PONE-D-23-19903Convergent and environmentally associated plastic chromatic polymorphism in Bryconops Kner, 1858 (Ostariophysi: Characiformes: Iguanodectidae)PLOS ONE

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Reviewer #1: Review of Gonçalves et al. “Convergent and environmentally associated plastic chromatic polymorphism in Bryconops”

The article aims to test whether color is plastic in Bryconops species or if phenotypic variation is related to cladogenetic events. The work is well written and can be a good contribution. However, it presents a problem that needs to be corrected (see below). Therefore, my overall recommendation is either a major revision and resubmit.

The biggest problem has to do with the identity of the species and the consequences for the article's assertions.

The main hypothesis of the work is linked to the supposed wide distribution of two species (B. caudomaculatus and B. melanurus), which could present polymorphism in the coloration pattern. My comments here do not cast doubt on the hypothesis presented by the authors, but rather on how they use the taxonomy of the group under study.

1) The COI seems like a good tool to demonstrate hidden diversity. However, does COI present sufficient levels of resolution for the recovery of relationships in any of the lineages?

2) Although many authors have mentioned these two taxa as widely distributed, specialists in the group point to the opposite. Chernoff and Machado-Allison (2005), for example, mentioned B. caudomaculatus as restricted to the Essequibo River and other similar species occurring in the Amazon basin. Chernoff et al. (1994) mentions that B. melanurus occurs only in coastal drainages of the Guianas. Silva-Oliveira (2020) in a broad taxonomic review of the genus, confirms the presence of the true B. melanurus only in coastal drainages of the Guianas and recognizes two species complexes (B. caudomaculatus and B. melanurus) with still underestimated diversity. Probably the authors are working with undescribed species and still without a clear taxonomic delimitation. Although Silva-Oliveira’s work has not yet been published, it is important that Gonçalves et al. seek more up-to-date information on the taxonomy of the species they are using as a model.

3) Bryconops rheoruber from the supplementary material looks like B. magoi or B. collettei. According to Silva-Oliveira et al. (2019), Bryconops rheoruber presents dorsal fin originates anterior to a vertical through the first pelvic fin ray.

4) Images of some of the species used in the study are missing. As coloration is the focus of the work, all species should be presented.

Other comments are in MS.

Reviewer #2: Gonçalves et al present an interesting study examining whether color patterns in the iguanodectid fish genus Brycanops exhibits convergence or has a strong phylogenetic signal and thus can be used as a character for taxonomic delimitation. The authors score colors from specimens collected from different water types and use sequences of COI to create a phylogeny. Species partitioning methods, tests of phylogenetic signal, and a test of convergence are then employed to test the hypothesis. Although a more robust phylogeny using more genes or genomic methods and the inclusion of more specimens would have been good, the methods seem robust to make the point the authors advocate. The point that phenotypic similarity of ecologically important traits may often be the result of convergence and may complicate phylogenetic inference or species delimitation using these traits is also an important one to make.

I do have some points that I think should be addressed, which are presented below.

-I was disappointed that there was not a figure in the paper itself with the characters scored that showed the traits and the color variation. Two figures are presented in the supplemental files but given how central they are to understanding the results, I strongly encourage the authors to combine these into one figure and include them in the manuscript itself. Putting them in the supplemental files just makes things more difficult for readers. In addition, PLoS one will have a general readership and many may have difficulty understanding the traits without seeing them.

-The abstract and some parts of the paper are written in a way that makes it seem like convergence and plasticity are synonyms, which is very misleading. For example, from the abstract (lines 26-28): “…To test

whether color is plastic in those species or if phenotypic variation is related to cladogenetic events…“. What is being tested is whether the trait is related to the phylogenetic relationships of the species or not. However, there is no way of knowing if it is phenotypically plastic, as the authors indicate themselves in the Discussion. Phenotypic plasticity refers to non-genetic changes in the phenotype, that is, changes in gene expression patterns that result in different phenotypes in fish that are identical for the underlying alleles. Convergence generally refers to convergent evolution, the independent evolution of similar phenotypic traits, often related to adaptation to similar environmental characteristics. They are different things. The text should be reviewed for how this is phrased, especially the abstract and introduction (although there may be problems in other sections). The authors should indicate that they are testing for phylogenetic independents and conducting a test of phenotypic convergence, with the plasticity text saved for the discussion, where they indicate possible causes and that it is not possible to distinguish between phenotypic plasticity and evolutionary convergence with these data.

Lines 55-57: Oliveira et al. (2011) is listed as the latest phylogenetic hypothesis of the Characiformes, but this is a phylogeny of the Characidae and there are more recent phylogenies of Characids (e.g., Mirande 2018). This should be reworded.

-Lines 102-104: I assumed that saying that traits are phylogenetically independent is the same as saying that they are evolving according to Brownian motion. If they are phylogenetically dependent, they would be more similar than null expectations under Brownian motion, right? What am I missing? Can this be explained more carefully since the topic can be confusing.

-Line 118: Indicate how many specimens from how many sites are included. The info is in the tables but would be good to make it explicit up front and not make the reader have to count.

-Lines 144-147: How was color standardized, was a color scale included in the photographs? If not, lighting under which the pictures were taken may interfere with the ability of the authors to score the colors accurately. This should be addressed in the text.

-Table 1: Please indicate how many individuals were collected from each water type. This information can easily be included given the structure of the table and will be helpful for the reader.

-Line 200 should be ABGD, not ABDG. Please check other parts of the manuscript for this typo.

-Lines 200-202, the ABGD method has several parameters one sets. What were the other values used and why? If the other methods have options for other parameters, these should be indicated as well. Otherwise, make sure to note that the defaults were used.

-Line 211 also talks about scoring colors, so addressing how color was standardized could be done here.

-Line 262: Has MOTU been defined already? It should be defined before it is used.

-Line 295: add a comma after “Brownian Motion models”.

-Lines 355-356: What if the water color leads to similar sexual preferences because similarities of how the fish colors contrast with particular water colors? Maybe some colors patterns stand out more in some water types. That could lead to a correlation between water types and sexual preferences for particular fin color patterns.

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Reviewer #1: No

Reviewer #2: No

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Submitted filename: PONE-D-23-19903_reviewer.pdf

PONE-D-23-19903R1Convergent and environmentally associated plastic chromatic polymorphism in Bryconops Kner, 1858 (Ostariophysi: Characiformes: Iguanodectidae)PLOS ONE

Dear Dr. Burlamaqui,

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.

 Note the comments below. One thing that needs special attention is why some statistics and P values were removed from Table 2. The results seem to have changed between the original version and the revision. This needs to be explained very carefully since it gets at the analysis that relates to the most important points made.  Please submit your revised manuscript by Dec 30 2023 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.

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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.

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We look forward to receiving your revised manuscript.

Kind regards,

Windsor E. Aguirre, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments:

The page and line numbers refer to the clean version of the manuscript in revision 1.

-For the title: I do not think that you have shown whether the pattern is the result of plasticity (non-genetic phenotypic change), so it seems misleading to include plasticity in the title. I think you can just remove the plastic from the title and leave it as: “Convergent and environmentally associated chromatic polymorphism in Bryconops Kner, 1858 (Ostariophysi: Characiformes: Iguanodectidae)”. The “environmentally associated” leaves the possibility of plasticity without actually using the word without showing that it is plasticity.

-Line 27: Add a comma “,” after cladogenetic events.

-Lines 34-37: For the last line of the abstract, it could be plasticity or convergent evolution. Either could cause problems for grouping or identifying species. I think you need to add the possibility of convergent evolution to the sentence and suggest modifying it to:

“This means that simple color characters cannot necessarily be relied upon for taxonomic revisions of the genus as local phenotypic variants may represent environmentally determined plasticity or convergent evolution. Further studies are required to determine the validity of these characters.”

-Lines 53-57: The modification to include “actually” does not work very well the way the word is used here. Although it is a long sentence, I suggest changing to:

“The genus includes Cis-andean small to medium sized tetras widely distributed in the Orinoco, Amazonas, Tocantins-Araguaia, Paraná-Paraguai, São Francisco rivers and several coastal basins draining from the Brazilian and Guiana shields [10-12]. Bryconops is currently included in the family Iguanodectidae along with Iguanodectes Cope, 1872 and Piabucus Oken, 1817 [13-14].”

-Line 80: Replace “demand” with “need”.

-Lines 112-113: Change to: “to test if such variation is independent of the evolutionary history and is correlated with water color from the specimen’s localities.”

-Line 148: Divide into two sentences. So: “photos, but was not available for all specimens. To avoid possible bias in using digital”

-Table 1 as currently formatted is confusing because the records are not clearly separated between species. Please make the separation between species easier to see.

-Line 203: Modify to: “(k80) model with TS/TV = 2.0. Default settings were used for the remaining parameters. bPTP was run via”.

-Line 211: Add a comma “,” after “For new samples,”

-Table 2: I assume that the lower and upper bounds and P are for the Wheatsheaf test. Please indicate so in the caption o there is no doubt. Also, not that for the Dorsal fin melanic patch, the numbers are on two different lines. The same happens for the two last traits. I assume this is not intentional and should be corrected.

-Table 2: Why were the P values removed from Abouheif’s C mean and Pagel’s test? Why were Moran’s I and Blomberg K removed? In the previous versions, these tests indicated strong significance. Now there is no indication of this. The actual numerical values have also changed. In the new version, only the color of the dorsal fin has a P value of less than 0.05. This needs to be explained carefully to ensure that the results still hold true since this involves the key component of the analysis and interpretation of convergence in these color traits.

-I would have liked to see figures S1 and S2 merged into one figure and included in the main manuscript. I think the pictures are beautiful and help the non-specialist understand the color characters better. However, I will not insist if the authors prefer not to make this modification for some reason. Just make sure to update the figure numbers if you do.

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Convergent and environmentally associated chromatic polymorphism in Bryconops Kner, 1858 (Ostariophysi: Characiformes: Iguanodectidae)

PONE-D-23-19903R2

Dear Dr. Burlamaqui,

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,

Windsor E. Aguirre, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Congratulations on the acceptance of the article. Please work with the editorial office to correct two minor details in the abstract:

Line 33: Should be "...with a similar trend"...". The "a" is missing in the abstract text.

Line 36: Remove "rather than local selection". This is not needed to understand the meaning of the text and local selection can lead to convergent evolution, so these are not really alternatives.

Reviewers' comments: