the results of the dual fluorescence assay and the traditional spore germination method for strain activity detection was performed.The results further confirmed that dual fluorescence assay can be used to detect pathogenic spore resistance to protective fungicides.