Fermentation Optimization For Bacillus Thuringiensis BtZ01 And Its Impact On Growth And Development In Embryonic And Post-embryonic Periods Of Mice

Bacillus thuringiensis (Bt) has a long use history as a biological pesticide, which has always been the focus of attention because of its high efficiency, safety, environmental friendly and other features. As a commercial strain, we must to ensure its efficiency, production and safety when use. BtZ01 is a strain preserved in our laboratory with its highly active insecticidal. In order to make it commercialization as soon as possible, we need to optimize its fermentation conditions and evaluate its safety, to provide more accurate data for its production and application.(1) The quadratic regression rotation combination design was used to optimize carbon, nitrogen and inorganic salts added amount of fermentation medium, while some fermentation conditions. Mathematical models of the relations between raw materials and bacteria amount was established, the single factor effect and interaction effects were analyzed. The most appropriate fermentative media was composed of corn meal 3.61%, soybean meal 3.46%, fish meal 1.03%, Yeast powder 1.13%, Peptone 0.18%、KH2PO4 0.65%、FeSO40.21%, CaCl20.21%, AL2(SO4)30.33% and MgSO4 0.23%. The optimized fermentation condition was determined as following:pH of medium was adjusted to 7.5 before sterilization, incubated at 37℃ for 44 h～50 h after inoculating 3% seed. Under this condition, the viable bacteria reach to 2.7 × 1010 CFU·mL-1 and Spores ratio reach to 80%.(2) To investigate the impact of Bacillus thuringiensis BtZ01 on growth and development in embryonic and post-embryonic periods of mice. Forty eight pregnant Kunming mice were randomly assigned into four groups. The control group was fed with sterilized water and the treated groups were fed with sterile saline containing 106 CFU·mL-1,108CFU·mL-1,5×109 CFU·mL-1 BtZ01, respectively. In the present study, the body weight and organ weight ratios were measured, and as well as bone malformations and microscopic pathology were examined. Results indicated that there were no significantly differences between the control and the treated groups for mice’s length and weight gain (P>0.05). The malformations ratios of the control and the treated groups were not different (P>0.05). However, the fontanels of high-dose group (5× 109 CFUmL-1 BtZ01) were wider and their kidneys were changed, compared with the control groups. There were no significant difference of intestinal morphology in 18 days mice (.P>0.05), but the intestinal villus height and villus length/crypts depth ratio of high-dose group and medium-dose group mice in 48days was significantly higher than other groups(P<0.05). The imbalance phenomenon of microflora was also not appeared. Generally, our results showed that the Bacillus thuringiensis BtZ01 has no adverse or toxic effects on embryonic and post-embryonic mice in a dose of 108 CFU·mL-1.