Various arrangements of mobile genetic elements among CC147 subpopulations of Klebsiella pneumoniae harboring blaNDM-1: a comparative genomic analysis of carbapenem resistant strains

Abstract

Background: Certain clonal complexes (CCs) of Klebsiella pneumoniae such as CC147 (ST147 and ST392) are major drivers of bla_NDM dissemination across the world. ST147 has repeatedly reported from our geographical region, but its population dynamics and evolutionary trajectories need to be further studied.

Methods: Comparative genomic analysis of 51 carbapenem-nonsusceptible strains as well as three hypervirulent K. pneumoniae (hvKp) recovered during 16-months of surveillance was performed using various bioinformatics tools. We investigated the genetic proximity of our ST147 strains with publicly available corresponding genomes deposited globally and from neighbor countries in our geographic region.

Results: While IncL/M plasmid harboring bla_OXA-48 was distributed among divergent clones, bla_NDM-1 was circulated by twenty of the 25 CC147 dominant clone and were mostly recovered from the ICU. The NDM-1 core structure was bracketed by a single isoform of mobile genetic elements (MGEs) [ΔISKpn26-NDM-TnAs3-ΔIS3000-Tn5403] and was located on Col440I plasmid in 68.7% of ST392. However, various arrangements of MGEs including MITESen1/MITESen1 composite transposon or combination of MITESen1/ISSen4/IS903B/IS5/ISEhe3 on IncFIb (pB171) were identified in ST147. It seems that ST392 circulated bla_NDM-1 in 2018 before being gradually replaced by ST147 from the middle to the end of sample collection in 2019. ST147 strains possessed the highest number of resistance markers and showed high genetic similarity with four public genomes that harbored bla_NDM-1 on the same replicon type. Mainly, there was a convergence between clusters and isolated neighboring countries in the minimum-spanning tree. A conserved arrangement of resistance markers/MGEs was linked to methyltransferase armA which was embedded in class 1 integron in 8 isolates of ST147/ST48 high-risk clones.

Conclusion: Our findings highlight the dynamic nature of bla_NDM-1 transmission among K. pneumoniae in Iran that occurs both clonally and horizontally via various combinations of MGEs. This is the first analysis of Iranian ST147/NDM + clone in the global context.

Keywords: High-risk clones; Iran; Klebsiella pneumoniae; MGEs; OXA-48; Phylogeny; ST147; Whole genome sequencing; armA.

Fig. 1

Phylogenetic tree of study 54 K. pneumoniae isolates. Maximum-likelihood phylogenic tree clustered different clones (STs) and CC147 strains (ST147, ST392) were identified as the dominant clone with the highest prevalence of varied resistance elements. The tree was constructed from the whole genome SNPs arising by mutation and labeling and visualization were done by using the iTOL web-interface. The data presented are the date of isolation for CC147, sequence types, capsular types, Integrative and conjugative Elements (ICEs), acquired resistance markers and amikacin/gentamicin susceptibility phenotypes. The hvKp strains have been shown in red

Fig. 2

Minimum spanning tree of CC147 based on the core genome MLST. The 25 CC147 strains were analyzed based on comparing 2365 alleles calculated in Ridom SeqSphere. Cluster was defined based on the maximum allelic distance of ≤ 15 alleles. Inside the bubbles the isolate ID numbers are shown, and the allelic distances between isolates are represented on the lines connecting them. Clusters 1 (ST392) and 2 (ST147) are shown by pink and pale blue zones, respectively

Fig. 3

Bar-plot displaying the prevalence of different replicon types and their association with STs. The Y-axis represents the number of positive isolates in each STs

Fig. 4

The virulence genes content of K. pneumoniae. A BLAST Ring Image Generator (BRIG) image shows the presence of virulence factors among different STs. The FFN format file of hvKp (strain No. 290 identified as ST86) was downloaded from VFDB website, and open reading frames (ORFs) are annotated based on the VFDB results and used as reference strains to draw the image. Aligning between the regions of interest and each genome are shown as Colored segments (indicate > 70% similarity), and gray segments (indicate > 50% similarity). From inside to outside of the figure: “deep sky blue “rings”; ST14, “blue” rings; ST40, “purple” rings; ST45, “yellow” rings; ST48, “aquamarine” rings; ST147, “red” rings: ST392, “blue-green [teal]” rings; ST377, “pale green” rings; ST1308. The other undefined colored rings are STs that include one or two strains

Fig. 5

Maximum-likelihood phylogeny of ST147/NDM + strains based on whole genome SNPs analysis. A phylogenetic reconstruction of 183 strains (including the 174 downloaded genomes and our 9 study strains) with K. pneumoniae strain 4/1–2 (GenBank ID: CP023839.1, NDM + strain), as the reference strain is shown. Colored strips surrounding the phylogram represent the country of origin of each strain, capsular types, ICE types, carriage of NDM/CTX-M-15, OXA-types, and genes involved in gentamicin and amikacin resistance

Fig. 6

Minimum Spanning Tree showing distance based on cgMLST of 1826 genes using the parameter ‘pairwise ignoring missing values for 130 ST147 genomes. Each circle is named with the geographical origin, including DTU: our ST147 strains, Ir: Iran, Tu: Turkey, In: India, P: Pakistan, Is: Israel, Le: Lebanon, Ru: Russia, UAE: United Arab Emirates. The data of carbapenemase genes is shown by the coloured rings inside each bubble

Fig. 7

Genetic context of resistance markers among study population. a contig harbouring bla_NDM-1 in ST392, b–d three forms of MGEs bracketing NDM-1, e, f two isoforms of MGEs arrangements in bla_CTX-M-15 harbouring contigs, g the integron 1 cassette structure harbouring the armA