Use of Scorpion Neurotoxins to Improve the Insecticidal Activity of Rachiplusia ou Multicapsid Nucleopolyhedrovirus

The nucleopolyhedrovirus of the mint looper, Rachiplusia ou (RoMNPV, also known as Anagrapha falcifera multicapsid nucleopolyhedrovirus), has previously been shown to kill the European corn borer, Ostrinia nubilalis, and to protect corn plants against this pest more effectively than the Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV). To further improve the performance of RoMNPV against O. nubilalis, we engineered it to express the insect-selective scorpion neurotoxins AaIT and LqhIT2 using the promoters of the AcMNPV p10 and p6.9 genes. RoMNPV recombinant virus expressing AaIT from the p10 promoter produced a reduced quantity of viral occlusions and polyhedrin protein and failed to occlude virions efficiently. Western blot analysis revealed that the amount of AaIT in hemolymph of recombinant virus-infected O. nubilalis was very low compared to that in the hemolymph of infected Heliothis virescens. In all but one case, insertion of the toxins did not alter the dose of virus required to kill three different hosts. Viruses expressing toxins from the p6.9 promoter killed larvae of O. nubilalis significantly faster than wild-type virus, but no significant difference in speed of kill was observed between AaIT- and LqhIT2-expressing viruses in this host. Expression of LqhIT2 reduced survival times of Helicoverpa zea and H. virescens to a significantly greater extent than expression of AaIT. An AcMNPV recombinant expressing LqhIT2 from the p6.9 promoter killed larvae faster than a recombinant utilizing the p10 promoter for expression, but recombinants using these promoters to drive AaIT expression did not exhibit any significant differences in speed of kill.