Dear Mr. Durán,

Thank you very much for submitting your manuscript "GenoVi, an open-source automated circular genome visualizer for bacteria and archaea uncovers genomic patterns for replicon identification in multipartite genomes" for consideration at PLOS Computational Biology.

As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. In light of the reviews (below this email), we would like to invite the resubmission of a significantly-revised version that takes into account the reviewers' comments.

The reviewers generally agree that the tool is useful and the manuscript is well written, but concerns remain. One important question (raised explicitly by reviewer #1, but also perhaps underlying some of the concerns of reviewer #3) is whether the example of Paraburkholderia (which covers most of the figures as well as the results section) is a good choice to demonstrate the value of GenoVi. For example, Fig. 3 does not seem particularly informative. The subsequent figures seem much clearer, but they also seem (as per the comment of reviewer #3) to have been generated by a different programme. While we don't find the use of companion tools to be objectionable per se (in fact, it can be salutary to not reinvent the wheel), in this case, the added value of GenoVi is unclear.

We cannot make any decision about publication until we have seen the revised manuscript and your response to the reviewers' comments. Your revised manuscript is also likely to be sent to reviewers for further evaluation.

When you are ready to resubmit, please upload the following:

[1] A letter containing a detailed list of your responses to the review comments and a description of the changes you have made in the manuscript. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out.

[2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file).

Important additional instructions are given below your reviewer comments.

Please prepare and submit your revised manuscript within 60 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email. Please note that revised manuscripts received after the 60-day due date may require evaluation and peer review similar to newly submitted manuscripts.

Thank you again for your submission. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments.

Sincerely,

Luis Pedro Coelho

Academic Editor

PLOS Computational Biology

Lucy Houghton

Staff

PLOS Computational Biology

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The reviewers generally agree that the tool is useful and the manuscript is well written, but concerns remain. One important question (raised explicitly by reviewer #1, but also perhaps underlying some of the concerns of reviewer #3) is whether the example of Paraburkholderia (which covers most of the figures as well as the results section) is a good choice to demonstrate the value of GenoVi. For example, Fig. 3 does not seem particularly informative. The subsequent figures seem much clearer, but they also seem (as per the comment of reviewer #3) to have been generated by a different programme. While we don't find the use of companion tools to be objectionable per se (in fact, it can be salutary to not reinvent the wheel), in this case, the added value of GenoVi is unclear.

Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #1: The manuscript describes a command line bioinformatics tool that is designed to take a genbank file and generate a circos plot to display the GC content and skew, as well as ortholog positions using the COGs database. Creating circos plots with the circos program is a non-trivial task with a steep learning curve. The advantage of this tool is in the automated processing and rendering of circos plots to display information that the authors believe would be most relevant to users when visualising bacterial genomes. There are other platforms and pipelines designed to help users create circos plots, but each have their own strengths and weaknesses and having more options available to end users is always a good thing. The manuscript is well written, and the tool performs well.

Main concerns:

GenoVi is sold as a visualisation tool, and an example is given for how GenoVi can help distinguish replicons from plasmids in a bacterial genus. The example uses the COG annotations generated by GenoVi as input for a gene composition-based analysis, but it does not indicate whether or not the GenoVi visualisations themselves were at all helpful in this endeavour. My impression is that the findings of the example could be achieved by simply annotating the genome with DeepNOG and then preforming the subsequent analyses presented in the results. It would be beneficial to better highlight the usefulness of the visualisations that GenoVi creates.

Figure 2

It’s impossible to tell what each of the tracks are. There are too many colours to simply rely on the colours alone to know what is what. I suggest using ideogram spacing and adding in labels for each track. http://www.circos.ca/documentation/tutorials/ideograms/spacing_breaks/lesson

Minor comments and suggestions:

For draft genomes, how does GenoVi lay out the contigs (order and orientation) and can this interfere with GC skew calculations at the boundaries?

For draft genomes, my understanding is that plasmid contigs and genome contigs are combined in the same circos plot. Is this correct? While this might not be ideal, there probably isn’t a neat or easy solution for this. Maybe it would be best to just make sure this is clear in the documentation.

“””Interactive web platform””” – this would be a great future addition.

“””GenoVi is the first open-source tool that uses a unique input file …”””

Genbank files are not unique, which is good because you don’t want unique (non-standard) file formats for your inputs.

Running the Program:

Do you need bioconda and conda-forge channels when installing circos?

Are there plans to add this to bioconda? This would make the installation of GenoVI and Circos a one-step process.

As the input file is always required, it could just be a bareword argument, e.g.

genovi input.gb …

Instead of specifying -status every time, consider having a -draft flag (and default to complete).

e.g. for complete genome:

genovi input.gb

e.g. for draft genome:

genovi input.gb -draft

I would call the output folder name “genovi”, rather than the name of someone else’s program.

consider support for gzipped genbank files

Reviewer #2: The authors have developed Python command-line application for the visualization circular genome representations of complete and draft genomes of bacteria and archaea. However, there are scope to impoverish the manuscript before acceptance.

Therefore, I recommend major revision as per the following suggestions.

1. Authors should include a comparison table highlighting advantages of their new tool over already exiting tools.

2. I suggest to containerize their software using docker or singularity for portability across machine.

Reviewer #3: It looks like the tool is useful for automatically creating circular genomics visualization. However, biggest concern I have is that the resulting circular visualizations themselves do not seem to enable complete analysis, and many parts of the analysis in the Results and discussion section actually seem to rely on externally-generated visualization, such as heatmaps and box plots.

- I wonder if the heatmaps and box plots illustrated in the manuscript can be generated automatically along with the circos plots. I think this will make the analysis process more seamless to users.

- One of the main limitation of the circular visualizations in term of their interpretation is that it is difficult to see the COG distribution patterns accurately due to the use of large number of similar colors. For example, in Fig. 2A, it is really hard to find what are the most representative COG categories. I wonder if it makes sense to enable users to filter COG categories, show only top-N categories, or etc.

- Many color palettes seem to be not color-blindness friendly. I suggest using color-blindness friendly palette as a default one (e.g., https://colorbrewer2.org/ or https://mikemol.github.io/technique/colorblind/2018/02/11/color-safe-palette.html).

- Sufficiently distinct colors for individual categories should be used whenever possible (e.g., currently, positive CDS, negative CDS, and rRNA are too similar which is difficult to differentiate in the visualization).

- Using the "complete" sequence option for the first tutorial example gives an error on my machine (i.e. "FileNotFoundError: [Errno 2] No such file or directory: 'circos-contig_1.png' -> 'circos/circos-contig_1.png'").

- Having more test datasets in the repository would be valuable.

Minor

- Color legends generated are sometimes too small to read.

- In general, I think interactive visualizations can make the analysis more effective (e.g., using Gos Python package, https://osf.io/yn3ce/), such as using mouse hover to see exact COG categories in tooltips.

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Have the authors made all data and (if applicable) computational code underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data and code underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data and code should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data or code —e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Michael J. Roach

Reviewer #2: Yes: Vikash K Singh

Reviewer #3: No

Figure Files:

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org.

Data Requirements:

Please note that, as a condition of publication, PLOS' data policy requires that you make available all data used to draw the conclusions outlined in your manuscript. Data must be deposited in an appropriate repository, included within the body of the manuscript, or uploaded as supporting information. This includes all numerical values that were used to generate graphs, histograms etc.. For an example in PLOS Biology see here: http://www.plosbiology.org/article/info%3Adoi%2F10.1371%2Fjournal.pbio.1001908#s5.

Reproducibility:

To enhance the reproducibility of your results, we recommend that you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

Attachments

Attachment

Submitted filename: Vikash_review_PCOMPBIOL-D-22-01291.docx

Dear Mr. Durán,

Thank you very much for submitting your manuscript "GenoVi, an open-source automated circular genome visualizer for bacteria and archaea" for consideration at PLOS Computational Biology. As with all papers reviewed by the journal, your manuscript was reviewed by members of the editorial board and by several independent reviewers. The reviewers appreciated the attention to an important topic. Based on the reviews, we are likely to accept this manuscript for publication, providing that you modify the manuscript according to the review recommendations.

We are in principle happy to accept the manuscript and ask only that the authors fix the small textual mistake pointed out by reviewer #1

Please prepare and submit your revised manuscript within 30 days. If you anticipate any delay, please let us know the expected resubmission date by replying to this email.

When you are ready to resubmit, please upload the following:

[1] A letter containing a detailed list of your responses to all review comments, and a description of the changes you have made in the manuscript. Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out

[2] Two versions of the revised manuscript: one with either highlights or tracked changes denoting where the text has been changed; the other a clean version (uploaded as the manuscript file).

Important additional instructions are given below your reviewer comments.

Thank you again for your submission to our journal. We hope that our editorial process has been constructive so far, and we welcome your feedback at any time. Please don't hesitate to contact us if you have any questions or comments.

Sincerely,

Luis Pedro Coelho

Academic Editor

PLOS Computational Biology

Lucy Houghton

Staff

PLOS Computational Biology

***********************

A link appears below if there are any accompanying review attachments. If you believe any reviews to be missing, please contact ploscompbiol@plos.org immediately:

We are in principle happy to accept the manuscript and ask only that the authors fix the small textual mistake pointed out by reviewer #1

Reviewer's Responses to Questions

Comments to the Authors:

Please note here if the review is uploaded as an attachment.

Reviewer #1: The authors have addressed our main concerns. The updated GitHub and user guide looks great!

Error in the Author summary: "difficulting its usability"

Ideas for future developments:

Consider support for alternative annotations to COGs, e.g. KEGG, Phrogs/pvogs for viral annotations etc.

Test dataset for Archaea

Reviewer #2: The authors have taken into account the previous suggestions and remarks. The present manuscript is suitable for publication.

Reviewer #3: Thank you for making changes. I do not have further comments.

**********

Have the authors made all data and (if applicable) computational code underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data and code underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data and code should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data or code —e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: Yes: Michael Roach

Reviewer #2: Yes: Vikash Kumar Singh

Reviewer #3: Yes: Sehi L'Yi

Figure Files:

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email us at figures@plos.org.

Data Requirements:

Please note that, as a condition of publication, PLOS' data policy requires that you make available all data used to draw the conclusions outlined in your manuscript. Data must be deposited in an appropriate repository, included within the body of the manuscript, or uploaded as supporting information. This includes all numerical values that were used to generate graphs, histograms etc.. For an example in PLOS Biology see here: http://www.plosbiology.org/article/info%3Adoi%2F10.1371%2Fjournal.pbio.1001908#s5.

Reproducibility:

To enhance the reproducibility of your results, we recommend that you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. Additionally, PLOS ONE offers an option to publish peer-reviewed clinical study protocols. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols

References:

Review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript.

If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice.

Dear Mr. Durán,

We are pleased to inform you that your manuscript 'GenoVi, an open-source automated circular genome visualizer for bacteria and archaea' has been provisionally accepted for publication in PLOS Computational Biology.

Before your manuscript can be formally accepted you will need to complete some formatting changes, which you will receive in a follow up email. A member of our team will be in touch with a set of requests.

Please note that your manuscript will not be scheduled for publication until you have made the required changes, so a swift response is appreciated.

IMPORTANT: The editorial review process is now complete. PLOS will only permit corrections to spelling, formatting or significant scientific errors from this point onwards. Requests for major changes, or any which affect the scientific understanding of your work, will cause delays to the publication date of your manuscript.

Should you, your institution's press office or the journal office choose to press release your paper, you will automatically be opted out of early publication. We ask that you notify us now if you or your institution is planning to press release the article. All press must be co-ordinated with PLOS.

Thank you again for supporting Open Access publishing; we are looking forward to publishing your work in PLOS Computational Biology. 

Best regards,

Luis Pedro Coelho

Academic Editor

PLOS Computational Biology

Lucy Houghton

Staff

PLOS Computational Biology

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