Study On Bacillus Subtilis Sporulation And Drying Temperature Optimization

Bacillus subtilis microbial ecological agents are widely used in animal breeding,plant disease prevention and water purification due to their strong adaptability and stress resistance through spores formation.At present,research on Bacillus subtilis additives mainly focuses on optimizing culture conditions to increase the effective biomass.However,study on changes of physical and chemical properties of medium and molecular mechanism of sporulation during preparation process has been relatively poor.In this study,dynamic changes of physical and chemical properties of medium and the expression of spore forming genes were studied.The relationship between medium properties,genes expression with the number of viable cells and spores were explored.Besides,drying temperature was optimized to improve the effective biomass of Bacillus subtilis additives.The results are as follows:（1）During solid-state fermentation process,viable cell number of Bacillus subtilis increased rapidly in the first 48 h,which reached 8.34×10⁹ cfu/g.Glucose in medium was consumed effectively during the fermentation time of 24-48h..Glucose content decreased from 13.6 mg/g to 6.38 mg/g The structure of medium was destroyed,so the T₂₁1 decreased.While during the fermentation time of 48-72h,the number of spores increased and the sporulation rate reached 72%.The utilization of nutrients in medium became weak and medium properties changed slightly.（2）After fermentation for 48 h,the number of spores continuously increased,whereas the number of vegetative cells decreased.Expression of sigF gene increased in the middle stage of fermentation while that of spo0A and sig E genes increased in the late stage of fermentation.Expression level of sigF gene after fermentation of 48h was 1.58 times of that of 24h.Expression level of spo0A gene after fermentation of72h increased by 1.92 times than that of 24h.Expression level of sigE gene increased by 89%than that of 24h.（3）Bacillus subtilis additives were dried by hot air at 60℃,70℃,80℃and90℃.Results indicated that effective biomass after drying at 80℃was significantly higher than that at other temperatures,which reached 4.85×10⁹ cfu/g.Within a certain range,glucose content increased with the drying temperature increase,but the glucose content began to decrease after drying at 90℃for 2 hours.Therefore,drying at 80℃for two hours is a suitable condition for preparing a Bacillus subtilis additives by hot air drying.