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Campylobacter fetus adheres to and enters INT 407 cells
1Department of Biology, St. Francis Xavier University, Antigonish, NS B2G 2W5, Canada. lgraham@stfx.ca
Abstract:
Campylobacter fetus is a Gram-negative bacterial pathogen of humans and ungulates and is normally transmitted via ingestion of contaminated food or water with infection resulting in mild to severe enteritis. However, despite clinical evidence that C. fetus infection often involves transient bacteremic states from which systemic infection may develop and the frequent isolation of C. fetus from extra-intestinal sites, this organism displays very poor invasiveness in in vitro models of infection. In this study, immunofluorescence microscopy and gentamicin protection assays were used to investigate the ability of six clinical isolates and one reference strain of C. fetus to adhere to and invade the human intestinal epithelial cell line, INT 407. During an initial 4-h infection period, all C. fetus strains were detected intracellularly using both techniques, though adherence and internalization levels were very low when determined from gentamicin protection assays. Microscopy results indicated that during a 4-h infection period, four of the five clinical strains tested were adherent to 41.3-87.3% of INT 407 cells observed and that 25.2-34.6% of INT 407 cells contained intracellular C. fetus. The C. fetus reference strain displayed the lowest levels of adherence and internalization. A modified infection assay revealed that C. fetus adherence did not necessarily culminate in internalization. Despite the large percentage of INT 407 cells with adherent bacteria, the percentage of INT 407 cells with intracellular bacteria remained unchanged when incubation was extended from 4 h to 20 h. However, microscopy of INT 407 cells 24 h postinfection (p.i.) revealed that infected host cells contained clusters of densely packed C. fetus cells. Gentamicin protection assays revealed that intracellular C. fetus cells were not only viable 24 h p.i. but also that C. fetus had increased in number approximately three- to fourfold between 4 and 24 h p.i., indicative of intracellular replication. Investigation of the role of the host cell cytoskeleton revealed that pretreatment of host cells with cytochalasin D, colchicine, vinblastine, taxol, or dimethyl sulfoxide (DMSO) did not impact upon C. fetus adherence or internalization of INT 407 cells. Microscopy indicated neither rearrangement nor colocalization of either microtubules or microfilaments in INT 407 cells in response to C. fetus adherence or internalization. Together, these data indicate that clinical isolates of C. fetus are capable of adhering, entering, and surviving within the nonphagocytic epithelial cell line, INT 407.
Insights
Campylobacter fetus, a bacterial pathogen, can adhere to and enter human intestinal cells. Despite poor in vitro invasiveness, clinical isolates replicate intracellularly, indicating potential for systemic infection.
Area of Science:
- Microbiology
- Infectious Diseases
- Cell Biology
Background:
- Campylobacter fetus is a Gram-negative bacterium causing human and ungulate infections, typically via contaminated food/water.
- Despite causing enteritis and sometimes bacteremia, C. fetus shows poor in vitro invasiveness.
- Clinical observations suggest C. fetus can establish extra-intestinal infections, contradicting in vitro findings.
Purpose of the Study:
- To investigate the adherence and invasion capabilities of Campylobacter fetus in a human intestinal epithelial cell line (INT 407).
- To determine if clinical isolates of C. fetus can survive and replicate within non-phagocytic host cells.
- To explore the role of the host cell cytoskeleton in C. fetus interaction with intestinal epithelial cells.
Main Methods:
- Utilized immunofluorescence microscopy and gentamicin protection assays to quantify C. fetus adherence and intracellular presence.
- Infected INT 407 cells with six clinical isolates and one reference strain of C. fetus.
- Investigated host cell cytoskeleton involvement using cytochalasin D, colchicine, vinblastine, taxol, and DMSO treatments.
Main Results:
- All tested C. fetus strains adhered to and were found intracellularly in INT 407 cells, though initial gentamicin assays showed low levels.
- Microscopy revealed significant adherence (41.3-87.3%) and internalization (25.2-34.6%) by clinical isolates.
- Intracellular C. fetus replicated threefold to fourfold within 24 hours post-infection, and host cytoskeleton disruption did not affect adherence or invasion.
Conclusions:
- Clinical isolates of Campylobacter fetus demonstrate the ability to adhere to, enter, and replicate within human intestinal epithelial cells (INT 407).
- The host cell cytoskeleton is not essential for C. fetus entry or survival within INT 407 cells.
- These findings highlight C. fetus's capacity for intracellular survival and replication, potentially explaining its ability to cause systemic infections despite poor in vitro invasiveness.