Introduction: Enterococcus spp is currently considered as one of the most important nosocomial pathogens. The antibiotic resistance of this group of bacteria is a particularly important problem in health centers. Objective: To characterize the antibiotic resistance of 50 Enterococcus spp strains isolated from hospitalized patients clinical samples. Materials and methods: We conducted a cross-sectional descriptive observational study in 50 clinical isolates of Enterococcus spp. Only one isolate per patient was analyzed. The identification and antibiotic susceptibility were studied by conventional and automated methods. The phenotypic analysis of glycopeptide resistance mechanisms was performed as recommended by the Spanish Society of Clinical Microbiology and Infectious Diseases. Results: Of 50 isolates obtained from clinical samples, 30 (60.0%) belonged to Enterococcus faecalis and 20 (40.0%) to Enterococcus faecium. The global resistance expressed by this genre was as follows: Ampicillin, 38/50 (76.0%); high-level gentamicin, 33/50 (66.0%); high-level streptomycin, 34/50 (68.0%); ciprofloxacin, 26/50 (52.0%); linezolid, 4/50 (8.0%); teicoplanin, 17/50 (34.0%); vancomycin, 25/50 (50.0%); minocycline, 31/50 (62.0%); tetracycline, 34/50 (68.0%); nitrofurantoin, 9/50 (18.0%). As regards glycopeptides, 25/50 (50.0%) showed a Van A mechanism and 10/50 (20.0%) a Van B mechanism. Conclusions: The isolates obtained at Hospital Hermanos Ameijeiras showed higher resistance rates than those reported in the consulted literature. The high percentage of vancomycin-resistant strains might have influenced the development of other Gram-positive bacteria resistant to this drug. This is the first report on Enterococcus faecium resistant to linezolid in a Cuban hospital.