Please use this identifier to cite or link to this item:
Optimization of culture conditions for production of Bacillus amyloliquefaciens strain WF02 and evaluation of its biocontrol efficacy
biocontrol,Bacillus amyloliquefaciens,optimization of culture conditions,response surface methodology (RSM),black rot of crucifers,
|Publication Year :||2019|
|Abstract:||液化澱粉芽孢桿菌WF02菌株分離自台中霧峰山區，已被證明具有防治Ralstonia solanacearum所引起之番茄細菌性青枯病以及Acidovorax avenae subsp. citrulli 所引起之瓜類細菌性果斑病的功效。在前期試驗發現WF02對於病原菌的拮抗能力與其活菌數呈正相關，因此本研究欲開發出能符合經濟效益，且在最短單位時間內可獲得最大生物量的培養基組成與發酵條件。初期利用三角搖瓶培養系統進行培養基組成與菌體生長條件的因子探討，篩選出糖蜜與硝酸銨作為理想的碳氮源組成，pH值的範圍則介於7～9之間，最適培養溫度則是落在37℃。使用桌上型全自動控制發酵槽針對pH、溶氧（DO）、攪拌速率等參數進行調節控制，以回應曲面法數學統計模型進行最佳發酵條件預測，藉由因子交互作用分析、因子交互作用最佳化等試驗使WF02能迅速達到生長增殖高峰。實驗結果發現WF02的生物量與糖蜜濃度、硝酸銨濃度、pH值間的關係與二階模型吻合，在最適條件下WF02活菌數可達109 cfu/ml以上。使用該培養基組成與發酵條件約提升了60％的WF02生物量，生產成本也降低了90％。此外，相較於先前試驗所使用的LB發酵產物，此優化發酵產物在對抗十字花科黑腐病之生物防治效率亦提升了17%。未來將結合本研究所建立之發酵技術與製劑化開發，提升菌株的儲架壽命與防治效果，將有助於WF02的商品化。|
Bacillus amyloliquefaciens strain WF02 was isolated from Wufeng mountain, Taichung city. It has been demonstrated that WF02 could control tomato bacteria wilt caused by Ralstonia solanacearum, as well as control bacterial fruit blotch caused by Acidovorax avenae subsp. citrulli. In my preliminary data indicated that the antagonistic activity of this bacterium was positively correlated with its biomass. Accordingly, the purpose of this study was to develop the optimal fermentation conditions that are economically viable, and that yield maximum biomass in the shortest unit of time. In the initial stage of optimization, the shake-flask culture system was used to investigate the factors associated with cell culture conditions. The molasses and ammonium nitrate were selected as the ideal carbon and nitrogen sources. The pH level ranged from 7 to 9, and the optimum culture temperature was 37°C. A desktop bioreactor was used to control the environment and culture parameters, including pH/ dissolved oxygen (DO)/ stirring speed etc. during fermentation. Then, I employed response surface methodology (RSM) to design the experiments for predicting optimal fermentation conditions. Optimize the WF02 growth to the maximum point with the variables interaction analysis and variables regression. In the results, I found that the relationship between biomass and the parameters, such as concentrations of molasses and ammonium nitrate, and pH were fitting a second-order model. Under the optimum conditions of fermentation, the maximum biomass productivity could be above 109 cfu/ml. The use of this medium composition and fermentation conditions increased the biomass of WF02 by approximately 60%, and the medium cost was also reduced by 90% in comparison with that used previously. Furthermore, the biocontrol effectiveness of the optimized cultural medium against black rot of Chinese cabbage caused by Xcc425 was enhanced (17% increased) in comparison with that of LB cultural medium. In the future, we would like to combine the fermentation technology established by this study with the formulation development to improve the shelf life and biocontrol efficacy for the commercialization of WF02 biocontrol agent.
|Appears in Collections:||植物病理與微生物學系|
Files in This Item:
|2.74 MB||Adobe PDF|
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.