Distinguishing bla _KPC Gene-Containing IncF Plasmids from Epidemiologically Related and Unrelated Enterobacteriaceae Based on Short- and Long-Read Sequence Data

Abstract

Background: Limited information is available on whether blaKPC -containing plasmids from isolates in a hospital outbreak can be differentiated from epidemiologically unrelated blaKPC-containing plasmids based on sequence data. This study aimed to evaluate the performance of three approaches to distinguish epidemiologically related from unrelated blaKPC-containing pKpQiL-like IncFII(k2)-IncFIB(pQiL) plasmids. Method: Epidemiologically related isolates, were short- and long-read whole genome sequenced. A hybrid assembly was performed and plasmid sequences were extracted from the assembly graph. Epidemiologically unrelated plasmid sequences were extracted from the GenBank. Pairwise comparisons were performed of epidemiologically related and unrelated plasmids based on SNP differences using snippy, phylogenetic distance using Roary and using a similarity index that penalizes size differences between plasmids (Stoesser-index). The percentage of pairwise comparisons misclassified as genetically related or as clonally unrelated was determined using different genetic thresholds for genetic relatedness. Results: The ranges in number of SNP differences, Roary phylogenetic distance, and Stoesser-index overlapped between the epidemiologically related and unrelated plasmids. When using a genetic similarity threshold that classified 100% of epidemiologically related plasmid pairs as genetically related, the percentages of plasmids misclassified as epidemiologically related ranged from 6.7% (Roary) to 20.8% (Stoesser-index). Discussion: Although epidemiologically related plasmids can be distinguished from unrelated plasmids based on genetic differences, blaKPC-containing pKpQiL-like IncFII(k2)-IncFIB(pQiL) plasmids show a high degree of sequence similarity. The phylogenetic distance as determined using Roary showed the highest degree of discriminatory power between the epidemiologically related and unrelated plasmids.

Small wards in the ICU: a favorable measure for controlling the transmission of carbapenem-resistant Klebsiella pneumoniae

“…Roary v3.13.0 [ 22 ] was used to perform a pan genome analysis and create core gene alignments on 95 isolates, and SNP alignments were extracted from the core gene alignments. Using K. pneumoniae (GCF_000240185) as a reference sequence [ 23 ], Snippy v4.6.0 ( https://github.com/tseemann/snippy ) was used to perform reference-based mapping and identify SNPs [ 24 ]. The alignment file was filtered from variants with elevated densities of base substitutions as putative recombination events by Gubbins version 2.4.1 [ 25 ].…”

“…Using K. pneumoniae (GCF_000240185) as a reference sequence [23], Snippy v4.6.0 (https:// github. com/ tseem ann/ snippy) was used to perform referencebased mapping and identify SNPs [24]. The alignment file was filtered from variants with elevated densities of base substitutions as putative recombination events by Gubbins version 2.4.1 [25].…”

Isolation and Characterisation of Human-Derived blaKPC-3-Producing Salmonella enterica Serovar Rissen in 2018

“…The description of the plasmid provides interesting confirmatory insights into the plasticity of pKpQIL-like plasmids, and their role in the ability to adapt to different bacterial species and genera by facilitating the spread of bla KPC-3 . These plasmids were isolated in clinically relevant Enterobacterales across the globe [ 11 , 40 ], but pKpQIL complete sequences are present in the NCBI database only in isolates of Klebsiella spp., E. coli , Enterobacter cloacae , and Raoultella ornithinolytica . The presence of bla KPC-3 in a pKpQIL-like plasmid is described in Salmonella spp.…”

Detection of NDM-1-Positive Aeromonas caviae from Bacteremia by Using Whole-Genome Sequencing

“…Snippy (rapid haploid variant calling and core genome alignment) was used to compare genomic differences between strains. 16 The alignment file was filtered from variants with elevated densities of base substitutions as putative recombination events by Gubbins version 2.4.1. 17 The filtered core-genome alignment file was used to construct a maximum likelihood tree using FastTree with the GTR+CAT model.…”