Gram-negative isolates (n = 84) from 71% of free-ranging Eastern box turtles (Terrapene carolina carolina) in Illinois and Tennessee, United States, demonstrated resistance to at least one antibiotic while 30% of isolates demonstrated resistance to two or more antibiotics. Resistance was observed against cefoxitin, amoxicillin-clavulanic acid, cefazolin, ampicillin, ticarcillin, cefovecin, and ceftiofur. Gram-positive bacteria were isolated from 49 turtles, and all were observed to be resistant to two or more antibiotics. Gram-positive isolate resistance was observed to penicillin, cefoxitin, oxacillin, clindamycin, amikacin, enrofloxacin, cefovecin, ceftiofur, cefazolin, marbofloxacin, gentamicin, erythromycin, trimethoprim-sulfamethoxazole, and chloramphenicol. Health parameters including packed cell volume, total white blood cell count (WBC), total solids (TS), and weight were not significantly different based on antibiotic resistance patterns; however, decreasing WBC and TS were observed when the number of antibiotic-resistant detections in Gram-positive bacteria increased.