Physiological Response And Multi-Omics Analysis Of Actinidia Arguta To Bacterial Canker(Psa)Infection

Pseudomonas syringae pv.actinidiae(Psa)is a highly destructive bacterial disease that has caused serious impact on kiwifruit industry,and there is no effective control method.Many studies have shown that the resistance of Actinidia arguta to Psa is significantly higher than that of Actinidia deliciosa and Actinidia Chinensis.Until 2019,the symptoms of canker disease were found for the first time in China on the branches of A.arguta,and it spread rapidly between adjacent lines.The speed of its infection is no less than that of Psa in A.chinensis.It has become the potential disease of A.arguta,which will cause great harm to A.arguta industry.In view of this problem,the specific research has been carried out and the related results as follows:Pathogens of suspected bacterial canker on A.arguta ‘Longcheng No.2’ were isolated and identified.The pathogenic bacteria were identified as Pseudomonas syringae pv.actinidiae(Psa)by the analysis of morphology,16 S rDNA and gyr B sequence,which belonging to Psa biovar 2 by Psa-specific primer sequence analysis.In order to evaluate the resistance on Psa ‘R12’,54 A.arguta germplasm resources were infected by artificial inoculation of stems,and their resistance levels were classified according to the disease index.In general,the germplasm were divided into five disease resistance categories,including 2 accessions with high resistance,accounted for 3.70 % of all the inoculated accessions;there were 11 resistant accessions,15 tolerant accessions,21 susceptible accessions,5highly susceptible accessions among them,accounted for 20.37 %,27.78 %,38.89 %,and 9.26 %,respectively.The resistant accession ’Cuiyu’ and susceptible accession ’TL02062’ were inoculated with Psa ’R12’by living branch inoculation method,and the activities of protective enzymes and the contents of salicylic acid(SA),jasmonic acid(JA),total phenol and lignin in plants at 0 d,2 d,7 d and 14 d after inoculation were detected and analyzed,and the correlation between 9 physiological indexes was analyzed to explore the response mechanism of different resistant accessions.The results show that after inoculation with Psa ’R12’,SOD,POD,PAL and PPO activities of resistant and susceptible accessions were significantly increased,but the extent of increase was different.After inoculation,the SA content of both accessions increased significantly,and the increase of resistant accession was significantly higher than that of susceptible accession.The JA content of disease-resistant germplasm increased gradually,while that of susceptible germplasm decreased firstly and then increased.The total phenolic content of resistant germplasm increased faster than that of susceptible germplasm.The results of correlation analysis showed that SA content was positively correlated with JA content,PAL and POD activities.PAL was significantly correlated with seven physiological indexes except PPO.The plant resistance was correlated with SA,JA,polyphenol content,PAL,SOD and PPO activities,and was closely related to SA content and PAL activity.SA and PAL play key roles in the resistance of A.arguta to Psa infection.Transcriptome sequencing analysis was performed on stem segments of resistant and susceptible germplasm before and after inoculation.Several genes involved in SA and ET/JA-mediated signaling pathways were up-regulated,and SA and JA/ET signaling pathways may be synergistically regulated to resist Psa invasion.The transcription factors involved in the response mainly included WRKY,MYB,Etc.,among which WRKY was the most responsive.The up-regulated expression of WRKY50 and WRKY51 in resistant germplasm was significantly higher than that in susceptible germplasm,suggesting that WRKY50 and WRKY51 may positively regulate SA signaling pathway to resist Psa infection.Resistance genes NL,TNL,N and T were differentially expressed,with TNL being the most active.KEGG pathway enrichment analysis showed that terpenoid backbone biosynthesis,sesquiterpenoid and triterpenoid biosynthesis were significantly enriched in all comparison groups;Flavonoid biosynthesis,phenylalanine metabolism,cyanoamino acid metabolism,and amino sugar and nucleotide sugar metabolism were more significantly enriched in resistant germplasm,which may play an important role in the process of plant resistance to Psa infection.Untargeted metabolomics analysis showed that the differential metabolites of resistant and susceptible germplasm gradually increased with the extension of inoculation time,However,the differential metabolites in susceptible germplasm are always more than those in resistant germplasm.Myrianthic acid,quillaic acid,ganolucidic acid B and tyromycic acid were significantly accumulated in resistant germplasm,while the triterpenoid antimicrobial-actinidic acid was also significantly accumulated in resistant germplasm,which may play an important role in inhibiting Psa diffusion.The enrichment pathways of resistant and susceptible germplasm were mostly uniform,but there were differences in the enrichment significance at different time after inoculation.Some pathways,such as oxidative phosphorylation,vitamin B6 metabolism,ascorbate and aldarate metabolism were only enriched in resistant germplasm.The transcriptome and metabolome association analysis showed that in the association network,the accumulation of alkaloids-geissospermine and triterpenoids was the most significant in resistant germplasm,and the pathways involved in phenylpropanoid biosynthesis,MAPK signaling pathway and so on.Weighted gene co-expression network analysis(WGCNA)indicated that DMR6,RGA3,AOX3 and transcription factors WRKY50,WRKY51,WRKY75 may be the key genes in response to ’R12’ infection,which needs further research.