Studies On The Formulation Of Biocontrol Agents Trichoderma Harzianum T4and Bacillus Subtilis B99-2and Biocontrol In The Green House

In order to solve the problems of short shelf life of Trichoderma harzianum T4and Bacillus subtilis B99-2formulations, new formulations of microbial pesticide is prepared for industrial production and agricultural application. This study aim to produce chlamydospores of Tirchoderma harzianum T4, develop the formulation of biocontrol agent T. harzianum T4and Bacillus subtilis B99-2, and investigate the biocontrol effect.In order to explore the T. harzianum T4liquid culture conditions to produce chlamydospores, application of response surface methodology (RSM) revealed that the optimum medium was carbon sourse A23.85g/L, calcium carbonate1.0g/L, carbon sourse one4.83g/L, ammonium sulfate2.0g/L, and potassium dihydrogen phosphate1.0g/L. By optimizing of dispersant dosage, inoculum and fermentation days, acquired T. harzianum T4fermentation process was Twain800.5%, inoculum of fermentation of2×107cfu/mL, and fermenting for6days. The maximum free chlamydospores could reach1.55×107cfu/mL, while the amount of chlamydospores was3.35×10cfu/mL.In this paper, according to experiments of biocompatible and water dispersible granules, select the suitable UV protection agents, stabilizer, binder, disintegrant for T. harzianum T4, and acquire the best formulation was silica10%, UV protection agent A0.4%, stabilizer B3%, EFW4%, disintegrant C4%, binder D5%. Preparation process as mix silica with UV protection agents, stabilizer, disintegrant, and then add mixture to fermentation broth, dry to a moisture content of about20percent under37℃, add5%aqueous binder solution, the final moisture content of solids was about25%, Then products was granulated and dried. Bacteria of Water dispersible granules of T4was8.5×108cfu/g, and suspension rate of that was43.8%, wetting time1.0s, disintegration time52s. Water dispersible granules (WDG) could rapidly spread in water and avoid dust with the advantages of both WP and aqueous suspension concentrate. This provided a basis for industrial production.This study was to optimize the method of encapsulated silica white-Bacillus subtilis B99-2mixture in alginate microspheres. It could not only improve the encapsulation efficiency of microbial cells, but also reduce the preparation and storage cost. The experiment was designed as follows, bacillus subtilis B99-2was adsorbed by carrier A, informing carrier A-B99-2mixture, and then microcapsules were prepared by emulsion method. This optimized method could improve encapsulation efficiency of microorganisms, and microcapsules could dry well at room temperature (22℃-25℃) during24h in stead of using costly freeze-drying method. Under UV (20W,8h),54℃(14days) and at room temperature (180days), the survival rates of microcapsules of Bacillus subtilis B99-2were60.82%,57.82%and97.56%respectively, higher than that of non-encapsulated Bacillus subtilis B99-2with38.27%,40.27%,36.22%. The release of encapsulated B99-2in sodium bicarbonate-sodium citrate solution was also assessed in this study.In the green house experiment, the water dispersible granules of T4chlamydospore showed40.72%reduction of Botrytis cinerea and the microencapsulation of Bacillus subtilis B99-2showed39.87%reduction of Botrytis cinerea, which was as good as50%procymidone wettable powder and40%pyrimethanil suspension.