The influence of MyD88-dependent innate immune signaling on intraepithelial microcolony formation.

<p>1-day-old MyD88^+/+ and MyD88^-/- mice were orally infected with 100 CFU <i>S</i>. Typhimurium WT. <b>(A)</b> 4 days after infection, small intestinal tissues were collected and analyzed by immunostaining. Three representative images showing <i>S</i>. Typhimurium (red) forming intraepithelial microcolonies in MyD88^-/- mice. Counterstaining with E-cadherin (green), WGA (white) and DAPI (blue). Bar, 10 μm. For wild type controls see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006925#ppat.1006925.g001" target="_blank">Fig 1D</a>. <b>(B)</b> Quantitative evaluation of the number of intraepithelial microcolonies per villus in MyD88^+/+ and MyD88^-/- mice at 4 days p.i.. <i>S</i>. Typhimurium microcolonies were quantified in 20–30 villi per animal (n = 6–8). Results represent the mean ± SD. <b>(C)</b> Transmission electron microscopy (TEM) images of intraepithelial <i>Salmonella</i> in MyD88^+/+ (left panel) and MyD88^-/- mice (right panel). Asterisks highlight bacteria. Bar, 2 μm. <b>(D)</b> Co-immunostaining for <i>Salmonella</i> (green) and LAMP1 (red) in small intestinal tissue sections at day 4 p.i.. Counterstaining with E-cadherin (white) and DAPI (blue). Bar, 5 μm. <b>(E)</b> Quantitative evaluation of the percentage of intraepithelial <i>S</i>. Typhimurium associated with LAMP1 staining. Four neonates were analyzed at day 4 p.i.. Results represent the mean ± SD. <b>(F)</b> Co-immumostaining for <i>Salmonella</i> (red) and the GFP expressing SPI2 reporter (pM973, green) in small intestinal tissue sections at day 4 p.i.. Counterstaining with E-cadherin (white) and DAPI (blue). Bar, 5 μm. <b>(G)</b> Quantitative analysis of the percentage of intraepithelial <i>S</i>. Typhimurium expressing the SPI2 reporter. Microcolonies from tissue sections from four neonates were analyzed at day 4 p.i.. Results represent the mean ± SD.