Antibiotic resistance is one of the main challenges worldwide due to the high morbidity and mortality caused by infections produced by resistant bacteria. In Colombia, this problem has been studied mainly from the clinical perspective; however, it is scarcely studied in the leachates produced in landfills. The objective of this study was to detect, identify and determine the antibiotic sensitivity profile of Enterobacterales isolated from a leachate treatment plant located in Cali, Colombia. Detection was performed using selective culture media, bacterial identification using Matrix-Assisted Laser Desorption/Ionization-Time-Of-Flight (MALDI-TOF, bioMérieux) and by sequencing the gene coding for the 16S ribosomal RNA subunit when discrepancies were observed between phenotypic characteristics and MALDI-TOF. Antibiotic sensitivity profiling was determined using the automated VITEK®2 system (bioMérieux). Twenty-one isolates were obtained, of which Klebsiella pneumoniae was the most frequent (23.8%), and 34% of the isolates showed decreased sensitivity to beta-lactam antibiotics such as cefoxitin, ampicillin/sulbactam and piperacillin/tazobactam. These findings suggest that leachates from landfills could be a reservoir of pathogenic bacteria carrying antibiotic resistance determinants, so periodic microbiological characterization of these effluents should be performed, promoting the One Health approach.