Detailed histopathological scores of all animals examined in this study.

Individual clinical scores for alveolitis, bronchiolitis, perivascular cuffing (PVC), vasculitis, interstitial pneumonia (IP), and pleuritis. Columns show data medians, symbols represent individual animals; 1-way ANOVA with Tukey’s post hoc test; p values are specified; n values are specified in Fig 4F. (TIF)</p

Urea-PAGE fractionation (uncropped) of RNA transcripts.

a) Template and primer used in the reaction. b-e) Independent repeats of primer extension assay by IAV polymerase. Dashed rectangle shows insert presented in Fig 1E. Replicates were included in quantitations presented in Fig 1E and 1F. (TIF)</p

All statistical analyses performed for this study.

All statistical analyses performed for this study.</p

Clinical signs of mice involved in mitigation of histopathology study.

a-b) Body weight measurements taken twice daily (a), and rectal body temperature determined once daily (b). Lines intersect, and symbols show, data means ± SD. Dashed line in (a) specifies predefined endpoint. (TIF)</p

Clinical signs in 4’-FlU-treated mice used in immune response-mitigation study.

a-b) Body weight measurements taken twice daily (a), and rectal body temperature determined once daily (b). Lines intersect, and symbols show, data means ± SD; n values are specified in Fig 4. Dashed line in (a) specifies predefined endpoint. (TIF)</p

Effect of 4’-FlU on the antiviral immune response and lung histopathology.

a)In vivo Bioplex and histopathology study schematic. b) Lung viral titers in animals treated as in (a). c-d) Changes in IL-6 (c) and TNF-α (d) levels present in BALF of animals treated as in (a), relative to levels at time of infection. Lines in (b) intersect, and symbols show, geometric means ± SD, lines in (c-d) intersect, and symbols show, data medians with 95% CI; 2-way ANOVA with Tukey’s post hoc test; P values are given. e) Representative photomicrographs of lung tissue extracted 5 days after infection of animals treated as in (a). Tissues of two individual animals per study arm are shown at 10× magnification; scale bar denotes 100 μm; Br, Bronchiole; Bl, Blood vessel. f) Histopathology scores of animals treated as in (a). Lungs were extracted 5 days after infection. Scores for each animal represents a mean of individual alveolitis, bronchiolitis, vasculitis, pleuritis, perivascular cuffing (PVC), and interstitial pneumonia (IP) scores. Columns represent data medians with 95% CI; symbols show mean scores for each individual animal; 1-way ANOVA with Dunnett’s post hoc test, P values and n values for each study arm are specified.</p

Clinical signs of mice infected and reinfected with pdmCa09.

a-d) Body weight measurements taken twice daily (a, c), and rectal body temperature determined once daily (b, d). Lines intersect, and symbols show, data means ± SD. Results are shown for animals after the original infection (a-b) and after homotypic reinfection (c-d). Dashed lines in (a,c) specify predefined endpoint. (TIF)</p

Clinical signs in pdmCa09-infected immunocompromised mice treated with 4’-FlU.

a-c) Shown are body weight measurements taken twice daily (a, c), and rectal body temperature (b, d) determined once daily. Lines intersect, and symbols show, data means ± SD; n values are specified in Fig 5. Results are shown for RAG1 KO (a-b) and IFNAR1 KO (c-d) animals. Dashed lines in (a,c) specify predefined endpoint. (TIF)</p

4’-FlU efficacy confirmation after infection with aerosolized inoculum virus.

a) Efficacy study schematic; n values are specified. b-c) Body weight measurements of animals from (a) taken twice daily (a), and rectal body temperature (b) determined once daily. Lines in (b-c) intersect, and symbols show, data means ± SD. d) Lung viral titers determined 4 days after infection. Columns represent geometric means ± SD, symbols show individual animals; 1-way ANOVA with Dunnett’s post hoc test; P values are specified. Dashed line in (d) specifies limit of detection. (TIF)</p

Reinfection of 4’-FlU-experienced animals with homotypic H1N1.

a) Schematic of the treatment and reinfection study. For infection and reinfection, mice received aerosolized pdmCa09. b) Survival study of animals infected, treated, and reinfected as shown in (a). Median survival time in days is specified; Kaplan-Meier simple survival analysis. c) Anti-H1N1 neutralizing antibody (nAb) titers developing in animals from (a). Lines intersect, and symbols show, geometric means ± SD; 2-way ANOVA with Tukey’s post hoc test; P values are specified. d) Survival of animals after re-infection. Median survival time in days is specified; Kaplan-Meier simple survival analysis. N numbers for animals in (a-d) are specified in (a).</p