The spread of KPC-producing Enterobacteriaceae in Spain: WGS analysis of the emerging high-risk clones of Klebsiella pneumoniae ST11/KPC-2, ST101/KPC-2 and ST512/KPC-3

Abstract

Objectives: We analysed the microbiological traits and population structure of KPC-producing Enterobacteriaceae isolates collected in Spain between 2012 and 2014. We also performed a comparative WGS analysis of the three major KPC-producing Klebsiella pneumoniae clones detected.

Methods: Carbapenemase and ESBL genes were sequenced. The Institut Pasteur MLST scheme was used. WGS data were used to construct phylogenetic trees, to identify the determinants of resistance and to de novo assemble the genome of one representative isolate of each of the three major K. pneumoniae clones.

Results: Of the 2443 carbapenemase-producing Enterobacteriaceae isolates identified during the study period, 111 (4.5%) produced KPC. Of these, 81 (73.0%) were K. pneumoniae and 13 (11.7%) were Enterobacter cloacae. Three major epidemic clones of K. pneumoniae were identified: ST11/KPC-2, ST101/KPC-2 and ST512/KPC-3. ST11/KPC-2 differed from ST101/KPC-2 and ST512/KPC-3 by 27 819 and 6924 SNPs, respectively. ST101/KPC-2 differed from ST512/KPC-3 by 28 345 SNPs. Nine acquired resistance genes were found in ST11/KPC-2, 11 in ST512/KPC-3 and 13 in ST101/KPC-2. ST101/KPC-2 had the highest number of virulence genes (20). An 11 bp deletion at the end of the mgrB sequence was the cause of colistin resistance in ST512/KPC-3.

Conclusions: KPC-producing Enterobacteriaceae are increasing in Spain. Most KPC-producing K. pneumoniae isolates belonged to only five clones: ST11 and ST512 caused interregional spread, ST101 caused regional spread and ST1961 and ST678 produced independent hospital outbreaks. ST101/KPC-2 had the highest number of resistance and virulence genes. ST101/KPC-2 and ST512/KPC-3 were recently implicated in the spread of KPC in Italy.

Figure 1.

Population structure of K. pneumoniae expressing bla_KPC genes. (a) Maximum-likelihood tree showing the relationship between the isolates sequenced in this study and similar strains whose sequences were obtained from the GenBank database. Branch length is indicative of the number of SNPs. (b) Minimum spanning tree of K. pneumoniae genomes as determined on the basis of the 694 core genome multilocus ST. Identification of isolates includes the ST; Spanish sequenced isolates are highlighted in boxes (a) or black circles (b).

Figure 2.

Distribution of multidrug resistance loci and virulence genes in the genomes of bla_KPCK. pneumoniae clones sequenced in Spain in comparison with strains isolated in other countries. The names of genes responsible for drug resistance (resistome) and virulence are found at the top of a heat map in which black rectangles indicate the presence of a gene and white rectangles indicate its absence. KPN1H1/ST258 was detected during an outbreak in the USA, ST512-K30BO was isolated during an outbreak in Italy and HS11286/ST11 was described in China.