Intensive care units (ICU) are the epicenter of antimicrobial resistance (AMR), and patients' infections are mainly caused by Gram-negative bacteria (GNB).To describe the frequency and trends in AMR of GNB deriving from the clinical samples of ICU patients at a tertiary care hospital in Mérida, Yucatán.Study which included the review of laboratory reports of all bacteriological samples collected from patients admitted to neonatal, pediatric and adult ICU from January 1 2019 to December 31 2021.433 GNB isolates were recovered, with Klebsiella pneumoniae being the most predominant isolate (n = 117; 27.02%). The majority of GNB were recovered from bronchial secretions (n = 163). Overall, GNB showed high resistance rates to ampicillin (89.48%), ampicillin/sulbactam (66.85%), cephalosporins (58.52-93.81%), tobramycin (58.06%), and tetracycline (61.73%). Among GNB, 73.90% and 68.53% exhibited multidrug-resistant, and highly resistant microorganisms' profiles, respectively, and 47.54% of Acinetobacter baumannii exhibited an extensively drug-resistant profile. A total of 80.33% of A. baumannii was carbapenem-resistant, and 83.76% of K. pneumoniae strains were ESBL-producing.Our data could be helpful to improve the empirical therapy and the infection-control program.Licencia CC 4.0 (BY-NC-ND) © 2023 Revista Médica del Instituto Mexicano del Seguro Social.