Insecticidal activity of recombinant baculovirus expressing both spider toxin isolated from Araneus ventricosus and Bacillus thuringiensis crystal protein fused to a viral polyhedrin

A novel recombinant Autographa californica nucleopolyhedrovirus (AcMNPV), ApPolh5-3006AvTox2, co-expressing two insecticidal toxins, one isolated from the spider Araneus ventricosus, and the other from Bacillus thuringiensis, was constructed to improve the insecticidal activity of AcMNPV. The recombinant virus was designed to express insect-specific spider toxin, Av-Tox2, under control of the early promoter from Cotesia plutellae bracovirus (CpBV). In addition, the B. thuringiensis cry1-5 crystal protein gene was introduced into the genome of this recombinant virus by fusing it with the viral polyhedrin gene, thus creating a hybrid polyhedrin-cry1-5 gene under control of the polyhedrin gene promoter. Reverse transcription-polymerase chain reaction (RT-PCR) analysis revealed that both Av-Tox2 and Polyhedrin-Cry1-5 fusion protein were successfully expressed in the infected cells. In addition, SDS-PAGE revealed that Polyhedrin-Cry1-5 fusion protein expressed by recombinant viruses occluded the polyhedra. ApPolh5-3006AvTox2 showed significantly reduced LD∧50 and ST∧50 values against both Plutella xylostella and Spodoptera exigua larvae. These results strongly suggested that coexpression of spider and B. thuringiensis insecticidal toxins could be successful in improving the insecticidal activity of baculoviruses.