Figure 3. Shigella effector OspC3 is critical for virulence in oral Shigella infection.
(A–H) Mice were treated orally with 25 mg streptomycin sulfate in water and infected 1 day later. B6.WT mice (co-housed wild-type [WT] and B6.Nlrc4+/–Casp11+/–) were orally challenged with 107 colony forming units (CFUs) of WT Shigella flexneri (n=7) and B6.Nlrc4–/– mice (co-housed B6.Nlrc4–/– and B6.Nlrc4–/–Casp11+/–) were challenged with WT (green, n=11) or ΔospC3 S. flexneri (blue, n=11). Mice were sacrificed at 2 days post-infection. (A) Representative images of the cecum and colon from B6.Nlrc4–/– mice infected with WT or ΔospC3 S. flexneri. The white arrow indicates clear but reduced inflammation in mice infected with the ΔospC3 strain. (B) Mouse weights from 0 through 2 days post-infection. Each symbol represents the mean for all mice of the indicated genotype. (C) Shigella CFUs per million cells from the combined intestinal epithelial cell (IEC) enriched fraction of gentamicin-treated cecum and colon tissue. (D) Quantification of cecum lengths normalized to mouse weight prior to infection; cecum length (cm)/mouse weight (g). (E) The ratio of fecal pellet weight when wet (fresh) divided by the fecal pellet weight after overnight drying. Pellets were collected at day 2 post-infection. (F, G) CXCL1 and IL-1β levels measured by ELISA from homogenized cecum and colon tissue of infected mice. (H) Blood scores from feces collected at 2 days post-infection. 1=occult blood, 2=macroscopic blood. (C–H) Each symbol represents one mouse. Data collected from two independent experiments. Mean ± SD is shown in (B, D–G). Geometric mean ± SD is shown in (C). Statistical significance was calculated by one-way ANOVA with Tukey’s multiple comparison test (B (day 2), C–G) and by Fisher’s exact test in (H) where data were stratified by presence (score = 1 or 2) or absence (score = 0) of blood. Data were log-transformed prior to calculations in (C) to achieve normality. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, ns = not significant (p>0.05).