Studies On Mutation Breeding And Antagonistic Substance Of Antagonistic Actinomycetes Agaist Alternaria Kikuchiana

Pear black spot caused by Alternaria kikuchiana is a serious air spreading disease which distributes widely in the world. 1t is a difficult problem to control the disease effectively. The research focuses on the biocontrol of pear black spot. Therefore, mutagenic breeding and inhibition effects were studied after an antibiotic actionmycete strain was screened and identified. It provided the theory basis and the technical support for the exploiting agricultural antibiotics for biocontrol.Three hundred and seventy two strains of actionmycetes were isolated from the rhizosphere soil samples using dilution method, from which six strains effectively inhibiting Alternaria kikuchiana were screened out by inhibitory zone method. After further screening on fruits, It was found that strain B105 was more effectivly than other antagonistic actinomycetes and its fermentation filtratethe inhibition rate got to 66.5% at 28℃. B105 strain also showed broad antagonism against pathogenic fungi in plant, such as: Physalospora piricola, Penicillium expansum Link, Venturia pirina, Botrytis cinerea et al.The morphological, cultural physiological, biochemical characteristics and 16S rDNA sequences of strain B105 were studied. Its substrate mycelium had no partition, the aerial mycelium were ramose, the spores were oval and the surface was smooth, which consistented with the characteristics of the genus Streptomyces. The analysis of 16S rDNA sequence of strain B105 suggested that it was most closely related to Streptomyces aureus and the sequence identity was 99%. From the polyphasic taxonomical view, the strain B105 falled into Streptomyces aureus.In order to improve the antibiotic-producing, UV, microwave and NTG were used to mut -agenize B105 spores, respectively. The results showed NTG has stronger mutagenic effect than UV and microwave. Under the optimum dosage of 1mg/ml, a high productive mutant strain named N2 was screened out. The diameter of antibiotic zone of N2 increased by 52.2% and the inhibitation effects to AlternAria kikuchiana increased by19.2% compared to B105 strain. Futhermore , its high capacity for producing antibiotic remained stable after 5 times of subcultures and its antifungal spectrum was not altered.The antagonistic substance produced by the strain was considered as one of its antagonistic mechanisms against Alternaria kikuchiana. The antagonistic substance was stable to heat, pH, and illumination, its inhibitory activity could remain 76.5% even incubating at 100℃for 120mins. Accoding to the results of Jack's Eight Solvent System Chromatography and pH Paper Chromatography of the fermentation extracts indicated that the antagonistic substance was basic and water solubility-alkaline antibiotic.The result of biocontrol test indicated that the fermentation filtrate had strong inhibition mycelium growth and spore germination of Alternaria kikuchiana, and the control effects on the diseased fruits and pear leaves were 85.7% and 82.3%, respectively, which were obviously better than 50% Duojunling (500 times dilution). The result indicated that strain B105 was really a potential biocontrol endophytic actinomycete.