Antimicrobial and Metal Resistance Genes in Bacteria Isolated from Mine Water in Austria

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

1. Introduction part is short, I suggest  to improve it and please describe clearly main idea of the work.
2.  I suggest to remove table 2 and submit it as supplementary file.
3. What do you mean while you titled method  ''In silico identification of antimicrobial resistance genes" . Please clarify it and describe it clearly.
4. Regarding to the sequences, I suggest to add them to the supplementary file.
5. Please describe primer designing also at the material methods.

Author Response

Reply to reviewer one:

Many thanks for your valuable comments. We changed the manuscript according to your suggestions as follows:

1. Introduction part is short, I suggest  to improve it and please describe clearly main idea of the work.

Many thanks for your suggestion. We rewrote the introduction part. Please find it line 40 to line 89 of the revised manuscript.

2. I suggest to remove table 2 and submit it as supplementary file.

Indeed both tables hold a lot of information that are difficult to describe in detail. Therefore we added the sequence homology in table 1 and submitted it as supplementary data.

3. What do you mean while you titled method  ''In silico identification of antimicrobial resistance genes" . Please clarify it and describe it clearly.

Based on whole genome sequencing, in silico identification of antimicrobial resistance genes is the use of computational tools (in this case ABRicate, coupled with some AMR databases) to detect genes associated with resistance to antibiotics and other antimicrobial agents, instead of conducting laboratory experiments including specific PCRs.

4. Regarding to the sequences, I suggest to add them to the supplementary file.

All sequences are now publicly available under PRJNA1224714

5. Please describe primer designing also at the material methods.

No primer design was done, data were extracted from whole genome sequencing, we outlined this more prominent in the M&M section.

 

 

Reviewer 2 Report

Comments and Suggestions for Authors

Comments on the manuscript entitled “Antimicrobial and metal resistance genes in bacteria isolated from mine water in Austria” authored by Jakob Prochask et al

The AMR topic is of great interests and data on its molecular characteristics are needed not only for medically important bacteria, but also for the environmental ones. The protocol includes advanced methods (the authors could name them in the abstract), the results are clearly presented (table, figure and text) and data are statistically analysed

Overall, the manuscript is well written, no major faults in my opinion, still certain aspects should be revised to underline and enrich the value of the results obtained

Specifically, 

1)    the novelty of the study

2)    integrate the results – One health is presented only in the introduction, while the authors obtained many and valuable results that deserve to be pointed out under One health approach as part of discussion section

It is the authors choice, but I strongly recommend adding One health in the abstract and conclusions 

3)    phrasing style for certain paragraphs of the manuscript and in particular of the abstract; the content is overshaded by the writing style, this is in fact a minor aspect, still consider revisions of the text: please avoid words repetition, correct English syntax and 

Abstract – the current form does not benefit from a line presenting the novelty and conclusion, please add accordingly. I strongly recommend the authors to use the lines 58-63 to produce a text for the abstract aim and conclusion; also the methods used to detect the bacteria could be added in the abstract (eg Mine microbiota was identified by MALDI-TOF and 16S rRNA gene sequencing with a further in silico identification of antibiotic resistance and virulence gene)

Introduction – well written, provides information regarding the mine waters heavy metal contamination, AMR and heavy metal resistance encoding genes connections and consequences.  By adding the One health concept paragraph, the study justification is very well presented. The aim of the study is also clearly and accurately presented. Please, add the study novelty 

Discussion – please integrate the results obtained using the One health, also add your study limitations

Conclusion – as mentioned above, the results should be highlighted using the One health

Author Response

Reply to reviewer two

Comments on the manuscript entitled “Antimicrobial and metal resistance genes in bacteria isolated from mine water in Austria” authored by Jakob Prochask et al

The AMR topic is of great interests and data on its molecular characteristics are needed not only for medically important bacteria, but also for the environmental ones. The protocol includes advanced methods (the authors could name them in the abstract), the results are clearly presented (table, figure and text) and data are statistically analysed

Overall, the manuscript is well written, no major faults in my opinion, still certain aspects should be revised to underline and enrich the value of the results obtained

Many thanks for your valuable respectful comments we changed the manuscript accordingly as follows:

Specifically, 

• the novelty of the study

We changed the conclusion and introduction accordingly. 

• integrate the results – One health is presented only in the introduction, while the authors obtained many and valuable results that deserve to be pointed out under One health approach as part of discussion section. It is the authors choice, but I strongly recommend adding One healthin the abstract and conclusions 

Thank you very much for this valuable suggestion. We added the "One Health" aspect at various parts of the manuscript including the abstract, introduction, discussion and conclusion.

• phrasing style for certain paragraphs of the manuscript and in particular of the abstract; the content is overshaded by the writing style, this is in fact a minor aspect, still consider revisions of the text: please avoid words repetition, correct English syntax and 

We tried to change the style of the abstract.

Abstract – the current form does not benefit from a line presenting the novelty and conclusion, please add accordingly. I strongly recommend the authors to use the lines 58-63 to produce a text for the abstract aim and conclusion; also the methods used to detect the bacteria could be added in the abstract (eg Mine microbiota was identified by MALDI-TOF and 16S rRNA gene sequencing with a further in silico identification of antibiotic resistance and virulence gene)

We added the conclusion and novelty in the abstract and also the methods used.

Introduction – well written, provides information regarding the mine waters heavy metal contamination, AMR and heavy metal resistance encoding genes connections and consequences.  By adding the One health concept paragraph, the study justification is very well presented. The aim of the study is also clearly and accurately presented. Please, add the study novelty 

Many thanks. We extended the "One Health" concept part and changed the studies aim including a conclusion of expected results.

Discussion – please integrate the results obtained using the One health, also add your study limitations

Please find the extended "One Health" part (which really improved the manuscript) at several parts of the manuscript including the discussion.

Conclusion – as mentioned above, the results should be highlighted using the One health

Changed accordingly.

Reviewer 3 Report

Comments and Suggestions for Authors

Title: Antimicrobial and metal resistance genes in bacteria isolated from mine water in Austria

This manuscript is well-written by the authors. I do believe that if they can improve the manuscripts following all comments. It might have a chance to publish in the journal.

Comments

1. Abstract: Please add the information of antimicrobial and metal resistance genes in bacteria.
2. It would be better if the authors add quantitative data in the abstract.
3. Please add a conclusion in the abstract.
4. The authors should explain or mention the relationship between antimicrobial and metal resistance genes. Why did the authors focus on these genes?
5. I suggest the authors to modify or re-write the introduction. What distinguishes your research between other? please give state of the art of your research

            -The first paragraph: describe the importance of antimicrobial and metal resistance genes in bacteria as well as the relationship between antimicrobial and metal resistance genes.

            -The second paragraph: describe the information of the area in Austria. What is the importance of the city. Why did the authors focus on this area?

-The third paragraph: describe One Health concept based on horizontal transferring of resistance genes?  

            -The fourth paragraph: describe the objective of this study. Why the authors are interested in this study?

6. Table 1 and Table 2 showed a different style. They should be design as the same pattern.
7. Table 1 is raw data. It should be a supplementary data. The authors should summarize the raw data. For example, numbers of isolates of Pseudomonas spp. How many percent?
8. Table 2: Why did the authors write metal resistance genes using capital alphabet?
9. Please modify Figure 1. It should be presented as an international pattern. All genes should be written in italic.
10. This manuscript presented the results only gene detection. It is not enough for publishing in Antibiotics. I suggest the authors determine the correlation between antimicrobial and metal resistance genes in bacteria.
11. Please add the information of the discussion. Try to compare the results (the author’s hypothesis) with other finding by other researchers.
12. Please delete some introduction and result sentences in the discussion part.
13. Line 204, 238, 269: There are 2 full stops.
14. Method- Isolation of bacteria: Please add more information on bacterial culture. Please describe types of media, and selected colonies.
15. Please write all scientific name, genes, in vitro, in vivo in italic.
16. Table 1 and Table 2 showed a different style. They should be design as the same pattern.
17. The references of 2020-2025 are suggested to be cited.
18. Please remove some old references.

 

 

 

 

 

 

 

Author Response

Reply to reviewer three

This manuscript is well-written by the authors. I do believe that if they can improve the manuscripts following all comments. It might have a chance to publish in the journal.

Many thanks for your valuable and extensive comments. We tried our best to advance the manuscript based on your comments.

Comments

1. Abstract: Please add the information of antimicrobial and metal resistance genes in bacteria.
2. It would be better if the authors add quantitative data in the abstract.
3. Please add a conclusion in the abstract.

We changed the abstract accordingly, added quantitative data and a conclusion.

1. The authors should explain or mention the relationship between antimicrobial and metal resistance genes. Why did the authors focus on these genes?

All genes identified by in silico analysis were described in the results section, and we focused on the most frequently identified genes.

1. I suggest the authors to modify or re-write the introduction. What distinguishes your research between other? please give state of the art of your research

            -The first paragraph: describe the importance of antimicrobial and metal resistance genes in bacteria as well as the relationship between antimicrobial and metal resistance genes.

            -The second paragraph: describe the information of the area in Austria. What is the importance of the city. Why did the authors focus on this area?

-The third paragraph: describe One Health concept based on horizontal transferring of resistance genes?  

            -The fourth paragraph: describe the objective of this study. Why the authors are interested in this study?

We modified the introduction according to your suggestions.

1. Table 1 and Table 2 showed a different style. They should be design as the same pattern.

We adopted the style

1. Table 1 is raw data. It should be a supplementary data. The authors should summarize the raw data. For example, numbers of isolates of Pseudomonas spp. How many percent?

Table 1 was extended with % in sequence homology using 16S rRNA gene sequence analysis and it will go into suppl. data.

1. Table 2: Why did the authors write metal resistance genes using capital alphabet?

We changed the style accordingly.

1. Please modify Figure 1. It should be presented as an international pattern. All genes should be written in italic.

Done

1. This manuscript presented the results only gene detection. It is not enough for publishing in Antibiotics. I suggest the authors determine the correlation between antimicrobial and metal resistance genes in bacteria.

We added several paragraphs in the results and discussion section about the species specific occurrence of genes and thus statistical calculations are not licit. Nevertheless we discussed the results and provided statistics where proper.

1. Please add the information of the discussion. Try to compare the results (the author’s hypothesis) with other finding by other researchers.

We added some paragraphs in the discussion.

1. Please delete some introduction and result sentences in the discussion part.

Done as suggested. Thank you.

1. Line 204, 238, 269: There are 2 full stops.

Thanks for spotting. Changed.

1. Method- Isolation of bacteria: Please add more information on bacterial culture. Please describe types of media, and selected colonies.

We added a sentence on the selection of colonies.

1. Please write all scientific name, genes, in vitro, in vivo in italic.

Done.

1. Table 1 and Table 2 showed a different style. They should be design as the same pattern.

Done and Table 1 goes to suppl. Data.

1. The references of 2020-2025 are suggested to be cited.

We tried to delete old references but despite one it was not possible to find equivalent literature from 2020-2025.

1. Please remove some old references.

Done.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

I would like to thank authors for reply and making revisions in the manuscrip

Reviewer 3 Report

Comments and Suggestions for Authors

The authors have edited and modified the manuscript according to the comments.