Dysglycemia and the airway microbiome in cystic fibrosis

Laura N. Brenner; Ching-Ying Huang; Minsik Kim; Lindsay Bringhurst; Christopher J. Richards; Leonard Sicilian; Isabel Neuringer; Melissa S. Putman; Peggy S. Lai

doi:10.1371/journal.pone.0331847

Peer Review History

Original SubmissionMarch 21, 2025
5 Jun 2025 Decision Letter - Vinod Yata, Editor PONE-D-25-12412Dysglycemia and the airway microbiome in cystic fibrosis.PLOS ONE Dear Dr. BRENNER, 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 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. 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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. 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 ****** 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 ****** 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: The authors attempt to associate cystic fibrosis related diabetes condition to changes in the airway microbiome using metagenomics which as the authors have evidenced in this study and analysis, an important factor in understanding this lung disease. The study design is sound with recruitment, sample and clinical data collection, group stratification based on glucose tolerance, etc. The statistical analysis performed is also very appropriate with the metrics used widely in ecology studies (diversity indices, species abundance, species richness, etc.). The preliminary conclusions seem to be supported by the data and overall this paper seems complete with a note in the discussion on future directions with more rigorous strategies on glycemic characterization, longitudinal follow-up, etc. Only a few minor comments follow: 1. I understand the logic behind use of inverse Simpson index to assess species diversity and Bray-Curtis distance for dissimilarity in species composition was very intuitive. It would be great if the data were also tested on another diversity index since in my experience the sensitivity of true diversity to rare vs abundant species varies in different indices. Although not crucial, but this would be good to verify the conclusions of the study. 2. The value of inverse Simpson indicating a lower alpha diversity associated with CFRD status in results section (-2.30) is different than the one given in the abstract (-1.98) although confidence intervals and p values agree. 3. In Figure 4 showing Antibiotic resistance gene abundance differences, the labels overlap a lot so hard to read. Also the figure uses BH corrected q values on y axis but the figure description mentions using raw p values instead. Also a small justification of why figure 2 used raw p values whereas in 3 and 4, BH q values were used. 4. In Figure 5a and 5b, is there a specific reason that the bars are different colors? They don't match other figures which have their own legends which makes complete sense if colors are different, but the lack of a legend in 5a and 5b suggests that the bars really don't need to be colored. 5. Table 2 which mentions association between microbiome and virome CFRD signatures and percent predicted FEV1 seems like the main table supporting the conclusion of the paper that virome and not microbiome being associated to lung function. This table was hard for me to really understand what the different regression models were in terms of which combinations of confounders were tested so a brief explanation on how models 1 through 5 are different would be nice. Apart from these, minor typos in the discussion sections and where Trimmomatic is cited in the methods section. Overall, a fun and exciting paper to review. ****** 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? 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Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0331847.r001
Revision 1
27 Jun 2025 Author Response We greatly appreciate the thoughtful review of our manuscript. We have made numerous revisions in response to these comments, which we believe have improved the quality of the manuscript. We describe these in detail below. Reviewer #1: The authors attempt to associate cystic fibrosis related diabetes condition to changes in the airway microbiome using metagenomics which as the authors have evidenced in this study and analysis, an important factor in understanding this lung disease. The study design is sound with recruitment, sample and clinical data collection, group stratification based on glucose tolerance, etc. The statistical analysis performed is also very appropriate with the metrics used widely in ecology studies (diversity indices, species abundance, species richness, etc.). The preliminary conclusions seem to be supported by the data and overall this paper seems complete with a note in the discussion on future directions with more rigorous strategies on glycemic characterization, longitudinal follow-up, etc. Response: Thank you for your generous comments and suggestions. Only a few minor comments follow: 1. I understand the logic behind use of inverse Simpson index to assess species diversity and Bray-Curtis distance for dissimilarity in species composition was very intuitive. It would be great if the data were also tested on another diversity index since in my experience the sensitivity of true diversity to rare vs abundant species varies in different indices. Although not crucial, but this would be good to verify the conclusions of the study. Response: We appreciate this thoughtful suggestion regarding the use of additional diversity indices. We have done so and the results remain consistent. We revised the results as below: In our linear model assessing the relationship between glycemic status and alpha diversity, CFRD status was associated with lower alpha diversity, as reflected by a decrease in the Inverse Simpson index (-1.98 [-3.80, -0.16], p = 0.033) and a nonsignificant reduction in species richness (-3.13 [-7.61, 1.34], p = 0.17). In the multivariate model, the effect size of the association between CFRD status and the Inverse Simpson index remained similar, with a beta estimate of -2.00 [-4.02, 0.02] (p = 0.052). IGT status was not significantly associated with alpha diversity. To test the hypothesis that glycemia affected overall microbial community structure, we assessed differences in beta diversity. CFRD status was associated with microbial community structure as measured by the Bray Curtis (PERMANOVA p=0.011, R2= 17%) (Figure 1) and Horn-Morisita (PERMANOVA p= 0.0067, R2 = 19%) dissimilarity in the multivariate model. 2. The value of inverse Simpson indicating a lower alpha diversity associated with CFRD status in results section (-2.30) is different than the one given in the abstract (-1.98) although confidence intervals and p values agree. Response: We apologize for the inconsistency. The results section reported an incorrect effect size and was updated to the correct one. 3. In Figure 4 showing Antibiotic resistance gene abundance differences, the labels overlap a lot so hard to read. Also the figure uses BH corrected q values on y axis but the figure description mentions using raw p values instead. Also a small justification of why figure 2 used raw p values whereas in 3 and 4, BH q values were used. Response: We have updated figure 4 to have less overlap and the updated the legend to use q-values. We have updated all the figures to use q values for consistency. 4. In Figure 5a and 5b, is there a specific reason that the bars are different colors? They don't match other figures which have their own legends which makes complete sense if colors are different, but the lack of a legend in 5a and 5b suggests that the bars really don't need to be colored. Response: We appreciate this helpful suggestion. We agree that the color variation in Figures 5a and 5b was unnecessary and potentially confusing. We have now updated both figures to use a single, consistent color for all bars. Additionally, we have included updated loading plots that incorporate covariates for clarity and completeness. These changes have improved the consistency and interpretability of the figure. 5. Table 2 which mentions association between microbiome and virome CFRD signatures and percent predicted FEV1 seems like the main table supporting the conclusion of the paper that virome and not microbiome being associated to lung function. This table was hard for me to really understand what the different regression models were in terms of which combinations of confounders were tested so a brief explanation on how models 1 through 5 are different would be nice. Response: Thank you for point this out. We have updated the table legend to better describe each models as below: "Univariate and multivariate linear regression models evaluating the association between CFRD-related microbiome and virome signatures and percent predicted FEV1. Model 1 includes only the virome score, while Model 2 includes only the microbiome score. Models 3 and 4 assess the virome and microbiome scores, respectively, with adjustment for age, sex, modulator use, CFRD status, and antibiotic use. Model 5 includes both the virome and microbiome scores in a single model, also adjusting for the same covariates." 6. Apart from these, minor typos in the discussion sections and where Trimmomatic is cited in the methods section. Overall, a fun and exciting paper to review. Response: Thank you! We have fixed the typos. Attachments Attachment Submitted filename: PLOS_Response_to_Review_20250625.docx https://doi.org/10.1371/journal.pone.0331847.r002
22 Aug 2025 Decision Letter - Vinod Yata, Editor Dysglycemia and the airway microbiome in cystic fibrosis. PONE-D-25-12412R1 Dear Dr. BRENNER, 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, Vinod Kumar Yata, PhD Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: https://doi.org/10.1371/journal.pone.0331847.r003
Formally Accepted
Acceptance Letter - Vinod Yata, Editor PONE-D-25-12412R1 PLOS ONE Dear Dr. Brenner, 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 Dr. Vinod Kumar Yata Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0331847.r004

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