Additionally, 81–176cj0596 (“”high”" inoculum, orange squares) was inoculated at an OD600 of ~0.2. Deletion of cj0596

increases the ABT-263 cell line motility of C. jejuni Because motility plays an important role in invasion of host intestinal cells JPH203 and is required for animal colonization, the motility of C. jejuni 81–176, 81–176cj0596, and 81–176cj0596 + was compared at 37°C (Figure 6). The average diameter of the zone of motility for the wild-type was 39.3 mm ± 3.7 at 48 h. The mutant was significantly more motile with a zone diameter of 66.0 mm ± 2.4 (p < 0.0001). The revertant returned to wild-type motility levels with a zone diameter of 42.5 mm ± 3.0. A similar increase in motility was seen when the assay was performed at 42°C (data not shown). Thus, Cj0596 is involved BIRB 796 in vitro in the expression of motility. Figure 6 Motility of C. jejuni strains at 37°C. MH motility plates (0.4% agar) were inoculated with strains 81–176 (black), 81–176cj0596 (red) and 81–176cj0596 + (blue) and the

zones of motility were measured after 48 hours. Statistical significance (p < 0.05) is represented by an asterisk. Deletion of cj0596 increases the ability of C. jejuni to invade INT407 cells, but does not affect adherence or intracellular survival The possibility that Cj0596 plays a role in interaction with host cells was studied by comparing the adherence and invasion abilities of C. jejuni 81–176, 81–176cj0596, and 81–176cj0596 + in an in vitro assay using INT407 intestinal epithelial cells (Figure 7). The mean percentages of the inoculum that adhered were 8.5 (± 1.4), 7.2 (± 0.7), and 4.7 (± 1.2) for the wild-type, mutant, and revertant, respectively, demonstrating that deletion of Cj0596 does not significantly affect

the ability of C. jejuni to adhere to INT407 cells (p > 0.05; Figure 7A). In contrast, mutation of cj0596 had a significant effect on the invasion ability of C. jejuni. While the percentages of the wild-type and revertant inocula invading INT407 cells were 0.041 (± 0.007) and 0.027 (± 0.005), respectively, the cj0596 mutant showed a nearly 20-fold increase in invasion (0.76 ± 0.11, p < 0.001; Figure 7B). The gentamicin and Triton X-100 sensitivities of the three strains were tested to ensure that the invasion results were not due to altered killing of a strain, and no significant difference was found for either compound. Figure unless 7 Abilities of C. jejuni strains to adhere to and invade INT407 cells. Strains 81–176 (black), 81–176cj0596 (red) and 81–176cj0596 + (blue) were grown to mid-log phase in biphasic culture. INT407 monolayers were inoculated with bacteria at an MOI of ~40. After 3 h, the cells were washed and bacteria adhered were enumerated (A). Gentamicin was added to another plate of cells and incubation was continued for an additional 2 h after which the cells were washed and bacteria invaded were enumerated (B). Statistical significance (p < 0.001) is represented by two asterisks.