PONE-D-21-11287

Demonstration of Laser biospeckle method for speedy in vivo evaluation of plant-sound interactions with argula

PLOS ONE

Dear Dr. RAJAGOPALAN,

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Academic Editor

PLOS ONE

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Reviewers' comments:

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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

Reviewer #2: Partly

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

Reviewer #1: No

Reviewer #2: No

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

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Reviewer #1: Yes

Reviewer #2: Yes

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

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Reviewer #1: Yes

Reviewer #2: No

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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 paper is good, but before publication I suggest authors to give more details about the CMOS used, including focal length of the lens, and the estimation of the speckle size produced by the imaging system. Finally, I suggest authors to consider more statistical analysis on the data to make more reliable their results.

Reviewer #2: Speckle imaging has been proved to be an effective way for structure measurement. In biology, it has been used to distinguish different cellular movement patterns and indicate dynamic processes. In this manuscript, authors used speckle imaging to study the interaction between plant leaves and various acoustic frequencies. They show that speckle imaging can perform real-time monitoring.

After reading this manuscript, several questions arise. I hope they can be answered appropriately.

1. Specifications of the speckle imaging system are required, including accuracy, stability, sensitivity, etc. For instance, Figure 6 claims the difference between leaves at different stages. However, it is possible that the authors merely measured the shot noise limit of the CMOS camera because all curves tend to overlap at BA=0.8. Different rising edges can be caused by different scattering ratio which may not relate to structural difference.

2. Phantom tests are also required to prove the performance of the system. Simple specimens, such as gel or metal, can be used to verify system performance before the biological tests. For instance, even though absorption boards are used to reduce vibration, system instability can also lead to a smaller BA when the stimuli frequency is 10 kHz. If the system can show a constant BA regardless of the stimuli frequencies when conducting phantom tests, BA values under different stimuli will be more trustworthy.

3. The reason for selecting 0 Hz, 100 Hz, 1 kHz and 10 kHz needs to be stated. As shown in the Introduction, meaningful phenomena were discovered within hundreds of Hz. Instead of using high frequencies, a high frequency resolution and a range within hundreds of Hz are preferred.

4. What is the necessity of creating BA? According to Eq. (2), it simply equals to 1-r, and r is the correlation coefficient.

5. The full name of an aberration needs to be listed when the aberration occurs first. In Line 42 and Line 95, there are missing full spells.

6. Grammar errors. The Abstract itself contains many grammar errors. Several errors in the Abstract are listed in the following. Line 25, it should be “are destructive” instead of “destructive”. Line 30, it should be “intensities of speckles change over time” instead of “intensity of speckles change in time”. Line 34, there is a missing article. Line 42, “a leaf” instead of “leaf” should be used.

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Reviewer #1: No

Reviewer #2: No

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PONE-D-21-11287R1Demonstration of Laser biospeckle method for speedy in vivo evaluation of plant-sound interactions with argulaPLOS ONE

Dear Dr. RAJAGOPALAN,

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 11 2021 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,

Xuejian Wu, Ph.D.

Academic Editor

PLOS ONE

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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 (if provided):

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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: All comments have been addressed

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

**********

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 addressed all the comments adequately. The paper is good, well organized, the results are interesting and novel and I suggest acceptance of the paper

Reviewer #2: The authors reply my concerns properly with newly-added details and improved English writing. Compared with the former version, the latest one is more convincible. However, there are still three questions I would like the authors to expand on.

1. For the phantom test shown in Fig. 5, it looks like that the BA increases a little along with the testing time. Is it necessary to subtract the phantom results from the Arugula results to correct the final BA values? Moreover, what are the reasons for the increase in the phantom test?

2. For the LSM results shown in Fig. 10, the control group shows ratios larger than 1. As there were no sound stimuli, what are the reasons for the ratios larger than 1?

3. The gray regions in Fig. 10 are too narrow to see. The resolutions of all the figures need to be improved.

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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

[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.

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Demonstration of Laser biospeckle method for speedy in vivo evaluation of plant-sound interactions with argula

PONE-D-21-11287R2

Dear Dr. RAJAGOPALAN,

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.

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Kind regards,

Xuejian Wu, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

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