| Phytophthora capsici Leonian is one of soil borne diseases, which is prevalently occur and seriously reduce pepper yield in conservation field. As a result of continuously cropping, flood irrigation and chemical abuse, the accumulation of pathogen source is increasing, the problem of pesticide pollution is aggravating and the micro ecology environment is liable to be broken. At present, the production of biocontrol agent is scarce and the effective biocontrol technology is absent. In the present study, the researches on genetic polymorphism, biocontrol strain and micro ecology biocontrol are carried out for pepper Phytophthora blight. The results are as follows:1. The analysis of morphology and rDNA-ITS homology for Phytophthora capsici showed:the various strains collected from different regions of Liaoning province are similar on the configuration. Most of mycelial branches are right or acute angle. The sporangiophore is colorless and filamentous. The sporangium is single, ovoid or pyriform, and the mastoid is single. The sequences of rDNA-ITS change from805bp to877bp. Clustering analysis show that those strains collected from different regions are obvious homology, and there are not regional characteristics.2. Using Sequence Related Amplified Polymorphism (SRAP) technology, the genetic polymorphism of Phytophthora capsici Leonian collected from different regions of Liaoning province is evaluated. The results show that the genetic similarity of Phytophthora capsici Leonian is very high and the strains have abundant polymorphism, while the regional diversity is not significant. The pathogenicity determination show that these strains are divided11pathogenic types, which of the intermediate type is primary. There are not significant correlations between the genetic similarity and pathogenicity.3. Fifty-six samples collected from different fields are separated by plate dilution method. The ASD strain is screened and has significant resistant to Phytophthora capsici Leonian by bacteriostatic test. The width of inhibition zone is11.7mm. The control efficiency of ASD strain reaches to75.12%against Phytophthora capsici in pot experiment, which is better than80%WP Mancozeb300mg/L and50%alkene acyl complexes of water dispersible granules20mg/L 4. The morphology and molecular determinations show that the ASD strain is velvet on PDA medium. The early myceliums vary from white to khaki, occasionally exude transparent liquid. The conidium is spherical, having small stems and arraying as catenulate type, which of top sporangium is radiate. The rDNA-ITS sequence of ASD strain is621bp, which of97%is similar to Aspergillus flavipes. According to the above result, the ASD strain is determined as Aspergillus flavipes.5. The study of resistance mechanism shows that the biological control ASD strain can produce antibacterial substances, which can cause mycelia abnormality and inhibit the pathogen growth. The activities of CMC and xylem enzymes are significant improved by the biological control ASD. With the increase of substrate concentration, the enzyme activities are improved. The main factors inhibiting the growth of Phytophthora are CMC and xylanase enzymes, while there is little relation with β-1,3glucanase and protease.6. The results of optimizing fermentation condition show that the order of carbon source induced ASD strain to produce antibacterial substances is monosaccharide, disaccharides and polysaccharides. The glucose is the most optimize carbon source. The organic nitrogen is significantly superior to the inorganic nitrogen. The best organic and inorganic nitrogen are glycine and ammonium nitrate, respectively. KCl, MgSO4·7H2O, FeSO4are benefit to produce the antibacterial substances when the concentrations are low. Combining with orthogonal test, the optimal condition is2%Glucose,0.1%Glycine,0.003%FeSO4,0.01%KCl, MgSO4·7H2O, pH6,180rpm,30℃, the Volume30mL/100mL, culturing for5days.7. The analysis of micro ecological system shows that the varieties of numbers of bacteria, actinomycetes and fungus are not significant, which collected from capsicum rooting zone soil of the healthy and infected plants in different regions and various years,. The numbers of bacteria and actinomycetes in capsicum rooting zone soil of healthy plant are more than those in rooting zone soil of infected plants, while the number of fungus is lower. The scales of bacteria/actinomycetes, fungus/actinomycetes and bacteria/fungus in rooting zone soil of infected plants are higher than those in rooting zone soil of healthy plants.8. By the morphological observation, physiological and biochemical test and16s rDNA sequence analysis, seven types of dominent bacillus are identified, which is43.75%of total bacteria. There are17soil funguses. The strains of B33、B40、B22and B44are only determined to genus level. The further study should be carried out in the future. 9. Research on the effect of micro ecological system in soil, the results show that the biocontrol strain ASD could significantly reduce the quantity of the bacteria and actinomycetes, and increase the number of the fungi in capsicum rooting zone soil of the healthy and infected plants. With the colonization time increase of the biocontrol strain ASD, the number of the bacteria is gradually increased in capsicum rooting zone soil of healthy plants, while it is not changed in the capsicum rooting zone soil of infected plants. The number of the actinomycetes is not changed in capsicum rooting zone soil of the healthy and infected plants, while the number of the fungi is gradually reduced.10. Research on the effect of soil enzyme, the activities of urease, acid phosphatase, sucrose enzyme and dehydrogenase are significantly increased by the biocontrol strain ASD in capsicum rooting zone soil of the healthy and infected plants, while the activity of catalase is not significant increased. With the colonization time increase of the biocontrol strain ASD, the activities of various enzymes are higher than the control and their variation trends are very similar, which firstly increased and decreased afterward. The responses of soil enzyme activities affected by the biocontrol strain ASD from high to low are as follows:urease, dehydrogenase, sucrose enzyme, acid phosphatase and catalase. The results showed that the biocontrol strain ASD could improve the micro ecological systems in soil. |
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