PONE-D-19-19788

Pru p 9, a new allergen eliciting respiratory symptoms in subjects sensitized to Peach tree pollen.

PLOS ONE

Dear Dr Maria Luisa Somoza,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

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Davor Plavec

Academic Editor

PLOS ONE

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Additional Editor Comments (if provided):

The manuscript covers an interesting topic but prior to sending it for peer review it needs a major revision in the methods section. All the methods used and the recruitment proces for patients should be described in significantly more detail. Also there is a significant mistake in Table 2 regarding the significance of difference between 2 tested groups regarding clinical presentation (conjuctivitis, rhinitis, asthma).

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Reviewers' comments:

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PONE-D-19-19788R1

Pru p 9, a new allergen eliciting respiratory symptoms in subjects sensitized to peach tree pollen.

PLOS ONE

Dear Dr Maria Luisa Somosa,

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.

We would appreciate receiving your revised manuscript by Jan 27 2020 11:59PM. When you are 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.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter.

To enhance the reproducibility of your results, we recommend that if applicable you deposit your laboratory protocols in protocols.io, where a protocol can be assigned its own identifier (DOI) such that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols

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). This letter should be uploaded as separate file and labeled 'Response to Reviewers'.
• A marked-up copy of your manuscript that highlights changes made to the original version. This file should be uploaded as separate file and labeled 'Revised Manuscript with Track Changes'.
• An unmarked version of your revised paper without tracked changes. This file should be uploaded as separate file and labeled 'Manuscript'.

Please note while forming your response, if your article is accepted, you may have the opportunity to make the peer review history publicly available. The record will include editor decision letters (with reviews) and your responses to reviewer comments. If eligible, we will contact you to opt in or out.

We look forward to receiving your revised manuscript.

Kind regards,

Davor Plavec

Academic Editor

PLOS ONE

Additional Editor Comments (if provided):

Dear Authors,

please revise your manuscript according to the reviewers' comments.

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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: (No Response)

Reviewer #2: (No Response)

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: Yes

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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

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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

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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: 1. Summary and overall impression

The manuscript represents a comprehensive analysis of the etiology of the allergic sensitization in a population directly or indirectly exposed to peach tree allergens in the regions where this pollen is substantially cultivated. Importantly, it brings identification of allergen(s) responsible for pollen sensitization and respiratory allergic symptoms. Although the study has been performed using the renewed methods and does not utilize a new technology, it adds a great value to the field of allergy sensitization and atopy.

The Discussion contains a concise summary and relevance of the results. The results are extensively elaborated in the context of similar publications and explain how the obtained results fit into the other research groups' findings. The last paragraph should be extended regarding the impact on the future work, i.e. steps and ideas for further research should be proposed.

The manuscript is well structured and the scope of the project is clearly presented. It enables a reader a complete and easy progression through the work. A coherent presentation of the results is consequently easy to grasp. The accompanying figures are simple and illustrative.

2. Evidence and examples:

Objective

The objective is nicely presented in a given theoretical background and argumented well enough to demonstrate its relevance and purpose. Given the size of the peach tree cultivated area and the prevalence of peach tree sensitization, the motivation is more than justified and the potential future application of the research is awaited.

Originality

The work uses established models of allergy study design and although it is not innovative, it builds up new knowledge to the topic.

Literature review

The literature review is nicely presented. The literature overview regarding the methodology of the study could be supported by a few more similar publications.

Proposed research

The proposed research is reasoned and well structured.

Methodology

A major drawback in the experimental design is the lack of the control population group. The drawbacks in the methodology and similar methodology-related issues should be transparently declared.

Minor revisions:

Paragraph 2.3: Peach Tree Pollen protein extract and Pru p 9 purification from peach pollen

• In the methodology should be clearly stated that the two chromatographic steps were used to obtain purified Pru p 9. First step - ion exchange chromatography using Waters AccellTM Plus QMA Sep-PakR cartridge should also include the size of the pores of the used cartridge. The second step - reversed-phase chromatography is described in detail and clearly.

Paragraph: Analytical procedures: SDS-PAGE analysis

• Tryptic digestion of bands excised from gel should include information regarding digestion mixture, concentration, period of incubation, temperature etc. or at least refer to a previous reference or established protocol. Although, in-gel trypsin digestion is a standard method it is relevant to state specific methodological details related to this work and type of sample.

• For MS analysis besides the type of used spectrometer for clarity the authors should add additional details related to the type of targeted plates, sample/matrix preparation (volume); type of laser with which is equipped mass spectrometer (wavelength, frequency). Need to add details regarding spectra recording: over which m/z range (linear mode?), at which accelerating voltage, average number of individual laser shots.

Optional additional figure: Mass determination spectra (figure of representative MALDI-ToF MS spectra to be added to the table with aminoacids residues verified by sequence analysis).

Reviewer #2: 1. More specific results and conclusions should be provided in the abstract

2. The criteria for patient selection and inclusion in the study need to be clarified, and indicate the number of respondents selected by which criteria

3. Provide a reference for the SPT, as well as a procedure for PT Pollen extracts

4. Clarify why respondents were done NPT

5. How do you explain that exposure to a particular allergenic component of PT is manifested by symptoms due to sensitization to that component and not because of cross-reactivity, because you did not provoke with the cross-reactive allergens

6. Indicate the most important weaknesses of your manuscript.

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Reviewer #1: No

Reviewer #2: Yes: Mirjana Turkalj

[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 to be viewed.]

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 us at figures@plos.org. Please note that Supporting Information files do not need this step.

Pru p 9, a new allergen eliciting respiratory symptoms in subjects sensitized to peach tree pollen.

PONE-D-19-19788R2

Dear Dr. Somoza,

We are pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it complies with all outstanding technical requirements.

Within one week, you will receive an e-mail containing information on the amendments required prior to publication. When all required modifications have been addressed, you will receive a formal acceptance letter and your manuscript will proceed to our production department and be scheduled for publication.

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If your institution or institutions have a press office, please notify them about your upcoming paper to enable them to help maximize its impact. If they will be preparing press materials for this manuscript, you must inform our press team as soon as possible and 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.

With kind regards,

Davor Plavec

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

The manuscript is acceptable for publication in its current form.

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

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2. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Yes

**********

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

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: Yes

**********

6. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: The authors have implemented the suggestions, especially regarding the methodology, and, thus, substantially improved their study. The study design should be kept in mind when planning future experiments.

Reviewer #2: The authors have adequately addressed my comments raised in a previous round of review and have adopted this manuscript acceptable for publication. All data underlying the findings described in the manuscript in a clear and correct way, and supports the conclusions.

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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: No

Reviewer #2: No