PONE-D-19-20091

Analysis of Virulence Potential of Escherichia coli O145 Isolated from Cattle Feces and Hide Samples Based on Whole Genome Sequencing

PLOS ONE

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: No

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data 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 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—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English?

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Reviewer #1: Yes

Reviewer #2: Yes

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5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Comments.

Author line: No affiliation for number 3 (Lance W. Noll3)

Abstract and elsewhere in the manuscript: the “average genome size” should be changed to say the “approximate average genome size” as these are just draft genomes.

Introduction, lines 58 and 63. Remove “have” from the sentence.

Materials and Methods, lines 82-85. I was surprise to see such few isolates used to make a major claim of cattle STEC O145:H28 isolates potential can cause human illness. There were only two human isolates used in this study. I don’t know how there can be any power to your statistical analysis with just two strains. I am not even sure the 16 cattle feces isolates would be enough to represent all the genetic diversity of STEC O145:H28 in cattle. There are sequenced STEC O145 that can be downloaded from Genbank to increase the number of human isolates and increase the diversity of the isolates used in this study.

Materials and Methods, lines 82-85. The two references sited (14 and 15) are both from cattle feces. I didn’t see where any of them were from an abattoir. How did you select which cattle fecal isolates to sequence? Between the two papers in the reference, there should have been well over 100 isolates you could sequence. Were the hide samples from the abattoir from the same or feed yard? They are closely related which makes me think they are epidemiologically related. How do you explain the clustering of cattle feces and hide isolates on the tree when the hide isolate more than likely came from feces?

Discussion, line 298. Remove “the” before ”fecal strains …”.

Reviewer #2: The manuscript describes the characterization of Shiga toxin-producing Escherichia coli O145 serogroup strains isolated from cattle feces, hide and human clinical samples. The paper is of interest, particularly due to the lack of information from the O145 serogroup, but some aspects need to be clarified.

1. I suggest using “big six non-O157 STEC” instead of “top-7 STEC”.

2. I suggest mention that your sample size is small (in the conclusion of abstract and discussion)

3. Lines 45 and 46 are not suitable for initiating of introduction, I suggest removing them.

4. Line 69: please mention the time of sampling for O145 collection? How many years? How many months? Since what year and month to what year and month?

5. Line 86 data is more appropriate for results section not for materials and methods.

9. Line 99: Correct the “))”

10. Line 144: "[260 (224-291)]" seems to be better than "(260 [224-291])". Check it.

11. Because of the small sample size, I disagree with percentage in the text and table 2. Consult with an epidemiologist.

12. in figure captions: What algorithm is used? (For example: UPGMA), what similarity coefficient is used? (For example: Dice)

13. In discussion, you can compare your stx subtypes with some other studies to show the importance of cattle as a disseminator of the subtypes:

For example: Virulence genes, Shiga toxin subtypes, major O-serogroups, and phylogenetic background of Shiga toxin-producing Escherichia coli strains isolated from cattle in Iran. Microbial pathogenesis. 2017 Aug 1;109:274-9.

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Reviewer #1: No

Reviewer #2: No

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Analysis of Virulence Potential of Escherichia coli O145 Isolated from Cattle Feces and Hide Samples Based on Whole Genome Sequencing

PONE-D-19-20091R1

Dear Dr. Nagaraja,

We are pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it complies with all outstanding technical requirements.

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With kind regards,

Baochuan Lin, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation.

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: (No Response)

Reviewer #2: Yes

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3. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: (No Response)

Reviewer #2: Yes

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4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data 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 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—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: (No Response)

Reviewer #2: Yes

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5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: (No Response)

Reviewer #2: Yes

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6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: (No Response)

Reviewer #2: (No Response)

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7. 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: No

Reviewer #2: No