Analysis On Population Structure Of Magnaporthe Oryzae In Jiangsu Province And Functional Analysis Of Melatonin Synthase MoAsmt2 In Magnaporthe Oryzae

Rice blast caused by Magnaporthe oryzae,is a fungal disease,which can infect rice of the whole growth stage.Rice blasts such as leaf blast neck blast is a consistent threat to crop production and quality.Currently,breeding and planting of resistant crop cultivars is the most effective,environmentally and friendly approach to control this disease.However,resistance gene often lost its resistant after it was applied for a while.The main cause for this is the high variability of M.oryzae.Thus,research on avirulence genes distributed and frequencies in M.oryzae will provide theoretical basis for resistant crop cultivars distribution.Meanwhile,further study to pathogenic mechanism of M.oryzae,which can be used for reference for the best prevention of rice blast.The avirulence genes including ACE1,PWL1,Avr-Pita,Avrl-C039,AvrPiz-t,Avr-Pib,Avr-Pia,Avr-Pii,Avr-Pik and Avr-Pi9 from 621 single spore strains of M.oryzae isolated from Jiangsu province from 2013 to 2017 were detected by 10 specific primers;Meanwhile,we identify the pathogenicity by applying Kiyosaw a’ s differential varieties to identify the composition and frequency of avirulence genes in Jiangsu province.Main results for this study are as follows:The PCR results indicate 10 avirulence genes are distributed in Jiangsu province and different frequencies appeared in different years.Among them,Avr-Pib and Avr-Pia show the highest and lowest frequency that is 74.56%and 9.34%;Respectively,Avr-Pib,Avr-Pik and AvrPiz-t show the highest frequencies and average frequencies are over 70.00%.In addition,it appears same result in regions analysis.We evaluated the distribution frequencies of 10 avirulence genes,and 10 avirulence genes are distributed in north,central,south Jiangsu region.However,the detection rate of north region is significantly higher than that of other regions.Moreover,the pathogenicity test utilizing Kiyosaw a’ s differential varieties reveals Piz-t,Pi-z,Pi-ta,Pi-ta2 showing low virulent,and the virulence frequency was lower than 20.00%.In sum,three avirulence genes Avr-Pib,Avr-Pik,AvrPiz-t are widespreading in Jiangsu crop production area.And resistance gene Pib,Pik and Piz-t still have the potential of disease resistance breeding in crop production.Melatonin can significantly improve plant resistance to fungi as a signal molecule.N-acetylserotonin methyltransferase as the key enzyme in the last step related to melatonin biosynthesis,which can transform tryptophan into melatonin.In this study,gene MoASMT2（MGG₀2120）related to N-acetylserotonin methyltransferase biosynthesis was obtained by sequence alignment.To evaluate the role of MoASMT2 on growth and pathogenicity in M.oryzae,we utilized gene knock-out strategy to getting a preliminary study on function of MoASMT2.Comparing to wild type Guy 11,the deletion mutant showed lower growing rate in complete medium（CM）,minimal medium（MM）,oatmeal medium（OM）,Rice decoction&corn medium（SDC）.Meanwhile,the conidia production was apparently reduced.The surface of mycelium appears fracture depression and the sensitivity to cell wall stress was enhanced.Conidial suspension（5 × 104 conidia ml-1）of the mutant,Guy11 and complemented strain of the susceptible rice seedling（Oryza sativa cv CO-39）and barley leaves by spraying.Guy 11 and complementary strain caused typical lesions,whereas The MoASMT2 mutant developed small and restricted lesions comparing to wild type.Meanwhile,we collected the conidial suspension of strains to inoculate on rice and barley health and wounded leaves,respectively.Wild type and complementary strain in health and wounded leaves showed typical lesions,similarly,The MoASMT2 mutant in wounded leaves appealed same lesions.In contrast,the MoASMT2 mutant only caused small lesions in health leaves.The above results indicate that MoASMT2 involve the invasion process of M.oryzae but it does not affect the expansion of invasion hypha.In sum,MoASMT2 regulated vegetative growth,cell wall integrity,and response to stress of M.oryzae,and then affect its function of pathogenicity.