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Additional Editor Comments (if provided):

Dear Authors,

The manuscript was evaluated by two expert reviewers, both of whom recognized the relevance of the research question, the robustness of the methodological approaches employed, and the overall quality of the results presented. Both reviewers considered the study to be an important contribution to the understanding of cryptic diversity and diversification processes within Astyanax, one of the most taxonomically challenging groups among Neotropical fishes.

After carefully considering the reviewers’ comments and conducting my own evaluation of the manuscript, I believe that the study has sufficient scientific merit for publication. However, before a final decision can be made, I ask that the authors address the few comments and suggestions provided by the reviewers.

Therefore, the editorial decision for this manuscript is Minor Revision. I kindly request that the authors submit a revised version of the manuscript together with a detailed response letter explaining how each reviewer comment has been addressed.

I congratulate the authors on their work and look forward to receiving the revised version of the manuscript.

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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?

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

Reviewer #1: Yes

Reviewer #2: Yes

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

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: The manuscript “Cryptic diversity in Astyanax (Characiformes: Acestrorhamphidae) from the Magdalena basin, Colombia: Insights from molecular and morphometric evidence” from Márquez et al. designed and evaluated a set of microsatellite loci to assess genetic diversity and population structure of Astyanax sp. in the middle and lower sections of the Cauca River. Additionally, COI and rag2 sequences were compared within a broad phylogenetic framework, to determine whether groups potentially identified through microsatellites correspond to divergent lineages within the genus, and additionally geometric morphometric analyses were conducted to investigate potential drivers of diversification in Astyanax. The MS by Márquez et al. presents the conditions and quality of literature review and adjusted the scope of this journal. Science is good and the MS is competently presented. The theoretical grounding as well as the proposed methodology are relevant and appropriate. I have made a few corrections throughout the text errors (pointed out in the attached document) that do not depreciate anywhere the results and discussion. In my opinion, the paper is worthily and should be accepted after adjustments.

In relation to the mitochondrial (COI) similarity between A. microlepis and Asp1, I suggest to the authors include in discussion this point of view/possibility. This would bring more robustness to their results.

Barcoding analyses for species identification have been widely used with effective results in several fish species; however, it does not always bring concise results, especially in morphologically cryptic species and complex groups like Astyanax and Psalidodon. Taxonomic uncertainties may result from a combination of aspects such as rapid speciation, species with a wide geographic distribution, lack of descriptions of local morphotypes, and inadequate phylogenetic analyses. Low genetic diversity using DNA barcoding (COI sequences) in Astyanax and Psalidodon (formerly referred to as Astyanax) may be the result of the recent evolutionary process of radiation. (see Ornelas-García et al. 2008 - https://doi.org/10.1186/1471-2148-8-340; Pereira et al. 2011 - https://doi.org/10.3109/19401736.2011.588213; Gavazzoni et al. 2023 - https://doi.org/10.1093/biolinnean/blac139; Gavazzoni et al. 2024 - https://doi.org/10.1089/zeb.2024.0; Silva et al 2024 - https://doi.org/10.1590/1982-0224-2024-0069)

Reviewer #2: That's an very interesting contribution, dealing with the genus Astyanax in the río Magdalena basin in Colombia. The genus Astyanax is one of the most taxonomically-challenging genera among Neotropical freshwater fishes, with a high diversity but relatively little morphological variation across Central and South America. The present MS showcases a particularly complex example, showing that phenotypic divergence in the genus may not be coupled with genetic divergence. The MS is overall very good and I have just very few suggestions to make, which follows below:

1) P. 11, l. 50: The genus does not occurs as far south as Patagonia. The southernmost record for the genus, in its present circunscripion, is the occurrence of Astyanax abramis and A. lacustris in the estuary of the Río de La Plata in Argentina. Some species formerly assigned to the genus occurs in the Pampasic area to the south, but these species were transferred to the genus Psalidodon (e.g., Psalidodon pampa).

2) P. 28, second paragraph (line 8): Change "configuration" for "shape"

3) P. 44, fig. 2: the quality of the figure is not good, it needs to be improved.

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what does this mean?). If published, this will include your full peer review and any attached files.

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Reviewer #1: No

Reviewer #2: No

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Attachments

Attachment

Submitted filename: PONE-D-26-14913_reviewer.pdf

Cryptic diversity in Astyanax (Characiformes: Acestrorhamphidae) from the Magdalena basin, Colombia: Insights from molecular and morphometric evidence

PONE-D-26-14913R1

Dear Dr. Márquez,

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,

Caio Santos Nogueira, PhD

Academic Editor

PLOS One

Additional Editor Comments (optional):

Dear Authors,

One of the original reviewers has evaluated the revised version of the manuscript and indicated that the manuscript is now suitable for publication. Therefore, I would like to congratulate you on the work you have done and on the improvements made to the manuscript.

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

Reviewer #1: All comments have been addressed

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

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

Reviewer #1: Yes

**********

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

Reviewer #1: Yes

**********

Reviewer #1: All suggestions made by the reviewers have been incorporated into the text. The manuscript is now more fluid and concise. Consequently, the manuscript is ready for publication.

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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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