Figure 1.

A mouse model of lung emphysematous infection: experimental design and characterization. (a) Experimental design: lung emphysema was induced by oropharyngeal instillation of PPE in CD1 mice (Day −17). Elastase instillation induces pulmonary damage compatible with emphysema-like lesions. Mice were instilled with PPE (E+/I-) or received physiological saline solution (V+/I-). Animals were infected with ~10⁸ CFU/mouse of H. influenzae Xen21 (Day 0), V+/I+ and E+/I+ groups. Assay types and sampling time points are indicated. (b) Longitudinal analysis of lung bacterial loads. Mice were infected as indicated in (a), euthanized 6, 12, 24, 30 and 48 hpi, and bacterial loads quantified in homogenized lungs. No significant differences were observed between V+/I+ (white) and E+/I+ (green) groups at any of the indicated time points. V+/I+ groups: bacterial counts were lower 24, 30 and 48- than 6 hpi (p < 0.005); bacterial counts were lower 24, 30 and 48- than 12 hpi (p < 0.05); bacterial counts were lower 48- than 24 and 30 hpi (p < 0.0001). E+/I+ animals: bacterial counts were lower 30 and 48- than 6 hpi (p < 0.0001); bacterial counts were lower 30 and 48- than 12 hpi (p < 0.005); bacterial counts were lower 48- than 24 and 30 hpi (p < 0.0001). Results are reported as log₁₀ CFU/lung and represented as box plot graphs (lines inside boxes represent median values). Statistical comparisons were performed using one-way ANOVA and Tukey’s multiple comparison test. *** p < 0.001; ** p < 0.01; * p < 0.05. (c) Bacterial bioluminescence was determined 1, 3, 6, 9, 12 and 24 hpi, representative animals are shown (bottom panel). Results are shown as mean ± SEM. Statistical comparisons were performed using one-way ANOVA and Tukey’s multiple comparison test. *** p < 0.001; * p < 0.05. (d) H. influenzae Xen21 light levels and bacterial numbers in vivo. Bacterial counts and animal luminescence determined at different time points post-inoculation (6, 12 and 24 h). Spearman’s correlation coefficient was determined