Enhancing the oral and topical insecticidal efficacy of a commercialized spider venom peptide biopesticide via fusion to the carrier snowdrop lectin (Galanthus nivalis agglutinin)

Abstract

BACKGROUND: Spear®-T sold as a contact foliar spray for the control of glasshouse pests such as aphids, thrips, spider mites and whiteflies, contains the recombinant spider venom peptide GS-ω/κ-HxTx-Hv1h (named as GS-ω/κ-HxTx-Hv1a by Vestaron) as the active ingredient. Here we investigate whether fusion of the peptide to snowdrop lectin, (Galanthus nivalis agglutinin; GNA) enhances the efficacy of this venom peptide towards aphid pests. Results: Recombinant GS-ω/κ-HxTx-Hv1h (HxTx-Hv1h) and an HxTx-Hv1h/GNA fusion protein were produced using the yeast Pichia pastoris. Purified proteins showed comparable toxicity when injected into lepidopteran (Mamestra brassicae) larvae, but significant differences in oral and contact activity towards aphids. HxTx-Hv1h had comparable acute oral toxicity to pea (Acyrthosiphon pisum) and peach potato (Myzus persicae) aphids with respective Day (2) median lethal concentration (LC50) values of 111 and 108 ∼M derived from diet assays. The fusion protein also showed comparable oral toxicity to both species but D2 LC50 values were >3-fold lower (35 and 33 ∼M for pea and peach potato aphids, respectively) as compared to HxTx- Hv1h. Topically applied toxin and fusion protein, but not GNA, caused significant reductions in pea aphid survival. Contact effects on mortality were significantly greater for aphids exposed to fusion protein as compared to toxin alone. Whole aphid fluorescence microscopy and immunoblotting suggest that improved efficacy is due to enhanced persistence of HxTx-Hv1h when fused to GNA following internalisation of ingested or topically applied proteins. Conclusions: This is the first study to report on the insecticidal activity of HxTx-Hv1h towards aphids and results suggest that a fusion protein-based approach offers opportunities to significantly enhance oral and contact efficacy of naturally derived toxins, such as HxTx-Hv1h, towards crop pests.