Fig 1.
Circular representation of plasmid pEC958.
The two outer rings show the coding sequences (CDSs) on the forward and reverse strand of the plasmid. Each CDS is colour-coded by its predicted function as shown in the figure. The grey ring depicts mobile elements identified on the plasmid. The two inner rings represent the GC plot and GC skew graph, respectively.
Fig 2.
Sequence comparison of pEC958 with other closely related plasmids.
Regions on plasmids are colour-coded as followed: red—replicon, blue—conjugation transfer, orange—mobile elements, dark pink—antimicrobial resistance genes.
Fig 3.
Organizational structure of mobile elements and antibiotic resistance genes on plasmid pEC958.
The colour coding is as followed: red—IS26 (8 copies IS26a to IS26h), yellow—ISEcp1, brown—Tn3 (partial), sky blue—Tn21 (partial), light green—IS1, dark pink—Tn5403, green—IS6100, dark blue—Tn1721, orange—ISEc23, dark pink—In54. Target duplication sites are indicated by triangle flags. The white blocks represent large regions not detailed in this figure.
Table 1.
Antibiotic resistance profiles (MIC, μg/mL) of EC958 and its pEC958 transformant in E. coli TOP10.
Fig 4.
Overview of the TraDIS screen for the identification of pEC958 essential genes.
(A) Graph showing the insertion index of each gene on pEC958 (top) in relation to the overall genetic organization of the plasmid (bottom). Nine essential genes (indicated in red) were identified that possessed an insertion index lower than 0.05. (B, C, D) Schematic showing the frequency of Tn insertions mapping to specific regions of pEC958. Essential genes required for the stable maintenance of pEC958 possessed a significantly reduced number of insertions.
Fig 5.
Maximum-likelihood phylogenetic tree showing the relationship of EC958_A0140 translated amino acid sequences.
EC958_A0140 sequenced are labeled by plasmid name. Also shown is the replicon nomenclature for each plasmid according to the FAB scheme and the parent organism.
Fig 6.
BRIG image depicting the presence of pEC958-like sequences in E. coli ST131 strains.
The strains are coloured according to their previously defined phylogenetic relationship: red (Clade A), orange (Clade B) and green (Clade C) [9]. The degree of coloured shading indicates the level of identity according to BLASTn between pEC958 (nucleotide position highlighted on the inner circle) and the draft Illumina assemblies of the E. coli ST131 strains [9]. BLASTn matches are coloured based on a nucleotide identity of between 70% and 90% (dark shading = high identity, light shading = low identity). Blank spaces in each ring represent BLASTn matches to pEC958 with less than 70% nucleotide identity, or pEC958 regions with no BLAST matches. Four strains originally characterised as ST131 but later shown to be ST95 are shown in black. Highlighted on the outer ring are the RepFII and RepFIA replicons, as well as antibiotic resistance genes, transposons and IS elements.
Table 2.
Prevalence of IncF plasmid types in E. coli ST131 strains.