Cell-mediated haemolytic activity of haemolymph from the Colorado potato beetle (Leptinotarsa decemlineata Say.)

Haemolytic activity was identified in cell-free haemolymph from larval and imago stages of Leptinotarsa decemlineata. The haemolytically active fraction of the haemolymph was active against human, sheep, bull, toad and mouse erythrocytes. There was no haemolysis in the presence of 0.001 M EDTA and 0.5% glutathione. The titre of haemolytic activity did not increase after injury or vaccination of the larvae with Microccocus lysodeikticus. Haemolysin, a heat-labile protein was partially purified by ammonium sulphate precipitation, gel filtration, and ion-exchange separation. SDS PAGE, electrophoresis and immunoblotting showed that the active factor was a protein with a molecular weight of approximately 55 kD. It was not bactericidal for various micro-organisms but the antibacterial activity of the lysozyme increased in the presence of haemolysin only when M. lysodeikticus were used as target cells. Spherulocytes synthesized and released the haemolytic protein in vitro. The haemolytic activity increased in the presence of lipopolysaccharide from Escherichia coli and Ca++ ions. The physiological role of the haemolysin is as yet unknown.