Recent studies evaluated the impact of dust exposure on pure and mixed cultures of Escherichia coli, Enterococcus faecalis, Klebsiella pneumoniae, and Pseudomonas aeruginosa, revealing increased biofilm formation and altered sensitivities to H2O2. In this study, we examined the impact of lead (Pb), house, road, and combined dust on K. pneumoniae and P. aeruginosa in pure, mixed, or eukaryotic co-culture with human alveolar basal epithelial (A549) cells. Although no impact on pure or mixed culture growth was observed when bacteria were exposed to Pb, house, or road dust, increased biofilm was produced by P. aeruginosa in the presence of 0.8g/mL of Pb, while P. aeruginosa and K. pneumoniae both exhibited increased biofilm production in the presence of 100g/mL of house, road, and combined dust. When co-cultured with eukaryotic A549 cells, both bacteria demonstrated increased proliferation 6h post-infection when challenged with house, road, or combined dust. However, when mixed bacteria were co-cultured with A549 cells, P. aeruginosa exhibited a significant similar to 1.5-fold increased proliferation in the presence of 100g/mL house, road, or combined dust. In sharp contrast, K. pneumoniae exhibited significantly reduced proliferation, when in mixed (with P. aeruginosa) A-549 co-culture, following exposure to 100g/mL house, road, or combined dust. To evaluate whether a host cell inflammatory response contributed to this disparity, NF-B activation was evaluated in each co-culture infection. K. pneumoniae-A-549 co-culture, treated with 100g/mL of combined dust, exhibited no alterations in NF-B translocation to the nucleus. Further, no differences in cytokine production were observed in the K. pneumoniae A-549 co-culture treated with 100g/mL of house dust. Taken together, these data suggest that within the lung environment, mixed infections exposed to dust or dust contaminants could benefit one organism at the expense of the other, independent of the activation of inflammatory pathways.