Campylobacter fetus translocation across Caco-2 cell monolayers
Introduction
Campylobacter’s are resident intestinal flora of many birds and mammals though several species including Campylobacter jejuni, Campylobacter coli, Campylobacter lari, Campylobacter upsaliensis and Campylobacter fetus are human pathogens [1]. In Canada, C. jejuni is the most frequent cause of human enteritis with reports of C. fetus infection being substantially lower [2]. C. fetus infection however, is more often associated with bacteremia and invasive disease or prolonged, relapsing illness, especially in compromised individuals [1]. Both C. fetus subspecies are also recognized animal pathogens. C. fetus subsp. venerealis induces venereal infection in cattle which can result in infertility in cows [3] whereas C. fetus subsp. fetus causes septic abortion in cattle and sheep [3], [4]. Though C. fetus supsp. fetus typically initiates infection at mucosal surfaces, transmission remains poorly defined. It is proposed that most animal and human infections are initiated following ingestion of C. fetus subsp. fetus-contaminated food or water [3], [5], [6], [7]. That infection is frequently associated with transient bacteremia [4], [5], and that this organism may be recovered from extra-intestinal locations, implies an ability to translocate across the intestinal epithelial cell barrier. A mechanism for C. fetus dissemination from the intestine has, however, not been forth coming.
Bacterial translocation across intact epithelial barriers has been described in Ref. [8]. Three translocation mechanisms are relevant to the intestinal epithelium: exploitation of M cells, specialized intestinal epithelial cells which transport luminal antigens across the epithelial barrier to underlying lymphoid tissue; transcellular translocation or transcytosis, in which pathogens invade epithelial cells, migrate across the cytoplasm and egress from the opposite surface, and; paracellular translocation or paracytosis, in which bacteria travel extracellularly through tight junctions between adjacent epithelial cells. Translocation across intestinal epithelial barriers via all 3 mechanisms has been reported for C. jejuni [9], [10], [11], [12].
The invasive nature of C. fetus is well established in clinical and veterinary situations and uptake into cultured human intestinal epithelial cells (INT 407) has also been demonstrated [13]. Caco-2 cells, a human colonic cell line, differentiate to form distinct apical and basolateral surfaces separated by tight junctions and express markers characteristic of human small intestinal epithelium [14]. In the present study, Caco-2 cells cultured on permeable supports were used as a model system to investigate C. fetus translocation across intestinal epithelial cell barriers.
Section snippets
C. fetus translocates across Caco-2 cell monolayers
One hour following apical infection of differentiated Caco-2 monolayers, C. fetus 11686 was detected in the basolateral medium (Fig. 1A). Although numbers of translocating C. fetus increased each hour during the subsequent 5 h, transepithelial electrical resistance (TER) values remained at 700–900 Ω/cm2 indicating that monolayer integrity was maintained for the duration of the 6 h assay (Fig. 1B). Salmonella typhimurium was also detected in the basolateral medium 1 h p.i. (Fig. 1A), however
Discussion
Numerous mucosal pathogens translocate across epithelial barriers enroute to establishing disease states. The present study characterizes C. fetus translocation across model intestinal epithelial cell monolayers. Initial assays comparing C. fetus 11686 to S. typhimurium and E. coli, as positive and negative control bacteria, respectively, validated the use of Caco-2 cell monolayers as a model for C. fetus translocation. Consistent with previous reports [14], [15], translocation of S. typhimurium
Bacterial strains and culture conditions
Bacterial strains used in this study are presented in Table 1. Stock bacterial cultures were maintained in peptone–glycerol broth (10 g peptone/L, 5 g NaCl/L, 25% v/v glycerol) at −80 °C and for use in all studies, bacteria were passaged five or less times. C. fetus and C. jejuni were cultured on Columbia agar (Oxoid, Nepean, Ontario) supplemented with 5% v/v sheep blood (sBAP; Oxoid) at 37 °C for 48 h under microaerobic conditions (5% O2; 10% CO2; 85% N2). Isogenic S layer-deficient C. fetus
Acknowledgements
This work was funded by a Natural Sciences and Engineering Research Council Discovery Grant to LLG.
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Present address: Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5.