Fed-batch culture of Bacillus thuringiensis for thuringiensin production in a tower type bioreactor

Fed-batch culture of Bacillus thuringiensis in a modified airlift reactor has been developed by using adaptive control of glucose concentration in the reactor. The glucose concentration was estimated via a correlation equation between carbon dioxide production rate and glucose consumption rate. The estimated glucose concentration as the output variable was fed back to computer for calculation of substrate addition. The modified reactor was an airlift reactor with a net draft tube. The airlift reactor had high oxygen transfer rate and low shear stress which were important factors for production of thuringiensin. Fed-batch culture of Bacillus thuringiensis in the modified airlift reactor provided significant improvement of thuringiensin production. (c) 1995 John Wiley & Sons, Inc.     

Recovery and purification of thuringiensin from the fermentation broth of Bacillus thuringiensis

Thuringiensin is a heat stable -exotoxin from Bacillus thuringiensis with a great potential for replacing the traditional chemical pesticides. A process using micellar-enhanced ultrafitration method to recover thuringiensin was significantly improved by the use of a spiral-wound membrane, which could be operated at a low transmembrane pressure drop. This method was performed by adding a surfactant cetylpyridinium chloride (CPC) into the fermentation broth. After the surfactant-thuringiensin conjugates were formed, the broth then passed through the ultrafiltration membrane and the retentate was collected. The results indicated the optimal concentration of CPC for producing a maximal recovery up to 99.3% is 4%. For purification, the centrifuged broth was further filtered by a membrane filter. The filtered solution then was mixed with 50% of activated carbon. The supernatant then was injected into a preparative HPLC. The eluate was collected during thuringiensin peak formation. This eluate was then concentrated by vacuum evaporation and dialysis using an electrodialyzer to remove the excess salts. The dialyzed solution was then crystallized by lyophilization. The purity of the thuringiensin crystal was identified by HPLC, capillary electrophoresis, and mass spectrometry.This revised version was published online in October 2005 with corrections to the Cover Date.     

苏云金素合成基因簇中非核糖体肽合成酶基因thu2功能的初步分析

对苏云金素生物合成基因簇中编码非核糖体肽合成酶基因thu2进行基因缺失插入失活的研究。用温敏型质粒pHT304-TS构建基因thu2的插入缺失质粒pEMB1434,电转化苏云金芽胞杆菌菌株CT-43后,通过抗性筛选和PCR验证得到thu2基因同源双交换基因敲除突变株CT-43-22。HPLC(高效液相色谱,High Performance Liquid Chromatography)检测发现CT-43-22没有苏云金素特征吸收峰;用pHT304构建得到含有完整thu2基因的回补质粒pEMB1435,电转化CT-43-22后得到互补重组菌CT-43-22b,发现其恢复了苏云金素的产生。显微镜观察突变株和互补重组菌均能产生正常的晶体和芽胞。thu2的基因敲除和基因互补实验证明,thu2基因为CT-43苏云金素生物合成的必需基因,但对晶体和芽胞的形成没有影响。     

Studies on Bacillus thuringiensis strains isolated from Swedish soils: insect toxicity and production of B. cereus-diarrhoeal-type enterotoxin

At moderate concentration, 23 of 40 strains of Bacillus thuringiensis isolated from Sweden were toxic to Trichoplusia ni and five were toxic to Aedes aegypti. Five of the strains were toxic to Diabrotica undecimpunctata at high concentration, two were toxic to Heliothis virescens at low concentration and five produced thuringiensin (formerly called -exotoxin). No strain was toxic towards the beet armyworm Spodoptera exigua at low concentration. Twenty-three of the strains produced a B. cereus-diarrhoeal-type enterotoxin.A. Abdel-Hameed is with the Department of Microbiology, Faculty of Pharmacy, Cairo University, Kasr-El-Aini Street, Cairo, Egypt. R. Landén is with the Department of Microbiology, Stockholm University, S-10691 Stockholm, Sweden. A. Abdel-Hameed's present address is the Department of Applied Chemistry and Microbiology, P. O. Box 27, Viikki, Building B, FIN-00014 University of Helsinki;     

A novel index for analysing the response of a microbial culture to changes in the environmental variables during cultivation

AIMS: To analyse the sensitivity of a microbial culture to variations in the cultivation conditions by using the motile intensity of the cells. METHODS AND RESULTS: Batch cultures of Bacillus thuringiensis were used to study the sensitivity of the cells to pulse changes in pH, temperature and oxygen supply. A droplet of the culture sample was visualized under an optical microscope and the image of the moving cells was captured with a computer controlled display camera attached to the microscope. Motile intensity was computed directly using an image analysis programme. The results showed that the different phases of cell growth exhibited different motile intensities. The motile intensity changed remarkably at the high level of the motile intensity, when the environmental variables are changed. CONCLUSIONS: The product formation was considerably reduced when a disturbance was applied at the high magnitude of motile intensity. SIGNIFICANCE AND IMPACT OF STUDY: Monitoring the motile intensity by image analysis is simple and makes it an attractive method for assessing the effect of environmental variables on the growth and product formation of microbial cultures.     

Production of thuringiensin from Bacillus thuringiensis using a net-draft-tube,modified airlift reactor

A net-draft-tube, modified airlift reactor and a stirred-tank reactor were used for thuringiensin production by Bacillus thuringiensis subsp. darmstadiensis growing with various concentrations of molasses. The optimum concentration of molasses for thuringiensin production in both reactors was 15 g/l. There was a 6 h delay in sporulation in the modified airlift reactor compared with that in the stirred-tank reactor. Thuringiensin yield in the modified airlift reactor (2.2 g/l) was consequently higher than that in the stirred-tank reactor (1.1 g/l).