The Responses of Sediment Bacterial Communities in Chinese Mitten Crab (Eriocheir sinensis) Culture Ponds to Changes in Physicochemical Properties Caused by Sediment Improvement
Round 1
Reviewer 1 Report
The work is very interesting, has a well-developed and executed experimental design, and uses relevant statistics to provide important information for the aquaculture industry. The level of field work, laboratory analysis, and data analysis performed by the authors is high and respectable.
However, the biggest problem is that the microbial community was analysed at the phylum level, which gives too low a resolution and one cannot obtain as much information about the effects of treatment on the ponds. Therefore, the weakest point of the work is the discussion and consequently the conclusion.
There are some problems with the language, but this seems to be mainly a result of autocorrections and deletion of text. There is also a problem with references that needs to be corrected.
Finally, I suggest that you check all your numbers again and work more on the Discussion. All my comments and suggestions are highlighted in yellow.
Comments for author File: 
Comments.pdf
Author Response
The question of why the abundance and changes at the class level of Proteobacteria have not been explored is because this study is a series of overall changes in the more common phylum in the culture environment, and does not focus on the specific changes of a particular phylum, so there is no lower level of Proteobacteria discussion. Regarding the other issues you mentioned, we have already revised them in the manuscript. We appreciate your valuable comments.
Reviewer 2 Report
Review for the paper "The responses of sediment bacterial communities in Chinese mitten crab (Eriocheir sinensis) culture ponds to changes in physicochemical properties caused by sediment improvement" by Tianheng Gao, Nannan Li, Wenlei Xue, Yuning Hu, Hai Lin submitted to "Fishes".
General comment.
Many of aquaculture farms, mostly located in tropical and subtropical countries, have been moved from low- to high-intensity culture, which has led to increases in the strain on the environment. However, this intensification increased stocking density and nutrient pollution, often leading to poor water quality issues. The combination of high density and poor water quality has been shown to increase the likelihood of pathogen outbreaks, which can, in turn, have negative implications for the quality and rate of production. An outcome of these higher disease rates in intensive farming is a reliance on antibiotics and other supplements, especially in countries where regulatory limits may not be clearly defined or monitored closely. There is a lack of information regarding the effects of sediment manipulation activities aiming at improvement of crab culture on nutrients and heavy metals and on the response of bacterial communities to the corresponding changes in the environment. The authors performed a study to reveal the effects of sediment improvement practices on the structure of bacterial communities. The authors found that Proteobacteria, Bacteroidetes, Acidobacteria, Chloroflexi, and Firmicutes were the most abundant taxa in sediments. Sediment improvement has been shown to affect ammonium, sulfide, total organic carbon, and heavy metals. The authors concluded that changes in nutrient composition and heavy metals resulted from sediment improvement further affected the structure and function of bacterial communities. The results of the study are illustrated with relevant figures and tables. Statistical methods are adequate and correctly used. The discussion gives a comprehensive interpretation of the main findings. The paper is of interest to many ecologists dealing with aquatic ecosystems. I some suggestions to improve the ms.
Specific remarks.
Citations are presented as (Authors, year) whereas according to the Instructions for Authors, these should be formatted as [number of source]. Please, change throughout the text.
The authors should indicate on the map the location of each sampling site (GC, PK…).
The authors should describe the procedure of sediment improvement in detail.
Figures 2-4, 7. The authors should increase the bold size and overall resolution.
Indicate the kind of error used in the captions.
The authors should check the numbering of figures and tables. There are tables and figures with the letter "S" (supplementary material).
Table S2 should be moved to the Materials and methods.
The authors should perform statistical comparisons of mean values presented in section 3.1, to support the trends they described.
The authors stated that "The Pearson correlation coefficient calculated based on Pearson correlation analysis was used to assess the statistical correlation between different variables" (P. 5) and "Spearman correlation analysis was used to analyze the relationship between bacterial communities and environmental factors in aquaculture pond sediments". They should clarify this concern.
Some text revisions are highlighted in the attached file.
Comments for author File: Comments.pdf
Author Response
Regarding the last question you mentioned: "The Pearson correlation coefficient calculated based on Pearson correlation analysis was used to assess the statistical correlation between different variables", this description is to get Figures 6B and 6C, and the other description is to get Figure 7. The other issues you mentioned have been revised in the manuscript. We appreciate your valuable comments.
Round 2
Reviewer 1 Report
Manuscript is sufficiently improved. I have just several minor observations, highlighted in pink color.
Comments for author File: Comments.pdf
Author Response
We have carefully revised the manuscript according to your requirements.
1.Reviewer: was
We changed "is" to "was" in abstract.
2. Without "a". Maybe is better to say "and other activities."
We modified the expression accordingly.
3. Did you mean Flavobacteria?
Yes, we corrected the word.
Thank you for your valuable comments.
Reviewer 2 Report
Second review for the paper "The responses of sediment bacterial communities in Chinese mitten crab (Eriocheir sinensis) culture ponds to changes in physicochemical properties caused by sediment improvement" by Tianheng Gao, Nannan Li, Wenlei Xue, Yuning Hu, Hai Lin submitted to "Fishes".
The authors did not provide text revisions to the following previous comments:
1. The authors should describe the procedure of sediment improvement in detail.
As the authors studied sediment bacterial communities before and after the improvement, the reader should be informed about what the improvement procedures were used.
2. Indicate the kind of error used in the figure captions.
Please, indicate which error was used: SD or SE?
3. The authors should perform statistical comparisons of mean values presented in section 3.1 to support the trends they described.
The authors must compare the values before and after the improvement using standard approaches. For example, they may perform t-tests, ANOVA or non-parametric analogs such as Mann-Whitney or Kruskal-Wallis tests for individual values or compare the whole data sets using multivariate approaches such as ANOSIM or PERMANOVA.
P5. Change "Table S2" to "Table S1". Also, I think that the authors should change "Table S1" and "Table S2" to "Table 1" and "Table 2" because the letter "S" is used for supplementary tables.
The authors should highlight with color all changes they made in the text because it is difficult to understand what was actually revised.
Author Response
We have revised the manuscript in response to your comments, thank you very much for your valuable comments.
Author Response File: Author Response.pdf
Round 3
Reviewer 2 Report
Third review for the paper "The responses of sediment bacterial communities in Chinese mitten crab (Eriocheir sinensis) culture ponds to changes in physicochemical properties caused by sediment improvement" by Tianheng Gao, Nannan Li, Wenlei Xue, Yuning Hu, Hai Lin submitted to "Fishes".
The authors have updated the paper with comparisons required using ANOVA, but they not updated the Materials and Methods and figure captions.
Please, include this information in the manuscript.
Author Response
- We added the sentence " One-way ANOVA using orgin6.0 was used to evaluated differences in physicochemical factors before and after sediment improvement " in 2.5.
- We add a sentence to the figure captions of Figures 2, 3, 4, and S1 respectively: " Values were given as mean ± SD, and different letters in the upper (a, b, c, d) indicate a significant difference (p < 0.05) before and after sediment improvement based on the analysis of variance.".
