Gender-related variations in hemolymph parameters of Carabus lefebvrei (Coleoptera: Carabidae): HPLC analysis and phenoloxidase activity

Abstract

We characterized the enzymatic activity of basal and total phenoloxidase and HLPC and SDS- PAGE profiles in hemolymph of Carabus lefebvrei males and females at different reproductive status. The phenoloxidase activity was activated by trypsin and inhibited by phenoloxidase activity specific inhibitor phenylthiourea. Our results demonstrated that both in males and females, there were no significant differences in the basal phenoloxidase activity between reproductive and virgin beetles, while the total phenoloxidase activity increased significantly in virgin specimens. Thus, resources seem to be invested to increase the humoral response in pre-reproductive phase forming a barrier against pathogens and preserving the fecundity and longevity of both sexes. The hemolymph DOPA-MBTH assay on polyacrilamide gel electrophoresis showed a high activity of monomeric form with an apparent molecular weight of 90 kDa and a dimer of about 170 kDa, also multimeric bands were present in both sexes. In the SDS-PAGE general protein pattern, specific bands were evident for reproductive and virgin males and females as biochemical markers of sexual difference in immunocompetence. Reproducible differences in peaks were recorded in HPLC analysis performed on virgin and reproductive males and females