Tracking the Emergence and Dissemination of a blaNDM-23 Gene in a Multidrug Resistance Plasmid of Klebsiella pneumoniae

Abstract

Since the discovery of bla_NDM-1, NDM β-lactamases have become one of the most widespread carbapenemases worldwide. To date, 43 different NDM variants have been reported but some, such as bla_NDM-23, have not been characterized in detail yet. Here, we describe the emergence of a novel bla_NDM-23 allele from a bla_NDM-1 ancestor and the multidrug resistance plasmid that has disseminated it through a Klebsiella pneumoniae ST437 clone in several Spanish hospitals. Between 2016 and 2019, 1,972 isolates were collected in an epidemiological survey for extended-spectrum-β-lactamase (ESBL)-producing Klebsiella pneumoniae in the Comunitat Valenciana (Spain). Three carbapenem-resistant strains failed to be detected by carbapenemase-producing Enterobacteriaceae (CPE) screening tests. These isolates carried a bla_NDM-23 gene. To characterize this gene, its emergence, and its dissemination, we performed antimicrobial susceptibility tests, hybrid sequencing with Illumina and Nanopore technologies, and phylogenetic analyses. The MICs of the bla_NDM-23 allele were identical to those of the bla_NDM-1 allele. The bla_NDM-23 allele was found in 14 isolates on a 97-kb nonmobilizable, multidrug-resistant plasmid carrying 19 resistance genes for 9 different antimicrobial families. In this plasmid, the bla_NDM-23 gene is in the variable region of a complex class 1 integron with a singular genetic environment. The small genetic distance between bla_NDM-23-producing isolates reflects a 5-year-long clonal dispersion involving several hospitals and interregional spread. We have characterized the genomic and epidemiological contexts in the emergence and community spread of a new bla_NDM-23 allele in a multidrug resistance (MDR) plasmid of Klebsiella pneumoniae. IMPORTANCE At a time when antimicrobial resistance has become one of the biggest concerns worldwide, the emergence of novel alleles and extremely drug-resistant plasmids is a threat to public health worldwide, especially when they produce carbapenem resistance in one of the most problematic pathogens, such as Klebsiella pneumoniae. We used genomic epidemiology to describe the emergence of a novel NDM-23 allele and identify it in a MDR plasmid that has disseminated through a K. pneumoniae ST437 clone in several hospitals in Spain. Using bioinformatic and phylogenetic analyses, we have traced the evolutionary and epidemiological route of the new allele, the hosting plasmid, and the strain that carried both of them from Pakistan to Spain. A better understanding of the NDM-producing K. pneumoniae populations and plasmids has made evident the spread of this clone through the region, enhancing the importance of genomic surveillance in the control of antimicrobial resistance.

Keywords: Klebsiella pneumoniae; MDR plasmid; XDR plasmid; complex class 1 integron; genomic epidemiology; novel NDM.

FIG 1

(A) Gene content and structure of plasmid p146KP-NDM23. The bla_NDM-23 embedded in the second variable part of a ISCR1 complex class 1 integron is highlighted in yellow. The middle ring shows the BLAST comparison between p146KP-NDM23 and p179KP-HG. The inner ring shows the GC skew of the plasmid. (B) ISCR1 complex class 1 integron where the bla_NDM-23 gene is embedded.

FIG 2

Structure comparison of plasmids pLK78, p146KP-NDM23, pKP179-NDM1, and pKDO1. pLK78 shares similarity with the integron sequence found in pKP146-NDM23 and pKP179-NDM1, while pKDO1 shares similarity with the backbone of those plasmids.

FIG 3

Whole-genome maximum-likelihood tree of ST437 KL36 isolates. Spanish isolates are highlighted in pink. Colors in the inner ring (1) indicate the country of origin of each sample. A star inside this ring indicates that the isolate was sequenced in this work. Colors in the center ring (2) represent carbapenem-resistant genes. In the outer ring (3), blue bars show the resistance score while purple bars show the virulence score obtained from Kleborate (30). The scale represents nucleotide divergence as substitutions per site.

FIG 4

Dissemination of the ST437 clone carrying the bla_NDM23 gene involves 5 different hospitals over 5 years. Isolates sequenced in this work are marked with a star inside the bla_NDM strip. For each isolate, the structural variants of plasmid p146KP-NDM23 are shown.