| Mosquitoes could cause severe illness of human and were the enemies of human health. In order to control them, a method with heat treatment was used to isolate Bacillus thuringiensis (Bt) with high toxicity against mosquitoes from many regions of Fujian province of China, including Wuyi Mountain. 71 isolates were obtained from 125 soil samples. The results showed that five isolates (QQ13, QQ17, QQ42, QQ66 and QQ92 ) were effective against Culex quinquefasciatus, three of which (QQ17, QQ66 and QQ92) showed high mortality. They might effectively promote the biocontrol of C. quinquefasciatusDuring sporulation, Bt synthesizes proteolytic enzymes. However, whether the protease profile of a Bt is related to the efficacy of the ICPs is still a subject of controversy. In order to investigate the role of specific protease of Bt in cell growth and in crystal protein stability and explore the possibility of the proteases to be used in the industry, PCR primers specific for the conserved regions of protease genes were designed and several PCR products were cloned and analyzed. The results showed that six protease genes, neutral protease A (nprA), tryptophan synthase beta chain, tryptophan synthase alpha chain, alkaline protease A (aprA), phosphohydrolase gene and glycogen phosphorylase, with known functions are identified from Bt subsp. kurstaki 8010, respectively, were obtained. In addition, a DNA sequence with unknown function, which may be Bacillus cereus group-specific gene, was also detected. A very simple and rapid method for extracting genomic DNA from Bt was also described.Recently, the ways to increase the efficacy of Bt have been hot spots. And research and development of new pesticide adjuvants plays a key role. Good pesticide adjuvants could improve physical and chemical characteristics of pesticide and increase the efficacy of Bt products. In current study, we had investigated 5 pesticide adjuvants for their potential usage in pesticide industries. Among the adjuvants, both BCM-101 and JFC could improve the wettbility of Bt preparation and enhance its activity to Heliotis armigera. In order to increase the efficacy of Bt, a new method of adjuvant-selection was proposed.In order to explore the effects of treatments after fermentation on Bt products, optimize the treatment procedures after fermentation, and design the regulation methods of fermentation, we have collected the data of Bt industrial treatment procedures after fermentation to lay a foundation on automatic fermentation. The results showed that as far as the indusrial Bt strain was concerned, the best air and media ratio was 1:1-1:1.2 and the good fermentation temperatures were 30-33℃. The pH values of fermentation liquid varied during the fermentation. In the treatment procedures after fermentation, membrane filtration methods were often used to produce 16000IU/mg Bt products while plate-frame pressure filtration methods were frequently applied in the production of 12000IU/mg Bt products. And the former plays a key role in the procedures. Many elements during spray-drying procedures, including membrane filtration and spray-drying time, could indirectly affect the efficacy of Bt products.The quality and yield of the tea were significantly improved after using TA-BR in combination with Bt. Tea yield could increase by 6.76%～26.90% (per mu), the proportion of the amino acids and the tea polyhenols was obviously reduced, the water extracts,the soluble sugar and Vitamin C of tea were higher than that of CK. In addition, the motability of TA-BR in combination with Bt on the Euproctis pseudoconspersa, Ectropis obliqua hypulina, Andraca bipunctata, which were the main pest damaging to tea plants, could reach above 90%. It is safe for natural enemies.In summary, the isolation and identification of Bt strains and toxin-related genes, the screen of pesticide adjuvants, the monitor of fermentation procedures and the research and development of TA-BR in combination with Bt had been characterized in this studies. In chapter 2, we obtained 3 Bt isolates with high toxicity against C. quinquefasciatus. This work might lay a foundation on biopesticides products. In chapter 3, we obtained 6 DNA fragments of protease genes and a gene fragment, which might be B. cereus group specific genes, by primer pairs specific for the conserved regions of Bt protease genes. In chapter 4, in order to improve the efficacy of Bt, several pesticide adjuvants had been tested in a Bt factory to find adjuvants with good physical and chemical characteristics, better wettability and adhesion ratio. In chapter 5, we developed a product, TA-BR in combination with Bt, to improve the insecticidal efficacy and promote the yield and quality of tea. This product could greatly promote the development of unharmful tea and make benefits of zoology, economy and society true. Since Bt plays a key role in the biocontrol of pests and plant diseases, the above finding might lay an important foundation in the further work. |
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