PONE-D-24-52670Effects of dietary lysine, methionine, and tryptophan on regulating GH-IGF system and modulation of inflammatory and immune response in Pangasius bocourtiPLOS 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

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

Reviewer #1: 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

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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

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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 idea of the study is interesting and the manuscript is well written. However, there is some issues that needs to be addressed. Additional comments can be found below.

Line 3-4: the name of the corresponding author is different in between line 3 and 4.

Line 25, 48: “Basa” lowercase

Line 30: “n = 15/replicate” It’s not clear what this number is. Number of fish per treatment?

Line 60: add “with” in “substituting with plant-based nutrients”

Line 63-65: “However, when fish meal is partially replaced with plant-based proteins, there is often a deficiency in total sulfur amino acids (TSAA), particularly methionine.”

Is this specifically for soybean meal or plant proteins in general?

Line 61-62: “A suitable alternative protein source for fish aquaculture is soybean meal, given its high protein content [9, 10].”

Although soybean meal was mentioned as an example of plant proteins, the rest of the introduction talks about plant proteins in general- you may want to be more specific.

Line 112: The description of the different amino acid supplementation is described in the methods reflects that they were completely missing/ not supplemented in diets A0, A1,…

But in reality, they were added to all diets and additional supplementation occurred in the diets.

So, the method section needs to be improved in a way to explain this.

Line 119: provide manufacturer information (city, state..etc)

Line 127: “a nearby hatchery” does this hatchery has a name? location?

Line 131-132: “To serve as the negative control (-ve A0) in the bacterial challenge study”

This part needs to be clarified further, there needs to be a further explanation that there were -ve and +ve A0.

Line 124, 158: “2.2 Growth experiment” and 2.4 Sample collection

I didn’t see a description of the growth indices at the end of the trial throughout section 2.2

Instead, I see this part “From each replicate of every treatment, five fish were randomly selected. Various parameters including total body weight, body length, feed conversion ratio, specific growth rate,”

For accurate representable measurement, the whole fish biomass should be measured and indices should be calculated based on the whole biomass, not a random sample due to differences between individuals that may lead to biased measurement.

Line 152: “((“ why there is 2 parenthesis?

Line 171: Lysine is missing the “e” in several rows in the table

Line 193, 195, 196-197: “DNase kit, SuperScript III kit, SYBR Green PCR Master Mix.” Manufacturer info?

Line 204: Remove “,”

Line 214: replace “measured” with “counted”

Line 216: “Haematocrit (Hct: %) was measured twice”

Explain further what do you mean by twice, two times from the same blood sample or 2 separate samples.

Line 243: “3.1 Growth” Growth what? Performance, indices,…etc??

Line 253-254: correct it to “significantly (P<0.05) increased among treatments.

Line 254: add “s” to “treatment”

Line 254: “increase” or “decrease”? If so, please check the rest.

Line 256, 257: add “the” before “end”

Line 257: add “the” before “growth”

Line 258: what is “(1 b).”? (Figure 1b)?

Line 262: remove “were”

Line 274-275: at the end of the bacterial challenge

Line 300: there should be a footnote explaining all the abbreviations in the table

Is TBW(g) the total body weight for the sample or the whole biomass? And why wasn’t the whole biomass provided?

Line 342, 412: “Basa” lower case

Line 363: replace “following” with “at the end of”

Line 365: replace “addition” with “additional supplementation”

Line 366-367: “lacking these amino acids.” I don’t think these AA were lacking- I believe they were supplemented in suboptimal levels, correct?

Line 400: add “fed diets” to supplemented with lysine, methionine, and tryptophan

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Reviewer #1: No

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<p>Effects of dietary lysine, methionine, and tryptophan on regulating GH-IGF system and modulation of inflammatory and immune response in Pangasius bocourti

PONE-D-24-52670R1

Dear Dr. Fatima,

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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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,

Ishtiyaq Ahmad, Ph.D

Academic Editor

PLOS ONE

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

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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

**********

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

Reviewer #1: 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

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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

**********

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 made all the necessary modifications. Please note that all the treatments' abbreviations should be included in the footnote of all tables (Line 174, 182, 334).

The manuscript is now in good shape for publication.

No further revisions are required.

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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

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