Tracking clonal and plasmid transmission in colistin- and carbapenem-resistant <i>Klebsiella pneumoniae</i>

The surveillance of mobile genetic elements facilitating the spread of antimicrobial resistance genes has been challenging. Here, we tracked both clonal and plasmid transmission in colistin- and carbapenem-resistant <i>Klebsiella pneumoniae</i> using short- and long-read sequencing technologies. We observed three clonal transmissions, all containing Incompatibility group (Inc) L plasmids and New Delhi metallo-beta-lactamase <i>bla</i>_NDM-1, although not co-located on the same plasmid. One IncL-<i>bla</i>_NDM-1 plasmid had been transferred between sequence type (ST) 392 and ST15, and the promiscuous IncL-<i>bla</i>_OXA-48 plasmid was likely shared between a singleton and a clonal transmission of ST392. Plasmids within clonal outbreaks and between clusters and STs had 0-2 single nucleotide polymorphism (SNP) differences, showing high stability upon transfer to same or different STs. The simplest explanation, without a comprehensive analysis with long-read sequencing, would be the spread of a single common IncL-<i>bla</i>_NDM-1 plasmid. However, here, we report <i>bla</i>_NDM-1 in five different plasmids, emphasizing the need to investigate plasmid-mediated transmission for effective containment of outbreaks.IMPORTANCEAntimicrobial resistance occupies a central stage in global public health emergencies. Recently, efforts to track the genetic elements that facilitate the spread of resistance genes in plasmids outbreaks, utilizing short-read sequencing technologies, have been described. However, incomplete plasmid reconstruction from short-read sequencing data hinders full knowledge about plasmid structure, which makes the exploration very challenging. In this study, we used both short- and long-read sequencing in clinical <i>Klebsiella pneumoniae</i> from University Hospital Centre Zagreb, Croatia, which was resistant to both last-resort antibiotics colistin and carbapenem. Our results show complex transmission networks and sharing of plasmids, emphasizing multiple transmissions of plasmids harboring carbapenem and/or colistin resistance genes between and within <i>K. pneumoniae</i> clones. Only full-length sequencing plus a novel way of determining plasmid clusters resulted in the complete picture, showing how future active monitoring of plasmids as a vital tool for infection prevention and control could be implemented.