The Role of Plasmid and Resistance Gene Acquisition in the Emergence of ST23 Multi-Drug Resistant, Hypervirulent Klebsiella pneumoniae

Multidrug-resistant (MDR) hypervirulent Klebsiella pneumoniae (hvKp) sequence type (ST) 23 (MDR-ST23-hvKp) is emerging in China. Despite its increasing importance, this pathogen has not yet been subject to detailed genomic interrogation. We identified 28 ST23 Kp isolated from three hospitals in China. The organisms were subjected to antimicrobial susceptibility testing and whole-genome sequencing (WGS). These novel genomic sequences were analyzed in combination with 218 publicly available genome sequences. We performed molecular serotyping and subtyping, assessed the composition of virulence-associated and antimicrobial resistance (AMR) genes, and determined mobile elements associated with horizontal gene transfer. Two MDR-ST23-hvKp were sequenced by long-read sequencing. The genetic characteristics of MDR and non-MDR isolates were compared. Among the 28 novel ST23 isolates, all were hvKp and 2/28 (7.1%) were MDR-hvKp. From the collection of 246 genomes, KL1 was the predominant serotype (224/246; 91.1%) and the siderophore combination of YbST46-CbST29-AbST1-SmST2 was dominant (101/246; 41.1%); 34/246 (13.8%) organisms belonged to MDR-ST23-hvKp. IncF and IncR plasmid replicons were significantly more prevalent in the MDR group (P < 0.05) than in the non-MDR group. IS26 was commonly involved in AMR acquisition. We observed that the acquisition of AMR genes within the ST23-hvKp was not associated with a loss of virulence genes. A 28-bp fusion site was highly conserved with two copies of the virulence-associated plasmid in ST23-hvKp, and we harbored by some of the IncFII plasmids of MDR-ST23-hvKp. Our data suggest that MDR-ST23-hvKp has undergone multiple independent genetic acquisition and recombination events within different sublineages. Notably, the acquisition of IncFII plasmids and/or IS26 contributed to the horizontal transfer of AMR genes within ST23-hvKp. Genomic surveillance is essential for further tracking of kMDR-ST23-hvKp. IMPORTANCE Hypervirulent Klebsiella pneumoniae (hvKp) has become the dominant pathotype in hospitals recently. The sequence type (ST) 23 hvKp, which are more commonly associated with the community-acquired infections previously, may have the capacity to acquire multidrug-resistant (MDR) phenotypes creating a new "superbug" (MDR-hvKp) in hospital. In the present study, we studied the associations of MDR and hypervirulence among ST23 K. pneumoniae from our strain collection and publicly accessible genome data. By comparative analysis of the carriage of resistance genes, virulence genes plasmid replicon types, and plasmid sequences, we found that IncFII plasmids were significantly more prevalent in MDR isolates and IS26 were commonly involved in resistance gene acquisition. We also discovered new MDR plasmids. These results provided an overview landscape of the genetic elements associated with MDR-ST23-hvKp based on currently accessible genome data and calling for further genomic surveillance and well-designed control studies of MDR-ST23-hvKp.