Genomic Characterization of Multidrug-Resistant and ESBL-Producing Klebsiella pneumoniae Isolated from Healthy Rabbits Intended for Human Consumption

<i>Klebsiella pneumoniae</i> is an important pathogen associated with multidrug resistance and virulence in both human and animal populations. While its prevalence and resistance patterns are well documented in clinical settings, data on <i>K. pneumoniae</i> in food-producing animals remain scarce. This study aimed to isolate and characterize multidrug-resistant <i>K. pneumoniae</i> strains from healthy rabbits raised for human consumption, with a focus on antimicrobial resistance genes, plasmid content, and associated mobile genetic elements. A total of 295 fecal samples were collected from rabbits across 20 commercial farms in northern Portugal. Isolates were confirmed using MALDI-TOF MS, tested for hypermucoviscosity, and subjected to antimicrobial susceptibility testing (EUCAST). Whole-genome sequencing (WGS) was performed to determine sequence types (STs), resistance genes, plasmids, and resistance determinants for metals and biocides. Six <i>K. pneumoniae</i> isolates were recovered, showing extensive antimicrobial resistance profiles, including ESBL genes such as <i>bla</i>_CTX-M-15, <i>bla</i>_SHV-28, and <i>bla</i>_TEM-1. The most frequent ST was ST307. Multiple genes resistant to heavy metals were identified. Plasmid analysis revealed the presence of IncFII, IncN, and ColRNAI types. Network analysis showed clusters of genetically related isolates and highlighted shared resistance mechanisms. The presence of multidrug-resistant <i>K. pneumoniae</i> in healthy rabbits destined for human consumption underscores the zoonotic potential of this species and the need for surveillance in the animal-food-human interface. These findings contribute to a better understanding of resistance ecology in the context of One Health.