Ecostoichiometry Reveals the Separation of Microbial Adaptation Strategies in a Bamboo Forest in an Urban Wetland under Simulated Nitrogen Deposition
, Haiyan Sheng 3, Yaoyao Liu 2, Rui Zhang 2, Desy Ekawati 4, Yifan Qian 5 and Yi Lou 5,*Round 1
Reviewer 1 Report
Dear Authors
you manuscript needs some improvement, spelling and gramar check and some points are missing within the Materail and Mehtods part.
Abstract
Line 8: Abbreviations SDN and LEfSe should be written out
Introduction
Line 36: Abbreviation for nitrogen should be given in this line instead of line 37
Line 46: “remains” instead of “remain”
Line 51: “are” helpful not “is”
Line 54: Why is a “new method” mentioned in this line? Is has no connection to any of the other mentioned points above and below.
Line 95: …our goal “was” ….. not our goal “is”
Material and Methods
Line 107: …site “was” not …..site “is”…..
Line 165: Demultiplexing of raw data, quality of demultiplexed reads, read quality filtering, processing of filtered reads, contig removal and so on is missing.
Author Response
You manuscript needs some improvement, spelling and grammar check and some points are missing within the Material and Methods part.
Abstract
- Line 8: Abbreviations SDN and LEfSe should be written out
Answer: Abbreviations SDN and LefSe have been written out.
Introduction
- Line 36: Abbreviation for nitrogen should be given in this line instead of line 37
Answer: We have changed the order of the abbreviation for nitrogen.
- Line 46: “remains” instead of “remain”
Answer: The word “remain” was revised accordingly.
- Line 51: “are” helpful not “is”
Answer: The word “is” was revised accordingly.
- Line 54: Why is a “new method” mentioned in this line? Is has no connection to any of the other mentioned points above and below.
Answer: The word “new” was omitted.
- Line 95: …our goal “was” ….. not our goal “is”
Answer: The word “was” was revised accordingly.
Material and Methods
- Line 107: …site “was” not …..site “is”…..
Answer: The word “was” was revised accordingly.
- Line 165: Demultiplexing of raw data, quality of demultiplexed reads, read quality filtering, processing of filtered reads, contig removal and so on is missing.
Answer: We added this information.
Reviewer 2 Report
This study describes a one-year application of N deposition (dissolved NH4NO3) to simulate elevated nitrogen deposition rates at 1-3 times the ambient (control) rate (already one of the highest rates globally at over 30 kg ha-1 yr-1) in an urban wetland moso bamboo forest in the Yangtze River delta. The study was designed specifically to evaluate the effects of SND (simulated nitrogen deposition at ambient, N30, N60, N90) on soil abiotic and biotic (microbial biomass C, N, P and bacterial and fungal community structures) properties. This is a well designed and described study that provides valuable insights into the effects of SND on soil physicochemical properties and ecostoichemistry, changes in microbial alpha- and beta-diversity, and the linkages between soil properties and microbial community structures.
For the most part, the manuscript is well written (the abstract does need to be tightened up – I have offered edits on the attached manuscript pdf), but, on occasion, there were some odd choices in verbs and sentence structure. Again, I have provided suggestions to tighten up the English writing. The one issue that I feel should be dealt with prior to acceptance in Forests is to better describe the “statistical” responses to the applied treatments – far too frequently, the authors describe non-significant response trends (i.e., applied treatments were not significantly different than the control). Some examples: L.187: “NH4 showed a decreasing trend with increasing SND” – but N30=N60=N90 (not significantly different from each other), L.188: “while NO3 increased” (only at the higher deposition rates: CK (control)=N30<N60<N90). For bacteria alpha-diversity, they note “Chao1 peaked at N30”, but do not highlight that N60 and N90 returned to ambient diversity levels. L.218-219: the authors do state that Shannon and Simpson indices were not significantly different, but then suggest there was an “upward trend” – Simpson’s index to two decimal places was 0.99 across the board (are differences at the third decimal point worth describing it as a trend?). L.222: “F/B declined with the increase of SND” – but N30=N60+N90 (no significant difference) and N60 was similar to the control. I have tried to highlight where these types of non-statistically valid trends are stated – these can be misleading. The results and subsequent discussion should focus on the statistically valid responses.
I enjoyed reviewing this article.
Comments for author File: 
Comments.pdf
Author Response
Reviewer: 2
- On occasion, there were some odd choices in verbs and sentence structure. Again, I have provided suggestions to tighten up the English writing.
Answer: Thank you for the advice, which has greatly improved our paper. We checked the whole paper and revised the language and sentence structure following these comments.
2. The authors describe non-significant response trends (i.e., applied treatments were not significantly different than the control). Some examples: L.187: “NH4 showed a decreasing trend with increasing SND” – but N30=N60=N90 (not significantly different from each other).
Answer: The text was revised accordingly.
3. Did you measure the actual inputs during the duration of the experiment to get a more precise estimate?
Answer: We did not measure the precise N deposition because of insufficient equipment, but we have cited several studies on the N deposition in West Lake and urban area of Hangzhou. After our wetland observation system in Tiaoxi Basin is established, those problems will be solved.
4. L.188: “while NO3 increased” (only at the higher deposition rates: CK (control)=N30<N60<N90).
Answer: It was revised accordingly.
5. For bacteria alpha-diversity, they note “Chao1 peaked at N30”, but do not highlight that N60 and N90 returned to ambient diversity levels.
Answer: It was revised accordingly.
6. L.218-219: the authors do state that Shannon and Simpson indices were not significantly different, but then suggest there was an “upward trend” – Simpson’s index to two decimal places was 0.99 across the board (are differences at the third decimal point worth describing it as a trend?).
Answer: The related description has been modified or deleted.
7. L.222: “F/B declined with the increase of SND” – but N30=N60+N90 (no significant difference) and N60 was similar to the control. I have tried to highlight where these types of non-statistically valid trends are stated – these can be misleading. The results and subsequent discussion should focus on the statistically valid responses.
Answer: The related description has been modified or deleted.
8. Figure quality is poor and needs to be improved, and fonts are too small for a journal publication.
Answer: This may have been a problem related to web formatting. We have provided our best figures in this revision.
9. How much variation was explained by RDA3&4? - 40.5% explained by RDA1&2 - is this for genus composition?This seems high if RDA1&2 explain 40.5% - I would have expected RD3&4 to explain less than 10% more of the variation.
Answer: We checked the dataset and found that the data shown were correct. The confusing sentence was revised. Fig. 5 A and B showed only the cumulative percentage variance (CPV) of bacterial genus data but not the CPV between the bacterial genus composition and the environment.
10. These correlation coefficients seem high compared to the bacteria ones above? Do these all refer to the genus composition portrayed in Fig. 5?
Answer: We agree with the reviewer’s point that correlation coefficients seem high compared to the bacterial coefficients. We understand that the reviewer may want to know the reason for the higher correlation coefficients and lower cumulative percentage variance in our paper. We think that there is no relationship between those two parts using RDA. We use the p value of the Monte Carlo test to determine whether or not the correlation is strong.
Round 2
Reviewer 1 Report
Abstract
Line 8: Abbreviations SDN and LEfSe should be written out
Introduction
Line 36: Abbreviation for nitrogen should be given in this line instead of line 37
Line 46: “remains” instead of “remain”
Line 51: “are” helpful not “is”
Line 54: Why is a “new method” mentioned in this line? Is has no connection to any of the other mentioned points above and below.
Line 95: …our goal “was” ….. not our goal “is”
Material and Methods
Line 107: …site “was” not …..site “is”…..
Line 165: Demultiplexing of raw data, quality of demultiplexed reads, read quality filtering, processing of filtered reads, contig removal and so on is missing.
Author Response
We are sorry for not indicating which line has been modified accordingly. Thank you for the advice, which has greatly improved our paper. We checked the whole paper and revised the language and sentence structure following these comments. We hope that the revised manuscript is now suitable.
Abstract
- Line 8: Abbreviations SDN and LEfSe should be written out
Answer: Abbreviations SDN and LefSe have been written out on lines 18 and 26.
Introduction
- Line 36: Abbreviation for nitrogen should be given in this line instead of line 37
Answer: We have changed the order of the abbreviation for nitrogen on line 45.
- Line 46: “remains” instead of “remain”
Answer: The word “remain” was revised accordingly on line 54.
- Line 51: “are” helpful not “is”
Answer: The word “is” was revised accordingly on line 59.
- Line 54: Why is a “new method” mentioned in this line? Is has no connection to any of the other mentioned points above and below.
Answer: The word “new” was omitted on line 62.
- Line 95: …our goal “was” ….. not our goal “is”
Answer: The word “is” was revised accordingly on line 102.
Material and Methods
- Line 107: …site “was” not …..site “is”…..
Answer: The word “is” was revised accordingly on line 114.
- Line 165: Demultiplexing of raw data, quality of demultiplexed reads, read quality filtering, processing of filtered reads, contig removal and so on is missing.
Answer: We added this information on lines 176-180. “Obtained DNA sequence reads were trimmed, and then chimeras were removed using USEARCH, and sample sequences were combined using QIIME_1.9.1 software. An operational taxonomic unit (OTU) was defined as a set containing DNA sequences with a similarity >97%. OTUs representing less than 0.005% of all sequences were removed before analysis. Diversity indexes were calculated using QIIME software.”
