Insulin/insulin-like peptide-binding protein (IBP) is abundantly found in venom of the solitary hunting wasp, Eumenes pomiformis (Hymenoptera: Eumenidae). E. pomiformis IBP (EpIBP) is most similar to insect IBP-like proteins that are known to inhibit insect growth and insulin signaling. To investigate the toxicity and target protein, EpIBP was in vivo expressed by Escherichia coli. Spodoptera exigua (Lepidoptera: Noctuidae) larvae injected with EpIBP showed a 20% lower pupation rate than the control larvae, although their body weight was not significantly different from the control when the larvae were provided artificial diet after the injection. EpIBP extended the larval stage without inducing paralysis of S. exigua larvae. To investigate the effects of EpIBP on caterpillar under a starvation condition, survivorship and body weight of the EpIBP-injected were evaluated without providing artificial diet until all the larvae died. The survivorship of the EpIBP-injected larvae was 24-36% higher than the control larvae at 4-5 d post-injection. The body weight of the control larvae reduced to 59% that is approximately 10% lower than the body weight of the EpIBP-injected larvae. These results suggest that EpIBP might inhibit the metabolism of the caterpillars, which is likely related with insulin-like peptide signaling pathway, suppress the loss of body weight and eventually extend the larval stage. An EpIBP-binding protein (EpIBPBP) isolated by immunoprecipitation was matched with a coiled-coil domain-containing protein of the fruit fly. The full-length sequence analysis of EpIBPBP is in progress.

To search for novel biologically active venom components, secretory venom proteins of two solitary hunting wasps, Orancistrocerus drewseni Saussure (1857) and Eumenes pomiformis Fabricus (1781), were identified by SDS-PAGE in conjunction with mass analysis with the aid of venom gland and sac-specific EST libraries constructed by suppression subtractive hybridization. Arginine kinase was the most predominant protein in both wasp venoms. Along with the full-length arginine kinase, a truncated form, which was known to have paralytic activity on a spider, was a common predominant protein in the two wasp venoms. Insulin/insulin-like peptide-binding protein was abundantly found only in E. pomiformis venom and the EST library, which might be due to its unique behaviors of oviposition and provision. It seemed that some venom proteins are secreted into venom fluid from venom gland cells via exosomes, not by signal sequence-mediated transport processes. Amphipathic α-helical peptides (10-15 amino acids) were predominantly transcribed in the venom gland/sac than protein components, and showed cell lytic activities against insect cells, mammalian cells, bacteria, and fungi. Phospholipase A2 and hyaluronidase, which are known to be the main components of wasp venoms, were found in both wasp venoms. In addition, a dendrotoxin-like peptide known to be a K+ channel blocker was also found in the venom of E. pomiformis.

To determine differential gene expression profiles in the venom gland and sac (gland/sac) of a solitary hunting wasp species, Eumenes pomiformis Fabricius (1781), a subtractive cDNA library was constructed by suppression subtractive hybridization. A total of 541 expressed sequence tags (EST) were clustered and assembled into 102 contigs. In total, 37 genes were found from the library by BLASTx search and manual analysis. A eumenitin-like venom peptide, EpVP1, occupied ca. 26% of the library. A novel venom peptide, EpDTX, shared sequence similarity with trypsin inhibitors and dendrotoxin-like venom peptides known as K+ channel blockers, implying it could be responsible for the paralysis of prey. As well as phospholipase A2 and hyaluronidase known to be main components of wasp venoms, several contigs encoding enzymes, including metalloendopeptidases and a decarboxylase likely involved in the processing and activation of venomous proteins, peptides, and neurotransmitters, were also isolated from the library. The presence of a gene encoding insulin-like growth factor binding protein suggests that solitary hunting wasps might control the prey to stay in larval stage by their venom. The abundance of these venom components in the venom gland/sac and alimentary canal was confirmed by quantitative real-time PCR.