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Dexamethasone (Dex), a synthetic glucocorticoid (GC), in feed has been shown to increase gut permeability via stress-mediated mechanisms, but the exact mode of action on gut barrier function is not fully understood. Stress has been reported to alter the profile and virulence of intestinal flora predisposing for opportunistic disease. This study aimed to evaluate the relationship between dietary Dex and recoverable intestinal microbial profile in broilers to better understand mode of action and refine future uses of the model. Three experiments were conducted that administered Dex-treated feed for one week in conjunction with the antibiotics BMD (bacitracin methylene disalicylate) or Baytril ® (enrofloxacin) to evaluate if enteric microbial mechanisms were important in Dex-induced permeability. Serum fluorescein isothiocyanate-dextran (FITC-d) and bacterial translocation (BT) have been reported to increase after Dex treatment and were used to assess gut epithelial leakage. Shifts in bacterial profiles were also measured on selective agar. Combining Dex with BMD or Baytril resulted in increased ( P < 0.05) serum FITC-d versus Dex-only. Additionally, Baytril did not reduce aerobic BT and bacterial profiles remained similar after Dex. These results suggest a minimal role of intestinal microbes in Dex-induced changes to intestinal barrier function.