Toxicity Test And Control Efficiency Of Bacillus Amyloliquefaciens Ba168Strain On Botrytis Cinerea

Tomato gray mould as a cosmopolitan fungal disease caused by B. cinerea that inflicsserious production losses and quality decline on tomato. At present, chemical control hasexposed a few defects and B. cinerea generated fungicides resistance commonly, and there isno any resistant variety for tomato gray mould was planted in the field, so it is a eximiouschoice for biological control of tomato gray mould.Strain Ba168was isolated and screened from the soil of Qinling virgin forest by ourlaboratory. With the following identifications of colony morphology, physiological andbiochemistry experiments and16S rRNA gene sequence as well, the strain was identified as B.amyloliquefaciens and showed B. cinerea antagonistic activity. This study consisted of twoparts:(1) Optimization of fermentation conditions for producting antimicrobial substances of B.Amyloliquefaciens Ba168strain. First, the growth curve was drawn that assured theinoculum’s age between20hour and23hour. Second, exploring the ralationship between thespores production and the antifungal activity and its result showed there were positivecorrelation of them. Then, taking spores production as measurement index, FermentationConditions of Ba168strain was optimized by comprehensive use of the single factorexperiment, the Plackett Burman Design, the method of steepest ascent and the Box-Behnkencenter-united experimental design as well as the method of response surface analysis. Theoptimum conditions were, corn meal1%, Soybean cake powder1%, KH2PO40.11%, MgSO40.05%, initial pH7, incubation age20hour, inoculation dosage5.77, reaction temperature32℃, bed rotation speed220r/min and the culture time of48hour. Finally, compared beforeand after the optimization of fermentation conditions to produce spore number and antifungalactivity. The results show that the strain Ba168optimized fermentation condition, improvedits antagonism activity of fermentation liquid on B. cinerea.(2) Toxicity test and control efficiency of B. amyloliquefaciens Ba168wet powder on B.cinerea. First, Inhibition rates of B. Amyloliquefaciens Ba168WP against fifteen pathogenicfungus were tested on potato dextrose agar, and the mean of antagonistic diameter betweenBa168strain WP and fifteen pathogenic Fungus were ranged from18.61mm to26.38mm.Second, five fungicides such as1×109CFU/g Ba168strain WP,1×109CFU/g Bacillus subtilis WP,40%carbendazim WP,50%iprodione WP and40%pyrimethanil WP for theinhibitory effect of growth of the hyphae were determinedby using hyphae growth ratemethod, and the value of50%effective concentration (EC50) were respectively946.28ug/mL,757.19ug/mL,75.36ug/mL,8.95ug/mL and5.53ug/mL. Third, five fungicides for theinhibitory effect of the germination of conidia were tested by using concave slide glass, andthe EC50were1665.56ug/mL,1416.16ug/mL,241.90ug/mL,37.32ug/mL and21.35ug/mLrespectively. Then the control effect of five fungicides in greenhouse experiment were tested,and the control effect of five fungicides(diluted750times solution) were68.47%,70.14%,74.18%,74.13%and80.63%respectively. Finally, the control effect of five fungicides in fieldexperiment were obtained, and the control effect of five fungicides(diluted750times solution)were63.37%,64.83%,69.56%,73.49%and75.55%respectively.Ba168strain WP haveextensive inhibitory spectrum of fungi, the results of indoor toxicity and the control effect offield experiment of Ba168strain WP were lower than that of selected three chemicalfungicides, but it is equivalent to that of B. subtilis WP. Therefore, Ba168strain WP can beviewed as assistant fungicides to control tomato grey mould.