PONE-D-24-25732Shell colour luminance of Cuban painted snails, Polymita picta and Polymita muscarum (Gastropoda: Cepolidae)PLOS ONE

Dear Dr. Gordillo Pérez,

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Hudson Alves Pinto, Ph.D

Academic Editor

PLOS ONE

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Additional Editor Comments:

Dear Authors,

I invite the authors to revise the MS, taken in count especially the comments presented by Reviewer 1.

I look forward to receiving a new version of this interesting work.

Best regards,

Hudson

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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?

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Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

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

Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #1: No

Reviewer #2: Yes

Reviewer #3: No

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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: Yes

Reviewer #3: 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 manuscript PONE-D-24-25732 aims to corroborate a basic prediction of evolutionary ecological explanations for the presence of shell color polymorphism in Polymita species from Cuba and other regions. Specifically, it suggests that color is influenced and maintained by the varying sunny environments in different populations and microhabitats. The study uses approximately 180 specimens from one species and about 150 from another, sourced from a museum collection. The authors photographed these specimens and used software to analyze the colorimetry and luminance value based on a methodology by Schilk (1995). The manuscript, however, lacks substantial scientific value, because study something obvious which do not add anything new to the system (see below) and uses very little experimental effort (see below). The authors should revise their objectives and methodology to meet international standards before attempting publication. Here are some general comments that may help the authors in the future:

Literature Review. Many classical references on Polymita (in Spanish) are of low scientific quality or are supposed to proof what they really proof. It might be beneficial to prepare an English review that objectively summarizes previous work and realistically presents the data. Currently, there is no clear proof that Polymita polymorphism is maintained by natural selection, despite some indirect suggestions. A comprehensive review could set a solid foundation for future research.

Use of Luminance. The methodology for using luminance as presented has little empirical value, especially with museum specimens that have been cleaned and prepared for conservation. It's well-known that the animals inside the shells can influence their ability to resist heat stress. Moreover, luminance can be significantly affected by the environment in which it is studied. The authors claim their preliminary experiment to check this was not significant (lines 137-145), but this might be due to insufficient statistical power. The preliminary checking used N=18, whereas the main experiment used N=180/150. A more robust experiment checking with at least N=30 per environmental treatment might yield significant results.

Prediction and Experimentation. The prediction that lighter shells will show higher luminance or brightness is obvious and adds nothing new. The critical question is whether darker live snails heat up more than lighter ones under natural conditions, and if these differences in heating capability influence individual fitness in nature. These experiments have not yet been conducted.

Reviewer #2: I am pleased to have the opportunity to review your manuscript entitled “Shell colour luminance of Cuban painted snails, Polymita picta and Polymita muscarum (Gastropoda: Cepolidae)”. In this study, the authors successfully demonstrated the shell colour measurement and showed the difference between colour morphs. I agree this study established the methodological baseline for the future study of Polymita snails. This research undoubtedly offers a substantial contribution to PLoS One. Comments which I pointed out are only a few minor points.

L53: Is this the correct way to number the references? Is it OK if it isn't in order of appearance?

L90: “we”

L123: I understood that the authors took photos with the shell apex facing up in your study. I wonder if there are snails with banding patterns that only appear on the bottom. If yes, would these banding patterns affect your result? For example, when a snail estivates on a tree, I was concerned that bands on the bottom could be seen. Also, I recommend you add the example photo to measure the luminance as the supplemental picture so that readers can easily understand how the authors took them.

L185: I think you're right, but it is better to compare the variances by doing the statistical test, such as the F-test or something?

L187: In Figure 2, the number of individuals is not written. I can read the numbers from the main text, but I think it would be kind if you added them to match the other figures.

L199: What did it mean? High density and homogenous?

L271: From the figures, it looks like P. muscarum is brighter, but did you compare luminance values using a statistical test? And, is it correct to recognise that you would like to compare between two species, including all colours from black to yellow or albino?

L274: Why did you introduce P. picta as a subspecies here?

Reviewer #3: This is a good paper reporting some interesting findings about the luminance of various colour morphs of the Cuban painted snails, Polymita picta and P. muscarum. I really enjoyed reading about this beautiful and less studied land snail, compared to e.g., C. nemoralis. I think the authors did a good job in investigating the link between colour and thermoregulation. The analyses are sound, and appropriate statistical analyses were carried out and the findings were well presented. There is a good understanding of the literature, and the authors adequately discussed the results in relation to previous literature, as well as suggesting avenues for future research. I do however have some minor reservations: Specifically, the raw dataset of measurements for each snail in both species should be made available to the reader beyond the tables of mean values provided in the manuscript. Additionally, there are some issues in the writing that needs addressing to make the text clearer and more understandable. Once that is completed, I am happy to suggest that only minor revisions are necessary before this manuscript is suitable for publication.

See below for specific minor issues that should be addressed.

Line 31: Remove the full stop between “using digital tools.” And “We were able”.

Line 39-41: The sentence is unclear, consider revising to something along the lines of: “Luminance differences support the hypothesis that colour possesses an adaptive value for thermoregulation, not only for the background colour, but also for the colour of the band, suggesting a complex pigment composition.”

Line 61:62: Recognised used twice with both American and British spelling. Consider revising to “-recognized indicators for climate change – are known threats for land snails”.

Line 70: Should be “has predominantly focused on non-Neotropical gastropods”

Line 88-90: It is not clear what “earlier methodology” the authors are referring to here. Consider specifying and elaborate on this in the text.

Line 90: “wee” should be “we”.

Line 131: The term “till the borders” makes little sense. Is it supposed to be “to the borders”?

Line 133: Is Histogram a function within a software or a standalone software? Would be good to specify.

Line 141: Consider revising to “Three conditions for imaging were tested”.

Line 150: “in three morphological types” should be “into three morphological types”.

Line 176: Species name should be in italics, the same is true for all species names in sub-headers.

Line 194: Consider revising to “The highest luminance values”

Line 199: “The black morph had the highest luminance values around the statistical mean” This statement is confusing: Firstly, It would be nice to state what the statistical mean is. Secondly, from looking at figure 3 and reading the text this might be a misstatement? The text prior to this statement clearly states that the lowest values were observed in black shells, which is also apparent from Figure 3. Consider revising to make this statement clearer and correspond to figure 3.

Line 235-238: The sentence flows strangely and is hard to understand, consider revising (e.g., add a comma after ‘albedo’)

Line 258: “According to us” consider revising to “According to the results presented here”

Line 267: Should read “measurements of oxidative stress”

Line 278: I don’t understand what the statement “mating longer” means. Please clarify.

Line 289: Consider revising to “This study shows that using a more solid statistical approach” for clarity.

Line 292: The sentence “Our findings suggest evolutionary mechanisms play an important role.” Is somewhat non-sensical. Played an important role in how and in what? Yes, evolutionary mechanisms play an important role in the development of phenotypes, this sentence needs more context. Consider revising to be more relevant to the research presented. E.g., something along the lines of “Our results indicate that thermal selection is a key evolutionary mechanism driving the contemporary distinct distribution of both subspecies.

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Reviewer #1: Yes: Emilio Rolán-Alvarez

Reviewer #2: No

Reviewer #3: No

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Shell colour luminance of Cuban painted snails, Polymita picta and Polymita muscarum (Gastropoda: Cepolidae)

PONE-D-24-25732R1

Dear Dr. Gordillo Pérez,

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,

Hudson Alves Pinto, Ph.D

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

I thank the Authors for their efforts and elegance in revising this MS. The text improved a lot and is now suitable for publication. Congratulations on this nice work.

Reviewers' comments:

None