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.

Three venom peptides, OdVP1, OdVP2, and OdVP3 were isolated from the venom of the solitary wasp Orancistrocerus drewseni (Hymenoptera: Eumenidae). The venom peptide amino acid sequences were determined by Q-TOF/MS/MS. The OdVP1, 2, and 3 with amidated C-terminals showed similar peptide sequences to the mastoparan from Vespula lewisii or the protonectin from Protonectarina sylveirae, suggesting that they adopt an amphipathic α-helix secondary structure. The amidation of C-terminal Leu of the venom peptides have been known to be required for their biological activities. The full-length open reading frame (ORF) sequences of the OdVP1, 2, and 3 were analyzed by 5’- and 3’-rapid amplification of cDNA ends (RACE). The overall gene structure of OdVPs showed a high homology to that of mastoparan B from Vespa basalis by containing signal sequence, prosequence, mature peptide and C-terminal glycine, but the mature peptide sequences were distinct from each other. The toxicological property and antimicrobial activity of OdVPs were characterized using synthetic peptides. This study on the venom peptides from O. drewseni should promote further studies on bioactive ingredients in the venom of solitary hunting wasps.