Listeria monocytogenes (LM; log-phase cells of strain LM2203) were cultured at 37◦C in Tryptic Soy Broth (Becton Dickinson, Sparks, MD). The LM strain was isolated from a cluster of listeriosis in pregnant women that led to fetal demise.38 (link) Each inoculum containing 1×108 colony forming units (CFUs) of LM was dissolved in 10mL whipping cream and delivered via oral gavage through a soft intragastric feeding tube under sedation (n=10), as previously described.38 (link),39 (link) Control inoculations (mock) consisted of 10mL whipping cream alone with no LM (n=3). Eight of the 13 animals were previously inoculated with LM following the same protocol. Those inoculations occurred 12–48 months prior to their terminal inoculation series. The timeline and number of previous inoculations per animal are detailed in Table 1 .
To assure a live dose of LM was given to the subjects, 500 μL of the 10mL whipping cream inoculum was diluted in phosphate-buffered saline (PBS; Catalog #P5368, Sigma-Aldrich, St. Louis, MO), plated on Trypticase soy agar with 5% sheep blood (Becton Dickinson, Sparks, MD), and incubated at 37◦C. Subsequent growth on the agar plates was quantified to confirm the dose of live LM inoculum delivered.38 (link)
To assure a live dose of LM was given to the subjects, 500 μL of the 10mL whipping cream inoculum was diluted in phosphate-buffered saline (PBS; Catalog #P5368, Sigma-Aldrich, St. Louis, MO), plated on Trypticase soy agar with 5% sheep blood (Becton Dickinson, Sparks, MD), and incubated at 37◦C. Subsequent growth on the agar plates was quantified to confirm the dose of live LM inoculum delivered.38 (link)
