Figure 4

Comparison of the invasive capacity of pre-inoculum and ex vivo obtained bacteria. HeLa cell invasion assay to evaluate the possibility of ex vivo Salmonella having its phagocytic activity influenced by some modulation due to host response. (A) Number of cells invaded and (B) number of bacteria per cell between aliquots of S_Typhi strain used in pre-inoculum conditions with ex vivo strain obtained from liver macerate. Pre-inoculum aliquot was obtained after growth of nine hours in LB broth and stored at − 80 °C for more than 15 days until the moment of the assay. The ex vivo aliquot was obtained from a macerated mouse liver coinfected on D4 and frozen at − 80 °C. The two ex vivo and pre-inoculum bacteria were thawed and set for invasion assay in HeLa cells. Pre-inoculum bacteria were diluted in a series of 10–10 and sown in SS-Agar medium to define the inoculum equivalence of ex vivo bacteria. The invasion time was 1, 3 and 6 h. After these periods, medium supplemented with gentamicin was added to the culture medium for 1 h in order to eliminate external bacteria. Slides were then washed, stained using the Romanowsky method and bacteria visualized. The data are shown in a box plot with mean and standard deviations. Asterisks: level of significance of *p < 0.05, **p < 0.005 and ***p < 0.0005.