The present study deals with the evaluation of the efficacy of amoxicillin bearing poly-lactic-glycolic acid (PLGA) microsphere formulation in treatment of experimental listeriosis in Swiss albino mice. Amoxicillin bearing PLGA microspheres were prepared by water-in-oil-in-water emulsion technique. PLGA microwspheres significantly regulated sustained release of encapsulated drug over extended time period. The rate of release increased in temperature dependent manner. Amoxicillin bearing PLGA microsphere successfully cleared bacterial burdens in vital organs (kidney, spleen, and brain) and also increased survival rate of treated animals in comparison to free form of the drug. The higher efficacy of microsphere based novel formulation of amoxicillin could be attributed to its targeted delivery to infected macrophages as well as sustained release over extended period of time.