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PONE-D-21-33119microRNA profile of Hermetia illucens (Black Soldier Fly) and implications on mass rearingPLOS ONE

Dear Dr. Christoffels,

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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: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: Yes

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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

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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: No

Reviewer #2: Yes

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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: Dear Prof. Falabella

The manuscript with ID PONE-D-21-33119 by DeRaedt and coauthors studied the microRNA profile of Hermetia illucens and its implications on mass rearing. The manuscript is interesting, tested hypothesis is well-defined and methodologies are well-described. However, I found numerous revisions that should be carefully revised by authors before the manuscript being considered for publication in PLOS ONE.

Please find the attached word file with some track changes.

Line 57: Support with The feasibility of using yellow mealworms (Tenebrio molitor): Towards a sustainable aquafeed industry. Animals, 2021, 11(3), pp. 1–38, 811

Line 58: Support with

Aquaculture, 2020, 522, 735136

Fish and Shellfish Immunology, 2021, 111, pp. 111–118

All scientific names present in the references should be written italic (Please see the attached word file).

Reviewer #2: In this paper authors analyzed the profile of microRNAs in all the developmental stages of Hermetia illucens. The aim was to relate them to possible functions in order to understand, in future works, the specific gene regulation to improve mass rearing techniques. I have some doubts that must be addressed, especially regarding the comparison among different life stages, it is not clear, and it seems that in many points sentences seem to be contradictory.

KEYWORDS

I suggest to change the keywords that are already in the title

INTRODUCTION

“Black soldier fly larvae (BSFL) contain 42% protein, 35% crude fat (7), and 8.72% chitin in prepupae (8).” This sentence is too generic, protein, lipid and even chitin content change according to the feeding substrates. I suggest to report some examples of different substrates and a range of protein, lipid and chitin content (for example https://doi.org/10.1038/s41598-019-46603-z; https://doi.org/10.1007/s12649-019-00924-2; https://doi.org/10.3390/su131810198)

Authors wrote “they are an ideal candidate for large scale rearing for food sustainability and waste reduction.” Although few studies are performed at industrial scale, I suggest to cite them (for example https://doi.org/10.1038/s41598-020-76571-8; https://doi.org/10.3390/ani10071243)

RESULTS

In the sentence “A total of 44 miRNAs (23% of the total microRNAs) showed differential expression between at least two life stage with an LFC” write “stages”

DISCUSSION

What does it mean “Focusing on arthropod species only”? Which species? I don’t understand as the authors said that they performed comparisons with arthropod species.

“Only three conserved microRNAs were found to be specific to a single life stage: hil-miR-6-2-5p (Egg), hil-miR-3884-3p (Larva), and hil-miR-12418-3p (Pupa).” In the adult stages there are exclusively common microRNAs? Moreover, as the authors identify stage-specific microRNAs, I suggest to perform also a comparison among common microRNAs in all the analysed stages and not exclusively Egg to Larva, Larva to Pupa, Pupa to Female, and Pupa to Male.

“The unique microRNAs identified had a higher number of life stage specific microRNAs: (Egg) hil-miR-m, hil-miR-p, hil-miR-r, hil-miR-s, hil-miR-u, (Pupa) hil-miR-ac, (Female) hil-miR-h, (Male & Female) hil-miR-a, hil-miR-b, and hil-miR-y.” This sentence is not really clear: did the adult stage have or not specific microRNAs?

“However, these 10 unique microRNAs provide insight into the egg, pupa, and adult life stages of the BSF and can be used to compare the developmental regulatory differences of microRNAs to those of other insect species (26)” and what about the larval stage?

“In all three cases the females had expression whereas the males had none.” What does this sentence mean?

“miRNA target prediction” why did the authors choose to explore in detail only 3 microRNAs?

This sentence needs a reference “The microRNA has been seen to increase in expression over the third larval instar, with highest expression levels in the pupa stage, in D. melanogaster.”

“Once validated, the microRNA can be harnessed in breeding protocols to improve mass rearing of BSF.” please provide the readers concrete ideas of using these microRNA to improve BSF mass rearing.

METHODS

Please specify the composition of “standardized proprietary diet”.

Why did the authors choose to perform specifically these comparisons? “Five different paired life stages were compared: Egg to Larva, Larva to Pupa, Pupa to Female, and Pupa to Male.”

FIGURE

“Finally, 21 microRNA were identified with an LFC of ±10 (11% of total miRNA). These included 18

miRNAs conserved across arthropods and 3 miRNAs unique to BSF (Figure 1; Table 3).” In figure 1 only 18 microRNAs were represented. Please fix this, moreover, looking at the image, I supposed a comparison among all the samples. As I wrote before, it is not clear in the main text.

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Reviewer #1: No

Reviewer #2: No

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Attachments

Attachment

Submitted filename: PONE-D-21-33119.docx

microRNA profile of Hermetia illucens (Black Soldier Fly) and implications on mass rearing

PONE-D-21-33119R1

Dear Dr. Christoffels,

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.

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Kind regards,

Patrizia Falabella

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

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: N/A

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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

**********

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 properly addressed the comments in the revised manuscript. The manuscript merits acceptance in its present form

Reviewer #2: I thank the authors for improving the manuscript according to my suggestions.

Regarding the keyword, generally they must not be equal to the words of title and this helps to improve the search of the manuscript during a website research. It was just a piece of advice, but it is not fundamental and the manuscript can be accepted in this form.

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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