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ABSTRACT: The occurrence of antimicrobial resistance in microorganisms isolated from eggshells, especially Enterococcus, might serve as a parameter to assess the selection of bacteria due to the use of drugs in the diet of laying hens. In order to evaluate the frequency and the antimicrobial resistance profile of Enterococcus spp. isolated from the eggshells, 225 Hy-line laying hens were submitted to a 25-d long trial. The treatments were the following: hens fed ration without antimicrobials (control) and groups that received oxytetracycline (10 mg kg−1), doxycycline (20 mg kg−1), lincomycin (50 mg kg−1), and enrofloxacin (10 mg kg−1) in the ration for 5 d. Six replications were analyzed per treatment, composed of a pool of 5 eggs each. They were collected before treatment and on days 3, 6, 15, and 25, totaling 150 samples. Eggshells were submitted to counts of Enterococcus spp., which were identified by proteomic analyses. Antimicrobial resistance was determined by the disk-diffusion test. It was observed that 97.3% (n = 146) of the samples were contaminated with Enterococcus spp. There were no differences (P > 0.05) in the bacterial counts between treatments on the same day of evaluation. E. faecalis and E. faecium were the most frequent on the eggshells of all treatments. Multiresistance to the four classes of antimicrobials was also verified in the isolated bacteria. A total of 83% of the Enterococcus isolates showed resistance to neomycin, which was not administered to the hens, demonstrating an environmental problem. Thus, feeding laying hens with diets added with antimicrobials induces drug resistance in Enterococcus spp., isolated form the eggshells.