Study Of Molecular Mechanism Of Diseases Resistance Induced By Burdock Fructooligosaccharide In Plants

Plant pathogens can cause considerable loss of crop yields.At present,the plant diseases mainly controlled by using of synthetic fungicides.But the use of fungicides to control plant diseases has been restricted due to their effects on food and possible side-effects on human health.Thus,it has become a worldwide trend to explore new alternatives to reduce the use of synthetic fungicidesPlant could generate various defense responses to protect themselves from pathogens.Many studies have shown that these defense responses could also be induced by a class of molecules called elicitors.Oligosaccharides are a well characterized class of elicitors that trigger a large array of defense responses.Burdock fructooligosaccharide(BFO) is a fructosan oligomer that was first isolated from the root tissue of Arcitum lappa by our group.Previous studies have shown that BFO induces various defense responses in plants.However,the systemic defense responses and the underlying molecular mechanism that mediated by BFO in plants have not been elucidated.Therefore,we used biochemical and molecular approaches to investigate BFO-mediated non-specific defense activation in plants.In addition,we also studied the defense responses to TMV and BFO in tobacco leaves at different ages.The mainly results are as followed:1.As an elicitor,chitosan oligosaccharide(CO) has been extensively researched, the results indicated that CO can induce resistance against multi-diseases in plants.In this study,the effects of BFO on the control of postharvest diseases in tomato fruit and the underlying mechanism were investigated with CO as a positive control and distilled water(DW) as a negative control.The results indicated that both BFO and CO could effectively inhibit natural postharvest diseases and reduce incidence of disease inoculated with Botrytis cinerea in tomato fruit.After treated 5 d by BFO the natural disease incidence and disease index were about 53.9%and 45.2%to DW treatment,and the natural disease incidence and disease index were still lower than DW treatment during the time after BFO treatment.In addition,inoculation with B. cinerea 4 d,the disease incidence and disease index were about 62.4%and 48.1%to DW treatment in BFO treated tomato fruit.BFO increased the mRNA level of genes encoding pathogenesis-related proteins(PRs),such as PR-1a,PR-2a(extracellularβ-1,3-glucanase),PR-2b(intracellularβ-1,3-glucanase),PR-3a(extracellular chitinase) and PR-3b(intracellular chitinase),and induced the mRNA accumulation of phenylalanine ammonia lyase(PAL) gene in tomato fruit.Furthermore,it evoked the activity of peroxides(POD) and enhanced the biosynthesis of phenolic compounds, but did not affect the activity of polyphenol oxidase(PPO).In this study,BFO and CO can not induce new isoenzyme bands of POD and PPO emerge,but the width of their isoenzyme bands were enhanced treated by BFO in tomato fruit.But interesting, treatment with CO could only strengthen the width the isoenzyme bands of POD,it has no effect on PPO in tomato fruit.These results suggest that controlling effects of BFO on both natural diseases and gray mould in tomato fruit might be attributed to enhance the transcript exprssion of defense related genes,increase the activity of POD and induce accumulation of phenolic compounds.2.The effects of BFO on the control of TMV in tobacco leaves and the underlying molecular mechanism were investigated.The results indicated that treating cells with BFO 48 h before inoculation with TMV significantly inhibited TMV multiplication. The level of TMV-CP transcripts in DW treated leaves was about 7.0-fold higher than that in the BFO treated leaves 24 h after inoculation with TMV.In addition,about 5 d later,the leaves treated with DW had the typical mosaic pattern,while the leaves treated with BFO did not.These results suggested that BFO treatment improved the level of resistance to TMV.Our results also indicated that the levels of transcription of PR protein genes,including acidic PRs(PR-1a,PR-2,PR-3,PR-4,PR-5) and basic PR-1 and defense-related enzymes[PAL and 5-epi-aristolochene synthase,(EAS)] genes were increased both locally and systemically by BFO treatment,in correlation with the induction of resistance against tobacco mosaic virus(TMV).In plants,H₂O₂ and salicylic acid(SA) act as signal molecules to induce systemic acquired resistance (SAR).And interestingly,BFO also induced accumulation of H₂O₂,SA and SA 2-O-β-D-glucoside(SAG) in tobacco leaves.In addition,the expression of PR genes is usually taken as a marker for systemic acquired resistance(SAR).Therefore,these results strongly suggested that BFO activated the defense responses and the mode of action of BFO might have been induction of SAR through H₂O₂ and SA signal molecules.3.The defense responses to TMV in tobacco leaves at different ages(young,adult and old) were studied.The results indicated that the content of TMV in old leaves was the most at 48 h after inoculation with TMV among of them,and about 8.1 and 35.1-fold higher than that in adult leaves and in young leaves at that time,respectively. Therefore,there is the strongest resistance against TMV in young leaves,followed by adult leaves,and old leaves have the weakest resistace.Followed,we investigated the response of defense related genes(PR-1a,PR-2,PR-3,PR-4,PR-5,PAL )to TMV in tobacco leaves at different ages.The results shown that the mRNA levels of above defense related genes all could be induced by TMV,but the responses intensity to TMV of the above genes was the weakest in young leaves,while it was the strongest in old leaves.In addition,we further studied the defense responses to BFO in tobacco leaves at different ages,and further investigated the effects on the resistance against TMV treatment with BFO in tobacco leaves at different ages.The results indicated that the response of the above defense relate genes was the strongest in young leaves, followed by adult leaves,and the old leaves is the weakest.But the effects of BFO on the resistance against TMV were the strongest in old leaves,followed by adult leaves, and the young leaves is the weakest.Inoculation with TMV after treated by BFO 48 h, the content of TMV was smaller than DW treated.At 48h inoculation with TMV,the content of TMV in DW treated tobacco young leaves,adult leaves and old leaves, were about 1.4,1.9 and 11.0-fold higher than BFO treated leaves,respectively.This result suggests,the effect of BFO treatment on the resistace against TMV was the strongest in old leaves,followed by adult leaves,and the young leaves is the weakest. In oppositing leaves at different ages,the transcripts level of the above defense related genes and the content of TMV in leaves treated by BFO both were smaller than the leaves treated by DW.Together above results,we come to the conclusion that there are different resistance to TMV in tobacco leaves at different ages,and the resistace against TMV was the strongest in young leaves,followed by adult leaves,and the old leaves is the weakest.In addition,the results indicated that treatment with BFO could inhibit the multiplication of TMV in tobacco leaves at different ages,and there is the best effect in old leaves,the young leaves are the worst.The results also indicated that there is different defense response between BFO and TMV in tobacco leaves.