To investigate evidence of cross-contamination and to determine patterns of antimicrobial drug susceptibility of Entero-coccus isolates in a commercial cattle processing system, samples were collected from 60 cattle shipped to a commercial abattoir. Enterococcus isolates were recovered from fecal and hide samples collected immediately before shipment from a feedlot to the abattoir, from postexsanguination hide samples at the abattoir, and from carcass samples collected after hide removal (preevisceration) and in the cooler. Of the fecal samples, 53.9% were culture positive for Enterococcus. Of hide samples collected at the feedlot, 77.8% were positive for Enterococcus, significantly lower (P < 0.01) than the proportion of hides that were culture positive at the abattoir (96.1%). For preevisceration carcass samples, Enterococcus was recovered from 58.3% of carcasses. Only 8.3% of the carcasses sampled in the cooler yielded Enterococcus. Resistance among Enterococcus isolates was common regardless of the type or location of sample from which the isolate was recovered. All 279 Enterococcus isolates were resistant to at least one antimicrobial drug, and 179 (64.2%) of these isolates were resistant to at least six drugs. The most common resistance was to chloramphenicol (100% of isolates) followed by flavomycin (90.3%), lincomycin (87.8%), tylosin (78.5%), erythromycin (76.3%), tetracycline (58.9%), quinupristin-dalfopristin (47.7%), bacitracin (17.9), streptomycin (9.0%), ciprofloxacin (1.4%), linezolid (0.7%), and salinomycin (0.4%). Enterococcus isolates also were characterized using pulsed-field gel electrophoresis to evaluate molecular similarities. Similar or indistinguishable electrophoresis patterns were found among isolates recovered at the feedlot and in the plant, providing evidence that feedlot-origin bacterial isolates are being transferred from cattle to carcasses within the processing environment through cross-contamination.