Whole-genome typing and characterization of blaVIM19-harbouring ST383 Klebsiella pneumoniae by PFGE, whole-genome mapping and WGS
TL;DR: This study demonstrates the application of WGM to understanding the epidemiology of hospital-associated K. pneumoniae and presents the first longitudinal genomic characterization of the highly dynamic carbapenem-resistant ST383 K.neumoniae clone that is rapidly gaining importance in Europe.
Abstract: Objectives We utilized whole-genome mapping (WGM) and WGS to characterize 12 clinical carbapenem-resistant Klebsiella pneumoniae strains (TGH1-TGH12). Methods All strains were screened for carbapenemase genes by PCR, and typed by MLST, PFGE (XbaI) and WGM (AflII) (OpGen, USA). WGS (Illumina) was performed on TGH8 and TGH10. Reads were de novo assembled and annotated [SPAdes, Rapid Annotation Subsystem Technology (RAST)]. Contigs were aligned directly, and after in silico AflII restriction, with corresponding WGMs (MapSolver, OpGen; BioNumerics, Applied Maths). Results All 12 strains were ST383. Of the 12 strains, 11 were carbapenem resistant, 7 harboured blaKPC-2 and 11 harboured blaVIM-19. Varying the parameters for assigning WGM clusters showed that these were comparable to STs and to the eight PFGE types or subtypes (difference of three or more bands). A 95% similarity coefficient assigned all 12 WGMs to a single cluster, whereas a 99% similarity coefficient (or ≥10 unmatched-fragment difference) assigned the 12 WGMs to eight (sub)clusters. Based on a difference of three or more bands between PFGE profiles, the Simpson's diversity indices (SDIs) of WGM (0.94, Jackknife pseudo-values CI: 0.883-0.996) and PFGE (0.93, Jackknife pseudo-values CI: 0.828-1.000) were similar (P = 0.649). However, the discriminatory power of WGM was significantly higher (SDI: 0.94, Jackknife pseudo-values CI: 0.883-0.996) than that of PFGE profiles typed on a difference of seven or more bands (SDI: 0.53, Jackknife pseudo-values CI: 0.212-0.849) (P = 0.007). Conclusions This study demonstrates the application of WGM to understanding the epidemiology of hospital-associated K. pneumoniae. Utilizing a combination of WGM and WGS, we also present here the first longitudinal genomic characterization of the highly dynamic carbapenem-resistant ST383 K. pneumoniae clone that is rapidly gaining importance in Europe.
Summary (1 min read)
- These can be utilized to develop in 70 silico restriction maps using the same enzymes as used for experimentally-generated WGMs, 71 allowing comparison of restriction patterns and eventual identification of the genetic content of the 72 variable regions in the strains of interest.
- 5 73 In this study, the authors demonstrate the utility of WGM in conjunction with WGS for typing, 74 characterizing, and dissecting the genomic features of carbapenem-resistant K. pneumoniae 75 isolated at the Tzaneio General (TG) hospital, Piraeus, Greece.
Strain collection 84
- Twelve K. pneumoniae (TGH1-TGH12), harbouring blaVIM and/or blaKPC, and isolated from 85 patients admitted to the intensive care unit (n=8) or surgical ward (n=4) at the TG hospital during 86 2010-2013 were studied.
- Clinical data and strain characteristics are outlined in Table 1. 87.
Antimicrobial resistance profiling 88
- All 12 strains were screened for resistance to 17 antibiotics, including β-lactams with and without 89 β-lactamase inhibitors, by disk diffusion (Table 1).
- MICs of carbapenems (ertapenem, imipenem 90 and meropenem) were determined by Etest (bioMérieux Inc., Durham, NC), and results 91 interpreted according to CLSI cut-offs.
- 11 Strains were screened for presence of extended-spectrum 92 β-lactamase (ESBL) and carbapenemase genes by PCR and Sanger sequencing as described 93 previously.
MLST and PFGE 95
- MLST was performed as described previously for seven marker genes (gapA, infB, mdh, pgi, 96 phoE, rpoB, tonB), 15 and sequence types were assigned using the Institute Pasteur database 97 (www.pasteur.fr/mlst).
- Both parameters showed 186 similar (sub) clustering of WGMs, identifying two clusters, C1 and C2, and two singletons, C3 187 and C4 , with C1 and C2 each divided into two sub clusters and a singleton .
Did you find this useful? Give us your feedback
Cites methods from "Whole-genome typing and characteriz..."
...tent with the PFGE and MLST data combined, especially in strains differing in less than 3 bands (37, 38)....
Cites result from "Whole-genome typing and characteriz..."
...This was not surprising because as others and we have previously observed comparing other K. pneumoniae strains (Deleo et al. 2014; Ramirez et al. 2014b; Sabirova et al. 2016), the genomes of this bacterium seem to be quite homogeneous....
...We propose that this zone of the K. pneumoniae genome can be used as a signature for epidemiology analysis....
Related Papers (5)
Frequently Asked Questions (1)
Q1. What contributions have the authors mentioned in the paper "Whole-genome typing and characterization of -harbouring st383 klebsiella pneumoniae by pfge, whole-genome mapping and wgs reference:" ?
In this paper, a whole-genome typing and characterization of harboring ST383 Klebsiella pneumoniae by PFGE, wholegenome mapping and WGS is presented.