| The fungus strain HXDC-1-2 Bipolaris yamadae was isolated from the infected Microstegium vimineum in Weeds Research Laboratory,Nanjing Agricultural University,which could be used to biocontrol common gramineous weeds such as Echinochloa crusgalli(L.)Beauv,Leptochloa chinensis(L.)Nees,Digitaria sanguinalis(L.)Scop,Setaria viridis(L.)Beauv,Alopecurus aequalis Sobol and annual Cyperus rotundus L.It has the potential to be developed into a bioherbicide.The technology of conidial production by liquid-solid fermentation has been developed at laboratory level,and it still suffered from the high cost of medium,and the uncertain technical parameters for mass-production fermentation at large industrial scale.In addition,the study on the shelf life of microcapsules had not been carried out.Hence,the liquid fermentation media were screened and optimized to reduce the cost and clarify the submerged fermentation processing parameter and the study on the shelf life of microcapsules was conducted in order to provide technical basis for the commercial development of Bipolaris yamadae as a bioherbicide.The submerged fermentation medium used by Bipolaris yamadae at the early stage was potato,glucose and water medium(PDB),which is costly at large-scale mass-production due to a larger amount of potato.Corn flour,soybean flour,rice flour,potato protein,potato starch and other materials were used to substitute for PDB medium to optimize fermentation medium.The mycelial and conidial production was used as selection criteria.The results showed that 50 g potato + 6g rice flour + 2g potato protein powder,50 g potato + 6g corn flour+ 2g potato protein powder,70 g potato + 6g potato starch + 2g potato protein powder could be used to substitute PDB as liquid fermentation medium.However,the cost of the latter two media was higher than that of the first one,so the formula of 50 g potato + 6g rice flour +2gpotato protein powder was the best.Submerged fermentation is an important step in the mass-production of Bipolaris yamadae,and the optimization of fermentation conditions may improve the production efficiency of the whole process.Through the liquid fermentation in 5 L fermentor,four factors and four levels of orthogonal experiment was designed to optimize the fermentation parameters such as time,temperature,rotatory speed and ventilation capacity.The effects of four parameters were evaluated by mycelial dry weight and pathogenicity index.The results showed that the optimum liquid fermentation conditions were as follows: fermentation time72 h,temperature 28 ℃,rotation speed 220 rpm and ventilation capacity 360 L / h。Shelf life is an important factor affecting the commercialization of biological herbicides.The shelf life of HXDC-1-2 microcapsule formulation was studied under the storage conditions at 4 ℃-atmospheric pressure,4 ℃-vacuum,room temperature-atmospheric pressure and room temperature-vacuum respectively.The germination rate and pathogenicity of the conidial of the microcapsules were detected every 15 days to determine the optimum storage conditions of the microcapsules.The results showed that the shelf life of the microcapsules was two and a half months.During this period,the germination rate of conidials decreased by 8.7% and 7% respectively under the storage conditions of 4 ℃-atmospheric pressure and 4 ℃-vacuum,and the pathogenicity was maintained at about 80%.However,the germination rate of conidial decreased by 28.8% and 26.7% under the conditions of room temperature-atmospheric pressure and room temperature-vacuum respectively,and the pathogenicity decreased to about 60%.Therefore,the microencapsulation of Bipolaris yamadae is more suitable for preservation at low temperature.After two and a half months,the conidial germination rate of the four storage conditions decreased significantly and maintained at about 30%.At this time,the conidial pathogenicity of the microcapsule was very low.In view of the fact that both low temperature and normal pressure and low temperature and vacuum can maintain high germination rate and high pathogenicity in two and a half months,and the difference is not significant,4 ℃-atmospheric pressure preservation method can be used as the market application storage method in its shelf life. |
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