The environmental bacterium Stenotrophomonas maltophilia is increasingly described as a multidrug-resistant pathogen of humans, being associated with pneumonia, among other diseases. But the degree to which S. maltophilia is capable of replicating in a mammalian host has been an issue of controversy. Using a model of intranasal inoculation into adult A/J mice, we now document that S. maltophilia strain K279a, the clinical isolate of S. maltophilia whose complete genome sequence was recently determined, is in fact capable of replicating in lungs, displaying as much as a 10-fold increase in c.f.u. in the first 8 h of infection. Importantly, as few as 104 c.f.u. deposited into the A/J lung was sufficient to promote bacterial outgrowth. Bacterial replication in the lungs of the A/J mice was followed by elevations in pro-inflammatory cytokines and also promoted resistance to subsequent challenge. We also found that DBA/2 mice were permissive for S. maltophilia K279a replication, although the level of growth and persistence in these animals was less than it was in the A/J mice. In contrast, the BALB/c and C57BL/6 mouse strains were non-permissive for S. maltophilia K279a growth. Interestingly, when five additional clinical isolates were introduced into the A/J lung, marked differences in survival were observed, with some strains being much less infective than K279a and others being appreciably more infective. These data suggest that the presence of major virulence determinants is variable among clinical isolates. Overall, this study confirms the infectivity of S. maltophilia for the mammalian host, and illustrates how both host and bacterial factors affect the outcome of Stenotrophomonas infection.