Study On Screening Of The Inhibitory Strains Against Potato Virus Y And It's Active Fractions

In order to screen biocontrol strains with inhibitory activity against potato virus Y?PVY?,the antiviral activities of moderate halophilic bacteria from Qrhan Salt Lake was determined by quantitative Real-time PCR technique.The results showed that among sixty eight moderately halophilic strains,there were fifty five strains with inhibitory activity against PVY.There are seven strains with inhibition rate higher than 90%,fifteen strains with inhibition rate between 70%and 90%,twelve strains with inhibition rate between 50%and 70%.In order to find potentially antiviral activity of moderate halophilic bacteria in Qrhan Salt Lake,the One-Factor-at-a-Time approach was used to screen the best carbon,nitrogen sources and inorganic salt of strain E40203a2 against PVY;the central combination experimental design?CCD?and Response Surface Methodology?RSM?were applied to optimize the optimal ratio of them.The results showed that the best carbon,nitrogen sources and inorganic salts of strain E40203a2 were glucose,beef extract,K₂HPO₄ and KH₂PO₄.The optimal compositions of medium were glucose 3.16 g/L,beef extract 16.35 g/L,MgSO₄·7H₂O 10 g/L,CaCl₂·2H₂O 0.4 g/L,KCl 5 g/L,KH₂PO₄ 10.51g/L and K₂HPO₄ 12.49 g/L.By optimizing of the culture medium,the inhibition rate of strain E40203a2 against PVY reached to 74.36%,which increased by 38.36%compared with the original medium.Furthermore,strain E40203a2 was identified as Virgibacillus pantothenticus by morphological and molecular identification.The strain E40203a2 was fermented massively in the optimized culture medium,and the inhibitory activity and mechanism of n-butanol and ethyl acetate extracts from fermentation were studied.The result of inhibitory activity showed that the inhibitory activity of n-butanol extracts against PVY was much higher than that of ethyl acetate extracts,with inhibition rate 99.70%and 96.43%,respectively.The result of inhibitory mechanism showed that after the treatment of both extracts on tobacco,the expression of total PVY nucleic acid and P1,HC-Pro,P3,6K1,VPg,N1a,N1b and CP protein-encoded genes in tobacco was down-regulation,the expression of PVY resistance-related genes eIF4E,NtTCTP,F-box and Cullin in tobacco was up-regulation,the expression of Pathogenesis-Related genes PSII,XTH,Cp and LHC in tobacco was down-regulation.Taken together,the moderately halophilic bacteria from Qrhan Salt Lake have biocontrol inhibitory effection on PVY.It is hopeful that the compounds with inhibitory activity against PVY will be isolated from these strains,which will provide the theoretical basis for the development of new type of microbial biological antiviral agents.