Towards Improved Grain Yield and Soil Microbial Communities of Super Hybrid Rice through Sustainable Management

Round 1

Reviewer 1 Report

Comments for author File: Comments.pdf

Author Response

Several minor corrections are needed, specifically:

line 6 and line 13 - have to be correctly written

line 36- rice - add the Latin name

line 377- the year should be entered

Response: Thanks. These errors have been corrected.

However, is the quality of the yield equally good? Suggestion: It would be good to examine the yield quality and compare among treatments. Sometimes a large yield is obtained, which is significantly lower in nutritional value.

Response: Thank you for this suggestion. In the Chinese study, it was found that there was no decrease in quality along with the increase in rice yield, so the effect of yield on quality was not specifically mentioned in this study.

Reviewer 2 Report

Dear authors,

I think that your paper can be published as submitted. I have only one question/comment at the par. 2.3.2. Soil Sample Collection.

With my best regards.

Comments for author File: Comments.pdf

Author Response

I have only one question/comment at the par. 2.3.2. Soil Sample Collection.

Response: Thanks. Soil samples were collected in two parts, namely (i) determination of soil physicochemical properties (ii) determination of soil microorganisms.

(i) Soil samples at the depth of 20 cm were randomly collected at 5 points in each treatment with a soil drill with 0-20 cm diameter. The soil samples of five soil samples were mixed, and after removing stone, root and plant residues with 2 mm mesh, each sample was divided into two parts: half plastic was used for physical and chemical analysis of fresh soil, and half air-dried was used for physical and chemical analysis. The treated soil samples were divided into three replicates for experimental analysis.

(ii) Before collecting the samples of soil microorganisms, the sampling apparatus should be disinfected and sterilized beforehand, the surface floating soil should be removed during sampling, and the 5-20 cm soil layer should be dug with an ethanol-fired shovel. After removing visible impurities, the soil was passed through a 2 mm sieve, and each sample was collected and mixed from 5 sampling points. The samples were stored in 5 ml centrifuge tubes after removing impurities, transported back to the laboratory at 0°C, and then stored at -80°C for DNA extraction.

Reviewer 3 Report

In this paper the effect of different fertilization practices on grain yield and soil microbial community in super hybrid rice was investigated. Fertilization practices were carried out over the period 2017-2022. In this period grain yield was determined at rice maturity, whereas in October 2022 soil sampling was performed to assess soil physico-chemical properties and the diversity of bacterial and fungal diversity. Results showed that the treatments HYHE and SHY led to a significant increase in rice yield and changed soil physico-chemical characteristics as NH4+-N, total P, available P, organic carbon, and available K. These changes affected the bacterial and fungal diversity. The authors concluded that optimizing fertilization management is crucial to improve the diversity and stability of the microbial community and the rice grain yield.

General comment:

 

The topic of the manuscript is interesting and in general the experimental design and results description are good; however, several points must be clarified. The discussion needs to be revised to make it more focused and clearer.

 

Specific comments:

 

Line 37. Please eliminate “and consumer”.

Line 105. Change “with” to “had”.

Line 115. Eliminate “each”.

Line 153. Please check the title.

Line 163. Please indicate the amount.

Lines 167-169. Please eliminate”microorganisms”and insert a comma after “soil”. If the procedure described was adopted in this study “should be” must be replaced by the appropriate temp of the verb.

Line 181. Please change “Determination of AP by molybdenum-antimony resistance colorimetry” to “Determination of AP was carried out by molybdenum-antimony resistance colorimetry.”

Line 185.  Please change “16s” to “16S”.

Line 189. Please eliminate “fusion”.

Lines 204-207. Please check the grammar. Why AVS instead of ASV?

Lines 240. The title of table 2 is not appropriate. Why rice varieties?

Line 246. Please change “contains” to “contained.”

Line 248. Why OTU? In material and methods AVS were indicated.

Please change the legend of figure 2 because is not clear.

Lines 345-346. Why fungal diversity with two different values of path coefficient? Could one of the two be referred to fungal community structure?

Figure 7 is not very clear.” Bacterial community” and “Fungal community” should be Bacterial community structure” and “Fungal community structure”, respectively. As soil properties refer to the different fertilization treatments I suggest changing in the figure "soil properties" with a more appropriate term that should be used also in other parts of the text where "soil properties" is mentioned.

Lines 365-366.  Please check the grammar of the sentence. The meanining of soil fertility should be better explained.

Lines 369-372. Please check the grammar.

Line 383. Add “healthy” after “environment.”

Line 384. Please eliminate “acidification”.

Line 400. Check the grammar to link this sentence to the following one.

Line 416. What do you mean for Acidobacteriota bacilli? To my knowlege it is not a phylum.

Line 419-429. I suggest reformulating this part reducing results description and giving more emphasis to results discussion.

Lines 433-434. Please check the meaning of the sentence. Chaetomium is a fungus of the Ascomycota phylum.

Line 435. Do you mean soil physico-chemical characteristics?

Line 441. Eliminate “FP”.

Lines 442-444. Please check the grammar of the sentence because the meaning is not clear.

Lines 448-459.  This part refers to conclusions.

Lines 461-462. Please change “Here we discovered optimizing management practices, specifically the HYHE and 461 SHY treatments, led to a significant increase in the yield of super hybrid rice.” “In this study we found that management practices, specifically the HYHE and SHY treatments, led to a significant increase in the yield of super hybrid rice”.

Line 466. Change “saw” showed.

 

A moderate revision of the english language is required. 

Author Response

Specific comments:

Line 37. Please eliminate “and consumer”.

Line 105. Change “with” to “had”.

Line 115. Eliminate “each”.

Line 153. Please check the title.

Line 163. Please indicate the amount.

Lines 167-169. Please eliminate”microorganisms”and insert a comma after “soil”. If the procedure described was adopted in this study “should be” must be replaced by the appropriate temp of the verb.

Line 181. Please change “Determination of AP by molybdenum-antimony resistance colorimetry” to “Determination of AP was carried out by molybdenum-antimony resistance colorimetry.”

Line 185.  Please change “16s” to “16S”.

Line 189. Please eliminate “fusion”.

Lines 204-207. Please check the grammar. Why AVS instead of ASV?

Lines 240. The title of table 2 is not appropriate. Why rice varieties?

Line 246. Please change “contains” to “contained.”

Please change the legend of figure 2 because is not clear.

Figure 7 is not very clear.” Bacterial community” and “Fungal community” should be Bacterial community structure” and “Fungal community structure”, respectively. As soil properties refer to the different fertilization treatments I suggest changing in the figure "soil properties" with a more appropriate term that should be used also in other parts of the text where "soil properties" is mentioned.

Lines 365-366.  Please check the grammar of the sentence. The meanining of soil fertility should be better explained.

Lines 369-372. Please check the grammar.

Line 383. Add “healthy” after “environment.”

Line 384. Please eliminate “acidification”.

Line 400. Check the grammar to link this sentence to the following one.

Line 419-429. I suggest reformulating this part reducing results description and giving more emphasis to results discussion.

Lines 433-434. Please check the meaning of the sentence. Chaetomium is a fungus of the Ascomycota phylum.

Line 441. Eliminate “FP”.

Lines 442-444. Please check the grammar of the sentence because the meaning is not clear.

Lines 448-459.  This part refers to conclusions.

Lines 461-462. Please change “Here we discovered optimizing management practices, specifically the HYHE and 461 SHY treatments, led to a significant increase in the yield of super hybrid rice.” “In this study we found that management practices, specifically the HYHE and SHY treatments, led to a significant increase in the yield of super hybrid rice”.

Line 466. Change “saw” showed.

Response: Thanks. These errors have been corrected. Also, we used professional English language editing services to improve the clarity and readability.

Line 248. Why OTU? In material and methods AVS were indicated.

Response: Thank you for this suggestion. Most studies use OTU to analyse the alpha diversity of bacterial and fungal communities, so this analysis is used here. In addition to this, other microbiological data in the manuscript were analysed using ASV analysis.

Lines 345-346. Why fungal diversity with two different values of path coefficient? Could one of the two be referred to fungal community structure?

Response: Thank you for this suggestion. Yes, one of them refers to fungal community structure, which has been modified in the manuscript.

Line 416. What do you mean for Acidobacteriota bacilli? To my knowlege it is not a phylum.

Response: Thanks. After reviewing the information, Acidobacteria is a newly separated family of bacteria, and there is still little research on them.

[1] Munk J K,Caitlin S,Ann L C, et al. High Diversity and Functional Potential of Undescribed “Acidobacteriota” in Danish Wastewater Treatment Plants . FRONT MICROBIOL,2021,12. https://doi:10.3389/FMICB.2021.643950.

Line 435. Do you mean soil physico-chemical characteristics?

Response: Thanks. Yes, Chaetomium breaks down cellulase and xylanase enzymes, which play an important role in the carbon cycle of natural ecosystems, altering soil physicochemical properties and indirectly improving soil health.

 

Comments on the Quality of English Language

A moderate revision of the english language is required. 

 

Response: Thanks. We used professional English language editing services to iA moderate revision of the English language is required.

Reviewer 4 Report

The experiments are well done, the analysis of data is correct. However, corrections and changes are to be realized.

 

Fig.1 and table 2 are not in accordance with the text. To be adapted.

Fig 1: 2nd figure and title to be changed

Table 2: title to be changed

 

Furthermore

Line 346

Bacterial diversity 346 (path coefficient = 0.89), fungal diversity (path coefficient = 0.58) and fungal diversity (path coefficient = 0.83) had a strong direct positive effect on yield, while bacterial community structure (path coefficient =-0.85) had indirect negative effect on grian yield.

Line 361

PLS-PM analysis showed that soil physical and chemical properties and soil microorganisms had a significant direct effect on crop yield, indicating that crop yield could be increased by enhancing soil microbial diversity and 363 community stability.

Changes are necessary.

(1)

There is not “a strong direct positive effect on yield”, it occurs not “that crop yield could be increased by enhancing soil microbial diversity and community stability “ as such interrelationships are not analyzed. In contrary, yields should occasion the consequence on microbial changes, so given the higher yield residues in HYHE and SHY, the higher root biomass, or the release of less root exudates in case of high availability of nutrients from fertilizers, as the support of microbes for the mobilization of soil nutrients is not necessary for plant development.

The authors may just show a correlation between yield and microbial parameters, but no deduce about a positive effect of microbial diversity etc on yield.

(2)

See the sentence in line 401

“This is because the appropriate fertilization ratios under different management practices can effectively regulate the microbial environment, provide energy and nutrition for the growth and reproduction of microorganisms, and increase the number and diversity of microorganisms” - should be completed by “may be in indirect way over the plant development and yield residues.”

So, the text lines 448pp and in conclusion is appropriate: correlations, but no expressed causality.

I propose the editor to accept the manuscript for publication in Agronomy after realization of the corrections.

Author Response

Fig.1 and table 2 are not in accordance with the text. To be adapted.

Fig 1: 2nd figure and title to be changed

Table 2: title to be changed

Response: Thanks. Amended.

Line 346

Bacterial diversity 346 (path coefficient = 0.89), fungal diversity (path coefficient = 0.58) and fungal diversity (path coefficient = 0.83) had a strong direct positive effect on yield, while bacterial community structure (path coefficient =-0.85) had indirect negative effect on grian yield.

Line 361

PLS-PM analysis showed that soil physical and chemical properties and soil microorganisms had a significant direct effect on crop yield, indicating that crop yield could be increased by enhancing soil microbial diversity and 363 community stability.

Response: Thanks. These errors have been corrected.

There is not “a strong direct positive effect on yield”, it occurs not “that crop yield could be increased by enhancing soil microbial diversity and community stability “ as such interrelationships are not analyzed. In contrary, yields should occasion the consequence on microbial changes, so given the higher yield residues in HYHE and SHY, the higher root biomass, or the release of less root exudates in case of high availability of nutrients from fertilizers, as the support of microbes for the mobilization of soil nutrients is not necessary for plant development.

The authors may just show a correlation between yield and microbial parameters, but no deduce about a positive effect of microbial diversity etc on yield.

Response: We really appreciate the anonymous reviewer for this great suggestion. The partial least squares path model analysis of the data did not show a direct positive effect of microbial diversity on yield, for example. Therefore, only the correlation between yield and microbial parameters was analysed here. In future work, we will also consider other methods to analyse similar issues.

See the sentence in line 401

“This is because the appropriate fertilization ratios under different management practices can effectively regulate the microbial environment, provide energy and nutrition for the growth and reproduction of microorganisms, and increase the number and diversity of microorganisms” - should be completed by “may be in indirect way over the plant development and yield residues.”

So, the text lines 448pp and in conclusion is appropriate: correlations, but no expressed causality.

Response: Thank you for this suggestion. Already added in the manuscript "may indirectly affect plant growth and development and residual yield"

Reviewer 5 Report

In this study, the authors Ye et al. want to reveal whether and how super hybrid rice affects the soil microbial community. Generally, it is a hot topic. However, the introduction and discussion are the weakest part of this paper. The author puts forward some general statements, but the introduction does not lead to scientific assumptions very well. The author needs to reorganize the manuscript to better demonstrate the necessity of this study. This study is not a hypothesis driven research. In addition, the whole manuscript seems more descriptive and cannot give readers more comprehensive knowledge of the importance of this study.  

English language should be polish by a native speaker, very hard to follow. 

Author Response

In this study, the authors Ye et al. want to reveal whether and how super hybrid rice affects the soil microbial community.Generally, it is a hot topic. However, the introduction and discussion are the weakest part of this paper. The author puts forward some general statements, but the introduction does not lead to scientific assumptions very well. The author needs to reorganize the manuscript to better demonstrate the necessity of this study. This study is not a hypothesis driven research. In addition, the whole manuscript seems more descriptive and cannot give readers more comprehensive knowledge of the importance of this study.

 

Response: Thank you for this suggestion. In the revised manuscript, we have revised the introduction and discussion, reduced the number of descriptive conclusions about the study in the manuscript, and restructured the paragraphs.

 

Comments on the Quality of English Language

English language should be polish by a native speaker, very hard to follow.

Response: Thanks. We used professional English language editing services to improve the clarity and readability.

 

Round 2

Reviewer 3 Report

Thanks to the authors for providing a revised version of the manuscript. After carefully reading the manuscript, I found several points that have been addressed. Below are the specific comments.

 

Lines 65-66. The sentence seems incomplete.

Lines 89-94. The aim is not clear. Please check the grammar.

Line 153. “Determination” of what? Please specify.

Lines 167-169. Please change “Before collecting the samples of soil, the sampling instruments are sterilized  beforehand, the top layer of floating soil is removed during sampling, and a 5-20 cm layer  of soil is dug with an ethanol-fired shovel” to Before collecting the samples of soil, the sampling instruments were sterilized beforehand, the top layer of floating soil was removed during sampling, and a 5-20 cm layer of soil was dug with an ethanol-fired shovel.”

Line 181. Add “method” after colorimetry.

Line 226. Change the legend of figure 1 to “Grain yield of super hybrid rice under different management practices of CK, FP, HYHE and SHY.

Line 248-249.  Please change “OUT” to “OTU”. Moreover, in the material and methods section only ASV were mentioned. I also saw your response to this point, and it does not clarify why in the results section only the OTUs were mentioned. You should explain which analyses were based on ASVs and OTUs. You should also explain why the sequences were analyzed by the two methods.

Please change the legend to “The alpha diversity of bacterial and fungal gene sequences under different management practices were Ace index (a), Chao1 index (b), Shannon index (c) and Simpson index (d), respectively. “to “The alpha diversity of bacterial and fungal gene sequences under different management practices measured by Ace (a), Chao1 (b), Shannon (c) and Simpson (d) indices, respectively.”

Figure 7 has not been corrected.” Bacterial community” and “Fungal community” should be Bacterial community structure” and “Fungal community structure”, respectively.

Line 345. Add “structure” after “community.”

Line 366. What does “Fertility of soil is the most intuitive characteristic of soil fertility” mean? Please see my previous comment on this point.

Line 375-378. Why reference 24 was included? I do not understand to which study the sentence refers. I understood the sentence was referred to this study

Lines 402-404. Please check the grammar.

Line 418. Please eliminate “and”.

Lines 446-449. Please change “The study showed that by comparing the physical and chemical properties of soil and microbial community, it was found that the contents of TN, NH4+-N, SOC and AP were important indexes affecting microbial community [41]. To “The study showed that by comparing the physical and chemical properties of soil and microbial community, the contents of TN, NH4+-N, SOC and AP were important parameters affecting microbial community [41]”.

Line 454. Please change “and then promote an increase in yield.” To “and increase the grain yield of super hybrid rice”.

Although the grammar of the manuscript was revised as stated by authors, several points must be checked to make the reading easier.

Author Response

Lines 65-66. The sentence seems incomplete.

Lines 89-94. The aim is not clear. Please check the grammar.

Line 153. “Determination” of what? Please specify.

Lines 167-169. Please change “Before collecting the samples of soil, the sampling instruments are sterilized beforehand, the top layer of floating soil is removed during sampling, and a 5-20 cm layer of soil is dug with an ethanol-fired shovel” to Before collecting the samples of soil, the sampling instruments were sterilized beforehand, the top layer of floating soil was removed during sampling, and a 5-20 cm layer of soil was dug with an ethanol-fired shovel.”

Line 181. Add “method” after colorimetry.

Line 226. Change the legend of figure 1 to “Grain yield of super hybrid rice under different management practices of CK, FP, HYHE and SHY.

Please change the legend to “The alpha diversity of bacterial and fungal gene sequences under different management practices were Ace index (a), Chao1 index (b), Shannon index (c) and Simpson index (d), respectively. “to “The alpha diversity of bacterial and fungal gene sequences under different management practices measured by Ace (a), Chao1 (b), Shannon (c) and Simpson (d) indices, respectively.”

Figure 7 has not been corrected.” Bacterial community” and “Fungal community” should be Bacterial community structure” and “Fungal community structure”, respectively.

Line 345. Add “structure” after “community.”

Line 366. What does “Fertility of soil is the most intuitive characteristic of soil fertility” mean? Please see my previous comment on this point.

Lines 402-404. Please check the grammar.

Line 418. Please eliminate “and”.

Lines 446-449. Please change “The study showed that by comparing the physical and chemical properties of soil and microbial community, it was found that the contents of TN, NH4+-N, SOC and AP were important indexes affecting microbial community [41]. To “The study showed that by comparing the physical and chemical properties of soil and microbial community, the contents of TN, NH4+-N, SOC and AP were important parameters affecting microbial community [41]”.

Line 454. Please change “and then promote an increase in yield.” To “and increase the grain yield of super hybrid rice”.

Response: Thanks. These errors have been corrected.

 

Line 248-249.Please change “OUT” to “OTU”. Moreover, in the material and methods section only ASV were mentioned. I also saw your response to this point, and it does not clarify why in the results section only the OTUs were mentioned. You should explain which analyses were based on ASVs and OTUs. You should also explain why the sequences were analyzed by the two methods.

Response: We really appreciate the anonymous reviewer for this great suggestion. The double-ended reads obtained by sequencing were spliced, filtered and noise-reduced (DADA2), followed by species annotation (ASV analysis and OUT analysis). In the ASV analysis, when some very low abundance species are present in the sample, they may be rejected as sequencing errors, therefore OTUs were used for alpha diversity analysis. Calculation of Shannon and Simpson indices based on OTU analysis, abundance-based coverage estimator (ACE) and Chao1. Abundance analyses, beta diversity analyses and correlation analyses were performed based on valid data obtained from ASV analyses. These have been added to the manuscript in 2.4. Sequence Processing.

 

Line 375-378. Why reference 24 was included? I do not understand to which study the sentence refers. I understood the sentence was referred to this study

Response: Thank you for this suggestion. Proofreading revealed that the study was on the effect of potash fertiliser on soil microorganisms, in agreement with the literature [24].

Round 3

Reviewer 3 Report

Thanks to the authors for providing a revised version of the manuscript. Most of the comments have been addressed. However, in my opinion,  there are still some points that needs to be clarified. One main point concern the analysis of sequences based on OTU and ASV pipelines. How can these two different methods infer the results obtained? The papers by by Chiarello et al. 2022 (Ranking the biases: The choice of OTUs vs. ASVs in 16S rRNA amplicon data analysis has stronger effects on diversity measures than rarefaction and OTU identity threshold. PLoS ONE 17(2): e0264443. https://doi.org/10.1371/journal.pone.0264443) and Jeske et al.2022 (Microbiome Analysis via OTU and ASV-Based Pipelines-A Comparative Interpretation of Ecological Data in WWTP Systems. Bioengineering (Basel). 2022 Mar 29;9(4):146. doi: 10.3390/bioengineering9040146. PMID: 35447706; PMCID: PMC9029325).could help the authors to clarify this point. To my knowledge only one of the two methods should be used to obtain more representative results or both methods could be used in the same study to compare the results.

 

Below other specific comments.

Lines 89-92. Please change ”In this study, the effects of different management practices on soil physicochemical properties and microorganisms of super hybrid rice were investigated by locational experiments through the analysis of soil physicochemical properties, microbial diversity and community structure.” to “In this study, a locational field experiment was conducted from 2017 to 2022 to investigate the effect of different management practices on soil physicochemical properties and soil microbial community diversity and structure of super hybrid rice.”

Line 151. Please change the title of chapter 2.3. “Sample Collection and analyses”. Moreover, the order of the subchapters should be “2.3.1. Soil Sample Collection”, 2.32. Determination of Soil Physicochemical Indexes,  2.3.3. Grain Yield and  2.3.4. Soil DNA Extraction and High-Throughput Sequencing Analysis, 2.5. Sequence Processing. The same order should be used for Results section.

Please change legend of figure 1 to “Changes of super hybrid rice yield under different management practices of CK, FP, HYHE and SHY”.

Please check throughout the text the term “OUT” and change to “OTU”.

Lines 380-382. As you wrote in the response you have to explain in the text that your results were in agreement with those observed by Kaiser et al. 2015 ( ref 24) specifying what these authors observed.

 

 

 

Some corrections of the english language are still required.

Author Response

11 August 2023

Re: revision to “Optimizing management practices for improved grain yield and soil microbial community in super hybrid rice” manuscript number agronomy-2524330-1

Dear editor,

Thank you for your assessment. The reviewers have done an excellent job in reviewing this manuscript, the comments and changes made have helped improve the clarity and scientific rigor of our manuscript. We thank you and the reviewers for spending so much time on this paper. Our responses to the reviewers’ comments point-by-point were included below as inserts marked in red.

I hope the revised version of the paper is acceptable and it is processed further. If there are any further queries/unclear points regarding the revised manuscript, please let me know.

 

 

Sincerely yours,

 

Yunbo Zhang

Hubei Collaborative Innovation Center for Grain Industry, Yangtze University

 

 

 

 

Responses to reviewer comments:

 

Thanks to the authors for providing a revised version of the manuscript. Most of the comments have been addressed. However, in my opinion, there are still some points that needs to be clarified. One main point concern the analysis of sequences based on OTU and ASV pipelines. How can these two different methods infer the results obtained? The papers by by Chiarello et al. 2022 (Ranking the biases: The choice of OTUs vs. ASVs in 16S rRNA amplicon data analysis has stronger effects on diversity measures than rarefaction and OTU identity threshold. PLoS ONE 17(2): e0264443. https://doi.org/10.1371/journal.pone.0264443) and Jeske et al.2022 (Microbiome Analysis via OTU and ASV-Based Pipelines-A Comparative Interpretation of Ecological Data in WWTP Systems. Bioengineering (Basel). 2022 Mar 29;9(4):146. doi: 10.3390/bioengineering9040146. PMID: 35447706; PMCID: PMC9029325). could help the authors to clarify this point.To my knowledge only one of the two methods should be used to obtain more representative results or both methods could be used in the same study to compare the results.

Response: We are very grateful to the anonymous reviewers for their valuable suggestions.

 

For the ASVs obtained, representative sequences of each ASV were species annotated to obtain the corresponding species information and species-based abundance distribution.At the same time, to obtain information on species richness and evenness within samples, as well as information on ASVs that are shared and endemic among different samples or subpopulations, the ASVs were analysed using abundance, alpha-diversity calculations, Venn diagrams and pedigree plots.The species composition and community structure of grouped samples can be tested for the significance of differences using dimensionality reduction analyses and sample clustering tree displays such as PCoA, PCA, NMDS, and selected statistical analyses such as LEfSe to test the significance of differences in species composition and community structure of grouped samples (Lines 233-236).

 

In the revised manuscript, we changed the OTU analysis to ASV analysis to ensure that the same analytical methods were used in this study to make the data more representative. The specific data additions and analyses are shown in Table 3, and we thank you again for your suggestions!

After quality control, each sample contained 62288-76393 sequences and 464-2188 ASVs numbers, and the sample sequencing coverage is 100%. Among the four treatments, the SHY treatment had the highest number of ASVs for both bacteria and fungi. In the FP treatment, fungi had the lowest number of ASVs (Lines 197-206).

 

Table 3. The alpha diversity of bacterial and fungal gene sequences under different management practices measured by Ace (a), Chao1 (b), Shannon (c) and Simpson (d) indices, respectively.

Microbe Type	Treatment	Richness Index		Diversity Index
		ACE	Chao1	Shannon	Simposn
Bacteria	CK	1827±46 b	1827±45 b	10.19±0.08 b	0.999±0.001 a
	FP	1934±9 ab	1977±93 ab	10.29±0.08 ab	0.999±0 a
	HYHE	2064±24 ab	2034±51 a	10.28±0.10 ab	0.999±0.001 a
	SHY	2188±81 a	2121±115 a	10.41±0.09 a	0.999±0 a
Fungi	CK	495±4 b	495±15 bc	6.16±0.34 a	0.958±0.015 a
	FP	464±6 c	464±20 c	5.68±0.18 b	0.917±0.049 a
	HYHE	511±8 b	511±27 b	6.34±0.10 a	0.969±0.007 a
	SHY	556±10 a	556±27 a	6.52±0.17 a	0.969±0.021 a

Note: Different letters for the same item indicate p < 0.05 (significant differences).

 

Lines 89-92. Please change ”In this study, the effects of different management practices on soil physicochemical properties and microorganisms of super hybrid rice were investigated by locational experiments through the analysis of soil physicochemical properties, microbial diversity and community structure.” to “In this study, a locational field experiment was conducted from 2017 to 2022 to investigate the effect of different management practices on soil physicochemical properties and soil microbial community diversity and structure of super hybrid rice.”

Please change legend of figure 1 to “Changes of super hybrid rice yield under different management practices of CK, FP, HYHE and SHY”.

 

Response: Thanks. These errors have been corrected.

 

Line 151. Please change the title of chapter 2.3. “Sample Collection and analyses”. Moreover, the order of the subchapters should be “2.3.1. Soil Sample Collection”, 2.32. Determination of Soil Physicochemical Indexes,  2.3.3. Grain Yield and  2.3.4. Soil DNA Extraction and High-Throughput Sequencing Analysis, 2.5. Sequence Processing. The same order should be used for Results section.

 

Response: The statements have been corrected. We will be happy to edit the text further , based on helpful comments from reviewers.

 

Lines 380-382. As you wrote in the response you have to explain in the text that your results were in agreement with those observed by Kaiser et al. 2015 ( ref 24) specifying what these authors observed.

 

Response: Thanks for your careful checks. Based on your comments, we have revised the references ( ref 23 ) to be more consistent with the analysis of the results (Lines 370-374).

It has been shown that the use of phosphorus fertilizer decreases significantly with the increase in the amount of phosphorus fertilizer, which is used at a rate of 15% during cultivation, with the rest remaining in the soil [23]. In the present study, it was found that the highest TP and AP contents were found in the soils of SHY treatment, which is in agreement with the results of previous studies.

 

Comments on the Quality of English Language: Some corrections of the english language are still required.

 

Response: Thanks for your suggestion. We have tried our best to polish the language in the revised manuscript

 

Author Response File: Author Response.pdf

Round 4

Reviewer 3 Report

Thanks to the Authors for providing a further revised version of the manuscript. The manuscript was improved; however, my main concern is still on the analyses based on OTUs and ASVs. Why in the subchapter 3.3. Beta Diversity of Bacterial and Fungal Communities, the OTUs were used to construct the VENN diagrams? As you stated in the response to the comment you changed the OTU analysis to ASV analysis to ensure that the same analytical methods were used in this study to make the data more representative.

Additional minor comments:

Lines 107-108. The sentence was repeated twice.

Lines 109-110. Please eliminate “developed by BioRice Co., Ltd. located in Changsha, Hunan. It is a representative variety of super hybrid rice in recent years, developed by BioRice Co., Ltd. in Changsha, Hunan,”.

Line 204. Please change “reduction (DADA2) were performed” to “were performed using DADA2 pipeline”.

Line 234. Please change “is” to “was”.

 

Minor corrections of the english grammar are required.

Author Response

Responses to reviewer comments:

 

Thanks to the Authors for providing a further revised version of the manuscript. The manuscript was improved; however, my main concern is still on the analyses based on OTUs and ASVs. Why in the subchapter 3.3. Beta Diversity of Bacterial and Fungal Communities, the OTUs were used to construct the VENN diagrams? As you stated in the response to the comment you changed the OTU analysis to ASV analysis to ensure that the same analytical methods were used in this study to make the data more representative.

 

Response: We thank the anonymous reviewers for their valuable suggestions. VENN plot construction using ASV in bacterial and fungal community Beta diversity. We apologise for the error in sequence units in the manuscript and have changed "OTU" to "ASV" in the latest manuscript.

 

Lines 107-108. The sentence was repeated twice.

Lines 109-110. Please eliminate “developed by BioRice Co., Ltd. located in Changsha, Hunan. It is a representative variety of super hybrid rice in recent years, developed by BioRice Co., Ltd. in Changsha, Hunan,”.

Line 204. Please change “reduction (DADA2) were performed” to “were performed using DADA2 pipeline”.

Line 234. Please change “is” to “was”.

 

Response: Thanks. These errors have been corrected.

 

 

Comments on the Quality of English Language: Minor corrections of the english grammar are required.

Response: Thanks for your suggestion. We have endeavoured to refine the wording in the revised manuscript.

Author Response File: Author Response.pdf