Fig 6. IAV-driven S. pneumoniae glucose and galactose metabolism modulates growth and virulence strategies.

(A) Growth curves for Spn grown alone in minimal media (THB 50%, black closed circles), or supplemented with glucose 1% w/v (red solid squares) or galactose 1% w/v (blue solid triangles) do not differ significantly. Growth of Spn challenged with IAV was blunted when grown in minimal media (black open circles) or with glucose (orange open squares), but not with galactose (purple open triangles) Data reflect n = 8 per condition (two-way ANOVA, ****p<0.0001 at t = 3h). Glucose supplementation in media rescues (B) expression of pneumolysin as measured by Western blot, (C) exacerbated cytotoxicity to A549 cells as measured by LDH assay (n = 8 per group, mean ± SEM, one-way ANOVA ****p<0.0001), and (D) cleaved caspase-3 expression by A549 cells infected with Spn+IAV. (E) Adhesion of bacteria to A549 cells was unchanged regardless of sugar supplementation, but Spn+IAV supplemented with galactose (F) improved invasion into A549 over Spn grown in glucose or un-supplemented media. Statistical analyses in E and F reflect n = 9 per condition, mean ± SD, analyzed by Kruskal-Wallis test with Dunn’s multiple-comparison post-test. Asterisks denote the level of significance observed: * = p ≤ 0.05; ** = p ≤ 0.01; *** = p ≤ 0.001.