Karyological Study of Acanthocephalus lucii (Echinorhynchida): The Occurrence of B Chromosomes in Populations from PCB-Polluted Waters
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsDear authors, first my congratulations for this interesting MS entitled “Karyological study of Acanthocephalus lucii (Echinorhynchida): the occurrence of B chromosomes in populations from PCB-polluted waters”
The manuscript in general is very clear and well presented, however I would like you to address some questions regarding some minor things:
Line 126, in the Carnoy solution, did you use “normal” acetic acid or was it glacial? In my expertise, when we use the normal one, chromosomes slides are very weak regarding chromosome integrity. Please modify it if needed.
In line 148, could you please add the name of the kit used for NT?
In line 153 the name of the vector is pGEM®-T Easy, please modify it. Also, could you please state how long did you ligate? It can be done at RT in 1h but also at 4ºC degrees ON. In my expertise, sometimes depending on the insert, the differences can be very high between the two options.
I am so excited to see this kind of results regarding Bs, I hope you will find some interesting things in the future if you continue working with them (maybe some NGS study to unravel the content of Bs or if they came from different chromosomes, maybe cut the Bs and perform FISH…)
Author Response
Responses to Reviewer #1 (in italics, indicated by asterisks)
Line 126, in the Carnoy solution, did you use “normal” acetic acid or was it glacial? In my expertise, when we use the normal one, chromosomes slides are very weak regarding chromosome integrity. Please modify it if needed.
*** We used “normal” acetic acid (98%). ***
In line 148, could you please add the name of the kit used for NT?
*** We did not use a kit for labeling the probes by nick translation (NT), but followed the procedure described in Hejníčková et al. 2019 (33). We have added “as previously described” to the text to clarify this. In References, we originally listed another citation by the same author (Hejníčková et al. 2021), which is now corrected. ***
In line 153 the name of the vector is pGEM®-T Easy, please modify it. Also, could you please state how long did you ligate? It can be done at RT in 1h but also at 4ºC degrees ON. In my expertise, sometimes depending on the insert, the differences can be very high between the two options.
*** The name of the vector has been corrected. Ligation was performed at 4°C overnight. This information is now added in the text. ***
Reviewer 2 Report
Comments and Suggestions for Authors
In this study, the authors performed a cytogenetic analysis of Acanthocephalus lucii specimens from three sites with different levels of environmental pollution. The study falls into the scope of Diversity and thus could be published. Before this can be done, some changes should be made as specified below.
In my opinion, the authors should avoid saying that they performed “a detailed cytogenetics analysis”, such as in the line 12. A detailed study would include additional cytogenetics approaches. Suggestion, delete the word “detailed”. Please, check the manuscript.
Introduction
It would be interesting to include a brief explanation about B chromosomes since the journal Diversity does not primarily focus on cytogenetics, and many of its readers might not be familiar with what these chromosomes entail.
Line 40 – The meaning of PCBs should appear early at line 33.
In the end of the introduction the authors said that they have analyzed for the first time the course of oogenesis in this INTERESTING species. These results were not discussed in the discussion section.
Methods
- The parasites were collected from which fish species?
Results
-In Figure 1A, an individual with three 'B' chromosomes is depicted. One of these 'B' chromosomes appears to be metacentric, displaying clear morphology. However, distinguishing the morphology of the remaining 'B' chromosomes is challenging. Nevertheless, it is evident that two of the 'B' chromosomes are smaller in comparison to the metacentric 'B'. Please review and confirm.
Discussion
- Line 425, use italic for Acanthocephalus;
- Line 365-366, reference is need;
-I general, the paragraphs in this section are enormous, please, try to summarize.
Author Response
Responses to Reviewer #2 (in italics, indicated by asterisks)
In my opinion, the authors should avoid saying that they performed “a detailed cytogenetics analysis”, such as in the line 12. A detailed study would include additional cytogenetics approaches. Suggestion, delete the word “detailed”. Please, check the manuscript.
*** Deleted. ***
Introduction
It would be interesting to include a brief explanation about B chromosomes since the journal Diversity does not primarily focus on cytogenetics, and many of its readers might not be familiar with what these chromosomes entail.
*** A short paragraph about B chromosomes has been added to the Introduction. ***
Line 40 – The meaning of PCBs should appear early at line 33.
*** Corrected. ***
In the end of the introduction the authors said that they have analyzed for the first time the course of oogenesis in this INTERESTING species. These results were not discussed in the discussion section.
*** We apologize for not including this in the discussion section. The requested information has been added to the text. ***
Methods
- The parasites were collected from which fish species?
*** From perch (Perca fluviatilis). This information has been added to the text. ***
Results
-In Figure 1A, an individual with three 'B' chromosomes is depicted. One of these 'B' chromosomes appears to be metacentric, displaying clear morphology. However, distinguishing the morphology of the remaining 'B' chromosomes is challenging. Nevertheless, it is evident that two of the 'B' chromosomes are smaller in comparison to the metacentric 'B'. Please review and confirm.
*** Yes, you are right, two of the three B chromosomes are smaller and have an unclear morphology. However, when we examined more metaphases from each individual, we found that all the B chromosomes examined were metacentric, although it was sometimes very difficult to define their morphology. B chromosomes are highly variable and are characterized by a considerable ability to vary in shape and size. In acanthocephalans, B chromosomes were always observed as the smallest chromosomes in the karyotype (present work, Špakulová et al. 2002, Orosová et al. 2023). Variation in the number and morphology of B chromosomes is a common chromosomal polymorphism found in natural populations of many eukaryotic organisms (Bakkali et al., Heredity 83, 428–434, 1999). ***
Discussion
- Line 425, use italic for Acanthocephalus;
*** Corrected. ***
- Line 365-366, reference is need;
*** The reference, [12], has been added to the text. ***
Round 2
Reviewer 2 Report
Comments and Suggestions for AuthorsI am happy with the new version of the manuscript.
