Acridine orange staining and viability of the coccoid form of Campylobacter upsaliensis.

Conversion of Campylobacter upsaliensis to the coccoid form during aerobic incubation at 37 degrees C was not prevented by treatment with chloramphenicol and was accompanied by severe decreases in isocitrate dehydrogenase activity and oxygen uptake. Although the coccoid forms fluoresced orange-red by acridine orange staining, agarose gel electrophoresis indicated an extensive degradation of the ribosomal RNA. This suggests that acridine orange staining may not be a good indicator of viability and that the coccoid form of C. upsaliensis at 37 degrees C is degenerative rather than part of the life cycle.

Invasion of human epithelial cells by Campylobacter upsaliensis.

Few data exist on the interaction of Campylobacter upsaliensis with host cells, and the potential for this emerging enteropathogen to invade epithelial cells has not been explored. We have characterized the ability of C. upsaliensis to invade both cultured epithelial cell lines and primary human small intestinal cells. Epithelial cell lines of intestinal origin appeared to be more susceptible to invasion than non-intestinal-derived cells. Of three bacterial isolates studied, a human clinical isolate, CU1887, entered cells most efficiently. Although there was a trend towards more efficient invasion of Caco-2 cells by C. upsaliensis CU1887 at lower initial inocula, actual numbers of intracellular organisms increased with increasing multiplicity of infection and with prolonged incubation period. Confocal microscopy revealed C. upsaliensis within primary human small intestinal cells. Both Caco-2 and primary cells in non-confluent areas of the infected monolayers were substantially more susceptible to infection than confluent cells. The specific cytoskeletal inhibitors cytochalasin B, cytochalasin D and vinblastine attenuated invasion of Caco-2 cells in a concentration-dependent manner, providing evidence for both microtubule- and microfilament-dependent uptake of C. upsaliensis. Electron microscopy revealed the presence of organisms within Caco-2 cell cytoplasmic vacuoles. C. upsaliensis is capable of invading epithelial cells and appears to interact with host cell cytoskeletal structures in order to gain entry to the intracellular environment. Entry into cultured primary intestinal cells ex vivo provides strong support for the role of host cell invasion during human enteric C. upsaliensis infection.