PONE-D-24-58988

Geography is a stronger predictor of diversification of monogenean parasites (Platyhelminthes) than host relatedness in characid fishes of Middle America

PLOS ONE

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Reviewers' comments:

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Comments to the Author

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Reviewer #1: Yes

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: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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5. Review Comments to the Author

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Reviewer #1: The manuscript numbered PONE-D-24-58988 and titled “Geography is a stronger predictor of diversification of monogenean parasites (Platyhelminthes) than host relatedness in characid fishes of Middle America” authored by Alda and colleagues. This study examines the relationship between characid fishes of the genus Astyanax and their monogenean parasites. The researchers constructed phylogeographic hypotheses for monogenean ectoparasites associated with four species of characid fishes from Panama, Nicaragua, and Mexico. They initially hypothesized that parasite diversity and community structure would correlate more strongly with host species than with geographic location. However, their findings concluded that geography or geographic location was a stronger predictor.

I was very pleased to read this paper, as it represents the first time this type of work has been done in the region. However, I have a number of major and minor concerns that the authors will need to address, and I hope these suggestions will help improve the manuscript.

Major concern:

In the Geographic analysis section of the methods, it is stated that PERMANOVA was used to test weather geographic location (Factor) or host identity (factor) better predicts parasite diversity and community structure (dependent variable). However, I believe that the way PERMANOVA test was set up, it is just telling if there are significant differences in parasite diversity (dependent variable) and community structure by geographic location (factor) or host identity (factor), so this needs to be fix.

Another important issue with the implementation of this PERMANOVAS is that no post-hoc test was applied to test for comparison between pairs of host or geographic locations. The result of the PERMANOVA was significant, but we don’t know if all comparisons are significant, remember that if a single comparison returns significant, we will have a significant p. value, but in reality, we don’t know how many comparisons are or are none significant.

The same comment applies to the second PERMANOVA in which authors try to elucidate if host species or geographic location predicted genetic similarity among individual parasites. I believe that here a simple mantel test would have given a better answer.

Minor concerns:

Line 101: Check journal citation style for (Choudhury et al. 2017). I believe it should be formatted as: “these biogeographical patterns [25]"

Line 103-4: "Lower Middle America (LMA)". This is more of a comment than a suggestion to change anything in the manuscript. My concern is that when one reads "Lower Middle America," it immediately evokes the southernmost part of Middle America, which suggests to me that the study area only includes Panama and Costa Rica. In many biological studies, the region encompassing Central America and southern Mexico is commonly referred to as Central America.

Line 121: The authors state that nine locations in Panama were sampled; however, the map in Figure 1a shows only seven locations.

Line 134: The reference to Matamoros et al. (2005) is not listed in the reference list. Instead, it is cited as: '28. Matamoros WA, McMahan CD, Chakrabarty P, Albert JS, Schaefer JF. Derivation of the freshwater fish fauna of Central America revisited: Myers’s hypothesis in the twenty-first century. Cladistics. 2015;31: 177–188.' Therefore, the correct citation should be: Matamoros et al. (2015).

Lines 134-7: The authors indicate in line 134 that this figure shows the results of a Maximum Likelihood analysis. However, in lines 136-137, they also mention providing Bayesian Posterior Probabilities, which suggests that this phylogeny is also the result of a Bayesian analysis. Please clarify this issue.

Table 1. The 'No. Parasite Species' column is not necessary, as the table is small enough for the reader to quickly discern the number of species per fish host. Additionally, the title 'Parasite Species' is causing some confusion as it is. I suggest centering this title for clarity.

Line 189: It is clear that the authors performed both Maximum Likelihood and Bayesian analyses; however, the way Figure 1 is presented suggests that only an ML analysis was conducted.

Table 2. Please explain the meaning of 'C', 'D', 'HS', 'L', and 'FD' in the table.

Line 247: “genetic distance matrix calculated in PAUP” authors need to specify which metric PAUP used in order to stimate distances¡

Line 274-5: Characithecium costaricensis (Price and Bussing 1967) Mendoza-Franco, Reina and Torchin 2009. I believe the authority for C. costaricensis is (Price and Bussing 1967), However, I do not understand what the role of Mendoza-Franco, Reina and Torchin 2009 citation is, please fix.

Line 296-7: This indicates that the ML and BI trees were not identical. However, earlier, you mentioned that the results were the same. Could you please clarify this discrepancy?

Lines 298-9: I suggest color coded the terminals in figure 2A the same way terminals were coded in figure 1B.

Line 368: The authors wrote: 'within Mexico'; however, I would recommend specifying 'southern Mexico.' Mexico is a large country, and the study's samples were collected from specific regions within the southern states. Using 'southern Mexico' would provide greater clarity and precision regarding the geographical scope of the study.

Line 406-409: Could the position of U. strombicirrus in these results be attributed to incomplete lineage sorting or any potential bias associated with relying solely on mitochondrial DNA data?"

Line 418-420: This section addresses the concerns I raised in my previous comment.

Other comments: The figure S1 is very helpful in understanding the results. I suggest inserting a smaller, color-coded map showing the areas of endemism. This addition would help readers geographically locate the tree terminals.

Reviewer #2: This is a high quality study using modern molecular tools to ask an interesting and well-defined question about host-parasite evolution, employing munificent and well-analyzed molecular data, with thorough scholarship and clearly-explained results. A model study illustrating the important conclusion that geographic location can be a stronger predictor of ecological association than the taxonomic identity of the host species.

Reviewer #3: The authors investigate and test factors driving the diversification of monogenean parasites in relation to their hosts, focusing on three species of the genus Astyanax and Eretmobrycon gonzalezoi. To achieve this, they analyzed mitochondrial genes—ATPase 6/8 from the fish and COI from several gill monogenean parasites—collected across multiple areas of endemism in Central America.

Using cophylogenetic analysis and modeling various scenarios involving the four host species and eleven geographic regions, they concluded that geography is a stronger predictor of parasite diversification than the genetic distance of the hosts. This finding challenges the traditional notion of parasite diversification through strict coevolution and expands our understanding of how parasites diversify in association with their hosts at a lower taxonomic level.

I find the article robust, and the conclusions are well-supported by the data. My primary comment concerns the use of the term “characid” throughout the text, which refers to an outdated classification of the family Characidae. I recommend revising this terminology to ensure clarity and avoid confusion. Additional minor edits are detailed below.

LINE 2: Characidae has been split recently and Astyanax has been reassigned to the family Acestrorhamphidae based on UCE. (Eschmeyer's Catalog of Fishes), but see Melo et. al. (2024) (https://doi.org/10.1093/zoolinnean/zlae101). May want to consider and use a different term. A potential alternative could be “Characin”, but I leave to the authors to decide what is more appropriate.

LINE 30: Update all instances of the term “Characids” as appropriate.

LINE 84-85: Update sentences to reflect latest classification of the genus Astyanax within Acestrorhamphidae: Acestrorhamphinae . Here could provide context highlighting old and recent classification.

LINE 101: Change in text citation format for consistency for “(Choudhury et al. 2017).”

LINE 120: Add parenthesis to “Eigenmann 1907” for consistency with other species authors.

LINE 121: Update “Characid”. Eretmobrycon gonzalezoi is now in Stevardiidae: Landoninae. See Melo et al. (2024).

LINE 134: Maybe you mean “Matamoros et al. 2015” instead of “Matamoros et al. 2005”. Add number of citation.

LINE 173: Typo “originally JB3 in 12/20/2024 6:16:00 AM”

FIG 1A: “A. ruberrimus (2)” appear in the figure but not mentioned in the text.

FIG 1A: For clarity could label the locations within the areas of endemism.

TABLE 2: Add the meaning of the event letters in the caption or text. E.g., Cospeciation (C), Duplication (D), etc…

LINE 242: Often Latin words such as “sensu” are italicized, however, may depend on the journal. Worth asking.

LINE 252: Add R version.

LINE 261: remove “characid”

LINE 303: Update family “characid”

LINE 311: Update family “characid”

LINE 364: Define the term “characids"

LINE 493: Typo. I think it should be “Fig 2B”

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Reviewer #1: No

Reviewer #2: No

Reviewer #3: No

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Geography is a stronger predictor of diversification of monogenean parasites (Platyhelminthes) than host relatedness in characin fishes of Middle America

PONE-D-24-58988R1

Dear Dr. Alda,

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,

Windsor E. Aguirre, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: