The present investigation used Galleria mellonella larvae as an infection model to describe the regulatory events associated with the induction of antimicrobial activity following injection of a sub-lethal dose of Escherichia coli into the larval hemocoel. The ability of larval hemolymph to generate this activity as natural antibiotics was examined at different time periods postinjection against both the target bacteria (Gram-negative, E. coli) and Gram-positive, Staphylococcus aureus, as well as against some selected fungi such as Candida albicans and Aspergillus fumigatus. The results indicated that the highest response was found against G -ve bacteria (E. coli) than any other pathogens tested, and the highest activity was obtained at 24 h post-treatment. This may indicate that the antimicrobial is specific to the inoculated bacteria. The presence of such activity in larval plasma was verified using microbiological and biochemical tests. E. coli-growth tests on solid agar were carried out, and the results showed drastic effects represented by bacterial mortality that exceeds 74.5% in bacteria-treated with immune plasma, while weak effects were evident in normal (untreated) and control (water-treated) plasma. These tests have confirmed this activity is bactericidal. Also, hemolysis tests were conducted using human erythrocytes as target cells, and the results indicated that all plasma samples possess a weak hemolysis activity, which confirms its safety on human and animals health. Additional biochemical tests of the mode of action were performed to determine the ability of the induced antimicrobial agent of the immune plasma to alter the cell membrane integrity of the E. coli bacterial cell by analysis of proteins. The results confirmed that the immune plasma is capable of inducing leakage of these intracellular molecules in the tested bacterial cells. Scanning electron microscopy (SEM) was also used to visualize the morphological effects resulting from treating the target bacterium with this antimicrobial agent and the results indicated a weak growth and an irregular pitted surface of the treated bacterial cells as compared to dense growth and regular smooth surface exhibited in the controls. These results may provide us with essential data for the development and production of new means of protecting humans and animals from infectious diseases.