Molecular Characterization of the Common Snook, Centropomus undecimalis (Bloch, 1792) in the Usumacinta Basin

Round 1

Reviewer 1 Report

The paper has improved and the only thing I ask is to include the meaning of the heterzygote deficiency in the MS. The reason given by the AA can be inserted in the MS paying attention to the different meaning of what they write. One of the hypotheses mentioned is to be discarded if the deficit is not present in all the populations analyzed. 

Author Response

We included the reviewer suggestion in line 615: “The heterozygote deficiency represents a deviation from the Hardy-Weinberg Equilibrium (HWE), when the observed heterozygosity (Ho) is less than the expected heterozygosity (He) (Rousset and Raymond, 1995). These deviations from the HWE proportions can be generated by the presence of null alleles and by an irregular system of inbreeding or by population structure (Rousset and Raymond, 1995; De Meeûs, 2017). In our study, the heterozygote deficiency observed could be explained by the presence of null alleles in some loci, but they were not shared across populations.”

Author Response File: Author Response.docx

Reviewer 2 Report

The ms is very interesting and brings interesting results about Common Snook in Mexico. All analyses were well done and the results are really not expected since this species has a wide distribution and is knew by their extensive moviments. I believe the results could be related to the number of fishes analyzed but it's not small at all... Thus, I guess the information obtained is useful and should be published but I also guess that it should be investigated again in the future using more specimens and other markers. 

Author Response

We thank the reviewer for the comments on our MS.

Author Response File: Author Response.docx

Reviewer 3 Report

This manuscript analyzed the population structure and genetic diversity of the Common Snook (Centropomus undecimalis) in the Usumacinta Basin. The results of this manuscript might be helpful for the analysis of population genetics and conservation of C. undecimalis. However, the manuscript should be improved in some parts. The detailed remarks are suggested below.

- Totally, 81 individuals were sampled from Rainforest (RZ), Floodplain (FZ) and River delta (RD). But low number of samples were used for analyzing genetic diversity using COI and CYTB gene. In COI analysis, eight haplotypes were found, in addition, all three zones had exclusive haplotypes. However, in CYTB analysis, four haplotypes were found and only FZ had an exclusive haplotype. This could be due to the low number of individuals were used for CYTB analysis. Therefore, more individuals should be analyzed for genetic diversity using CYTB gene, or authors should explain why only 34 out of 81 samples were used for CYTB sequencing.

- Authors used microsatellite markers previously designed. The informativeness of microsatellite markers for C. undecimalis examined in this study should analyze (e.g. PIC).

- AMOVA test was conducted with K=3 from DACP and K=3 from STRUCTURE. However, authors mentioned the most likely number of clusters was K=2 in analysis by STRUCTURE. Thus, AMOVA test should be conducted with K=2 from STRUCTURE.

- Authors analyzed the population structure using mitochondrial genes and nuclear microsatellite. However, the results from these two group markers were not consistent. Therefore, authors should compare and discuss these two different results.

- Small point: For kits and machines used in this study, authors did not mention city and country names. It is better if authors can add this information to the manuscript.

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Round 2

Reviewer 3 Report

The revised version was corrected well according to comments. 

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

Intro

Second paragraph:

Change “barcodes” to “DNA barcodes”

Can’t say genetic barcodes “monitor and explore biological diversity…like never before”, not with the advent of next-generation sequencing data.

Define mtCox1

Change “combine” to “combined”

Third paragraph:

“…main hydrographic basins”, do you mean “river basins”.

Fifth paragraph:

“…sampling in Mexican side”, revise, poor sentence structure

Materials & Methods

Section 2.1

Change “locality” to “location” throughout this paragraph

Please indicate how many samples from each sampling location were collected.

Section 2.2

Why were there only 34 samples that amplified at CytB? Were these 34 samples distributed across all the sampling sites? If the results of the CytB analysis will be reported on, the authors will need to address these questions.

Check TM in this paragraph, was this auto-corrected? If not authors need to define.

The authors need to indicate on which sequencing machine/platform the samples were run.

The authors need to report how sequences were assembled; de-novo, or was a reference sequence used?

Is Table 1 necessary given these msat markers are defined in another paper?

Section 2.3

First sentence not complete, please revise.

For sentence starting “Alignments were performed using the BIOEDIT…”, place a period after the reference and start a new sentence with “We checked…”.

Please clarify whether the ML tree and modeltest was performed separately for COI1 and CytB.

The authors need to include an FST test (or PhiST) for the mitochondrial data, as it was also done with the microsatellite data.

Section 2.4

The authors need to explain why STRUCTURE was run with a K ranging from 1 to 16, while DAPC was run with a K from 1 to 6.

Section 2.5

Please indicate how geographical distances were measured? In-river distance? Over land distance? What platform was used?

Results

The authors need to report how many individuals had each haplotype. It seems like Cox1 Hap1, 4-8 have only one individual with that haplotype, in which case, there are not many conclusions that can be drawn.

Section 3.1

Please explain what the rationale was for choosing Cun-10 over Cun-09?

The authors need to present the graph of log likelihood values outputted from STRUCTURE, not just the EVANNO graph presented in S1. In addition, why does Figure S1 only have a K to 5? The methods say a K to 16 was tested.

When the authors describe which “populations” are in which cluster, it’s a bit mis-leading to say Tzendales, Lacantun, Chacamax, San Pedro and Canitzan are all in one cluster since Lacantun, San Pedro, and Chacamax all have shared ancestry with the second cluster (K=2, Fig 3A). This is true even at K=3. The authors need to re-think these STRUCTURE findings and maybe look more into why Canitzan and Tzendales are in distinct clusters. I would not interpret these STRUCTURE results to imply there are two clusters and that one is exclusive to the rainforest/floodplain.

Figure 3, provide a color key for the plots. In addition, it is hard to tell what sampling locations are included in each of the 3 zones, please fix.

For the DAPC results, I’m interpreting this section to imply the authors performed multiple DAPC runs, however, there is only one result presented (Fig 3c). It is also confusing because in subsequent paragraphs, the authors say that both K=2 and K=3 was the result from the Evanno test, this needs to be clarified. In addition, this section has many instances of parentheses that are not closed.

Where are the results for the FST test? Please report these. In addition, how was FST calculated? Among samples sites, sample sites grouped based on zone?

Discussion

I am having a hard time being convinced that there are notable genetic differences between the upper and lower parts of this river system based on the genetic data. I would like to see the FST results, as I would suspect they are similar to the AMOVA in which no differentiation is observed.

If there are genetic differences between the upper and lower reaches of this river system, how do the authors explain this biologically when these fish are known to migrate and spawn in the same area?

The authors need to address how sampling date could also bias results. For instance, is there an elevated genetic diversity in the floodplain zone because sampling occurred in this region during a mass migration to or from spawning grounds (or if food is more plentiful here, hence more fish congregating), in which by chance you will sample more haplotypes?

The authors need to watch their word choices and broad sweeping conclusions used in the discussion. For example, the first paragraph in section 4.1: “…existence of geographic structure for the freshwater fauna in the basin…”. First, you can only comment about the genetic structure observed, there are no landscape/seascape measures being used here and second, all freshwater fauna may not fit this observation, surely not all have been tested.

The authors should incorporate the results from the mt and nucDNA and actually discuss what results were contradicting, etc, not keep them in separate sections.

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 2 Report

Martinez et al paper studied the variation and the genetic structure of Centropomus undecimalis through the use of nuclear and mitochondrial markers. The paper is well written and the objectives described are quite clear even if there are some aspects that should be clarified.

I have some observations that I report below:

1) insert the name of the species in the keywords

2) It seems that the authors did not use the same number of sequences of the two molecular markers to generate the distribution frequencies. Please clarify

2) I would like to see the combined haplotype network for the two mitochondrial markers.
3) Why wasn't a Baesian analysis done for phylogenetic inference?
4) Check Tab 2 there is an offset in the first line between n and the number of individuals

5) In the microsats paragraph change tab 2 with tab 3.

6) I suggest to the authors to insert a tab with the paired values of Fst and significance after Bonferroni corrections. Also I suggest to calculate the number of migrants.

7) Why did you not discuss the significance of the heterozygote deficiency and the haplotype network for mtCox1 (star-like shape)?

8) Finally I suggest to use the software COLONY 2.0.6.4. to detect possible groups in each of the populations analyzed.

Author Response

Please see the attachment

Author Response File: Author Response.docx