MOSQUITO LARVICIDAL ACTIVITY OF ESCHERICHIA-COLI WITH COMBINATIONS OFGENES FROM BACILLUS-THURINGIENSIS SUBSP ISRAELENSIS

The genes cryIVA and cryIVD, encoding 134- and 72-kDa proteins, respec tively, and the gene for a regulatory 20-kDa polypeptide of Bacillus t huringiensis subsp. israelensis (serovar H14) were cloned in all seven possible combinations by the Escherichia coli expression vectors pT7 and pUHE. The four combinations containing cryIVA (cryIVA alone, with cryIVD, with the 20-kDa-protein gene, and with both) displayed high le vels of mosquito larvicidal activity in pUHE. The toxicity of the comb ination of cryIVA and cryIVD, with or without the 20-kDa-protein gene, was higher than has ever been achieved with delta-endotoxin genes in recombinant E. coli. Fifty percent lethal concentrations against third instar Aedes aegypti larvae for these clones decreased (i.e., toxicit y increased) continuously to about 3 x 10(5) cells ml(-1) after 4 h of induction. Larvicidal activities, obtained after 30 min of induction, were lower for clones in pT7 and decreased for an additional 3.5 h. I nduction of either cryIVD or the 20-kDa-protein gene alone resulted in no larvicidal activity in either pT7 or pUHE20. Cloned together, thes e genes were slightly toxic in pT7 but not in pUHE20. Five minutes of induction of this combination (cryIVD with the 20-kDa-protein gene) in pT7 yielded a maximal mortality of about 40%, which decreased rapidly and disappeared completely after 50 min. CryIVD is thus apparently de graded in E. coli and partially stabilized by the 20-kDa regulatory pr otein. Larvicidal activity of the combination of cryIVA and cryIVD was sevenfold higher than that of cryIVA alone, probably because of the c ross-stabilization of the polypeptides or the synergism between their activities.