| At present,the fruit and vegetable reduction in production because of the disease.The traditional and effective method is to spray chemical reagents,but it can cause a lot of harm,such as pollution environment,human health,plant disease resistance and so on.Therefore,it has become a trend to find an effective method to replace chemical control.In this study,we studied the control effect of berksteria monobacterium on strawberry fruits after harvesting.By separating and purifying the antibacterial active substance of this biobacterium,and studying its characteristics,the antibacterial mechanism of B-1 was preliminarily studied.The results are as follows:(1)We study the influence of the postharvest storage of the fruit by naturally decompose and in wound inoculate method.Including the different fermentation times,different treatment methods and different treatment solutions on strawberry.And we study the character of the strawberry on the postharvest storage by sprinkling the B-1 on the fruit in the preharvest period.This experiment showed that the fermented liquid of inhibition rate of 24 hours was decreased by 55.04% compared with the CK,the liquid of 3 minutes immersing was decreased by 46.74% compared with the CK.The inhibition of gray mold of fermentation solution,diluted 5 times fermentation broth and bacterial suspension on strawberry were 100%.And the character of strawberry was no influence before sprinkling B-1 on the preharvest period.(2)The separation and purification of the B-1.We split the bacterium suspension and treat with saturated ammonium of 80%.We separated and purified the protein,which has antimicrobial activity,by anion exchange chromatography and G-100 dextran gel electrophoresis.The SDS-PAGE analysis indicated the protein was single stripe and its molecular mass was 17.6KDa.We studied the antifungal protein by the MS and the possible peptide fragment is M.AHTLPPLPYAEDALAPTISLETIQYHY GK.H.(3)We conduct an experiment about antifungal stability.The results indicated that it has good tolerance to acid.And it is not sensitive to some organic reagent and protease.The protein still has antifungal effect under UV for 2 hours.The protein is sensitive to the reducing agent,for example the DTT and mercaptoethanol.The 50 mg/mL of twain-80 can increase antifungal activity by 12.5%.(4)The antifungal protein has bacteriostasis to the most of bacteria that cause disease.With the decrease of concentration,the antibacterial activity decreased.The inoculation experiments on the fruit in vitro showed that,the antibacterial protein also has good bacteriostatic effect on the gray mould on the fruit.When storage was 12 d,the incidence of fruit was less than half.By scanning electron microscopy and laser confocal electron microscopy to observe the effect of antimicrobial protein on the grey mycelia,and we found that the hyphae treated with antibacterial protein,with CK ratio,its appearance was destroyed and its DNA content,mitochondrial membrane potential and active fluorescent substance were all reduced under the dye of fluorescent substance From the microscopic perspective,the antibacterial decoction can have an effect on the growth and development of gray mould silk.(5)Toxicological safety evaluation of B-1.We used animal experiments to in experiments on the safety of the strain’s virulence.The strain of this strain showed that when the maximum dose was reached,the animal still did not die,indicating that it was a low-toxic substance.In the experiment of cystoscopy,the stimulus intensity of the skin was 0 and did not produce sensitization,and the sensitization intensity was I.In the eye stimulation experiment,the stimulus score of the eye was 0,and the stimulation of the eyes after 48 h was still 0.The study indicated the safety of B-1,indicating the value and significance of the further development of B-1. |
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