The mechanisms responsible for the increasing prevalence of colistin-only-sensitive (COS) Pseudomonas aeruginosa isolates in a Spanish hospital were investigated. Pulsed-field gel electrophoresis revealed that 24 (50%) of the studied isolates belonged to the same clone, identified as the internationally spread sequence type 235 (ST235) through multilocus sequence typing. In addition to several mutational resistance mechanisms, an integron containing seven resistance determinants was detected. Remarkably, the extended-spectrum β-lactamase GES-1 and its Gly170Ser carbapenem- hydrolyzing derivative GES-5 were first documented to be encoded in a single integron. This work is the first to describe GES enzymes in Spain and adds them to the growing list of β-lactamases of concern (PER, VIM, and OXA) detected in ST235 clone isolates. Copyright © 2009, American Society for Microbiology. All Rights Reserved.