Whole-genome sequencing resolves a polyclonal outbreak by extended-spectrum beta-lactam and carbapenem-resistant in a Portuguese tertiary-care hospital.

has emerged as an important nosocomial pathogen, with whole-genome sequencing (WGS) significantly improving our ability to characterize associated outbreaks. Our study sought to perform a genome-wide analysis of multiclonal isolates (=39; 23 patients) producing extended spectrum beta-lactamases and/or carbapenemases sourced between 2011 and 2016 in a Portuguese tertiary-care hospital. All isolates showed resistance to third-generation cephalosporins and six isolates (five patients) were also carbapenem resistant. Genome-wide-based phylogenetic analysis revealed a topology representing ongoing dissemination of three main sequence-type (ST) clades (ST15, ST147 and ST307) and transmission across different wards, compatible with missing links that can take the form of undetected colonized patients. Two carbapenemase-coding genes were detected: , located on a transposon, and on a novel class 3 integron. Additionally, four genes coding for ESBLs (, , and ) were also detected. ESBL horizontal dissemination across five clades is highlighted by the similar genetic environments of gene upstream of IS on a -like transposon. Overall, this study provides a high-resolution genome-wide perspective on the epidemiology of ESBL and carbapenemase-producing in a healthcare setting while contributing for the adoption of appropriate intervention and prevention strategies.