This study aimed to determine the frequency of colistin resistance in Pseudomonas aeruginosa isolates obtained from three healthcare facilities in Lima and cryopreserved at the Laboratorio de Resistencia Antimicrobianos e Inmunopatología of the Universidad Peruana Cayetano Heredia (UPCH). The co-listin broth disk elution method was used for the phenotypic identification of colistin resistance. We detected the expression of the mcr-1 gene by using the phenotypic diffusion method with combined colistin and ethylenediaminetetraacetic acid (EDTA) disks; and polymerase chain reaction (PCR) was used for molecular identification of the gene. Of the 97 isolates, 7 (7.2%) were resistant to colistin; howe-ver, none carried the mcr-1 gene. This is the first report from Peru on clinical isolates of colistin-resistant Pseudomonas aeruginosa, which suggests the need for implementation of appropriate methodologies for the epidemiological surveillance of colistin-resistant pathogens.