Bioluminescence determination of antibacterial activity of Bombyx mori and Galleria mellonella haemolymph

Abstract: We describe an antibacterial assay based on bioluminescence of two Gram negative bacteria Photorhabdus luminescens and transformed Escherichia coli, which can be used for a real-time measurement of antibacterial activity. We observed a significant dose-dependent decrease of bioluminescence using both bacterial species during one hour after exposure to Bombyx mori or Galleria mellonella haemolymph. The humoral origin of the antibacterial activity observed in whole haemolymph was confirmed in haemolymph plasma without haemocytes. Antibacterial activity operating against Gram negative bacteria was measured in unaffected insect larvae as well as after septic injury; increased antibacterial activity in haemolymph was detected in the latter case which confirms inducibility of antimicrobial agents. This method can be widely used for determination of antibacterial activity in insects and supposedly in other invertebrates.