PONE-D-21-38427Seroprevalence of brucellosis and molecular characterization of Brucella spp. from slaughtered cattle in Rwanda

PLOS ONE

Dear Dr. Ntivuguruzwa,

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.

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In addition to the comments provided by the reviewers please address the following:

Line 26-27: parallel interpretation means positive in at least one test. So, what the authors mentioned is not parallel but the result of serial interpretation i.e, positive in both tests.

Line 56: It was not clear what the authors wanted to mean by “As brucellosis is a herd disease”

Line 105: 0.5% should be5% ; what was the basis to consider a 5% expected prevalence? "The expected prevalence was estimated at 0.5%." Here the word "estimated" is not correct.

Line 200: Brucellosis seroprevalence among slaughter cattle in Rwanda; this subsection can be replaced by “Descriptive statistics” and lines 201 to 205 could be placed under this subsection.

Then the remaining lines can be placed under another sub-section like Brucellosis seroprevalence.

Please insert another table [preferably Table 2] summarizing all test results

Please define brucellosis case status [preferably if culture or PCR positive] and only show the distribution of the brucellosis according to the different variables presented in Table 2 deleting the distribution of brucellosis based on RBT and i-ELISA [now it will become Table 3]. Please recheck the odds ratio as also suggested by one reviewer.

Finally, please evaluate the performance of RBT and i-ELISA considering culture/PCR the as gold standard.

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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: Yes

Reviewer #2: Yes

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

Reviewer #1: No

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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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: Dear Authors,

This is an interesting piece of work that will have an important implication on policy in Rwanda. I must congratulate the team for a well-conducted and written manuscript. However please double check odds ratio and its confidence interval which seems incomplete.

All the best for your future.

“Thank You”

Reviewer #2: Seroprevalence of brucellosis and characterisation of Brucella spp. from slaughtered cattle in Rwanda is well written paper about a very important zoonotic disease of human and animal health

Following are suggestions for improvement of paper before accepted for publication

line 36-37. is test and slaughter policy applicable in Rwanda

line 62-63. remove underline sign from species name of Brucella

line 148, have you performed microscopy or biochemical testing for Brucella susceptable isolates

line 149, there is a need to add details of positive and negative controls used in genus specific PCRs

line 179-180. what awas product length of B. abortus and melitensis in case of AMOS and Bru-ladder PCR assays

-it will be a good addition, if you can add sequencing results too (if possible, otherwise you can try it next time)

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Reviewer #1: No

Reviewer #2: No

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PONE-D-21-38427R1

Seroprevalence of brucellosis and molecular characterization of Brucella spp. from slaughtered cattle in Rwanda

PLOS ONE

Dear Dr. Ntivuguruzwa,

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.

==============================

ACADEMIC EDITOR:

1. Please ignore my previous comment regarding the evaluation of RBT and i-ELISA considering culture/PCR as gold-standard as it was not performed properly and also it may not be possible to evaluate efficiently due small size of culture /PCR positive samples.
2. Please estimate odds ratio appropriately. There should have one reference for every variables and usually the category with lowest prevalence is considered as reference. For example, Low throughput for abattoirs, eastern for provinces, etc.
3. Please replace "univariate" with "univariable" from Table 3 title.
4. Other than reference category, every category of a variable should have an odds ratio. What did you mean by "undetermined"?
5. Please add before line 177: The brucellosis case status was defined based on the results of culture and ITS PCR tests in series.
6. Lines 177-180: Please add the following: The chi-square test was used to evaluate the univariable association between the explanatory variables and brucellosis case status. Any explanatory variable associated with brucellosis at a P≤0.20 was included in the multivariable logistic regression analysis to identify risk factors.
7. Lines 222-223: The brucellosis case status was considered as prevalence based on bacterial growth and  confirmation by ITS PCR. Please delete this sentence.
8. Please delete lines 249-250
9. Please delete lines 334-346 as these results are not statistically significant.
10. The authors could add a discussion about the 70 samples which were culture positive but PCR negative
11. Please elaborate ITS and AMOS in the Tables 2 and 3 foot note
12. Line 348: Please replace "rate" with "prevalence"

==============================

Please submit your revised manuscript by Oct 27 2022 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.

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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.

We look forward to receiving your revised manuscript.

Kind regards,

A. K. M. Anisur Rahman, Ph.D.

Academic Editor

PLOS ONE

Journal Requirements:

Please 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.

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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: Yes

Reviewer #2: Yes

********** 

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

Reviewer #1: Yes

Reviewer #2: 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 #2: 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 #2: 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: The authors have addressed all the comments as suggested and it is a good piece of work which will be useful in the policy implications in Rwanda. Congratulations to the team for the great work.

Thanks and Regards

Reviewer #2: Authors have addressed all of my comments adequately

Hopefully this paper will be a nice piece of research work for brucellosis researchers across the globe.

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Reviewer #1: Yes: Dr Ritik Agrawal

Reviewer #2: No

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PONE-D-21-38427R2Seroprevalence of brucellosis and molecular characterization of Brucella spp. from slaughtered cattle in RwandaPLOS ONE

Dear Dr. Ntivuguruzwa,

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.

==============================

ACADEMIC EDITOR:I still need clarification about one of my previous comment: The authors could add a discussion about the 70 samples which were culture positive but PCR negative

Response: All samples that presented growth on the CITA agar plate were subjected

to ITS PCR for confirmation. The 70 samples were not culture-positive for Brucella and

therefore can not be discussed.If this is the case then please clarify the figure 89 (29%) under the column 'culture'.Please remove Table 4 as it is not based on the results of a multivariable model. Multivariable model must contain at least two variables. As only one variable is associated with a p value <0.20 in the univariable screening there is no need to run the multivariable model. Please include odds ratio and their 95% Confidence interval in the Table 3. ==============================

Please submit your revised manuscript by Dec 04 2022 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.

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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.

We look forward to receiving your revised manuscript.

Kind regards,

A. K. M. Anisur Rahman, Ph.D.

Academic Editor

PLOS ONE

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Please 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.

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Seroprevalence of brucellosis and molecular characterization of Brucella spp. from slaughtered cattle in Rwanda

PONE-D-21-38427R3

Dear Dr. Ntivuguruzwa,

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.

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A. K. M. Anisur Rahman, Ph.D.

Academic Editor

PLOS ONE

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