Next Article in Journal
Emergence of cfr-Mediated Linezolid Resistance in Staphylococcus aureus Isolated from Pig Carcasses
Next Article in Special Issue
Antimicrobial Resistance of Escherichia coli and Pseudomonas aeruginosa from Companion Birds
Previous Article in Journal
Role of Artificial Intelligence in Fighting Antimicrobial Resistance in Pediatrics
Previous Article in Special Issue
Molecular Characterization and Comparative Genomics of IncQ-3 Plasmids Conferring Resistance to Various Antibiotics Isolated from a Wastewater Treatment Plant in Warsaw (Poland)
Open AccessCommunication
Peer-Review Record

First Report of an Escherichia coli Strain Carrying the Colistin Resistance Determinant mcr-1 from a Dog in South Korea

Antibiotics 2020, 9(11), 768; https://doi.org/10.3390/antibiotics9110768
Reviewer 1: Anonymous
Reviewer 2: Rosario Gil
Antibiotics 2020, 9(11), 768; https://doi.org/10.3390/antibiotics9110768
Received: 29 September 2020 / Revised: 30 October 2020 / Accepted: 31 October 2020 / Published: 2 November 2020
(This article belongs to the Special Issue Antibiotic Resistance Genes: Spread and Evolution)

Round 1

Reviewer 1 Report

The manuscript by Moon and co-workers describes the colistin resistance determinant mcr-1 carrying by E. coli strain isolated from a dog. Colistin resistance determinants are getting attention due to the growing evidences that they spread in bacteria isolated from humans and animals. This short communication describes the identification of mcr-1 gene encoded on conjugative plasmid from IncI2 incompatibility group. On the basis of susceptibility testing, pulsed-field gel electrophoresis (PFGE) of mcr-1 positive isolates, conjugation assay, plasmid sequencing and comparative analysis the mcr-2 gene is characterised.

I have a couple of remarks which in my opinion should be addressed before publication:

The major comments :

  • What is the mechanism of colistin resistance encoded by mcr-1 gene? The information could be added in introduction section.
  • As I understand correctly from 1202 E. coli isolates two demonstrated colistin-resistance, what with the resistance to other antibiotics in this collection?
  • I could not find the Table S1 maybe the above information is included there?
  • Short characterisation of IncI2 group would be appreciated in the introduction or results and discussion section.
  • A multiplex PCR assay is mentioned in materials and methods but no results is presented in the manuscript?

The minor concerns:

  • The photo of PFGE in Figure 1 could be bigger.
  • The source of plasmid next to its name could be provided in Figure 2.
  • There is no description of CP050290 in Figure 2 and 3.
  • 3, line 108 – add KY to 471145 in second brackets; the same p. 4, line 121;
  • 3, line 111 – add 7 to KY65747 in first brackets; the same p.4, line 124;
  • The gene names on the top of the Figure 3 should be bigger and clearly visible.

Author Response

We deeply appreciate the positive feedback on our manuscript. Your comments and suggestions are important to improve the quality of our manuscript. Therefore, we have carefully revised the manuscript. The manuscript is double-checked for typographic and grammatical errors. The introduction and methods are revised, taking the type of article (short communication) into consideration. The figures are also replaced by those with better resolution. Additionally, the mistakes on the citation that occurred during copy-editing were corrected. We have highlighted the modified sections of the manuscript. The responses to your comments and suggestions are summarized as follows:

The major comments:

Point 1. What is the mechanism of colistin resistance encoded by mcr-1 gene? The information could be added to the introduction section.

Response 1: The mechanism of colistin resistance encoded by the mcr-1 gene is briefly described in lines 26-29.

Point 2. As I understand correctly from 1202 E. coli isolates two demonstrated colistin-resistance, what with the resistance to other antibiotics in this collection?

Response 2: We do not have the antimicrobial resistance profiles (other than colistin) of all the 1202 E. coli isolates. However, we determined the susceptibility profiles of the two colistin-resistant isolates to other antimicrobials. One of the colistin-resistant isolates (mcr-1-negative) was susceptible to all the tested antimicrobials (Line 101-105). Additionally, the susceptibility profiles of the colistin-resistant and mcr-1-positive isolate is presented in figure 1.

Point 3. I could not find Table S1 maybe the above information is included there?

Response 3. We apologize for the inconvenience and the table is included.

Point 4. Short characterization of IncI2 group would be appreciated in the introduction or results and discussion section.

Response 4. A brief description of IncI2 plasmids is included in lines 120-122.

Point 5. A multiplex PCR assay is mentioned in materials and methods but no results are presented in the manuscript?

Response 5. We apologize for the mistake and this section is deleted from the methods section.

The minor concerns:

Point 6. The photo of PFGE in Figure 1 could be bigger.

Response 6. Corrected as suggested.

Point 7. The source of plasmid next to its name could be provided in Figure 2.

Response 7. Based on your suggestion, general information about the plasmids is presented in Table 1.

Point 8. There is no description of CP050290 in Figures 2 and 3.

Response 8. The description is included in Table 1.

Point 9. 3, line 108? add KY to 471145 in second brackets; the same p. 4, line 121;

Response 9.  Correct accession numbers are displayed in Table 1 and Line 115-118.

Point 10. 3, line 111? add 7 to KY65747 in first brackets; the same p.4, line 124;

Response 10. Correct accession numbers are displayed in Table 1 and Line 115-118.

Point 11. The gene names on the top of Figure 3 should be bigger and clearly visible.

Response 11. Corrected as suggested.

Reviewer 2 Report

See attached file

Comments for author File: Comments.pdf

Author Response

We deeply appreciate the positive feedback on our manuscript. Your comments and suggestions are important to improve the quality of our manuscript. Therefore, we have carefully revised the manuscript. The manuscript is double-checked for typographic and grammatical errors. The introduction and methods are revised, taking the type of article (short communication) into consideration. The figures are also replaced by those with better resolution. Additionally, the mistakes on the citation that occurred during copy-editing were corrected. We have highlighted the modified sections of the manuscript. The responses to your comments and suggestions are summarized as follows:

Point 1. The introduction is too short to focus on the problem. It will be interesting to know if similar efforts have been made to detect E. coli (or other Enterobacteriaceae) strains carrying mcr-1 from companion animals in other countries, for comparison.

Response 1. Thank you for the constructive comments on our manuscript. The introduction is revised, and additional information on mechanisms of colistin resistance is elaborated. Additionally, previous studies on the detection of mcr-1 in E. coli isolated from humans, food animals, and, most importantly, companion animals were cited (Line 24-36). A comparison of the mcr-1 gene in E. coli from companion animals is included in results and discussion (Line 88-91).

Point 2. Line 26: The statement is incorrect. Even though the first reported cases of resistance to colistin were due to chromosomal mutations in otherwise susceptible bacteria (reviewed in 2015, reference [1]), soon after, plasmid-mediated mobile colistin resistance (mcr) was detected by many research groups working independently (several of which are cited in the manuscript), and examples accumulated rapidly (see Matamoros et al. 2017 as an example. https://www.nature.com/articles/s41598 017 15539 7).

Response 2. The chromosomal mutation in this section was referring to the first reported cases of colistin resistance. However, as you emphasized, plasmid-mediated mobile colistin resistance is reported by many researchers. To avoid confusion this section is rephrased (line 24-26).

Materials and Methods

Point 3. Line 39: I tried to check the procedure for E. coli isolation and identification in reference [11]. But that same expression (“…as described previously”) is cited in the Tamang et al. 2013 paper, referring to a previous paper by the same group which is only available to me as pay per view. Therefore, I think a short sentence to describe the procedure should be included here.

Response 3. Thank you for the comment and this section is revised as follows” Isolation and identification of E. coli isolates were performed using Eosin methylene blue agar (EMB, Becton Dickinson, Sparks, MD, USA) and MacConkey agar plates (MAC, BD, Spark, MD, USA).” (line 44-46)

Point 4. Line 46: [14] refers to the analysis of mcr-8 in Klebsiella pneumonie. Are the PCR primers common to these two mcrs?

Response 4. Thank you for the comment. We apologize for the mistakes and it is replaced with an appropriate reference (Line 166, Reference 1). Some of the mistakes in the citation occurred during copy-editing and changing the citation style. We have double-checked all the references in the text as well as in the lists of references.

Point 5. Line 49. [15] refers to the selection of transconjugants in the case of a plasmid encoding a beta-lactamase. Is the selection of transconjugants made the same way in this case?

Response 5. Thank you for the comment. We apologize for the mistakes and it is replaced with an appropriate reference (Line 212, Reference 18).

Point 6. Line 57: I could not accede to the protocols in the cited website.

Response 6. An updated website is provided (Line 216, Reference 20)

Point 7. Line 60: A plasmid is not a genome. Remove “The genome of” at the beginning of the sentence.

Response 7. Removed as suggested (Line 71).

Results and Discussion

Point 8. I could not accede to the Supplementary material.

Response 8. We apologize for the inconvenience and the table is included in this submission.

Point 9. Lines 70-75: Is one case out of 1,202 isolates from 1,815 samples analyzed enough to consider it a problem? To evaluate the potential risk, it would be also interesting to compare with the incidence found in other countries, in other domestic animals, in food…. Maybe this is just an isolated case, and the human owner or the dog food could be the original focus.

Response 9. The identification of one mcr-1 carrying isolates (two colistin-resistant) from the total of 1202 isolates seems very low. But we should be worried about the plasmid-mediated transfer of mcr genes and thus, the resistance gene could be transferred to any of the Enterobacteriaceae. Considering the close interaction between humans and dogs in the country there is a high probability for this isolate to disseminate into humans. As indicated in Line 88-91, the prevalence of the mcr-1 gene among E. coli isolates in this study (0.84%) is lower compared to the reports in isolates from companion animals from other countries but almost comparable to our previous report in isolates from food animals in Korea (0.1%). As you suggested out, we have also considered the possibility of transfer from food animals or humans or the environment. Thus, we selected specific isolates (from chickens) that demonstrated identical phenotypic and molecular characteristics (MLST) for detailed comparison. The dog isolate exhibited a different PFGE profile compared to chicken isolates. In addition, a comparison of mcr-1 carrying plasmids showed close similarity with those described in human isolates. Therefore, agreeing with your comment, we commented on the possibility of transmission from humans and/or environmental sources (as reported in previous reports) (Line 137-143). However, further studies might be needed to precisely determine the source of E. coli by focusing on the owner or the dog food.

Point 10. Line 100: The IncI2 plasmids, which have a broad host range, are----

Response 10. Corrected as suggested (Line 120)

Point 11. Figure legends 2 and 3: Instead of describing each sample in both figures, it would be clearer to present a table with the relevant data: name of the plasmid, GenBank Acc. No., bacterial host, animal host, country, work of reference. Please, be careful with the GenBank Acc. No.

Response 11. Corrected as suggested (Table 1, Page 3).

Point 12. Line 117 (Figure 3 legend): “isolated” better than “originated”.

Response 12. Corrected as suggested (Line 130).

 

Round 2

Reviewer 1 Report

p.4, line 126 – tree not three

Author Response

Dear Reviewer,

We are grateful for the meticulous revision and constructive comments on our manuscript. We have revised the manuscript according to your comments.  We have highlighted the revised section (2nd revision) of the manuscript in blue.

Point. p.4, line 126? tree not three

Response. Thank you for the correction and we have corrected it accordingly (line 126).

Reviewer 2 Report

The database in reference 20 is not spelled correctly. It should be "http://enterobase.warwick.ac.uk/species/ecoli/allele_st_search"

(allele instead of a1lele)

Author Response

Dear Reviewer,

We are grateful for the meticulous revision and constructive comments on our manuscript. We have revised the manuscript according to your comments.  We have highlighted the revised section (2nd revision) of the manuscript in blue.

Point. The database in reference 20 is not spelled correctly. It should be http://enterobase.warwick.ac.uk/species/ecoli/allele_st_search.

Response: Thank you for the correction and we apologize for the mistake and we corrected it, as suggested. (line 217)

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.


Back to TopTop