Study On The Physiological Resistance Mechanism Of Walnut(Juglans Regia) To Bacterial Blight

Juglans regia L.is an important nut tree that has a wide range of economic value cultivated all over the world.In some walnut orchards,walnut blight can become a problematic disease that affects seriously the growth of walnut trees and the quality of nuts.Although walnuts are rich in phenolic compounds,the potential for disease resistance by phenolic chemicals is not fully understood.Hence,Xanthomonas arboricola PV.Juglandis（Xaj）,the pathogen of walnut blight,was isolated and identified in this experiment.The fruits of four walnut varieties,and‘Xiangling’,‘Xiluo 2’,‘Yuanfeng’and‘Xifu 2’with 60days fruits after pollination were selected as experimental materials to analyze the biochemical reaction of Walnut in response to Xaj.Specifically,we determined the composition and content of phenolic compounds among different varieties in the selected walnut fruits,as well as the content of total phenols（TPC）,total flavonoids（TFC）,phenolic compounds of different monomers after inoculation with bacterial pathogens including gallic acid,catechin,p-coumaric acid,chlorogenic acid,quercetin,ferulic acid,protocatechuic acid,myricetin quercetin as well as juglone.Besides,phenylalanine ammonia lyase（PAL）and polyphenol oxidase（PPO）related phenylpropane metabolism enzymes were determined.At the same time,we measured the activities of superoxide dismutase（SOD）,catalase（CAT）as well as peroxidase（POD）related to active oxygen scavenging system.This study has a certain reference significance for the in-depth study of the resistance mechanism of walnut to Xaj,and has a certain guiding role in the breeding of disease resistant varieties.The main results are as follows:1.The pathogen of walnut blight,a gram-negative bacterium,Xanthomonas arboricola PV.Juglandis was identified by molecular identification and morphological identification through separating and purifying the fruit suspected of blight disease in our experiment.2.After inoculation with Xaj,we preliminarily concluded that‘Xiluo 2’was the most resistant variety and‘Xifu 2’was the most susceptible variety according to the phenotypic observation and incidence rate calculation.3.After infection with pathogen,we found that SOD,CAT and POD may be involved in the interaction between walnut and Xaj by measuring the activities of active oxygen scavenging enzymes in this research.Firstly,after 4 days of inoculation,the SOD activity in all varieties was significantly higher than that in the control samples.And the activity of SOD in resistant variety‘Xiluo 2’was the highest;Besides,activity of CAT in resistant variety‘Xiluo 2’was significantly higher than that of the control fruits,but there was no significant difference between other varieties after 8 days of inoculation.Lastly,the activity of POD in the resistant variety‘Xiluo 2’first increased rapidly and significantly higher than that in the control fruits in the later stage,but the remaining varieties began to increase significantly on the 8th day after treatment and was similar to that of the control.4.We speculated that PAL and PPO may also be involved in the defense response of walnut blight through measurement of the activities of phenylpropane metabolism related enzymes after treatments in this research.First of all,the PAL activity of all varieties decreased first and then increased.The resistant variety‘Xiluo 2’increased significantly on the 8^thday of inoculation and presented a distinct difference with the control samples.However,the PAL activity of the remaining varieties changed little in the whole process of disease development.On the other hand,the activity of PPO showed an upward trend after inoculation,which was significantly higher than that of the control samples on the 16th day.We also found that the activity of PPO maintained the highest activity in‘Xiluo 2’and maintained the lowest activity in‘Xifu 2’respectively in our article.5.The phenolic chemicals in four walnut varieties were detected by liquid chromatography in our experiment.Gallic acid,catechin,p-coumaric acid,chlorogenic acid,quercetin,ferulic acid,protocatechuic acid and myricetin were detected in different wavelength.Specifically,our results showed that the composition of phenolic compounds in the fruits of all varieties was similar,but the content was different.Among the four varieties,the content of chlorogenic acid was the highest,followed by catechin.The contents of gallic acid and myricetin were similar,and the contents of them were dependent on the different varieties.At the same time,the contents of protocatechuic acid and ferulic acid were relatively small in the four varieties.6.After inoculation with pathogen,on the one hand,we concluded that the total phenol content showed an upward trend in the four varieties,but there was a significant difference between the two varieties‘Xiangling’and‘Xiluo 2’and the control samples,indicating that the defense mechanism of walnut against Xaj may be realized through the early accumulation of phenolic compounds.However,the content of total flavonoids had no significant difference in the whole disease development process.In addition,we also found that protocatechuic acid and ferulic acid may participated in the interaction between walnut and Xaj.on the 4^thday,the content of protocatechuic acid increased significantly in the four varieties,and there was a significant difference between the resistant variety‘Xiluo2’and the control fruits,while the remaining varieties changed little compared with control fruits.In the early stage of disease development,the content of ferulic acid increased significantly in‘Xiluo 2’and decreased significantly in‘Yuanfeng’.