This study investigates the impact of imidacloprid insecticide, on the flesh fly Sarcophaga ruficornis, (Diptera: Sarcophagidae), a medically and veterinary significant species associated with myiasis. Lethal (0.02%) and sub‐lethal (0.004%) doses were administered over 24 and 48 h, focusing on midgut histopathological, ultrastructural and biochemical alterations. Imidacloprid‐induced disruptions in physiological and metabolic functions, leading to adult fly mortality. Histological and cytological analyses revealed significant cellular changes, including cell degeneration, cytoplasmic vacuolization, chromatin condensation, irregular epithelial borders, disrupted peritrophic membrane and the release of cell components into the midgut lumen. Biochemical analyses demonstrated a decrease in total protein, carbohydrate and lipid contents, correlating with insecticide concentration and duration. Antioxidant enzymes, particularly glutathione S‐transferase (GST) and superoxide dismutase (SOD), resulted in a significant increase compared to the control, proportional to time and concentration. This research provides a comprehensive understanding of imidacloprid‐induced toxicity in the midgut of Sarcophaga ruficornis, emphasizing its potential for effective pest population management.