Study On Resistance To Sclerotinia Sclerotiorum In Rapeseed (Brassica Napus L.) Induced By Bacillus Thuringiensis

Bacillus thuringiensis produce many types of insecticidal crystal proteins,which have insecticidal activity against many important pests of agriculture.Bt strains are most widely used insecticides,and have strong antifungal bioactivity against many plant pathogens.Recent studies reported that Bt strains could induce systemic resistance in tomato against Ralstonia solanacearum,but the induction mechanism was not clear.In this study,165 standard and 5 common Bt strains kept in our lab were testified for their activity to induce systemic resistance in rapeseed against Sclerotinia sclerotiorum.Meanwhile signaling pathway of rapeseed activated by Bt strains was identified by transcriptome sequencing and Real-time PCR to primarily reveal the mechanism of systemic resistance induced by Bt strains.This study lay the theoretical foundation for the application of Bt strains in plant disease control.1.A total of 12 Bt strains with antifungal activity against S.sclerotiorum were obtained by the plate confrontation test.Disease resistance related genes of 4 higher activity strains were analyzed and all these strains contained chit gene.Chitinase activity of 4AP1 strain cell culture was 672.4 U/mL.2.Excluded 12 Bt strains with antifungal activity against S.sclerotiorum,Bt strains which can induce systemic resistance in rapeseed against S.sclerotiorum were selected from the remaining 158 Bt strains.A total of 18 Bt strains with induced activity was obtained by the method of S.sclerotiorum fungus cake leaf inoculation,and 7 days after the inoculation the rapeseed leaves were not wilt.While rapeseed leaves or the whole plant were wilt treated by Bt strains without induced activity.4I4?4F5?4AZ1?4M1?4BM and 4CC1 strains still have stable induction activity,while the other 12 strains lost induction activity after the treatment of heating at 100? for half an hour.Exopolysaccharide was isolated from 6 Bt strains(4I4,4F5,4AZ1,4M1,4BM1 and 4CC1),which showed the same induction activity as the original Bt strains cultures.These preliminary results suggest that exopolysaccharide of Bt stains induced systemic resistance in rapeseed against S.sclerotiorum.3.After the treatment of 4F5?4BM1 culture,the expression of genes of salicylic acid(SA),jasmonic acid(JA)and ethylene(ET)signal transduction pathway were analyzed by qRT-PCR.The result showed that the gene expression level of WRKY70(SA)and PDF1.2(ET)were up-regulated,while HEL(JA)was down-regulated.Signaling pathway of the rapeseed leaves activated by Bt strains was further analyzed by transcriptome sequencing.In 4F5 strain-treated rapeseed leaves,total 86 genes were up-regulated and 217 genes were downregulated,while total 64 genes was up-regulated and 62 genes was down-regulated in 4BM1 strain-treated rapeseed leaves.SA?JA?ET signaling pathway related genes were up-regulated among two treatments.4.The 5 days rapeseed leaves induced by 4F5?4BM1 strain were ate by newlyhatched larvae of Plutella xylostella were fed with rapeseed leaves 5 days after inducing by 4F5 and 4BM1 strain.There were no significant differences between the induced rapeseed and the negative control in damage area of leaves,weight and hatch ability of Plutella xylostella.The results showed that there were no obvious effects on insect resistance of rapeseed induced by Bt strains.A total of 18 Bt strains were found to induce systemic resistance in rapeseed against S.sclerotiorum.It was also found that SA-dependent signaling pathway ? JA-dependent signaling pathway and ET-dependent signaling pathway were involved in the induction process and exopolysaccharide was primarily identified as the induced substance.This study provided new ideas for construction and screening of engineering Bt strains with insecticidal activity and disease-resistance.It is of great significance to broade the application scope of Bt strains,increase the market share and reduce the use of chemical pesticides and pesticide.