Analyses of biological activities of CDT. (A) Cytolethal distending effect by OMVs on HCT8 cells. HCT8 cells without treatment (a, b), HCT8 cells treated with 50 μl of OMVs (total protein concentration was 60 μg/ml) from wild type C. jejuni strain 81-176 (c, d), or from the cdtA::km mutant (e, f). After 72 hours of treatment the actin filaments and nuclei were stained with phalloidin and DAPI, respectively, as described in materials and methods. Upper panels (a, c, e) show merged images from staining with both dyes and lower panels (b, d, f) show images from DAPI staining only. Bars represent 40 μm. (B) Effect of thymidine uptake on HCT8 cells after treatment with OMVs from wild type C. jejuni strain 81-176 and the cdt::km mutant strain DS104 for 48 h. Cells were grown in 96-well plates and 10 μl of OMVs were added to the wells. The results are from triplicate wells and two independent experiments. Data are expressed as mean percentage (± SE).