Molecular characterization of antimicrobial resistance related genes in E. coli, Salmonella and Klebsiella isolates from broilers in the West Region of Cameroon

Jude Fonbah Leinyuy; Innocent Mbulli Ali; Karimo Ousenu; Christopher B. Tume

doi:10.1371/journal.pone.0280150

Peer Review History

Original SubmissionMarch 8, 2022
3 Oct 2022 Decision Letter - Dwij Raj Bhatta, Editor PONE-D-22-06329Characterisation of antimicrobial resistance genes and assessment of penetrance in plasmid-mediated resistance genes in E coli, Salmonella and Klebsiella in the West Region of Cameroon.PLOS ONE Dear Dr. Ali, 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 submit your revised manuscript by Nov 17 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If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. Additional Editor Comments (if provided): Country specific publication on circulating antibiotic resistant isolates of E.coli,Klebsiella Salmonella and molecular charecterization of resistance related genes has been reviewed and the manuscript require minor revision as suggested by reviewers! Answer all queries from reviewer! Revise as suggested! TItle change suggested may be "Molecular charecterization of AMR related genes in E.coli,Klebsiella and Salmonella isolates from Cameroon [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 ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: 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. 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(Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: This study highlights the importance of molecular methods in studying the dissemination of a particular resistance determinant in both resistant and susceptible isolates. I have only minor comments Major comments: 1.In the Financial disclosure section, the authors mentioned that “The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript”. Please provide the details related to the fund as per the author’s guideline. 2.In ethics statement section, the authors need to mention the approval number, type of consent –oral or written, etc. 3.In abstract, the methods part is missing. The main methods should be mentioned in the abstract section. Minor comments: Please correct the grammatical and typing errors in the text such as 1.In introduction section, last paragraph: “We focused on 3 genera o……………………….. which known for the extensive colonisation of their guts by a myriad of Enterobacteriaceae” Please rearrange the sentence 1.Please italicize the genus and species of bacteria throughout the text. For eg, in methods section, E.coli should be italicized. ****** 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. 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Attachments Attachment Submitted filename: review report.docx https://doi.org/10.1371/journal.pone.0280150.r001
Revision 1
21 Oct 2022 Author Response Reviewer comments This study highlights the importance of molecular methods in studying the dissemination of a particular resistance determinant in both resistant and susceptible isolates. I have only minor comments Major comments: In the Financial disclosure section, the authors mentioned that “The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript”. Please provide the details related to the fund as per the author’s guideline. The authors declared at submission while requesting for country-based waiver of publication fees, that the research was not externally funded. Resources for the research were provided by the authors. So we attached the letter from the Head of Department to that effect. The financial disclosure statement selected might have been a submission error which has now been corrected in the resubmission. In ethics statement section, the authors need to mention the approval number, type of consent –oral or written, etc. In Cameroon, the only well-established institutional review board deals with ethical clearance of research on human subjects (the National Ethical Committee for Research involving Human Subjects – CNERSH/SP – Yaoundé, Cameroon). However, at the level of the universities, research projects dealing with animals are reviewed by experts and the protocol is submitted for administrative authorisation from the Ministry of Livestock, Fisheries and Animal Husbandry with a recommendation from the Head of Department concerned. In addition, Law number 2021/014 of 09/07/2021 regulating access and use of genetic material and its derivatives in the Cameroonian territory in its article 4 is not binding to biological resources whose usage is not aimed at exploiting the genetic resources, as the case with our research (https://www.prc.cm/en/news/the-acts/laws/5293). A statement on the consent from farmers to sample broilers at farms has been included in the method section lines 101 to 104. In abstract, the methods part is missing. The main methods should be mentioned in the abstract section. The methods part in the Abstract has been updated to be explicit from lines 19 to 25. Minor comments: Please correct the grammatical and typing errors in the text such as In introduction section, last paragraph: “We focused on 3 genera o……………………….. which known for the extensive colonisation of their guts by a myriad of Enterobacteriaceae” The sentence has been rearranged to bring out the intended meaning, see lines 71 to 74. Please italicize the genus and species of bacteria throughout the text. For eg, in methods section, E.coli should be italicized. The manuscript has been thoroughly reviewed and scientific names italicised. Additional remarks 1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The manuscript has been thoroughly updated to meet journal’s specifications 2. Please include your full ethics statement in the ‘Methods’ section of your manuscript file. In your statement, please include the full name of the IRB or ethics committee who approved or waived your study, as well as whether or not you obtained informed written or verbal consent. If consent was waived for your study, please include this information in your statement as well. We refer the Reviewer to previous comments in response to this concern (number 2 above) 3. We note you have included a table to which you do not refer in the text of your manuscript. Please ensure that you refer to Table 1,2,5,6,7,9,10 & 11 in your text; if accepted, production will need this reference to link the reader to the Table We have made sure In-text table citations have been included as recommended 4. We suggest you thoroughly copyedit your manuscript for language usage, spelling, and grammar. If you do not know anyone who can help you do this, you may wish to consider employing a professional scientific editing service. The manuscript has been thoroughly proofread and language usage, spelling, and grammar errors corrected 5. Please review your reference list to ensure that it is complete and correct. The references have been re-checked to ensure that they match the journal’s specifications Editor recommendation for title change Title change suggested may be "Molecular characterization of AMR related genes in E. coli, Klebsiella and Salmonella isolates from Cameroon” The authors feel that changing the title may in part mask the main aim of the research which not only pointed to the problem of resistance but raises the hypothesis of a mutated (resistance) gene (which does not necessarily translate to being resistant as detected in in-vitro AST) to contribute to the fate of the isolate (penetrance) and hence the ability to silently circulate in the study area. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0280150.r002
15 Nov 2022 Decision Letter - Dwij Raj Bhatta, Editor PONE-D-22-06329R1Characterisation and assessment of penetrance in plasmid-mediated antimicrobial resistance genes in E coli, Salmonella and Klebsiella in the West Region of Cameroon.PLOS ONE Dear Dr. Ali, 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 submit your revised manuscript by Dec 30 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. 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, Dwij Raj Bhatta, PhD 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. Additional Editor Comments: Revision 1 manuscript has been reviewed by editor :Abstrct section ,methodology, poorly written, recheck spelling, simplify language, cut shot abstract section! In result section, catagorize,clearly mention number of sucestible isolates by disc diffusion method and minimum inhibitory concentration of each antibiotics used during susceptibility detetmination! Mention name of Salmonella serovars ,was all Typhi?plasmid en coded AMR genes in Salmonella ,E.coli,and Klebsiella are not charectrized with propoer protocol! Revise result presentation! conclusion : poorly written provide result based conclusion Change title as sugested previously by editor , as editor is not convicend with conclusion of penetrance! remove the section [Note: HTML markup is below. Please do not edit.] [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. https://doi.org/10.1371/journal.pone.0280150.r003
Revision 2
7 Dec 2022 Author Response Response to Editor Comments Abstract section, methodology, poorly written, recheck spelling, simplify language, cut short abstract section! The methodology in the abstract has now been rewritten clearly, however much could not be done to contract the section without leaving out essential information. Lines 21 – 27. In result section, categorize, clearly mention number of susceptible isolates by disc diffusion method and minimum inhibitory concentration of each antibiotic used during susceptibility determination! The result section has been clearly categorised and the presentation of the tables simplified. In addition, The MIC of the antibiotics used can be found in line 151, Table 1. Mention name of Salmonella serovars, was all Typhi? The Salmonella serovars used have been specified. Line 112. Plasmid encoded AMR genes in Salmonella, E. coli, and Klebsiella are not characterized with proper protocol! Authors noted some commissions in the protocol and thank the reviewer for bringing this to our attention. We have now corrected this, mentioning clearly the necessary conditions, quantities and concentrations required for independent reproducibility of the resistance genes characterisation. Lines 161 – 171. Revise result presentation! The result section has been thoroughly revised. Conclusion: poorly written provide result based conclusion The conclusions have been reworded to reflect the primary findings of this work. This can be found in Lines 341 – 348. Change title as suggested previously by editor, as editor is not convinced with conclusion of penetrance! remove the section. The title has been modified to “Molecular characterization of antimicrobial resistance related genes in E. coli, Salmonella and Klebsiella isolates from the West Region of Cameroon” as suggested by the Editor. Authors used the notion of penetrance here to made adequacy between genotype and phenotype. Though the expression of antibiotic resistance can vary in intensity depending on prevailing conditions, we used the notion of penetrance by extrapolation based the interpretations of disc diffusion AST only to give an idea of the proportion of isolates considered susceptible that will carry resistance genes. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0280150.r004
12 Dec 2022 Decision Letter - Dwij Raj Bhatta, Editor PONE-D-22-06329R2Molecular characterization of antimicrobial resistance related genes in E. coli, Salmonella and Klebsiella isolates from the West Region of Cameroon.PLOS ONE Dear Dr. Ali, 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 submit your revised manuscript by Jan 26 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, Dwij Raj Bhatta, PhD 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. Additional Editor Comments : Revision 2 manuscript is well written! However, in the abstract section word treat be replaced by word threat! Salmonella serovars names either S.Tphi or S.ParatyphiA or S.Paratyphi B or S.Paratyphi C and their sucesptibility to tested antibiotics be presented separately not as Salmonella spp. [Note: HTML markup is below. Please do not edit.] [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. https://doi.org/10.1371/journal.pone.0280150.r005
Revision 3
20 Dec 2022 Author Response Response to reviewers Journal Requirements • 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. The financial disclosure provided remains unchanged. • 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. No special protocol was developed by the authors in this manuscript. • 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. The reference list has been thoroughly reviewed and none of the articles, web pages, and videos cited has been retracted. No modification has been made on the references. • 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. The manuscript does not contain images. Additional Editor Comments • In the abstract section word treat be replaced by word threat! We thank the Editor for pointing out this spelling error which escaped out notice despite several revisions. Line 14. To fish out such errors, the manuscript has been submitted to a spelling and grammar check web site (https://quillbot.com) and necessary corrections have been made. • Salmonella serovars names either S.Tphi or S.ParatyphiA or S.Paratyphi B or S.Paratyphi C and their sucesptibility to tested antibiotics be presented separately not as Salmonella spp. We have revised the research data to comply with the revision requested. Lines 113, 119, 181, and 196. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0280150.r006
21 Dec 2022 Decision Letter - Dwij Raj Bhatta, Editor Molecular characterization of antimicrobial resistance related genes in E. coli, Salmonella and Klebsiella isolates from broilers in the West Region of Cameroon. PONE-D-22-06329R3 Dear Dr. Ali, 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, Dwij Raj Bhatta, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): The revision 3 manuscript be accepted for publication as it provides important information on AST pattern of emerging &circulating multidrug resistant E.coli , Klebsiella and Salmonella serovars in Camroon! Reviewers' comments: https://doi.org/10.1371/journal.pone.0280150.r007
Formally Accepted
29 Dec 2022 Acceptance Letter - Dwij Raj Bhatta, Editor PONE-D-22-06329R3 Molecular characterization of antimicrobial resistance related genes in E. coli, Salmonella and Klebsiella isolates from broilers in the West Region of Cameroon. Dear Dr. Ali: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. 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. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Professor Dwij Raj Bhatta Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0280150.r008

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