Unraveling Cefiderocol Resistance in NDM- and OXA-48-Like Co-Producing Klebsiella pneumoniae Isolates Through Integrated Genomic and Phenotypic Analysis

Background/Objectives: The co-production of New Delhi metallo-β-lactamases (NDM) and OXA-48-like carbapenemases in Klebsiella pneumoniae represents a major therapeutic challenge due to extensive drug resistance and limited treatment options. This study aimed to investigate the molecular epidemiology, resistance profiles, and mechanisms associated with reduced susceptibility to cefiderocol in clinical isolates co-producing NDM and OXA-48-like carbapenemases. Methods: A total of 45 clinical K. pneumoniae isolates collected in healthcare settings in Northern Italy were analyzed. Antimicrobial susceptibility testing, including cefiderocol and aztreonam/avibactam, was performed according to EUCAST guidelines. Whole-genome sequencing was used to characterize sequence types, resistance determinants, virulence factors, plasmid replicons, and phylogenetic relationships. Mutations in iron uptake and transport genes were investigated in cefiderocol-resistant isolates. Results: Most isolates belonged to the high-risk clone ST147 (44/45) and were grouped into three main phylogenetic clusters. The isolates exhibited extensive multidrug resistance, with universal susceptibility only for aztreonam/avibactam. Cefiderocol resistance was observed in 42.2% of isolates and was unevenly distributed across the phylogeny. Mutations in iron uptake genes, particularly cirA and chrA, were identified in the majority of resistant isolates, although several strains retained wild-type sequences, indicating heterogeneous resistance mechanisms. Comparative phylogenetic analysis demonstrated close relatedness to international isolates, suggesting the global dissemination of related lineages. Conclusions: NDM- and OXA-48-like carbapenemase co-producing K. pneumoniae isolates are characterized by clonal dissemination, complex resistance profiles, and emerging cefiderocol resistance driven by multifactorial mechanisms. The preserved activity of aztreonam/avibactam highlights its potential as a key therapeutic option against these high-risk pathogens.