Stiudies On High Toxic Bacillus Thuringiensis Strain (TS16),Fermentative Media And Additives

This study mainly concerns the control of vegetable's important lepidopteran pest (Plutella xylostella, Spodoptera litura, Spodoptera exigua) of south china. Eliminating plasmids of Bt strains (WB2, WB3, WB6, 8010) by high temperature treatment or by complex treatment with SDS and high temperature, a highly toxic Bacillus thuringiensis TS 16 was obtained. The electrophoresis profiles of plasmid DNA indicated that the plasmid which is less than 2Kb was eliminated, and the profile of plasmid of 6.6 Kb became thin. The insecticidal activity of TS 16 to Plutella xylostella was noticeably higher (1.5 fold increase) than that of the original strain WB6. Media which was mainly composed of dextrose and peptone were screened with shake flask fermentation and orthogonal test. Results indicated that spore numbers were mainly influenced by carbon and nitrogen instead of inorganic salt. Among inorganic salts, the influence of FeSO4.7H20 is more noticeable than others. So composition of the medium was divided into three factors carbon sources (xi), nitrogen sources (x2) and inorganic salts. With fixed inorganic salts content, the relationship between fermentation media with different carbon and nitrogen sources and spore numbers was demonstrated by using the double factors Orthogonal-rotation-combination-test. A response-plane equation was formed as follows: y1 55.200+1 3.380x1+22.866x2-24.097x12-22.682x22-i-17.575x1x2 It's easy to acquire the best combinatorial medium from this equation. Bacillus thuringiensis was grown in culture media in which different agricultural by-products as the main carbon and nitrogen sources. These products included corn meal, starch, defatted soy four, fishmeal, defatted peanut flour and yeasts. Of this several products tested, fishmeal and defatted peanut flour were the most efficient substrates for Bt. With orthogonal experiment, A good combinatorial medium (defatted soy four 32g/L, corn meal 14 g/L, defatted peanut flour 32 g/L, fishnieal 14 g/L, peptone 4 g/L, yeasts 6 g/L, FeSO4.7H2O 0.04 g/L and ZnSO47H2O 0.08 g/L) was obtained, in which the Bt strain TS 16 can produce high toxicity from the fermentative liquid. Fermentative condition tests indicated that air content, pH value and temperature all effected spore mumbers and efficacy of fermentative liquid. In the medium which mainly consisted of dextrose and peptone, the optimal pH value was 7.2, while composed by agricultural by- products, the optimal pH value was 8.5. As far as temperature concerned, TSL6 grew and developed well at 25~C?0C. Effects of 82 additives on Bt activity were tested. Different mass concentration of 14 additives and increase efficacy were determined. It indicated that many additives were able to increase efficiency of Bt fermentative liquid for killing diamondbacks in some degrees. Different mass concentration of 14 additives influenced effect of Bt in controlling diamondbacks remarkably. Addtives enhanced the insecticidal activity of Bt by 0.4378 2.7743 fold.