Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2017 Aug 24;61(9):e00995-17.
doi: 10.1128/AAC.00995-17. Print 2017 Sep.

Characterization of a Large Antibiotic Resistance Plasmid Found in Enteropathogenic Escherichia Coli Strain B171 and Its Relatedness to Plasmids of Diverse E. Coli and Shigella Strains

Affiliations
Free PMC article

Characterization of a Large Antibiotic Resistance Plasmid Found in Enteropathogenic Escherichia Coli Strain B171 and Its Relatedness to Plasmids of Diverse E. Coli and Shigella Strains

Tracy H Hazen et al. Antimicrob Agents Chemother. .
Free PMC article

Abstract

Enteropathogenic Escherichia coli (EPEC) is a leading cause of severe infantile diarrhea in developing countries. Previous research has focused on the diversity of the EPEC virulence plasmid, whereas less is known regarding the genetic content and distribution of antibiotic resistance plasmids carried by EPEC. A previous study demonstrated that in addition to the virulence plasmid, reference EPEC strain B171 harbors a second, larger plasmid that confers antibiotic resistance. To further understand the genetic diversity and dissemination of antibiotic resistance plasmids among EPEC strains, we describe the complete sequence of an antibiotic resistance plasmid from EPEC strain B171. The resistance plasmid, pB171_90, has a completed sequence length of 90,229 bp, a GC content of 54.55%, and carries protein-encoding genes involved in conjugative transfer, resistance to tetracycline (tetA), sulfonamides (sulI), and mercury, as well as several virulence-associated genes, including the transcriptional regulator hha and the putative calcium sequestration inhibitor (csi). In silico detection of the pB171_90 genes among 4,798 publicly available E. coli genome assemblies indicates that the unique genes of pB171_90 (csi and traI) are primarily restricted to genomes identified as EPEC or enterotoxigenic E. coli However, conserved regions of the pB171_90 plasmid containing genes involved in replication, stability, and antibiotic resistance were identified among diverse E. coli pathotypes. Interestingly, pB171_90 also exhibited significant similarity with a sequenced plasmid from Shigella dysenteriae type I. Our findings demonstrate the mosaic nature of EPEC antibiotic resistance plasmids and highlight the need for additional sequence-based characterization of antibiotic resistance plasmids harbored by pathogenic E. coli.

Keywords: EPEC; Escherichia coli; mosaic; plasmid.

Figures

FIG 1
FIG 1
Circular plot of pB171_90. The outermost track of the plot contains the protein-coding genes on the positive strand, while the adjacent track contains the protein-coding genes on the negative strand. The predicted protein functions are indicated by colors as follows: replication (blue), virulence (purple), antibiotic resistance (red), conjugative transfer (orange), plasmid stability (green), transposition (yellow), and unknown (gray). The red line is the GC content along a 100-bp sliding window. The blue line is the GC skew along a 100-bp sliding window, with light blue indicating a positive skew and dark blue indicating a negative skew. The circular plot was generated using Circos 0.69-3 (80).
FIG 2
FIG 2
Phylogenomic analysis of E. coli genomes that carry genes with similarity to those of pB171_90. A SNP-based phylogeny was generated using ISG as previously described (35, 70). There were 149,150 conserved SNP sites identified among all of the E. coli genomes analyzed relative to the reference E. coli strain IAI39 (GenBank accession number NC_011750.1) that were used to construct a maximum-likelihood phylogeny with 100 bootstrap values using RAxML v.7.2.8 (71). The phylogeny was midpoint rooted and labeled using iTOL v.3 (72). Bootstrap values of ≥80 are designated by a gray circle. The scale bar indicates the approximate distance of 0.01 nucleotide substitutions per site. The E. coli phylogroups B1, A, B2, D, E, and F are denoted by bold letters. Symbols designate the presence of pB171_90 genes in each genome with tetA and tetR both represented by green stars (both genes, solid green star; one gene, open green star). Colors of the genome labels indicate the presumptive pathotype that was determined by in silico detection of canonical virulence genes.
FIG 3
FIG 3
In silico detection of pB171_90 genes in diverse E. coli genomes. The presence or absence of all predicted protein-coding genes of pB171_90 in the 134 E. coli genomes that contained the pB171_90 genes csi and/or traI is indicated by the BSR values in the heatmap (see Table S1 in the supplemental material). The clustered heatmap was generated using the heatmap2 function of gplots v.3.0.1 in R v.3.3.2. The genomes were clustered using the complete linkage method with Euclidean distance estimation. The pB171_90 genes are represented by the rows, while each column represents a different E. coli genome. The predicted pathotype of each E. coli genome is indicated by the color of the rectangle at the top of the heatmap (see legend for detail).
FIG 4
FIG 4
Phylogenetic analysis of csi. The csi nucleotide sequences of pB171_90 and other E. coli genomes were aligned using ClustalW. The alignment was used to construct a maximum-likelihood phylogeny with the Kimura 2-parameter model and 1,000 bootstraps using MEGA7 (79). The scale bar represents the approximate distance of 0.001 nucleotide substitutions per site. Bootstrap values of ≥50 are indicated by a gray circle. The predicted E. coli pathotypes are indicated by the color of their genome label (see legend), and the phylogroup and MLST of each genome are denoted in parentheses.
FIG 5
FIG 5
Phylogenetic analysis of traI. The traI nucleotide sequences of pB171_90 and other E. coli strains were aligned using ClustalW. The alignment was used to construct a maximum-likelihood phylogeny with the Kimura 2-parameter model and 1,000 bootstraps using MEGA7 (79). The scale bar represents the approximate distance of 0.02 nucleotide substitutions per site. Bootstrap values of ≥50 are indicated by a gray circle. The predicted E. coli pathotypes are indicated by the color of their genome label (see legend), and the phylogroup and MLST of each genome is denoted in parentheses. The blue vertical lines indicate the two clades identified in the csi phylogeny (Fig. 4).
FIG 6
FIG 6
In silico detection of pB171_90 genes in other human enteric pathogens. The predicted protein-coding genes of pB171_90 were identified in additional human enteric pathogens that contained the pB171_90 gene csi and are listed in Table S1 in the supplemental material. The clustered heatmap was generated using the heatmap2 function of gplots v.3.0.1 in R v.3.3.2. The genomes were clustered using the complete linkage method with Euclidean distance estimation. The pB171_90 genes are represented by the rows, while each column represents a different genome. The species are designated by the color of the rectangle at the top of the heatmap (see legend).

Similar articles

See all similar articles

Cited by 4 articles

Publication types

MeSH terms

Feedback