Bacteriological analysis and antibiotic resistance in patients with diabetic foot ulcers in Dhaka

Poulomi Baral; Nafisa Afnan; Maftuha Ahmad Zahra; Baby Akter; Shek Rabia Prapti; Mohammed Muazzam Hossan; Fahim Kabir Monjurul Haque

doi:10.1371/journal.pone.0301767

Peer Review History

Original SubmissionJanuary 12, 2024
21 Feb 2024 Decision Letter - Shailesh Kumar Shahi, Editor PONE-D-24-01408Bacteriological Analysis and Antibiotic Resistance in Patients with Diabetic Foot Ulcers in DhakaPLOS ONE Dear Dr. Haque, 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 by reviewer 1 during the review process. Please submit your revised manuscript by Apr 06 2024 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, Shailesh Kumar Shahi, PhD 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. PLOS ONE now requires that authors provide the original uncropped and unadjusted images underlying all blot or gel results reported in a submission’s figures or Supporting Information files. This policy and the journal’s other requirements for blot/gel reporting and figure preparation are described in detail at https://journals.plos.org/plosone/s/figures#loc-blot-and-gel-reporting-requirements and https://journals.plos.org/plosone/s/figures#loc-preparing-figures-from-image-files. When you submit your revised manuscript, please ensure that your figures adhere fully to these guidelines and provide the original underlying images for all blot or gel data reported in your submission. See the following link for instructions on providing the original image data: https://journals.plos.org/plosone/s/figures#loc-original-images-for-blots-and-gels. In your cover letter, please note whether your blot/gel image data are in Supporting Information or posted at a public data repository, provide the repository URL if relevant, and provide specific details as to which raw blot/gel images, if any, are not available. Email us at plosone@plos.org if you have any questions. [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: Partly Reviewer #2: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A 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: Yes Reviewer #2: Yes ****** 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: Reviewer responses: The authors carried out this study on an important and most studied disease like diabetes. They have conducted this research on people who have backgrounds in Bangladesh. The author's main study focus was on the diabetic ulcers and the bacterial isolates that are colonized, and the study was conducted more than the required relevance to socioeconomic and educational backgrounds. The authors give a lot of examples and more over frequently they are trying to justify their findings with the previous data and depend on survey studies conducted by the national and international diabetic organizations/ foundations. After reading, understanding and looking at their research results and discussions, I have a few major and minor comments on this manuscript. Major concerns: 1. The authors could have taken up a larger demographics rather than a small number of cohorts. That could help to generate a wider bacterial colony to identify and test their antibiotic resistance in vitro conditions. 2. The authors could have focused on the host and the identified bacterial species and their specific relation in the background of diabetes, as the authors claimed they recruited people from the urban and rural areas which might include hygienic and unhygienic lifestyles might become non-specific hosts for several bacteria for a short period. Precisely identifying the diabetic ulcerative bacterial populations will provide a valuable clue for the treatment. 3. The authors seem to have the same kind of growth media to culture different species of bacteria such as gram-positive and gram-negative and also we cannot rule out the bacterial adaptations to the available nutrients that potentially cause response or resistance to different antibiotics. 4. I would suggest the authors conduct more deliberate experiments to see the bacterial responses to an antibiotic, the current study involved basic experimental settings instead of deep investigations. Minor concerns: 1. The authors have missed a great option to show their all data and importantly they have limitedly presented their results and focused to discuss on the already available data from various sources to justify their study. 2. The figures have a very poor resolution and are not able to read the Figure 1 contents at all. Impression: This study will help as a little reference for further studies whoever wants to investigate deep into the diabetic ulcerative bacteria in connection with antibiotic resistance. Reviewer #2: Authors did an excellent job. The manuscript is well written. It has good research with innovative aim. I will recommend for publication. Some minor grammatical mistakes are in the manuscript that needs to correct before the publication. ****** 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: No Reviewer #2: Yes ******** [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: Reviewer Responses_PLOS ONE.docx https://doi.org/10.1371/journal.pone.0301767.r001
Revision 1
29 Feb 2024 Author Response Reviewer #1: Reviewer responses: The authors carried out this study on an important and most studied disease like diabetes. They have conducted this research on people who have backgrounds in Bangladesh. The author's main study focus was on the diabetic ulcers and the bacterial isolates that are colonized, and the study was conducted more than the required relevance to socioeconomic and educational backgrounds. The authors give a lot of examples and more over frequently they are trying to justify their findings with the previous data and depend on survey studies conducted by the national and international diabetic organizations/ foundations. After reading, understanding and looking at their research results and discussions, I have a few major and minor comments on this manuscript. Major concerns: 1. The authors could have taken up a larger demographics rather than a small number of cohorts. That could help to generate a wider bacterial colony to identify and test their antibiotic resistance in vitro conditions. Author’s Response: We value the feedback from the reviewers. Despite the constraints posed by limited resources, facilities, and the challenges associated with conducting clinical studies in Bangladesh, we made concerted efforts to optimize our sample size. It is important to note that our study is only the third of its kind in Bangladesh (1,2), with the most recent one conducted nearly a decade ago (1). This scarcity of research underscores the significance and timeliness of our contribution to the existing body of knowledge in this domain. While we do recognize the potential benefits of a larger population and the inclusion of a broader range of bacterial species to enhance thoroughness, nonetheless, our study serves as a catalyst for further, extensive research on this topic. 2. The authors could have focused on the host and the identified bacterial species and their specific relation in the background of diabetes, as the authors claimed they recruited people from the urban and rural areas which might include hygienic and unhygienic lifestyles might become non-specific hosts for several bacteria for a short period. Precisely identifying the diabetic ulcerative bacterial populations will provide a valuable clue for the treatment. Author’s Response: We appreciate the reviewer’s comments and acknowledge them. An unhygienic lifestyle can heighten the susceptibility of sores to infections. Nevertheless, due to the prolonged persistence of infections, the patients were hospitalized. All participants in our study were hospitalized, ensuring adherence to rigorous hygienic guidelines. Moreover, our study centered on patients from Ibrahim Diabetic Foot Care Hospital and Shaheed Suhrawardy Medical College and Hospital, institutions renowned for their advanced diabetic treatment facilities. Samples were obtained from deep pus, and the infections were categorized based on Wagner's grade, indicating active infections. This strongly implies that the identified bacteria are the causative agents of these infections. Our findings guided physicians in prescribing antibiotics. Nevertheless, we recognize a limitation in our focus on hosts. Despite our efforts to prioritize patients, time constraints and follow-up challenges hindered a more in-depth exploration of host-bacteria relationships. We concede that a more extensive investigation into the host factors could have yielded additional insights. Due to this, our study emphasized analyzing bacterial species and their antibiotic resistance profiles. 3. The authors seem to have the same kind of growth media to culture different species of bacteria such as gram-positive and gram-negative and also we cannot rule out the bacterial adaptations to the available nutrients that potentially cause response or resistance to different antibiotics. Author’s Response: We acknowledge the reviewer's feedback. In our study, only one gram-positive bacteria, specifically Staphylococcus aureus, was isolated. The standard mannitol salt agar (MSA) was employed as the selective medium for its cultivation. For other gram-negative bacterial species, we utilized well-established culture media: Cetrimide Agar for Pseudomonas aeruginosa, HiCrome KPC Agar for Klebsiella pneumoniae, HiCrome UTI Agar for Escherichia coli (confirmed later using EMB Agar), and MacConkey Agar for Proteus spp. These culture media are routinely employed in the growth of their respective bacteria. Information about the culture media used for growing these bacteria is provided in S2 Table. It is noteworthy that none of our culture media incorporated antibiotic supplements, mitigating the likelihood of bacterial adaptations to available nutrients potentially influencing responses or resistance to different antibiotics. 4. I would suggest the authors conduct more deliberate experiments to see the bacterial responses to an antibiotic, the current study involved basic experimental settings instead of deep investigations. Author’s Response: We appreciate the reviewer's suggestion. The in vitro antibiotic testing of the isolates was conducted in triplicate. The primary focus of our research was to assess resistance of bacterial isolates to the recommended antibiotic dosages. Our study did not delve into the effects of antibiotics during follow-up; this was beyond the scope of our investigation. Minor concerns: 1. The authors have missed a great option to show their all data and importantly they have limitedly presented their results and focused to discuss on the already available data from various sources to justify their study. Author’s Response: We appreciate the reviewer's comments. Considering the reviewers concern we have made adjustments to ensure a greater focus on our results. All the data in the revised manuscript has been thoroughly presented. This modification aims to prevent our findings from being overshadowed by data from other studies. 2. The figures have a very poor resolution and are not able to read the Figure 1 contents at all. Author’s Response: We appreciate the feedback from the reviewers. The photos have been formatted in accordance with PLOS One guidelines. However, we acknowledge that the resolution may have been compromised during the conversion of the photos into a PDF file. We concur with the observation that the contents of Figure 1 are challenging to read, and consequently, we have implemented modifications to enhance its legibility. Reviewer #2: Authors did an excellent job. The manuscript is well written. It has good research with innovative aim. I will recommend for publication. Some minor grammatical mistakes are in the manuscript that needs to correct before the publication. Author’s Response: We are appreciative of the reviewer’s comments. We have thoroughly rechecked our manuscript and corrected all the grammatical mistakes. References: 1. Karmaker M, Sanyal SK, Sultana M, Hossain MA. Association of bacteria in diabetic and non-diabetic foot infection - An investigation in patients from Bangladesh. J Infect Public Health [Internet]. 2016;9(3):267–77. Available from: http://dx.doi.org/10.1016/j.jiph.2015.10.011 2. Paul S, Barai L, Jahan A, Haq JA. A Bacteriological Study of Diabetic Foot Infection in an Urban Tertiary Care Hospital in Dhaka City. Ibrahim Med Coll J. 2009;3(2):50–4. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0301767.r002
21 Mar 2024 Decision Letter - Shailesh Kumar Shahi, Editor Bacteriological Analysis and Antibiotic Resistance in Patients with Diabetic Foot Ulcers in Dhaka PONE-D-24-01408R1 Dear Dr. Haque, 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. If you have any questions relating to publication charges, 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, Shailesh Kumar Shahi, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): Dear Dr. Fahim Kabir Monjurul Haque, Thank you very much for properly revising the manuscript. I would appreciate it if the resolution of the figures could be improved. High-quality visuals are crucial to effectively convey our findings and ensure clarity for the readers. Additionally, I recommend a thorough review and correction of the English grammar and language usage throughout the manuscript. I believe that these adjustments will significantly enhance the overall quality of the publication. Best Wishes, Shailesh Shahi 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 ******** 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: Partly 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: (No Response) Reviewer #2: (No Response) ****** 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: Chakrapani Vemulawada Reviewer #2: Yes: Avinash Singh ******** https://doi.org/10.1371/journal.pone.0301767.r003
Formally Accepted
30 Apr 2024 Acceptance Letter - Shailesh Kumar Shahi, Editor PONE-D-24-01408R1 PLOS ONE Dear Dr. Haque, 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 If revisions are needed, the production department will contact you directly to resolve them. If no revisions are needed, you will receive an email when the publication date has been set. At this time, we do not offer pre-publication proofs to authors during production of the accepted work. Please keep in mind that we are working through a large volume of accepted articles, so please give us a few weeks 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. 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 Dr. Shailesh Kumar Shahi Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0301767.r004

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 .