PONE-D-22-29703Decrease in the prevalence of antimicrobial resistance in Escherichia coli isolates of Canadian turkey flocks driven by the implementation of an antimicrobial stewardship programPLOS ONE

Dear Dr. Varga,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

Please respond to all comments by the reviewers.  To cater to readers that are not that statistical savvy, in the materials and methods please include a short 1 or 2 sentence explanation on the use of negative binomial mixed effect regression and then mixed effect multivariable logistic regression models for analysis of the data.  Please keep the name of the statistical technique consistent throughout the manuscript. In the Data Management, include the basis of considering 'intermediate' as susceptible rather than 'resistant. In the Discussion, include discussion of the study findings in relation to the variable AMR findings from countries that have had long term experience of controlling antibiotic usage in food animals such as Scandinavian countries.

Please submit your revised manuscript by Jan 12 2023 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.

Please include the following items when submitting your revised manuscript:

• A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Latiffah Hassan

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at 

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and 

https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. In your Data Availability statement, you have not specified where the minimal data set underlying the results described in your manuscript can be found. PLOS defines a study's minimal data set as the underlying data used to reach the conclusions drawn in the manuscript and any additional data required to replicate the reported study findings in their entirety. All PLOS journals require that the minimal data set be made fully available. For more information about our data policy, please see http://journals.plos.org/plosone/s/data-availability.

""Upon re-submitting your revised manuscript, please upload your study’s minimal underlying data set as either Supporting Information files or to a stable, public repository and include the relevant URLs, DOIs, or accession numbers within your revised cover letter. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories. Any potentially identifying patient information must be fully anonymized.

Important: If there are ethical or legal restrictions to sharing your data publicly, please explain these restrictions in detail. Please see our guidelines for more information on what we consider unacceptable restrictions to publicly sharing data: http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions. Note that it is not acceptable for the authors to be the sole named individuals responsible for ensuring data access.

We will update your Data Availability statement to reflect the information you provide in your cover letter.

[Note: HTML markup is below. Please do not edit.]

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: Yes

Reviewer #2: Partly

**********

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

Reviewer #1: Yes

Reviewer #2: Yes

**********

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: No

Reviewer #2: No

**********

4. 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: Yes

Reviewer #2: Yes

**********

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: Rima D. Shrestha（PONE-D-22-29703）and colleagues, examined the AMU and AMR surveillance data over six years (2016-2021), collected by CIPARS from turkey farms across three major turkey-producing Canadian provinces/regions to evaluate the impact of a stepwise AMU reduction strategy on the prevalence of AMR in E. coli isolated from turkey flocks. The topic is highly important in the field for mitigating AMR crisis. The overall study is well organized and analyzed, and I have a few minor comments in the below:

1) considering the data transparency and reproducible result, the raw data or so-called source data should be provided. This is the core part in this study. Therefore, excel sheet with detail information regarding all E. coli AMR data, as well as their metadata, should be provided as the supplemental document. This is the source data needed for further evaluation.

2) Even though the table 2 is informative, I would suggest a few figures based on the data from the table could deliver more attractive trend from the study, I would provide this figures as the supplemental documents.

3) Line 304. It would be nice to group the strains with their regions/provinces, for flocks in this heatmap.

4) Additionally, more informative analysis sould be focus on certain antimicrobials that some flocks use and other flocks do not use (a comparison between two groups). A pari-wise comparison would be more informative, side-by-side comparison, the outcome (decreasing trend) of the use and not use certain antimicrobials. I would like to see these results, which might further support AMU has a direct impact on AMR.

Reviewer #2: The manuscript evaluates the implementation of an antimicrobial stewardship strategy in the Canadian turkey industry. The strategy consist on several stepwise withdrawals of antimicrobial uses; in 2014 3rd generation cephalosporins and fluoroquinolones were withdrawn; in 2018, aminoglycosides, macrolides, penicillins, and streptogramin, and between 2019 and 2020, elimination of preventive use of bacitracin and tetracyclines was implemented. Documenting the effectiveness of antimicrobial stewardship is not a particular novel area, but certainly an important one. The major criticism of this paper is that the claims that the implementation was highly effective is not always supported by the data and/or analysis. The effectiveness of the interventions seems to be based on the year effects using 2016 as referent, but the major implementations took place from 2018 to 2020, so comparing against 2016 does not seem the most useful approach to evaluate interventions (L322-L332). For example, based on your figure 2, it looks like tetracycline resistance started to decrease in 2017 before the step 3 strategy was implemented. What other factors could explain the reduction in resistance?. As tetracycline, gentamicin also indicated a sharp decrease from 2017 to 2018 although the implementation was done at the end of 2018.

If implementation finishes in 2020, is it reasonable to only use data from 2020-2021 as a basis to evaluate the post-implementation period?

Reporting the AMU use (currently in Table 1) as a figure (e.g. histogram of AMU by year and by antibiotic category would help to put the results of AMR in perspective. I.e., Did the reductions in AMU really precede the changes in AMR?

**********

6. 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: Min Yue

Reviewer #2: No

**********

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment 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. Registration is free. 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 PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Decrease in the prevalence of antimicrobial resistance in Escherichia coli isolates of Canadian turkey flocks driven by the implementation of an antimicrobial stewardship program

PONE-D-22-29703R1

Dear Dr. Varga,

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

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Latiffah Hassan

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 #3: All comments have been addressed

**********

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: Yes

Reviewer #3: Yes

**********

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

Reviewer #1: Yes

Reviewer #3: Yes

**********

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: Yes

Reviewer #3: Yes

**********

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: Yes

Reviewer #3: Yes

**********

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: none, I feel that great improvements from current version, which is ready for conditionally acceptance.

Reviewer #3: The authors have satisfactorily addressed comments from the reviewers. However, the following issues should be addressed to improve the clarity of the manuscript:

-Line 204: Include the percentages for the proportions of the farms that used antimicrobials by region (Western region 204 (n=157/209,?), followed by Ontario (n=100/153, ?) and Québec (n=75/148,?) so that the reporting formats are consistent with those presented before ( see Lines 201-203).

-Use commas (,) to separate thousands in figures with four (4) digits and above so that these figures are not confused with years in reference. For example, use 1,976 in place of 1976. Do this for all 4-digits and above figures throughout the manuscript.

-Line 396: insert a space between ‘Quebec’ and ‘and’ to show that they are two separate words.

**********

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: Yes: Min Yue

Reviewer #3: Yes: Hassan Ismail Musa

**********