Sediment Carbon Sequestration and Driving Factors in Seagrass Beds from Hainan Island and the Xisha Islands

Round 1

Reviewer 1 Report

The paper looks at the role of seagrasses in carbon fixation in two marine areas in southern China. The authors conducted multivariate analyses on both abiotic and biotic factors including bacteria and fungi. In this respect, the paper seems to be very interesting. However, I have doubts about the choice of study area. The study analysed results from a large (relatively) island and its various fragments and then compared this with results from small, scattered islands in the open sea. In this case, it is questionable whether comparable elements were compared. After reading the text, the authors have not dispelled my doubts. The sample size is also not very impressive, as a total of 20 samples were analysed. Obviously, the multithreaded nature of the study causes some limitations, so this comment is not crucial. In other methodological aspects, I am unable to deduce which forms of nitrogen compounds were analysed, i.e. nitrate or nitrate nitrogen etc.. The authors take a casual approach to this issue. 

The term eutrophic/oligotrophic refers to the water and not the land/island. It is  wrong use of the term in title of article. The title of the study absolutely needs to be corrected.

Other specific comments:

Lines 65-66 the sentence adds nothing 

Line 70 what is the genesis of these islands because the term natural island defines their origin. Is Hainan island artificial? What is hidden after the enigmatic term intensively used by man

Lines 71-71 it is not clear what has been compared as a consequence

Line 90 the accuracy of the recording of values

Lines 98-99 The map is not developed properly and provides more doubt than clarification. Please describe in the title which part of the graphic relates to Hainan Island and which to Xisha Islands. The small letters do not make it easy to discern the study area. 

Due to the poor quality of the graphic , one has to believe the authors that Hainan Island is indeed an island.

Line 111 please specify the company producing the device

Line 112 Which form of nitrogen was actually determined nitrate nitrogen or nitrates? - note indicated in main comments

Line 124 Total organic carbon is TOC? The term TN is missing the term "total"

Line 165 How was the data converted?

Line 166 when were the different tests used

Line 176 again ambiguity with the forms of nitrogen (in the table there is a different notation of the form)

Line 187 the table caption does not explain anything. What type of data distribution was obtained for this table

Line 192 what test is presented here

Fig.2 Used t-student test and LSD so what was the data distribution?

Line 259 Where is the differential analysis described in the methodology? For what reason were 3 measures used. 

Conclusions should be revised to remove elements relating to the research carried out and to highlight the main achievements of the work relevant to scientific development.

Author Response

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Reviewer 2 Report

Han et al. investigated the sediment carbon sequestration and driving factors in sea grass beds from eutrophic and oligotrophic zones in China
The topic is interesting, especially in the current scenario of increasing shift toward the importance of carbon sequestration by marine vegetated habitats.

However, I believe the manuscript can be improved with a better presentation of the hypotheses, clarification of sampling design, and a better review and use of the literature on the topic.

The statistical analysis is not clear at all. There are few analysis that mentioned in the manuscript but different usage of each test is not obvious for readers. I would strongly suggest to rewrite the section. In the Results I see few other test such as Nmds and Anosim as well. There are not mentioned in the M and M and the results are not discussed.

As a statistician I think I could not understand how metabolic functions of bacterial communities could be related to carbon sequestration by sea grasses. More importantly, how this functions are connected to carbon sequestration via analysis? I think we need more evidence form authors that this could be linked to carbon sequestration, which I doubt.

It is important also to note that the authors used only person correlation index to find so called drivers in their study. I would recommend to use robust tests such as CCA, Distlm, BIOENV, RDA and any other test to related this two important factors.

Author Response

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Reviewer 3 Report

COMMENTS FOR THE AUTHORS

Manuscript Title Sediment carbon sequestration and driving factors in seagrass  beds from eutrophic Hainan Island and oligotrophic Xisha Islands.

I found the manuscript very interesting. Many efforts must presumably have been made to collect and analyze all data.  But several issues should be addressed, before the manuscript can be published.

 

1. Raw metagenomic data should be publicly available (please upload them to GenBank).

2. It is not clear why the Authors dealt only with microbial phyla and did not analyze lower taxa (orders, families). Otherwise it is quite difficult to explain why r-strategists (Proteobacteria and Bacteroidota) dominated in oligotrophic seagrass beds of Xisha Islands. Perhaps key players were members of other phyla.

3. As for fungi participating  in degrading organic matter in marine sediments, I would be careful in pointing out their role in such processes unless data on their total numbers are available. Relative abundance did not contain such information.

4. I seems that Section Materials and Methods does not contain all necessary information. For example it is not explained how enzyme unit (U) for each enzyme is defined.   If U means the amount of the enzyme that catalyzes the conversion of one micromole of substrate per minute under the specified conditions of the assay method, the results obtained for urease and invertase seems far to high. Moreover, one cannot compare the activity of a certain enzyme to that of another since they act on different substrates. Therefore the sentence (line 251): “Thereinto, the activity of urease was the highest, followed by invertase, and alkaline phosphatase activity was the lowest (Table S4)” should be removed.

5. As for functional prediction of bacterial communities related to carbon cycle (3.6. ), it is quite strange why no results concerning carbon autotrophy (especially phototrophy) are presented, considering quite substantial relative abundance of  Cyanobacteria (up to 8.50 %). I would also expect the presence of photoheterotrophy.

6. There are some sentences in Discussion which are difficult to understand.  For example (line 397): “Invertase could promote the mineralization of carbohydrates into CO₂,  H₂O, NH⁴⁺ and inorganic components” How come? Or (line 404): “In addition, the famous ROC degrading bacteria Bacteroidota (Bacteroidetes)…….” Famous?

7. Please kindly check if all references are cited correctly. For example reference 47 is not.  Some of cited references concern terrestrial ecosystems. I am not sure whether all which is true for soil is also true for marine sediments,  I guess both striking similarities and sharp contrasts can be observed.

Author Response

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Round 2

Reviewer 1 Report

Many thanks for the comments given and the changes made to the text. One key point remains. The authors indicate that a Student's t-test was used to compare results. But, in order to apply this test one has to specify the type of distribution of the data. Was such a procedure carried out and what results did it give?

Once I have this information, I will suggest further editorial steps.

Author Response

One key point remains. The authors indicate that a Student's t-test was used to compare results. But, in order to apply this test one has to specify the type of distribution of the data. Was such a procedure carried out and what results did it give?

Response: thank you for your questions which were of great help to improve the quality of our manuscript. Student’s t-test is used when comparing the means of two groups (i.e., pairwise comparison), and the data need to follow a normal distribution. So, in this study, all data were firstly checked for normality and homogeneity of variance using Shapiro-Wilk's test and Levene's test, respectively, and the raw data were log-transformed if they did not pass the normality test. Please see line 176-178 of the revised text.

Reviewer 2 Report

The authors implemented the revisions I requested.

Author Response

Thank you very much.

Reviewer 3 Report

Dear Authors,

Thank you for addressing all issues. In one case I was not specific enough. Namely (line 450), invertase is responsible only for the hydrolysis of sucrose into glucose and fructose, therefore it cannot mineralize carbohydrates into CO₂, H₂O and especially other compounds. Please kindly correct that sentence.

Please also kindly remove the word "cyanobacteria" from lines: 342, 345, 354 and 474 since all information contained there refer to metabolic functions. 

 

Author Response

In one case I was not specific enough. Namely (line 450), invertase is responsible only for the hydrolysis of sucrose into glucose and fructose, therefore it cannot mineralize carbohydrates into CO₂, H₂O and especially other compounds. Please kindly correct that sentence.

Response: Thank you very much for your suggestion. We have corrected that sentence. Please see line 426 of the revised text.

 

Please also kindly remove the word "cyanobacteria" from lines: 342, 345, 354 and 474 since all information contained there refer to metabolic functions.

Response: Thank you. We have deleted the word "cyanobacteria" in lines 342, 345, 354 and 474.

Round 3

Reviewer 1 Report

I have no more comments.