Extracorporeal shockwave therapy (ESWT) is applied successfully in various orthopedic disorders. Since shockwaves have demonstrated significant bactericidal effectiveness in vitro, safety and effectiveness of ESWT in vivo were evaluated in a rabbit model of osteomyelitis. Chronic osteomyelitis was induced by injecting sodium morrhuate and Staphylococcus aureus into the proximal tibia of 12 New Zealand white rabbits. Four and five wk after the initial operation, soft focused ESWT was applied twice to the infected limbs. Clinical parameters and laboratory values were followed and blood samples were taken for culture before and 30 min after ESWT. Following sacrifice after 8 wk, lungs, spleen and kidneys were studied histologically for signs of sepsis and secondary infection. Tibial osteomyelitis was assessed clinically, and by radiologic, microbiologic and histologic procedures. Signs of bacterial spreading were not detectable after ESWT, neither in blood cultures nor in histologic analyses of representative organs. Temperature, body weight, C-reactive protein and white blood cell levels also remained unchanged after ESWT. Of particular interest, histologic scores of osteomyelitis were significantly decreased in the ESWT-group compared to the untreated control (p = 0.019). However, S. aureus was still detectable in tissue samples of all animals. This is the first study investigating the effects of ESWT applied to infected target areas. ESWT of infected bone did neither induce bacterial spreading nor worsening of infection, and the results suggest the reported treatment protocol of ESWT to be beneficial in the treatment of chronic bone infections.