Galleria mellonella is a Lepidopteran pest insect infesting the honeycombs, and the last instar larva of the insect is also a valuable model organism for evaluating drug efficacy, pathogenicity of infectious agents, and environmentally sound chemical insecticides. Levamisole is an anthelmintic drug that is used for the treatment of parasitic infections in animals. In this study, we investigated the oxidative and genotoxic effects of dietary levamisole on the hemolymph of the G. mellonella last instars to evaluate levamisole as an alternative chemical for traditional insecticides. In this context, first instar larvae were reared up to the last instars in artificial diets containing levamisole (w/w) and a control diet without levamisole. Malondialdehyde (MDA), and protein carbonyl (PCO) contents, and the activity of antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), glutathione-s-transferase (GST) and glutathione peroxidase (GPx) were determined to evaluate oxidative stress in the hemolymph of G. mellonella last instars. Furthermore, the comet assay was performed on hemolymph to evaluate DNA damage levels. The dietary levamisole led to a significant decrease in MDA and PCO contents, and antioxidant enzyme activities (SOD, CAT, GST and GPx) when compared to control diet. However, dietary levamisole significantly increased tail moment, tail %DNA, and tail length, which are strong indicators of DNA damage. We infer from these that levamisole induced DNA damage and crippled antioxidant enzymatic defense in correlation with gradually increasing dietary concentration. The study would also provide valuable insights into the multifunctional aspects of levamisole, including its use in pest management.