Effect of Cothurnia variabilis and Epistylis gammari (Ciliophora: Peritrichia) on Metabolic Rate of the Crayfish Cambarellus (Cambarellus) montezumae
Round 1
Reviewer 1 Report
Dear authors,
I read the manuscript with interest. Congratulations, this is a fine addition to our knowledge about crayfish symbionts and their relationship with the hosts. Although I have some comments and suggestions (highlighted directly in the PDF file), these are easy to follow. After this revision, I recommend this paper to be accepted for publication in the Diversity journal.
Sincerely
Comments for author File: 
Comments.pdf
Author Response
Dear Reviewer, thank you very much for your comments, we address the changes according to your proposals. We have already read all the articles you kindly suggest, and we love them, and they will serve us for future publications. Here are the answers.
- I guess that this is true for adult crayfish, am I right? If yes, add this note here.
Significant differences concerning the effect of oxygen consumption were found with more than 50 ciliated zooids attached to the crayfish gills, for adults or juveniles, so we did some redaction changes.
- Add here a note that the process of respiration in decapod crustaceans takes place by drawing the water stream into the branchial chamber as it is circulated over gill surfaces.
Done. We read the articles about the function and morphology of the gills of crayfish and added a note about how respiration is carried out in the introductory section to give an overview of respiration in decapods.
- I suggest to cite these references here: Ďuriš…..
Done. We read the article about the epibiosis of mussels attached to crayfish bodies, and it is interesting to know that this relationship is frequent in aquatic environments.
- Add a coma, 1,531
Done.
- Here I suggest to add a note on possible overlap to biological invasions due to transport of symbiont s via pet trade (Cambarellus crayfish are popular in pet trade).
Done. It is essential to know the distribution of the species since if they are invasive, they can considerably affect the environment.
Reviewer 2 Report
This is an interesting study that reports physiological effects of two epibiont ciliates on a Cambarellus crayfish. The study would contribute to the literature of interactions between decapod crustaceans and their epibionts, which have been rarely reported from freshwater ecosystems. The manuscript is generally well written. Nevertheless, the study has room for improvement, as the statistical analysis is too simple at present. Furthermore, some issues need clarifications to better understand and interpret the results of the study.
Main issues
- What were the size (e.g. carapace length) and sex of the crayfish used in the experiment? If the authors recorded these information, please describe in the Method section.
- Provided that the authors have information about crayfish size, what is the consequence of ciliate infestations on the relationship between crayfish size and weight? This is probably important to interpret the consequences of heavy ciliate infestations on crayfish growth-related consequence. For example, if heavily infested crayfish show relatively large weight in relation to its size (or positively dispersed from the predicted weight from a length–weight regression line), crayfish–ciliate relationship may be considered mutualistic.
- If the authors have recorded the sex of crayfish, please show whether sex-specific ciliate infestations were evident. This is better accompanied by a statistical test.
- The authors only showed the overall compositions of the two ciliates. Please show details about whether ciliate compositions differed according to ciliate numbers or infestation rate. This is better accompanied by a statistical test on the compositional difference of ciliates according to crayfish weight or size.
- At present, ciliate numbers and crayfish body weight were independently used in simple regression analyses to explain oxygen consumption (Figs. 2 and 3). It may be better to perform multiple regression analysis by using ciliate numbers, crayfish body weight/size and their interaction term as independent variables to explain oxygen consumption.
Based on the #1-5, Results and Discussion parts may be annotated.
Minor issues
- L69 Camaronina
Is this a crayfish pellet? Please specify the type of food.
- L79 OHAUS 07932-0900
Is this an electronic balance? If so, it may be better to state “electronic balance” and enclose the model name, company name and manufacturer city name in parenthesis. Same is true for other terms that only indicated the model name or registered name (e.g. Camaronina).
- L216-217
The last sentence of Conclusion may be better improved to reflect what could be suggested, implied or further clarified based on the present study. At present, the concluding sentence is too general, and the benefits and disadvantages of ciliate infestations on crayfish could not be inferred from the present study per se.
- I don’t see the point in separating the first two paragraphs of Introduction. The second paragraph may be combined in the first paragraph.
Author Response
Dear Reviewer , Thank you very much for your comments, we address the changes according to your proposals. Here are the answers.
- What were the size (e.g. carapace length) and sex of the crayfish used in the experiment? If the authors recorded these information, please describe in the Method section.
Did not record the total length and sex of the crayfish, since we had in mind the objectives of the work at the beginning of the experiment, relying on the literature about the measurements of oxygen consumption in crustaceans, we understood that the necessary data to carry out these measurements, it was only the weight of the organisms and the quantity of oxygen consumption, and of course the variable of the number of ciliates in the gills. So, unfortunately, we do not have this information.
- Provided that the authors have information about crayfish size, what is the consequence of ciliate infestations on the relationship between crayfish size and weight? This is probably important to interpret the consequences of heavy ciliate infestations on crayfish growth-related consequence. For example, if heavily infested crayfish show relatively large weight in relation to its size (or positively dispersed from the predicted weight from a length–weight regression line), crayfish–ciliate relationship may be considered mutualistic.
As we mentioned, we do not have this data, but thanks to your comments.
- If the authors have recorded the sex of crayfish, please show whether sex-specific ciliate infestations were evident. This is better accompanied by a statistical test.
The sex of the crayfish was not recorded. So, unfortunately, we do not have this information.
- The authors only showed the overall compositions of the two ciliates. Please show details about whether ciliate compositions differed according to ciliate numbers or infestation rate. This is better accompanied by a statistical test on the compositional difference of ciliates according to crayfish weight or size.
We found that the crayfish that had more than 50 cells of ciliates on the gills began to increase in oxygen consumption. This was checked with a statistical test and the significant differences are mentioned in lines 162-165, and also in Figure 3, where the colors of the circles show the number of attached ciliates (the darker the circle, greater number of ciliates) . The figure also shows the relationship between weight, oxygen consumption and the number of ciliates in the gills.
- At present, ciliate numbers and crayfish body weight were independently used in simple regression analyses to explain oxygen consumption (Figs. 2 and 3). It may be better to perform multiple regression analysis by using ciliate numbers, crayfish body weight/size and their interaction term as independent variables to explain oxygen consumption.
We performed the statistical test of multiple regression analysis as suggested; the test results were those that had already been shown in the paper. In addition, however, we added the data from the trial, lines 157-162.
- L69. Camaronina. Is this a crayfish pellet? Please specify the type of food.
Done. We specify the type of food and fill in brand information.
- L79. OHAUS 07932-0900. Is this an electronic balance? If so, it may be better to state “electronic balance” and enclose the model name, company name and manufacturer city name in parenthesis. Same is true for other terms that only indicated the model name or registered name (e.g. Camaronina).
Done. We specify the suggested data about the balance.
- L216-217. The last sentence of Conclusion may be better improved to reflect what could be suggested, implied or further clarified based on the present study. At present, the concluding sentence is too general, and the benefits and disadvantages of ciliate infestations on crayfish could not be inferred from the present study per se.
Done. We improved the last paragraph.
- I don’t see the point in separating the first two paragraphs of Introduction. The second paragraph may be combined in the first paragraph.
Done. We combine the first two paragraphs of the introduction section.
Reviewer 3 Report
The submitted manuscript is constructed properly and meets criteria of Diversity editorial board. The introduction grounds the research in the literature quite well.
Methods: I recommend complete the body weight and body length crayfish.
Was the eperiment performed in repetition?
Line no. 85-92. Describe the analyses in the details.
Discussion: The discussion section is repetitive, when compared to the results section. This duplication should be eliminated.
Conclusion: Add suggestion and the real necessities for future study.
Author Response
Dear Reviewer , Thank you very much for your comments which substantially improve this work, we address the changes according to your proposals. Here are the answers.
- Methods: I recommend complete the body weight and body length crayfish.
Done. We completed the information with the suggested data.
- Was the experiment performed in repetition?
We did not repeat the experiment, but we did perform pre-experiment tests to minimize errors.
- Line no. 85-92. Describe the analyses in the details.
Done. We add more details about the identification of the ciliates.
- Discussion: The discussion section is repetitive, when compared to the results section. This duplication should be eliminated.
Done. We erased data already mentioned in the Results Section.
- Conclusion: Add suggestion and the real necessities for future study.
Done. We add a section of the analysis of the prospects of the study.
Round 2
Reviewer 2 Report
The revisions of the manuscript were incomplete in some places. Here are some additional comments and follow-up comments that were unanswered in the authors’ response letter. The Conclusion section needs a rework, as the section comprise awkward sentences and poorly structured paragraphs. As a general rule, please show the line numbers where the changes had been made in response to review comments.
- L73-75
After this sentence, I recommend the authors to show the mean, standard deviation and range of crayfish biomass (weight) used in the experiment.
- L108 vs. L115-117, L136-138 vs. L139-141 Statistical tests
Does the linear regression analysis between crayfish oxygen consumption rate and ciliate numbers on crayfish gills partly overlap with the multiple regression analysis using ciliate numbers and crayfish weight as independent variables and crayfish oxygen consumption rate as the dependent variable? If so, these analyses are largely redundant so only the result of multiple regression should be shown.
In the previous review, I pointed out to check whether the interaction term between ciliate numbers and crayfish weight/size shows statistical significance because of the variability in oxygen consumption rate in small crayfish. The results of multiple regression analysis may be better shown in a table.
- L160-162 Discussion
The first sentence of Discussion had better start with the significant findings of the present study. I suggest reverting back the original sentence.
- L121-125 Ciliate compositions according to crayfish weight
In the previous review, I pointed out the following.
“The authors only showed the overall compositions of the two ciliates. Please show details about whether ciliate compositions differed according to ciliate numbers or infestation rate. This is better accompanied by a statistical test on the compositional difference of ciliates according to crayfish weight or size.”
This comment seems to have been misinterpreted.
I pointed out to see whether numerical dominancy in Epistylis gammari change according to crayfish weight/size or ciliate infestation numbers.
This time, I suggest to show the compositional difference of the two ciliates according to crayfish weight. Crayfish weight may be arbitrarily separated into 2-3 weight classes. Contingency table analyses (e.g. chi-squared test, Fisher's exact test) on compositional difference between the two ciliates may be performed for these crayfish weight classes.
- Model names and manufacturer’s City names
Manufacturer’s City names are only shown for shrimp pellet and electronic balance. Other equipment or device used in the present study only show the model names. Whether or not Manufacturer’s City names are shown should be consistent across the manuscript.
- L226-231
This sentence is too long. The sentence needs to be separated into two or more sentences.
- L241-243 This change responds to the interaction’s context, as the biotic conditions, abundance, and identity of the interacting organisms and the abiotic conditions like the space availability.
This sentence is awkward, and the meaning is unclear. It needs to be rewritten.
- L244 Therefore, ciliate-crayfish epibiosis can change depending on several factors.
This sentence may be moved to the last sentence of the preceding paragraph - don’t change the paragraph
- L244 For example, the availability of food intensifies reproduction and abundance.
This sentence is not understandable. The availability of food intensifies reproduction and abundance of what? The sentence needs a follow-up explanation.
- L256-257 The epibiosis is a dynamic and continuous process where the benefits and disadvantages for the organisms vary according to the ecological context.
This is not the right place for this sentence to appear. This sentence may be omitted as the meaning is not much different from that in L239-241.
- L258-263 We studied the effect of ciliated epibiosis --- showing the importance of this type of researchs.
The first three lines are largely redundant to that already appear.
Below is a suggested revision for the last paragraph (correct typographic errors also).
In future studies, it would be necessary to determine the energy balance using the complete bio-energetics equation, and to estimate the global effect of ciliates attached to crustaceans. This study presents the basis of the effect of ciliates on oxygen consumption by crayfish, showing the importance of this type of researches.
- L261
Regarding the bio-energetics equation, what could be identified or inferred from determination of energy balance (of what?) using complete bio-energetics equation?
Author Response
Dear reviewer
Thank you very much for your comments, which undoubtedly improved this manuscript. We attach a pdf file with the answers point by point for your review.
Author Response File: Author Response.pdf
Reviewer 3 Report
The submitted manuscript is constructed properly and meets the criteria of Diversity editorial board.
The authors have corrected all reviewers’ suggestions to the manuscript. I'm found no deficiencies and errors in the manuscript. I my opinion, the manuscript is acceptable for publication in Diversity.
Author Response
Thank you very much for your comments; they undoubtedly improved this manuscript.
