A Drosophila model of systemic LPS delivery via dorsolateral thoracic injection reveals conserved mechanisms of infection-associated tissue damage.

Lipopolysaccharide (LPS) activates innate immunity in Drosophila melanogaster, but optimal delivery methods and systemic impacts remain unclear. We compared six LPS administration routes and identified dorsolateral thoracic injection as the most effective, enabling dose-controlled immune activation with negligible procedural mortality. Using this approach, LPS induced widespread apoptosis in the ovary, midgut, and testis, along with severe sperm defects-including disrupted axonemes and failed individualization-and tissue-specific organelle stress. It also triggered systemic oxidative stress and senescence-like phenotypes, evidenced by elevated ROS and SA-β-gal activity. Mechanistically, LPS activated the Immune Deficiency (IMD) and JAK-STAT pathways in a time- and dose-dependent manner, with relish expression increasing up to 7.22-fold and cleavage confirmed by Western blot. Critically, co-injection of an IKK inhibitor significantly rescued LPS-induced lethality, demonstrating that immune hyperactivation drives organismal death. Our work establishes dorsolateral injection as the optimal LPS delivery method and reveals that systemic LPS challenge causes organismal collapse via coordinated immune overactivation, oxidative damage, organelle dysfunction, and cellular senescence.