Construction The Multiple Screening System Of An Antagonistic Bacillus Sp.Against Banana Fusarium Wilt And Research On Its Control Effects

Fusarium wilt of banana was a kind of soil-borne disease caused by pathogenic fungi Fusarium oxysporum f.sp. cubense (FOC), resulting a great threat to the banana industry and affecting the production of banana seriously. However, the prevention and control against the disease was still a problem to solve. The characteristics of the biological control technology is environment safety, pathogens did not obtain the resistance easily, improving soil quality, promoting crop growth, therefore it becomes the research hot spot. However, the evaluation of bio-control strains mainly depended on traditional screening methods, and screening culture medium with LB, PDA and NA containing rich nutrition mostly, resulting in making mistakes easily.Based on the resource of the laboratory, our purpose was to condunct a comprehensive evaluation and identification on the Bacillus subtilis which had antagonism effect against Fusarium oxysporum f.sp.cubense; after the four tests,the objective strain was identified; finding the suitable liquid fermentation medium for industrial production; researching on the characteristics of the antimicrobial substance and its effect on FOC; evaluating the effect of the treatment with compound composed of fermentation broth containing bio-control strain D34 and organic matrix by pot experiment. The main results listed as follows:1?Taking the nutrition adaptability, compatibility of bio-control fungi, the lipopeptides of the biosynthesis genes PCR and bacteriostatic rate of FOC in the soil as four standards, evaluating 22 strains preserved in the laboratory. The result showed that the strain D34 meet these four requirements.The strain D34 can keep antagonistic activity under the condition of relatively poor nutrition, showing a low antagonist activity on biocontrol strains of Trichoderma and Paecilomyces lilaeimls, having the lipopeptides of the biosynthesis genes (ItuB?ItuD?Bam?FenD?Sur) and the bacteriostatic rate of FOC in the soil is 20.49%.2?Bio-control strain D34 can produce ?-glucan enzyme?pectinase, cellulose and proteolytic enzymes which can make an destruction on fungal cell wall structure, and can produce ACC ammonia-lyase?siderophores and indoleacetic acid which can promote plant growth.3?Combined with microscopic morphology?physiological and biochemical characteristics and the 16s rRNA sequence, identified the bio-control strain D34 as Bacillus amyloliquefaciens.4?Comparing the effect of different carbon and nitrogen source of culture medium on the antagonist activity of the strain D34. Bacillus sp. D34 showed the strongest antimicrobial activity in 9 orthogonal combination A3B3C2(Corn Flour 2%, Bean Flour 1%, Dibasic Sodium Phosphate 0.1%). The fermentation liquor of Bacillus sp. D34 had destructive effects on the pathogen hyphae by microscopic observation.5?The research on the fermentation of Bacillus sp. D34 found that the antagonism active substance can resist high temperature?proteolytic enzymes?ultraviolet irradiation and organic solvents. After hydrochloric acid precipitation combined with methanol, the spore germination inhibition rate and the mycelial growth inhibition rate of the pathogenic fungi of the extraction active substances were 83.87% and 51.29% respectively. The extraction active substances had inhibiting effect on different phytopathogenic fungus, such as F. oxysporum f.sp. niveum?F. oxysporum f.sp. cucumerinum?F. oxysporum f.sp. benincasae?F. oxysporum f.sp. melonis. Using preparative liquid chromatograph combined with the HPLC-ESI-MS, speculating that extraction active compounds contained the homologous of lipopeptides of Iturin(1028.5)? BacillomycinD(1030.5)?Surfactin(1007.6)?Fengycin(1448.8).6?Using rifampicin marked Bacillus sp. D34, no significant differences were found between the mutant strain RD200(Rif 200 ug/mL) and the wild-type strain D34 in colony morphology?growth regulation and the stability of the antagonism activity.7?Compared with water control treatments, the incidence of the treatment with compound composed of fermentation broth containing bio-control strain D34 and organic fertilizer reduced by 60.14%, the number of pathogenic fungus (FOC) reduced by 42.06%, the bananas grew well, the soil respiration rate increased 2.26 times, the number of the soil culturable fungus, bacteria, actinomyces, increased by 51%?27% and 19% respectively.