The complete genome sequence of five pre-2013 Escherichia coli sequence type (ST)1193 strains reveals insights into an emerging pathogen
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Fluoroquinolone-resistant Escherichia coli sequence type (ST)1193 is a profound, emerging lineage associated with systemic, urinary tract and neonatal infections. Humans, companion animals and the environment are reservoirs for ST1193, which has been disseminated globally. Following its detection in 2007, ST1193 has been identified repeatedly amongst fluoroquinolone-resistant clones in Australia. However, despite the growing importance of ST1193, only three complete genomes are published in the literature, none of which are from Australia. Here we expand on the available ST1193 resources with the complete genomes of five ST1193 strains sequenced using Oxford Nanopore Technologies and Illumina. Using in silico genotyping, we found that all strains were multi-drug resistant, including resistances to fluoroquinolones and cephalosporins. In vitro antibiotic susceptibility testing mostly correlated with individual genotypes. The exception was MS8320, which had additional in vitro resistance to piperacillin/tazobactam, ampicillin/sulbactam, cefazolin and doripenem (carbapenem). Further investigation identified seven additional copies of an IS26 transposable unit carrying a bla TEM-1B beta-lactamase gene, suggesting this tandem amplification is associated with extended resistance phenotypes. Uropathogenicity factors, including three separate siderophore-encoding loci, were conserved in chromosomal and plasmid regions. Using all complete genomes, we further elucidated the recombination events surrounding the previously described K5/K1 capsular locus switch. Phenotypic confirmation of differing capsules in Australian ST1193 strains, coupled with genetic analysis revealing insertions downstream of the capsular locus, underscored the genetic distinctions between K5 and K1 capsule encoding strains. This study provides five new reference ST1193 genomes from Australia. These include the earliest complete K5-capsule ST1193 genomes on record (collected 2007), alongside our reference genome (MS10858), a clinical isolate obtained early during the ST1193 expansion and representative of the predominant K1-associated clade. These findings lay the foundations for further genomic and molecular analyses that may help understand the underlying reasons for the rapid global expansion of ST1193.
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Thank you for the careful revision of the manuscript in response to the reviewers.
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Please check the reviewers comments carefully and address them, particularly those from Reviewer 1, who is asking for some clarifications and further details. The questions towards additional analysis (points 7-9 from Reviewer 1), I would leave up to the authors whether or not they want to expand the paper with further analyses or not. I can see a benefit for these, but consider the current state of the paper suitable already for the scope of the Access Microbiology Platform.
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Comments to Author
Here, the authors describe the hybrid WGS of five strains from the ST1193 group of E. coli from both the K1 and K5 phylogroups. The authors have deposited high-quality assemblies in to the relevant databases, and have carried out in-depth genotypic and phenotypic analysis including virulece/amr gene annotation and MIC assays. This work will provide an excellent resource as previously, reference genomes for ST1193 were limited. The paper is well written, with highly detailed methodology and annotation. The images represent the comparative genomics elements of the manuscript, and mutations and plasmid variation are depicted well in supplementary figure. I believe the manuscript is ready for publication in it's current state.
Please rate the manuscript for methodological rigour
Very good
Comments to Author
Here, the authors describe the hybrid WGS of five strains from the ST1193 group of E. coli from both the K1 and K5 phylogroups. The authors have deposited high-quality assemblies in to the relevant databases, and have carried out in-depth genotypic and phenotypic analysis including virulece/amr gene annotation and MIC assays. This work will provide an excellent resource as previously, reference genomes for ST1193 were limited. The paper is well written, with highly detailed methodology and annotation. The images represent the comparative genomics elements of the manuscript, and mutations and plasmid variation are depicted well in supplementary figure. I believe the manuscript is ready for publication in it's current state.
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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Comments to Author
The manuscript by White et al. proposes to provide insight into the genomics of ST1193 E. coli strains by the obtention of five complete genomes using hybrid assemblies. Authors also claim that they have sequenced one of the oldest ST1193 strains causing urinary tract infection. Even though I believe that those complete genomes are an important resource that should be made available for future reference, the text that accompanies them could be extensively summarized without loss of content. I also suggest additional analysis (points 7-10) that would add value to the present manuscript, if no follow-up project is planned at this stage. 1. Title: the use of the word "historical" can be misleading, considering that the sequenced strains were collected between 2007 and 2012. Consider revising. 2. Item 7.6 …
Comments to Author
The manuscript by White et al. proposes to provide insight into the genomics of ST1193 E. coli strains by the obtention of five complete genomes using hybrid assemblies. Authors also claim that they have sequenced one of the oldest ST1193 strains causing urinary tract infection. Even though I believe that those complete genomes are an important resource that should be made available for future reference, the text that accompanies them could be extensively summarized without loss of content. I also suggest additional analysis (points 7-10) that would add value to the present manuscript, if no follow-up project is planned at this stage. 1. Title: the use of the word "historical" can be misleading, considering that the sequenced strains were collected between 2007 and 2012. Consider revising. 2. Item 7.6 - "Compiling a high-quality…": How were the 616 genomes identified as belonging to ST1193 prior to downloading from SRA? Was a screening performed on EnteroBase before recovering the reads? If that was the case, consider reorganizing the paragraph for clarification. In lines 233-235, authors mention that the genomes belonged to previously published datasets, yet no references are provided regarding their origin. Please consider adding this information, together with their metadata and accession numbers on a Supplementary Table. 3. Lines 260-263: The additional nine E. coli genomes mentioned in the text are not present in Figure 1, but in Supplementary Figure S1. Please rephrase to avoid confusion. 4. Consider removing the genomic descriptions and AST data of the three isolates not sequenced in this study (i.e., MCJCHV-1, 09-02E, and AVS0096) from Tables 1 and 2, as these results were not produced with the same methodologies described in the Methods section and are not the main focus on this manuscript. 5. There is a discordance between the number of SNPs used to build the phylogeny of ST1193 genomes depicted in Figure 3. In the Figure legend, authors mention that it was based on a 4,640 bp alignment, while on the main text this number corresponds to the SNPs that were removed from the final alignment because they were found in recombinant regions, and the phylogenetic tree was built based on a 9,687 non-recombinant SNP alignment. Please revise and adapt the text accordingly. Also notice that bacterial genomes are not biallelic and, therefore, the SNPs cannot be considered biallelic as mentioned in the text. 6. In the main text (Lines 363-364), authors only describe the origin of 30/42 strains of human origin belonging to Clade 2. Was there no metadata available for the remaining 12? Please clarify. 7. Did authors consider performing additional experimental work to confirm that the phenotypic resistance to piperacillin/tazobactam, ampicillin/sulbactam, cefazolin, and doripenem observed in isolate MS8320 was indeed due to the presence of multiple copies of blaTEM-1B? Please clarify and/or consider adding this analysis on a revised version of the manuscript. 8. It would be interesting to use the circularized F-:A1:B10 and F-:A1:B20 plasmids obtained in this study as references to identify their presence in the >600 ST1193 genomes included in this study. Is each type of plasmid exclusive of a given clade, i.e., plasmids F-:A1:B10 and F-:A1:B20 can only be found in Clade 1 and 2, respectively? Please consider adding this analysis to a revised version of the manuscript. 9. Building a dated phylogeny to confirm author's claim that K5/Clade 2 strains evolved into K1/Clade 1 strains would be an important addition to a revised version of the manuscript. 10. I disagree with the final claim from the authors, presented in the last paragraph of the Discussion. Even if they only sequenced five new genomes belonging to ST1193, they could have performed the so called "further large-scale phylogenetic analyses using more appropriate, closely related genomes" (Lines 487-488) with the additional 616 curated genomes used to build the phylogeny presented in Figure 3. Will that be object of a follow-up (maybe current) project? Please clarify.
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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