Screening,Identification And Biocontrol Effect Of Antagonics Strains Against Tobacco Wildfire

Tobacco wildfire is one of the bacterial diseases occurring on tobacco,and the pathogen is Pseudomonas syringae pv.tabaci.The disease is relatively serious in the northeast and northwest tobacco areas.It mainly occurs in the prosperous period,harming tobacco leaves.When it is serious,it will burn like fire,causing adverse effects on tobacco production.Currently,chemical control is mainly used,but wildfire pathogens are prone to develop drug resistance and cause pesticide residues on tobacco,affecting the ecological environment.Biological control is becoming increasingly important in tobacco disease control.Among them,Bacillus sp.is widely distributed,easy to isolate,highly resistant,and induces plant resistance to pathogen infection,making it the best choice for biocontrol agents.In this study,Bacillus velezensis CDY8-11 was selected from the rhizosphere soil of healthy tobacco plants in diseased tobacco areas,which has a significant inhibitory effect on tobacco wildfire pathogens,and its fermentation studies and control effects were conducted to provide new ideas for the control methods of tobacco wildfire disease.The research results are as follows:1.A total of 48 samples of healthy tobacco rhizosphere soil were collected from the tobacco epidemic area of Panzhihua City,Sichuan Province.A total of 286 strains with different characteristics were isolated by dilution coating method.Six strains with good inhibitory effects on Pseudomonas syringae were obtained through preliminary screening and re-screening using plate confrontation method,among which strain CDY8-11 had the best antagonistic effect on Pseudomonas syringae,and the inhibition circle was 22.29 mm.Combining morphological characteristics,fatty acid content,API 50CH detection,16S r RNA,and gyr B gene sequence,the strain CDY8-11 was finally identified as Bacillus velezensis.Strain CDY8-11 has inhibitory effects on various pathogenic bacteria,and could produce siderophore,cellulase,protease,and also dissolve phosphorus and fix nitrogen.2.The single factor experiment and response surface optimization method were used to optimize the liquid fermentation medium and fermentation conditions of B.velezensis CDY8-11.The optimal fermentation medium components were 28 g/L maltose,20 g/L yeast,and 1%KCl.The optimal fermentation conditions were:temperature 27℃,incubation time 36 hours,p H 7.0,liquid content 30 m L/250 m L,and inoculation amount1%.After optimization,the strain OD₆₀₀was 3.419,which was two times as much as that before optimization.The inhibition circle of the optimized fermentation broth against tobacco wildfire pathogen was increased from 22.29 mm to 24.54 mm.3.The filter paper confrontation experiment indicated that there was no inhibitory effect between the six strains of antagonistic bacteria,and the affinity was good.The inhibitory effect of mixed antagonistic fermentation broth on Pseudomonas syringae was better than that of single antagonistic bacteria.Strain CDY8-11 has strong biofilm forming ability.The experiments showed that the bacterial content in rhizosphere soil appeared a trend of first increasing and then decreasing,with a bacterial content of 9.3×10⁶cfu/g on the 3rd day.4.The results of dish experiments showed that the fermentation broth of strain CDY8-11 promoted tobacco seed germination,with a seed germination rate of 89.2%.The results showed that compared to spraying the fermentation broth of wildfire pathogen alone,spraying the fermentation broth of CDY8-11+wildfire pathogen increased the activities of POD,CAT,PAL,and decreased the content of MDA,indicating that strain CDY8-11 can induce tobacco resistance.The agronomic traits of tobacco plants treated with fermentation broth of strain CDY8-11 were significantly higher than those of the control,and the fresh weight of a single leaf increased by 93.97%.The pot experiment showed that strain CDY8-11 had a good biological control effect,with a control effect of 69.41%against tobacco wildfire disease.In summary,strain CDY8-11 has application value in controlling tobacco wildfire and promoting tobacco growth.