Targeting of an expressed neurotoxin by its recombinant baculovirus

AaIT, an insect-selective neurotoxic polypeptide derived from scorpion veno m, has recently been used to engineer recombinant baculoviruses for insect pest control. Lepidopterous larvae infected with an AaIT-expressing baculov irus; reveal symptoms of paralysis identical to those induced by injection of the native toxin. However, the paralyzed larvae treated by the recombina nt virus possess an approximately 50-fold lower hemolymph toxin concentrati on than insects paralyzed by the native toxin. The mechanism of this potent iation effect was studied using immunocytochemistry, electrophysiology and toxicity assays. (i) Light microscopy, using peroxidase-conjugated antibodi es, revealed the presence of toxin in virus-susceptible tissues, including tracheal epithelia located close to the central nervous system and beyond i ts lamellar enveloping sheath. (ii) High-resolution immunogold electron mic roscopical cytochemistry clearly revealed the presence of recombinant AaIT toxin inside the thoracic and abdominal ganglia on neuronal cell bodies and axonal membranes. (iii) Ventral nerve cords dissected from silkworm larvae infected with the recombinant baculovirus exhibited a high degree of excit ability, expressed as enhanced frequency and bursting mode of their spontan eous activity, when compared to nerve cords infected with the wild-type vir us. We conclude that the recombinant-virus-infected tracheal epithelia, out branching in the body of an infected insect, (i) locally supply a continuou s, freshly produced toxin to its neuronal receptors and (ii) introduce the expressed toxin to the insect central nervous system, thus providing it wit h critical target sites that are inaccessible to the native toxin.