Whole-Genome Analysis of Antimicrobial-Resistant Salmonella enterica Isolated from Duck Carcasses in Hanoi, Vietnam

Round 1

Reviewer 1 Report (New Reviewer)

 

In this well written manuscript, the authors investigate Salmonella contamination in duck meat in Hanoi, Vietnam. The authors use antibiotic susceptibility testing and whole genome sequencing to reveal the presence of antibiotic resistance genes in Salmonella enterica isolates from duck meat. Overall, the study is well designed, the data is clearly presented and the conclusions are supported by the data.

 

I have one minor suggestion:

Line 59-60: “In recent years, Salmonella also has been significantly concerned about its antibiotic resistance characteristics”. Do the authors mean that there has been a concern about antibiotic resistance characteristics of Salmonella ? Please rephrase

 

Author Response

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Author Response File: Author Response.docx

Reviewer 2 Report (New Reviewer)

The study presented by Nguyen et al. on the analysis of antimicrobial resistance in Salmonella isolated from duck is of great interest given the importance of salmonellosis and the increase in duck consumption worldwide.

The study and the methodology used are well-planned, and the results are presented in a clear and simple way. However, the discussion is quite poor, since only 3 references have been included in the discussion. Specifically, between lines 344 and 390 no scientific reference is included. In this section, the authors discuss their own results suggesting some theories, but they do not contrast the results obtained with those of similar studies. If there are no similar studies in ducks, the results should be compared with those obtained in related species. Nor is there a clear conclusions section... On the other hand, the results of an MLST are presented, but the "Material and Methods" section does not describe the MLST in any subsection, nor the serotyping of the Salmonella strains. Neither is it indicated which antimicrobial susceptibility test is used to obtain the phenotypic profile of the strains (Kirby-Bauer or Disk Diffusion). It would be interesting to include a reference to it. When mentioning the criteria for the classification of MDR strains, a reference to this should be included.

Also, there are some details that should be changed:

- The format of the references is [6], [6, 9], or [6-12]. Please update it.

-L49. Please write Salmonella subspecies epitopes in italics accordingly.

- L59 and later: please use the acronym ARG for antimicrobial resistance genes.

- L78, 115. Please write Salmonella in italics.

-L83. What is the meaning of AR?

-L98. Please change 05 to "five". If the number is less than 10, it is recommended to write it down.

- L149, 155. Please include the details of the software (commercial company, version, city and country of origin).

-L156-157. Please indicate the meaning of CARD and ARG-ANNOT.

- L168. Include here the results of the isolated serotypes (L262-272).

-L222. Please change 3^rd to third.

- L274-276. This sentence corresponds to the material and methods section, please remove it from here.

-L335. Comment on the results obtained in 45/46 studies of Salmonella in poultry. I am very sure that there are many more studies... where does that figure come from?

-L346. Please change the semicolon between study and However to a period.

Finally, I strongly recommend the authors to check the grammar well to improve the comprehension and quality of the article.

Author Response

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Author Response File: Author Response.docx

Reviewer 3 Report (New Reviewer)

The manuscript entitled " Whole Genome Analysis of Antimicrobial Resistant Salmonella enterica Isolated from duck carcasses in Hanoi, Vietnam" represents a considerable amount of work. The following comments need to be addressed before the manuscript is suitable for publication in Curr. Issues Mol. Biol. Journal.

 

 

 

 

-          What is the state of Salmonella in Vietnam? please, add background about Salmonella in Vietnam.

-          Line 60: I propose adding this reference (Hagag, A., Naguib, D., Mohamed, A., & Elgohary, A. (2022). Prevalence, virulence genes, and antibiotic resistance of Escherichia coli and Salmonella spp. isolated from pigeons and humans. Mansoura Veterinary Medical Journal, 23(1), 24-30.

-          Line 78: Salmonella should be italic throughout the manuscript.

-          Line 88: Please, add the number of references to Pornsukarom et al.

-          In material and methods: I observed that authors did not statistically analyze their results. Please, statistically analyze your results.

-          Line 164: Please, add the total number of positive samples/47.

-          Line 302: 72 should be changed to 72.34%.

-          Line 303: The authors said that their results are significantly higher in comparison to previous studies. What are these studies? Please add a reference.

-          Lines 381-390: Please rewrite this paragraph.

 

-           

Author Response

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Author Response File: Author Response.docx

Reviewer 4 Report (New Reviewer)

Authors presented a study on antibiotic resistant Salmonella enterica from duck carcasses using whole genome analysis approach. They found predominance of serovar Muenster and a plethora of genetic determinants of antibiotic resistance in analyzed genomes, indicating high extent of multidrug resistant Salmonella contamination in duck meat. Methodology used in this study is state-of –the art and in compliance with modern trends for analysis of antibiotic resistance. However there are several issues that I will point out and that should be addressed that will improve overall presentation of the results and conclusions based on findings in this study.

1. First of all, when we are dealing with a burden of contamination of the food with antibiotic resistant bacteria and their epidemiology we tend to have as recent data as we can. Authors presented study based on the samples from 2019, which is almost four years ago. It is my opinion that this is major drawback of the manuscript since these results may not correspond to current epidemiology. Also, were the samples collected during different seasons (it is important because of temperature and humidity variations that are of importance for bacterial load variations), in order to answer the question is contamination present during entire year or just during one season. In abstract authors’ claim that they sampled during October 2019, but in materials and methods section they claim that sampling was done during the year 2019.

2. Authors should include resistance to colistin in antibiotic susceptibility testing.

3. Line 181. How authors determined the presence of ESBL strains since they did not describe it in materials and methods section (double disk diffusion test?).

4. Results section concerning in silico prediction of antibiotic resistance genes should be rewritten in order to make it comprehensive and to avoid redundancy. For example, please avoid referring to general knowledge (line 210. Gentamicin which is antibiotic belonging to the aminoglycoside… or discussion of the results (line 213. Enzyme frequently found..

Please be aware of the terminology

Line 214. Genes that have the ability of resistance… Genes do not have the ability of resistance they encode for the effectors that through their activity provide resistance to the cells

Line 219. Is resistant to all aminoglycoside drugs… this is something that you should address in the discussion section, since you can discuss on potential resistance that can arise from the presence of this gene, but you cannot state this in the results section since you did not analyze susceptibility to all aminoglycosides

Line 223. Delete antimicrobial

Line 224. Well-known gene… please avoid non-scientific terminology

Line 237. Please rephrase this sentence

Line 242. These mutations resulted… please delete this sentence this is not discussion section

Line 244. Please write IsKpn19 according to the nomenclature, delete mobile genetic factor and designate it as an insertion sequence

Line 248. The same as previous

Line 251. English usage.

5. Please match presented data for plasmids. Line 276. Three plasmids were detected. Line 294. You found four different plasmids (based on replication origin)

6. Line 284. English usage, non-comprehensible sentence

7. Line 288. The type 3 secretion systems include…. This is obsolete for the Results section. Lines 289-291. Please delete this

8. Lines 298-299. Where is the evidence, since this is a Results section you should present only results from your study

Discussion

Line 349. This sentence is obsolete

Line 365-367. Please rephrase this is non-comprehensible

Line 369. What is motor genetic factor? Please avoid non-scientific nomenclature

General comment

Authors should perform comparative genomics analysis of the sequenced genomes (it is not complicated) in order to reveal epidemiological connection between Muenster isolates.

 

 

Author Response

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Author Response File: Author Response.docx

Round 2

Reviewer 2 Report (New Reviewer)

After the changes introduced, the scientific quality of the article presented by Nguyen et al. has greatly improved. In my opinion it can already be published in present form. 

Reviewer 4 Report (New Reviewer)

Authors adequately addressed and resolved all relevant issues.

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.

Round 1

Reviewer 1 Report

I have reviewed the manuscript submitted by Trung Thanh Nguyen et al. titled “Whole Genome Analysis of Antimicrobial Resistant Salmonella enteritica Isolated from duck carcasses in Hanoi, Vietnam” (cimb-1966262). The authors analyzed the whole genome sequence of salmonella collected from ducks in Vietnam. They found that 72% of ducks they tested had Salmonella and the most of salmonella strains isolated had several antibiotic-resistant characteristics. They also identified several genes for antibiotic-resistant.

 

However, this manuscript was a list of results and lacks conclusions. It is necessary to note how this result is important for public health, and/or for improving antimicrobial usage. In addition, the manuscript was not well written. For example, Table S1 was not refereed in the text, and there is no explanation of how to draw the phylogenic tree. Abbreviations should be written in full at first appearance. This manuscript has to be brushed up more.

 

 

The points to be noted are as follows.

 

1.       Abstract

Please write a conclusion not a list of results.

 

2.       lines 20 and 25,

05 and 08 is not common, please change to 5 and 8.

 

3.       Line 36

What do authors mean? Potential for what?

 

4.       Lines 47-61

Please write the source. Which one was from meat and which one was from eggs?

 

5.       Lines135

Have the authors deposited sequences to the databank? Please write the accession numbers.

 

6.       Materials and methods

Please write how to draw the phylogenic tree presented in Figure 2.

 

7.       Lines 162-165

Where can we confirm these values? Table S1? Please refer the Table S1 in the text.

 

8.       Lines 163-189, Figure 1

Please revise values to take into account the significant digit.

 

9.       Figure 1

The maximum value of Y axis must be 1.

 

10.   Line224

 

In table 2, 51_S13 have 11 genes for beta-lactam. Please check.

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

The article entitled “Whole Genome Analysis of Antimicrobial Resistant Salmonella enteritica Isolated from duck carcasses in Hanoi, Vietnam” is of moderate interest and requires extensive English grammar editing. Some of the major issues are:

1-    Salmonella as a genus should always be italicized and capitalized the S. When discussing salmonella as generic, it should be without capitals and not italicized. 

2-    The correct name for the bacterium in this study is Salmonella enterica sbsps enterica. 

3-    CLSI methods for disk diffusion require MullerHinton media, no BHI.  The phenotypic study needs to be repeated with the appropriate media and controls. 

4-    No information is provided on the use of controls for the Disk Diffusion tests performed.

5-    It is not clear why the authors chose to select eight isolates randomly for whole genome sequencing and did not use epidemiological criteria to do so. 

6-    In general, the material and methods section lack specificity, and it is most obvious when describing the Illumina sequencing and bioinformatics section.

7-    The introduction is long and does not clearly articulate the need for the study. 

8-    The discussion is vague and lacks findings that are relevant to the audience of this journal. 

9-    The citations are minimal, and the literature on this topic is extensive. 

Author Response

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Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

I have reviewed the manuscript resubmitted by Trung Thanh Nguyen et al. titled “Whole Genome Analysis of Antimicrobial Resistant Salmonella enterica Isolated from duck carcasses in Hanoi, Vietnam” (cimb-1966262).

They have revised the manuscript enough. However, there are some minor points to be considered. After minor revisions, I think this manuscript will be accepted for publication.

 

 

The minor points were listed as follows.

 

1.       Lines 20 and 147

It would be better to use the same words, “whole duck samples” or “duck carcass samples”.

 

2.       Line 8

The “08” is written. Please change to 8 from 08.

 

3.       Lines 170-189

Is xx μg correct instead of concentration?

 

4.       Lines 220-224

 

I think 72 or 72.3% (69 or 69.2) are good significant figures.

Author Response

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Author Response File: Author Response.pdf

Reviewer 2 Report

This reviewer is still concerned in regards to the appropriate susceptibility testing. The quality of Mueller-Hinton agar used for the tests is critical as variations in the concentration of divalent cations in Mueller-Hinton agar can significantly influence the results. Quality should be tested not once with control but multiple times, and certainly every time a batch is prepared for in-house media. The results seemed to have been the same, but we do not know the diameter sizes considered as S or R. Table S1 should include the diameter sizes and the results of the E. coli control (diameter sizes) on every batch of media. 

Author Response

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Author Response File: Author Response.pdf