Application of various mixtures of medicinal herbs in the diet of laying hens: Evaluating preventive approach of fatty liver syndrome

Mohammadreza Khodaei; Mehran Torki; Fariborz Khajali; Iraj Karimi

doi:10.1371/journal.pone.0330363

Peer Review History

Original SubmissionApril 4, 2025
5 Jun 2025 Decision Letter - Ewa Tomaszewska, Editor Dear Dr. Torki, Please submit your revised manuscript by Jul 20 2025 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. 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, Ewa Tomaszewska, DVM Ph.D 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. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information . [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? 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 Reviewer #1: Yes Reviewer #2: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English??> Reviewer #1: No Reviewer #2: No ****** Reviewer #1: While the manuscript presents a well-designed experiment exploring the use of herbal additives to prevent FLHS in laying hens, the overall quality of English language and expression requires significant improvement. Numerous grammatical errors, awkward phrasings, and formatting inconsistencies—especially in the introduction and methods sections—may hinder reader comprehension. For instance, several sentences are either incomplete or improperly constructed, such as line 76–77, and citation formatting is occasionally unclear or broken. I recommend that the authors undergo thorough professional English language editing to enhance clarity, readability, and overall presentation before further consideration. Reviewer #2: Dear Authors, Strictly follow my comments/corrections in the attached file. 1. In the introduction section, need for the study should be highlighted at the end followed by the hypothesis of the study 2. In methodology, clearly elaborate the sample size determination 3. add appropriate citations 4. In the results, improve readability and add actual p-value of each result 5. In the discussion, add logical reasoning of each result before discussing with the previous studies. 6. In the conclusion, briefly describe and add limitation of the study Regards ****** 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: Yes: Sohail Ahmad ******** [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. Attachments Attachment Submitted filename: Review of Article-20250523.docx Attachment Submitted filename: PONE-D-25-15950_reviewer.pdf https://doi.org/10.1371/journal.pone.0330363.r001
Revision 1
1 Jul 2025 Author Response Reviewer #1: While the manuscript presents a well-designed experiment exploring the use of herbal additives to prevent FLHS in laying hens, the overall quality of English language and expression requires significant improvement. Numerous grammatical errors, awkward phrasings, and formatting inconsistencies—especially in the introduction and methods sections—may hinder reader comprehension. For instance, several sentences are either incomplete or improperly constructed, such as line 76–77, and citation formatting is occasionally unclear or broken. I recommend that the authors undergo thorough professional English language editing to enhance clarity, readability, and overall presentation before further consideration. We have thoroughly reviewed the entire manuscript and carefully revised the text to address all grammatical errors, awkward phrasings, and formatting inconsistencies, particularly in the Introduction and Methods sections as noted. The sentence on lines 76–77 has been restructured for clarity. The citation formatting throughout the manuscript has been corrected. We have also sought assistance from a professional English language editing service to ensure the manuscript meets high standards of readability and presentation. Line 37, the author could change “background” to “introduction”, because “background” may be confused as the same one in Abstract. The heading “Background” was changed to “Introduction”. Lines 38–42: The point that the liver synthesizes lipids and is heavily burdened in hens is repeated. The author could try to combine them with a single sentence. We have revised the text to eliminate redundancy and merged the information into a concise sentence (page 2, lines 40-44). The methods mentioned in the manuscript should be cited formally. Such as Line 51-54. We have carefully reviewed the entire Methods section to ensure all procedures, including those mentioned in lines 51–54, are properly cited with relevant and appropriate references. Lines 55–64: The paragraph listing all plants and their bio-actives is dense and hard to read. The author could Use a table or bullet points for readability or briefly summarize: "These herbs are rich in flavonoids, alkaloids, sulfur compounds, and dietary fibers, with demonstrated antioxidant, anti-inflammatory, and lipid-lowering properties." Thank you for your helpful suggestion. We have replaced the dense paragraph with a table summarizing the medicinal plants used and their key bioactive components (Table 1). Lines 37-69, the author could use clearer topic sentences to guide the reader through each section logically, including liver function, disease description, economic impact, and herbal solutions. We have revised lines 37–69 to include clearer and more distinct topic sentences at the beginning of each major section. Lines 88–90, the author use informal phrasing “running into the experimental trial", which can be corrected to "A two-week adaptation period was implemented to acclimate hens to the diets prior to the 12-week experimental trial." We have revised the sentence to eliminate informal phrasing. For animal study design, the randomization or blinding details were missing. Please mark those in the experiment design part. Thank you for highlighting this important point. In response, we have updated the experimental design section to include details regarding randomization and blinding (page 5, lines 89-90). No mention of sample size calculation or statistical power. The author could revise as "Sample size was determined based on expected effect sizes from previous studies to achieve 80% power at α = 0.05." We have revised the Methods section to include a statement on sample size calculation. The text now reads: “Sample size was determined based on expected effect sizes from previous studies to achieve 80% power at α = 0.05” (page 5, lines 88-89). Reviewer #2: Dear Authors, Strictly follow my comments/corrections in the attached file. 1. In the introduction section, need for the study should be highlighted at the end followed by the hypothesis of the study Thank you for your insightful comment. We have revised the final part of the Introduction to clearly emphasize the need for the study and to better establish the context and significance of our research. Additionally, we have now explicitly stated the hypothesis of the study to guide the reader and clarify our research objectives. 2. In methodology, clearly elaborate the sample size determination We have revised this section to provide a clear explanation of how the sample size was determined (Page 5, lines 88-89). 3. Add appropriate citations We have thoroughly reviewed the entire Methods section and ensured that all procedures are now properly cited with relevant and appropriate references. 4. In the results, improve readability and add actual p-value of each result Thank you for your valuable comment. We have thoroughly revised the Results section to enhance clarity and improve readability. Additionally, we have included the actual p-values for all statistical comparisons rather than simply stating significance levels (e.g., p < 0.05). 5. In the discussion, add logical reasoning of each result before discussing with the previous studies. Thank you. We have revised the Discussion section to provide a clear and logical interpretation of each key result before comparing our findings with those of previous studies. These revisions aim to enhance the scientific depth and coherence of the discussion. 6. In the conclusion, briefly describe and add limitation of the study. We have revised the Conclusion section to include a brief summary of the main findings, emphasizing the significance of our results. In addition, we have acknowledged the limitations of the study, such as the relatively short duration, the use of a single dose per herbal mixture, the lack of individual herb evaluation, and the restriction to one hen strain and age group. This addition provides a balanced perspective and suggests directions for future research. Ad libitum feed was provided? at this age? why? The CD was intentionally formulated to predispose hens to fatty liver hemorrhagic syndrome (FLHS). However, due to its imbalanced nutrient profile, feed intake could be influenced as hens attempt to compensate for nutrient deficiencies. By allowing ad libitum access, we enabled the hens to express natural compensatory feeding behavior, which is crucial for observing true diet-induced metabolic and hepatic changes. https://doi.org/10.1371/journal.pone.0330363.r002
12 Jul 2025 Decision Letter - Ewa Tomaszewska, Editor Dear Dr. Torki, 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 Aug 26 2025 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. 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 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, Ewa Tomaszewska, DVM Ph.D Academic Editor PLOS ONE Journal Requirements: If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise. 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. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author Reviewer #1: All comments have been addressed Reviewer #2: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions??> Reviewer #1: Yes Reviewer #2: Partly ****** 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 Reviewer #1: Yes Reviewer #2: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English??> Reviewer #1: Yes Reviewer #2: Yes ****** Reviewer #1: The manuscript is well written and addresses an important question in the field. The authors have responded thoroughly to all previous comments and have made the necessary revisions to improve the clarity, rigor, and impact of the work. I have no further concerns regarding scientific content, ethics, or publication practices. I recommend the manuscript for publication in its current form. Reviewer #2: Dear Authors, Follow my comments in the attached file, in the materials and methods section, give details about cage manufacturer and stocking density per bird. Also remove the model used for THI by Marai et al. as this study focuses on Rabbits and this model is applicable to different species. Remove this model and add the appropriate model suitable for chickens. Improve the logical reasoning of the results. in conclusion, add limitations of the study Regards ****** 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: Yes: Sohail Ahmad ******** [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 Attachments Attachment Submitted filename: PONE-D-25-15950_R1_reviewer.pdf https://doi.org/10.1371/journal.pone.0330363.r003
Revision 2
25 Jul 2025 Author Response Reviewer #1: The manuscript is well written and addresses an important question in the field. The authors have responded thoroughly to all previous comments and have made the necessary revisions to improve the clarity, rigor, and impact of the work. I have no further concerns regarding scientific content, ethics, or publication practices. I recommend the manuscript for publication in its current form. Thank you very much, indeed. Reviewer #2: Dear Authors, Follow my comments in the attached file, in the materials and methods section, give details about cage manufacturer and stocking density per bird. Thank you. The stocking density has been already provided in the Materials and Methods section, as stated in the following sentence: “The hens were housed in wire cages (Machine Toyour, Co., Juybar, Mazandaran, Iran) measuring 45 × 45 × 45 cm, with three birds per cage, corresponding to a stocking density of 675 cm² per bird.” The name of cage manufacturer has now been added (Page 5, Lines 91-92). Also remove the model used for THI by Marai et al. as this study focuses on Rabbits and this model is applicable to different species. Remove this model and add the appropriate model suitable for chickens. Thank you for bringing this up. We have replaced the previous THI model by a new THI model specifically used for poultry (Boonkum et al., 2025). Improve the logical reasoning of the results. Thank you for your comment. Based on the available literature, we have improved the logical reasoning of the results by clarifying the arguments, aligning our findings with previous studies, and strengthening the supporting evidence. Please see the changes in the revised manuscript. In conclusion, add limitations of the study Thank you for your comment. The limitation of the study had already been acknowledged in the conclusion section (Page 31, Lines 563-566). Attachments Attachment Submitted filename: Respone to Reviewers.docx https://doi.org/10.1371/journal.pone.0330363.r004
31 Jul 2025 Decision Letter - Ewa Tomaszewska, Editor Application of various mixtures of medicinal herbs in the diet of laying hens: Evaluating preventive approach of fatty liver syndrome PONE-D-25-15950R2 Dear Dr. Mehran Torki, 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 will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. For questions related to billing, please contact billing support . 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, Ewa Tomaszewska, DVM Ph.D Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: Reviewer's Responses to Questions Comments to the Author Reviewer #2: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions??> Reviewer #2: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? -->?> Reviewer #2: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available??> The PLOS Data policy Reviewer #2: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English??> Reviewer #2: Yes ****** Reviewer #2: (No Response) ****** 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 #2: No ******** https://doi.org/10.1371/journal.pone.0330363.r005
Formally Accepted
Acceptance Letter - Ewa Tomaszewska, Editor PONE-D-25-15950R2 PLOS ONE Dear Dr. Torki, I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now being handed over to our production team. At this stage, our production department will prepare your paper for publication. This includes ensuring the following: * All references, tables, and figures are properly cited * All relevant supporting information is included in the manuscript submission, * There are no issues that prevent the paper from being properly typeset You will receive further instructions from the production team, including instructions on how to review your proof when it is ready. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few days to review your paper and let you know the next and final steps. Lastly, 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. You will receive an invoice from PLOS for your publication fee after your manuscript has reached the completed accept phase. If you receive an email requesting payment before acceptance or for any other service, this may be a phishing scheme. Learn how to identify phishing emails and protect your accounts at https://explore.plos.org/phishing. If we can help with anything else, please email us at customercare@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 Ewa Tomaszewska Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0330363.r006

Open letter on the publication of peer review reports

PLOS recognizes the benefits of transparency in the peer review process. Therefore, we enable the publication of all of the content of peer review and author responses alongside final, published articles. Reviewers remain anonymous, unless they choose to reveal their names.

We encourage other journals to join us in this initiative. We hope that our action inspires the community, including researchers, research funders, and research institutions, to recognize the benefits of published peer review reports for all parts of the research system.

Learn more at ASAPbio .