The Isolation Of Antagonistic Microbe For Controlling Fusarium Wilt Of Strawberry And Study On Its Fortified Bio-organic Fertilizer

Fusarium wilt of strawberry（FWS）caused by Fusarium.oxysporum f.sp.fragariae（FOF）is one of the most serious soil-borne fungal diseases in the strawberry production,which threat to the sustainable and healthy development of the strawberry production worldwide.At present,management of fusarium wilt is principally achieved through the use of chemical pesticides and solar heating of soil pasteurization.However,the efficacy of those traditional control managements are limited and those traditional methods have caused some problems,such as pesticide residue,food safety and environmental pollution.Therefore,the efficient and sustainable method is needed to control fusarium wilt of strawberry.To solve this problems,the objective of this study is to develop a bio-organic fertilizer（BOF）for controlling fusarium wilt of strawberry and investigate its possible mechanism of action for providing a theoretical basis.The main results are as following:（1）In this study,two antagonistic strains X-1 and Z-1 were screened from the rhizosphere soil of healthy strawberry plants by confrontation culture method.The inhibition rate of strains X-1 and Z-1 were 75.7%and 70.2%,respectively.According to the results of morphological,physiological,biochemical characteristics and analysis of 16S rRNA sequences,X-1 and Z-1 were identified as Bacillus licheniformis and Bacillus methylotrophicus,respectively.The results of inhibition spectrum showed that two strains had wide range of antibacterial effect.The effects of different culture conditions on the growth of X-1 and Z-1 strains were also studied in this paper.（2）An aerobic composting system was constructed using pig manure as material and adding maize straw as conditioner.The temperature above 50℃of aerobic composting system was maintained for 9 days in the process of composting.The physiological,biochemical and biologic characteristics of aerobic composting in the thirtieth days were examined as follows:the pH value was 7.9,the moisture content was 26.47%,the EC value was 4.102 ms·cm^-1,the carbon and nitrogen ratio was 11.8,the germination index was86.55%.Those above characteristics met the standard on evaluation of compost maturity.The suspension of X-1 and Z-1 were mixed（1:1）and stirred with manure at the rate of 10%（v/w）for secondary fermentation.The component indexes of BOF reached the production standards of bio-organic fertilizer in China.（3）To evaluate the control and growth efficacy of BOF fortified with antagonist against fusarium wilt of strawberry,pot experiments were conducted.The results of control pot experiment suggested that the application of BOF was significantly effective to control FWS with the 75.39%biocontrol efficacy.The biocontrol efficacy BOF treatment were increased by 71.20%and 52.56%,compared to the chemical fertilizer and organic treatment.Meanwhile,it was shown that BOF can not only significantly increased the plant height,root length,fresh weight and dry weight,but also raise the chlorophyll,soluble sugars and soluble proteins content of strawberry leaves and enhance root activity.（4）The possible mechanism of BOF was investigated.Five antibiotic biosynthesis genes（ituA,ituB,ituD,bamB and sur）were detected in strain Z-1 and bamB gene involved in Bacillomycin biosynthesis was also detected in the strain X-1 based on PCR-assays.Therefore,it was possible to synthesize lipopeptide antibiotics in antagonistic bacteria X-1and Z-1 for the suppression of FOF growth.Under scanning electron microscopy,noticeable morphological changes,such as hyphal swellings and hyphal collapse,were found in the hyphae of FOF in the presence of the antagonistic bacteria X-1 or Z-1.Strawberry seedling leaves treated with BOF exhibited significantly higher activities of SOD,PPO,CAT and POD,which suggested that applying BOF enhanced the systemic resistance in strawberry seedlings through induction of defense enzymes.Bacterial and fungal community structure and composition analysis showed that an application of BOF could increase the abundance of beneficial bacterial groups and decrease in the abundance of Fusarium.There was a significant difference in microbial community structure of rhizosphere soil in BOF treatment,when compared with CK,CF and OF treatments.