Co-expression and Synergism Analysis of Vip3Aa29 and Cyt2Aa3 Insecticidal Proteins from Bacillus thuringiensis

Vegetative insecticidal protein (Vip3) from Bacillus thuringiensis shows high activity against lepidopteran insects. Cytolytic δ-endotoxin (Cyt) also has high toxicity to dipteran larvae and synergism with other crystal proteins (Cry), but synergism between Cyt and Vip3 proteins has not been tested. We analyzed for synergism between Cyt2Aa3 and Vip3Aa29. Both cyt2Aa3 and vip3Aa29 genes were co-expressed in Escherichia coli strain BL21 carried on vector pCOLADuet-1. Vip3Aa29 showed insecticidal activity against Chilo suppressalis and Spodoptera exigua, with 50% lethal concentration (LC(50)) at 24.0 and 36.6 μg ml(-1), respectively. It could also inhibit Helicoverpa armigera growth, with 50% inhibition concentration at 22.6 μg ml(-1). While Cyt2Aa3 was toxic to Culex quinquefasciatus (LC(50): 0.53 μg ml(-1)) and Chironomus tepperi (LC(50): 36 μg ml(-1)), it did not inhibit C. suppressalis, S. exigua, and H. armigera. However, the co-expression of Cyt2Aa3 and Vip3Aa29 showed synergistic effect on C. suppressalis and S. exigua, and the individual activities were strengthened 3.35- and 4.34-fold, respectively. The co-expression had no synergism against C. tepperi and H. armigera, but exerted some antagonistic effect on Cx. quinquefasciatus. The synergism between Cyt2Aa and Vip3Aa was thus discovered for the first time, which confirmed that Cyt toxin can enhance the toxicity of other toxins against some non-target insects. By synergism analysis, the effectiveness of microbial insecticides can be verified.