Ex Vivo Translocation of Escherichia coli (E. coli) F18 in Pig Ileum That Was Challenged with Norepinephrine (NE) or Adrenocorticotropic Hormone (ACTH) Compared with Controls

Rearing stresses induced by pathogens, weaning, or out of feed events may make pigs more susceptible to bacterial infections. One proposed mechanism is through the increased activation of the hypothalamic-pituitary axis and the production of ACTH or NE. Previous studies have shown that luminal catecholamines have modulated bacterial growth and expression of virulence associated factors. To study the effect of ACTH and NE, we used an F18 E. coli transfected with a red florescent protein (RFP)-plasmid which carries ampicillin resistance to assess bacterial growth in vitro and tissue translocation in an ex vivo intestinal model. We hypothesized that NE or ACTH would increase pathogenic E.coli proliferation and translocation in the ileum of pigs. We evaluated E. coli translocation in ex vivo ileum explants when the tissue was incubated with Krebs buffer (Control), 10 µM NE, or 0.01 µM ACTH in modified Ussing chambers. Prior to our ex vivo studies, 6 well plates containing MacConkey agar with ampicillin containing 0 and 0.01 µM ACTH or 0 and 10 µM NE each inoculated with 300 colony forming units (CFU) of RFP-E. coli. After 24 hour incubation CFU where assessed. There was an increase (P<0.01) in CFU due to NE (253 CFU) treated wells compared with Control (227 CFU), but due to ACTH (233 CFU) treatment. Next we studied the effect of 0.01 µM ACTH and 10 µM NE in ex vivo conditions utilizing 8 pigs at approximately 8 weeks of age that were fed a common diet for 5 weeks. Pigs were euthanized and ileum sections were flushed and collected. Treatments were applied to serosal chamber in modified Ussing chambers: 1) Control, 2) 0.01 µM ACTH, and 3) 10 µM NE. Each pig ileum section was represented in each of the 3 treatments. After a 20 min incubation period, 3 x 10⁸ CFU/ml RFP-E. coli was administered to each mucosal chamber. After 60 minutes, aliquots were taken from serosal chamber, plated, and CFU were determined after 24 hours. Ileum samples that were incubated with ACTH (7012 CFU) or NE (2883 CFU) had greater (P = 0.003) translocation of RFP-E. coli compared with Controls (207 CFU), but were not different from each other. These data indicate that ACTH or NE can increase ex vivo translocation of RFP-E. coli in the ileum and may partially explain the mechanism in which stress increases susceptibility to bacterial infections.