Stenotrophomonas maltophilia is a pathogen that causes infections mainly in immunocompromised patients. Despite increased S. maltophilia isolation from respiratory specimens of patients with cystic fibrosis (CF), the real contribution of the microorganism to CF pathogenesis still needs to be clarified. The aim of the present study was to evaluate the pathogenic role of S. maltophilia in CF patients by using a model of acute respiratory infection in DBA/2 mice following a single exposure to aerosolized bacteria. The pulmonary bacterial load was stable until day 3 and then decreased significantly from day 3 through day 14, when the bacterial load became undetectable in all infected mice. Infection disseminated in most mice, although at a very low level. Severe effects (swollen lungs, large atelectasis, pleural adhesion, and hemorrhages) of lung pathology were observed on days 3, 7, and 14. The clearance of S. maltophilia observed in DBA/2 mouse lungs was clearly associated with an early and intense bronchial and alveolar inflammatory response, which is mediated primarily by neutrophils. Significantly higher levels of interleukin-1beta (IL-1beta), IL-6, IL-12, gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), GROalpha/KC, MCP-1/JE, MCP-5, macrophage inflammatory protein 1alpha (MIP-1alpha), MIP-2, and TARC were observed in infected mice on day 1 with respect to controls. Excessive pulmonary infection and inflammation caused systemic effects, manifested by weight loss, and finally caused a high mortality rate. Taken together, our results show that S. maltophilia is not just a bystander in CF patients but has the potential to contribute to the inflammatory process that compromises respiratory function.