The Research On The Extraction Of Crude Toxin From Colletotrichum Gloeosporioides In Citrus And It's Pathogenicity

Anthracnose is an disease that affects the postharvest storage of citrus seriously.Colletotrichum gloeosporioides is the main pathogenic bacterium that belong to Colletotrichum of Deuteromycotina,which has strong pathogenicity.It can cause huge economic costs by make storage citrus decay.The study of pathogenic mechanism of toxin from C.gloeosporioides is few,most of the researchers are focus on the growth characteristics and screening antagonistic bacteria of C.gloeosporioides.According to foreign research,p H of the environment,secretion protein of pathogens and pathogenic genes are the virulence factors of C.gloeosporioides.Pathogenicity will be change when the virulence factors were changed.However,these study are focus on the production conditions of toxin,structural analysis and whether it has a host specificity.In this research,the optimum culture medium of C.gloeosporioides was studied to explore the optimal toxicant factor,the crude extract of toxins was used to study the effects of crude toxin on the physiology metabolism in post-harvest citrus to explore the effect of toxins,and the induced resistance to postharvest citrus by p-Hydroxybenzoic acid is to find a new way for citrus storage.The main conclusions were as followed:1.The C.gloeosporioides were cultivated in vitro,types and compositions of the medium were changed to study the optimum culture medium.The study shows: toxins produced by Czapek's medium can form larger lesions on citrus.Changing the type of carbon,nitrogen and inorganic salts in the Czapek's medium can affect the formation of toxins,the best carbon source is fructose,the nitrogen source is sodium nitrate,and the inorganic salt is dipotassium hydrogen phosphate.The optimal culture conditions were 30 ??19 days?100 r /min?p H 8 through the study of four factors and four levels orthogonal experimental,and the lesion diameter is 5.39 ± 0.22 mm under this condition.2.The crude extraction of toxin was studied by comparing the ammonium sulfate fractionation and organic solvent extraction of different polar organic solvent,supernatant obtained after methanol extraction can produce the largest lesion in citrus.The main components of crude toxin contains ketoses,protein or amino acids and other substances bases on the analysis of chemical properties.High temperature and changes of p H will affect the activity of crude toxins.3.The crude toxin was prepared at different concentrations to study the effect of crude toxin on the microstructure of citrus peel.It was concluded that the crude toxin can bring greater damage to citrus fruit cells as the concentration was increased or the incubation time was prolonged,peel cells showing shrinkage and rupture in this condition,the reason may be that the crude toxin contains enzymes that can hydrolyze the cell wall and some other pathogenic ingredients to form large lesion in citrus.4.The results in the study of the effects of crude toxin on the physiological metabolism of citrus can be shown that typical lesion were observed in the toxin test,the content of malondialdehyde were significantly increased,while the content of cell wall compounds were decreased.When treated with a low concentration of crude toxin,there was a positive correlation between the content of flavonoids compounds,lignin,the activities of polyphenol oxidase,peroxidase and lesion diameter,the correlation between cell wall compounds and lesion diameter were negative.However,the defense enzyme system and citrus fruits were damaged when the concentration of crude toxin was higher,that made the activities of enzymes rapid decreased.This demonstrate that physiology metabolism of citrus fruits can be influenced by the crude toxin of C.gloeosporioides,and that improves toxins plays an extraordinary important role in the pathogenicity of C.gloeosporioides.5.p-Hydroxybenzoic acid can induce the resistance of postharvest citrus fruits and prolong the shelf life of citrus.Through the study we can know that p-HBA displayed strong antifungal activity to C.gloeosporioides,when the concentration was higher,the effect was increased.The activity of PPO,POD,SOD and PAL was increased when inoculated with p-HBA and C.gloeosporioides,while the content of MDA was decreased.p-Hydroxybenzoic acid has a strong antimicrobial activity,it can induce the increase of defending enzyme's activity and decrease the peroxidation of membrane lipid,the significance obviously is there for citrus fresh-keeping.