Contrasting Effects of Moso Bamboo Expansion into Broad-Leaved and Coniferous Forests on Soil Microbial Communities

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Author,

Please find my recommendations for the current version of manuscript "Contrasting Effects of Moso Bamboo Expansion into Broad-leaved and Coniferous Forests on Soil Microbial Communities"

1. The abstract should consider to be less narrative and include also numerical values 
2. The introduction section should better highlight the current knowledge gaps and challenges/divergence evidenced from literature related to Moso Bamboo expansion in forest soil microbiome; Here the authors should be more explicit.
3. Please consider to evidence also the relevance for addressing this gaps
4. L122: Please clarify this coordinates 27°40′–26°53′ N, 117°09′–117°18′ E
5. L152: Please replace "-20°Cprior" with "-20 °C prior"
6. L157-158: Please remove "Hanna-Kunath-Str. 11, 28199 Bremen, German" and insert the brand of equipment as "Thermo Fisher Scintific" followed by town and country of production "Bremen, Germany"
7. L212, L234: Please check figure 1 and 2. I suppose that there are bar plot but invisible in the manuscript format available for me
8. L219, L240: In case of table 3 and 4 the manuscript should present also the F value (not just p-value)
9. The results should better consider results seasonal variability as "Soil sampling was performed in March, June, and September 2022" (L146)
10. L226-233: Please check these statements. Please have in view that total microbial biomass is the sum of fungal and bacterial PLFA...
11. L267-272: Such statement should be sustained by nitrification or denitrification rates in my opinion
12. Same in case of exogenous carbon case (L273-276) where at least enzymatic assessments are needed
13. L259: The manuscript should focus also to better interpret the results related to the mixed forests also as there are statistically significant variabilities according to results section
14. When comparing results the manuscript also should consider differences in climate, soil, and bamboo management among studies 
15. Please consider to present also the limitation of the current study and if case further recommendations 

Author Response

Comment 1: The abstract should consider to be less narrative and include also numerical values. 

Response: Thanks for your suggestion. We have added specific numerical values (percentages) of relevant indicators showing their changes to the abstract to enhance its readability. Details please see L26-31 in the revised version.

Comment 2~3: The introduction section should better highlight the current knowledge gaps and challenges/divergence evidenced from literature related to Moso Bamboo expansion in forest soil microbiome; Here the authors should be more explicit. Please consider to evidence also the relevance for addressing this gaps.

Response：Thank you for your suggestion. The introduction section of this paper has fully demonstrated that the differential impacts of Moso bamboo expansion on soil microbial communities are associated with the expanded forest types and study regions. However, to better understand the mechanisms by which Moso bamboo expansion into different forest types affects soil microbial communities, it is necessary to eliminate the interference of different sampling sites. We have added the sentence "when the potential impacts of different sampling sites are overlooked" in L112-113 in the revised version to make the research objectives more explicit, hoping that the revised version enhances readability.

Comment 4: L122: Please clarify this coordinates 27°40′–26°53′ N, 117°09′–117°18′ E

Response: We have corrected the miswritten coordinates.

Comment 5: L152: Please replace "-20°C prior" with "-20 °C prior"

Response: Done.

Comment 6: L157-158: Please remove "Hanna-Kunath-Str. 11, 28199 Bremen, German" and insert the brand of equipment as "Thermo Fisher Scintific" followed by town and country of production "Bremen, Germany"。

Response：We have changed the sentence as you suggested, details please see L166 in the revised version.

Comment 7: L212, L234: Please check figure 1 and 2. I suppose that there are bar plot but invisible in the manuscript format available for me。

Response:We have changed the format of the figures to make them more convenient for reviewers to review, details please Fig.1 and Fig.2 in the revised version.

Comment 8: L219, L240: In case of table 3 and 4 the manuscript should present also the F value (not just p-value)

Response: Thanks for your suggestion. The F values have added in the table 3 and 4 in the revised version.

Comment 9: The results should better consider results seasonal variability as "Soil sampling was performed in March, June, and September 2022" (L146)

Response: Thanks for your suggestion. We have emphasized the impact of seasons in the result description section concerning changes in soil microbial community structure, that is, the influence of sampling time on soil microbial PLFAs. Details please see L221-222and L242-243 in the revised version.

Comment 10: L226-233: Please check these statements. Please have in view that total microbial biomass is the sum of fungal and bacterial PLFA...

Response: Thanks for your suggestion. We have carefully checked the data of soil microbial PLFAs, and corrected the data with descriptive errors. Hopefully the revised version will be more readable. Details please see L210-212, and L230-232.

Comment 11: L267-272: Such statement should be sustained by nitrification or denitrification rates in my opinion.

Response: We have changed the sentence “altering soil nitrogen transformation pathways” to “thereby decreasing organic acid input and soil nitrogen net nitrification and mineralization rates” in L288-289 in the revised version.

Comment 12: Same in case of exogenous carbon case (L273-276) where at least enzymatic assessments are needed

Response: Thanks for your suggestion. We have changed the sentence “especially oligotrophic bacterial” to “especially oligotrophic bacterial that are adept at secreting chitin-degrading enzymes”, L291-292 in the revised version. Hopefully the revised version will be more readability.

Comment 13: L259: The manuscript should focus also to better interpret the results related to the mixed forests also as there are statistically significant variabilities according to results section

Response: Thanks for your suggestion. In the present study, mixed forests represent a stage in the process of Moso bamboo expansion. Both Coniferous forests or broad-leaved forests will eventually be replaced by Moso bamboo forests. When Moso bamboo invasion at different stages leads to inconsistent effects on soil microbial community structure or other relevant factors, it is necessary to focus on the results of mixed forests. When the results are consistent, the results of Moso bamboo expansion effects presented in this study include the combined effects of Moso bamboo forests and mixed forests.

Comment 14: When comparing results the manuscript also should consider differences in climate, soil, and bamboo management among studies 

Response: Thanks for your suggestion. In the introduction section of this study, we discussed that Moso bamboo expansion may have diverse impacts on soil microbial

community structure due to varying research regions, forest types, and sampling sites. To investigate these impacts on soil microbial communities during Moso bamboo invasion into two typical vegetation types in subtropical regions, this study selected sampling sites within the same climate zone, with identical soil types, and under similar bamboo management practices. This approach allowed us to exclude potential confounding effects from climate variability, soil heterogeneity, and different bamboo management strategies.

Comment 15: Please consider to present also the limitation of the current study and if case further recommendations 

Response: Thank you for your suggestion. As you suggested, we have added discussions on the limitations of this study and prospects for further research in the conclusion section. The specific content is as follow: “In addition, given the limitations of the sampling sites in this study, it is suggested that further research should be conducted across a larger spatial scale (e.g., different climate zones) and include more vegetation types to verify the universality of the “forest type-dependent responses” to Moso bamboo expansion. ”

Reviewer 2 Report

Comments and Suggestions for Authors

• Clearly state the objectives of the study.

• Avoid overly long paragraphs to improve readability and logical flow.

• The soil sampling and processing methodology is well described; however, the section would benefit from improved structuring. Consider breaking long paragraphs into smaller, logically grouped subsections (e.g., sampling, chemical analysis, microbial analysis) to enhance readability.

• Please ensure consistency in reporting units and symbols throughout the section. For example, use standard spacing (e.g., “−20 °C” instead of “-20°C”) and report microbial biomass as “nmol g⁻¹ dry soil” for clarity.
• It would strengthen the methodology to clearly state how randomization was achieved when selecting the nine sampling points. Also, were the samples taken as true biological replicates or pooled before analysis?
• The PLFA method is appropriately cited, but a bit more detail could be helpful for readers unfamiliar with the technique. For instance, mentioning the type of GC-MS or GC-FID used, and the criteria for biomarker selection (gram-positive, gram-negative, AM fungi, etc.) would improve reproducibility.

• Please clarify how PLFA data were processed before statistical analysis. Were the values log-transformed or standardized prior to ordination or multivariate analyses?

• While the use of the 0–10 cm soil layer is common, consider briefly justifying this choice, especially if deeper soil layers may also be ecologically relevant in the studied forest types.

  • Avoid use of “etc.” in scientific writing (e.g., "removing debris such as fine roots, stones, etc."), as it appears vague.

  • Define abbreviations at first mention (e.g., PLFA, TN, TC) even if they are common in soil science literature.

  • Consider moving equipment manufacturer details (e.g., for analyzers) to a footnote or supplementary table if they disrupt flow.

  • Please revise the unit format consistently throughout the manuscript as Guidelines. (mg/kg or mg kg-1 )...

• The discussion is generally well-aligned with the stated hypotheses, but the transition between the two hypotheses could be clearer. You may explicitly restate both hypotheses before interpreting the contrasting responses in MF and CF.
• Use clearer paragraph breaks to separate interpretations related to broad-leaved forests (MF) vs. coniferous forests (CF) for better readability. 

• The discussion is very informative but presented in overly long and dense paragraphs. For improved comprehension, please divide key points into shorter, focused paragraphs. For example:

  • One paragraph for changes in microbial biomass in MF.

  • One for nutrient cycling mechanisms (SOC, NO₃⁻-N, NH₄⁺-N).

  • One for community composition shifts (F:B ratio).

  • One for CF-specific findings.

  • One for possible mechanisms (e.g., pH, litter quality, shading effects).

  • Line 263: “...in theses ecosystems” → should be “these ecosystems.”

  • Line 282: “...thereby enhancing microbial biomass-predominantly via bacterial proliferation” → revise for clarity. Consider: “...thus enhancing microbial biomass, primarily due to bacterial proliferation.”

  • Line 312–313: Sentence ends abruptly with "underscoring the imp" → appears to be an incomplete sentence. Please revise or complete the thought.

  • While changes in pH and litter quality are discussed, it would be helpful to explore functional implications, such as potential impacts on nutrient cycling efficiency, carbon sequestration, or forest ecosystem resilience.
  • The conclusion summarizes key findings effectively. However, some sentences are overly condensed. Consider briefly restating the ecological implications of the changes in microbial community composition (e.g., nutrient cycling, soil health, or forest function). Consider adding 1–2 sentences about the practical or ecological relevance of the findings. 

Author Response

Comment 1: Clearly state the objectives of the study.

Response: Thanks for your suggestion, we have revised the expression in the abstract to make the research objectives more explicit. Details please see L 20-21 in the revised version.

Comment 2: Avoid overly long paragraphs to improve readability and logical flow.

Response: Thanks for your suggestion. Following the reviewers’ comments, we have optimized the language throughout the manuscript, making efforts to avoid overly long sentences. Hopefully the revised version will have better readability and logical flow.

Comment 3: The soil sampling and processing methodology is well described; however, the section would benefit from improved structuring. Consider breaking long paragraphs into smaller, logically grouped subsections (e.g., sampling, chemical analysis, microbial analysis) to enhance readability.

Response: We have split the originally long paragraphs into shorter ones, hoping that the revised version will have better readability. Details please see L161-169 in the revised version.

Comment 4: Please ensure consistency in reporting units and symbols throughout the section. For example, use standard spacing (e.g., “−20 °C” instead of “-20°C”) and report microbial biomass as “nmol g⁻¹ dry soil” for clarity.

Response: Thank you for your detailed comments regarding the consistency of units and symbols in the manuscript. We have thoroughly checked and revised the entire document to ensure uniformity.

Comment 5: It would strengthen the methodology to clearly state how randomization was achieved when selecting the nine sampling points. Also, were the samples taken as true biological replicates or pooled before analysis?

Response: We have revised sentence “and nine random points were selected for soil collection at 0-10 cm depth” to “and nine random points were selected using a coordinate-based randomization method for soil collection at 0-10 cm depth” to more clearly explain the randomization method for soil collection at the nine sampling points. Details please see L154-155 in the revised version. In addition, soil from the nine sampling points was mixed into one sample, which served as a true biological replicate. We have revised the description of the soil sample collection method to make it more readable. Details please see L156-157 in the revised version.

Comment 6: The PLFA method is appropriately cited, but a bit more detail could be helpful for readers unfamiliar with the technique. For instance, mentioning the type of GC-MS or GC-FID used, and the criteria for biomarker selection (gram-positive, gram-negative, AM fungi, etc.) would improve reproducibility.

Response: Thanks for your suggestion. We have supplemented the information on the instruments used for PLFAs analysis as follows: The treated soil samples are analyzed by an Agilent 6890GC/5973 MS gas chromatography - mass spectrometry instrument (L171-172 in revised version). The PLFA method can distinguish the biomass of Gram-positive bacteria, Gram-negative bacteria, and AM mycorrhizal fungi in soil based on different markers. However, this study focuses more on community changes rather than functional specificity changes, and no further functional differentiation was performed to avoid misleading the results.

Comment 7: Please clarify how PLFA data were processed before statistical analysis. Were the values log-transformed or standardized prior to ordination or multivariate analyses?

Response: Thanks for your suggestion. In the data analysis section of the study, we state that all data were subjected to normality tests and homogeneity of variance tests, and no logarithmic or other transformations were performed on the data.

Comment 8: While the use of the 0–10 cm soil layer is common, consider briefly justifying this choice, especially if deeper soil layers may also be ecologically relevant in the studied forest types.

Response: Deep soil microbial community structure may also respond to changes in forest types. However, given that the scientific hypothesis of this study is based on the impacts of Moso bamboo expansion on the quantity, quality, and decomposition rate of litter, which in turn affect soil nutrient elements and microbial community structure, focusing on changes in the surface soil microbial community is more conducive to verifying the scientific validity of the hypothesis.

Comment 9: Avoid use of “etc.” in scientific writing (e.g., "removing debris such as fine roots, stones, etc."), as it appears vague.

Response: Thanks for your suggestion, we have deleted etc. in the revised version. 

Comment 10: Define abbreviations at first mention (e.g., PLFA, TN, TC) even if they are common in soil science literature.

Response: Thank you for your suggestion regarding the definition of abbreviations. I have carefully reviewed the manuscript and confirmed that all abbreviations (e.g., PLFA, TN, TC) are clearly defined at their first appearance. This ensures clarity for all readers, regardless of their familiarity with soil science literature.

Comment 11: Consider moving equipment manufacturer details (e.g., for analyzers) to a footnote or supplementary table if they disrupt flow.

Response: Thank you for your suggestion. The equipment manufacturer details are already included in the text as necessary for clarity and reproducibility.

Comment 12: Please revise the unit format consistently throughout the manuscript as Guidelines. (mg/kg or mg kg^-1 ).

Response: Thanks for your suggestion, we have changed mg/kg to mg kg^-1 through out the study in the revised version.

Comment 13: The discussion is generally well-aligned with the stated hypotheses, but the transition between the two hypotheses could be clearer. You may explicitly restate both hypotheses before interpreting the contrasting responses in MF and CF. Use clearer paragraph breaks to separate interpretations related to broad-leaved forests (MF) vs. coniferous forests (CF) for better readability. 

Response: Thanks for your suggestion, we have added subheadings in the discussion section to clearly distinguish the impacts of Moso bamboo expansion into different forest types. Additionally, we have split lengthy sentences into shorter ones to make the structure clearer and more readable. We hope the revised version will be more readable.

Comment 14: The discussion is very informative but presented in overly long and dense paragraphs. For improved comprehension, please divide key points into shorter, focused paragraphs. For example: One paragraph for changes in microbial biomass in MF. One for nutrient cycling mechanisms (SOC, NO₃⁻-N, NH₄⁺-N). One for community composition shifts (F:B ratio). One for CF-specific findings. One for possible mechanisms (e.g., pH, litter quality, shading effects).

Response: Thanks for your suggestion. As suggested, we have split the overly long and dense paragraphs into shorter ones concerning different possible mechanisms. Details please see the discussion section in the revised version (L273-329).

Comment 15: Line 263: “...in theses ecosystems” → should be “these ecosystems.”

Response: Done

Comment 16: Line 282: “...thereby enhancing microbial biomass-predominantly via bacterial proliferation” → revise for clarity. Consider: “...thus enhancing microbial biomass, primarily due to bacterial proliferation.”

Response: Thanks for your suggestion. We have changed the sentence as you suggested. Details please see L290-291 in the revised version.

Comment 17: Line 312–313: Sentence ends abruptly with "underscoring the imp" → appears to be an incomplete sentence. Please revise or complete the thought. While changes in pH and litter quality are discussed, it would be helpful to explore functional implications, such as potential impacts on nutrient cycling efficiency, carbon sequestration, or forest ecosystem resilience.

Response: Thanks for your suggestion. We have changed the sentence to “Collectively, our results demonstrate that PF expansion into CF influence soil microbial biomass and composition through changes in plant litter input-driven soil carbon and nutrient cycling processes, as well as soil pH, underscoring the importance of context-dependent mechanism in mediating Moso bamboo expansion effects.”. Hopefully the revised version will be more readable.

Comment 18: The conclusion summarizes key findings effectively. However, some sentences are overly condensed. Consider briefly restating the ecological implications of the changes in microbial community composition (e.g., nutrient cycling, soil health, or forest function). Consider adding 1–2 sentences about the practical or ecological relevance of the findings. 

Response: Thank you for your suggestions. In accordance with your comments, we have optimized the sentences in the conclusion section and added discussions on the limitations of the current study as well as future research prospects. Details please see L340 - 343 in the revised version. Hopefully the revised version will be more readable.

Reviewer 3 Report

Comments and Suggestions for Authors

The topic of the study is significant because it explores the impact of the invasive spread of Moso bamboo on soil microbial communities. This research provides crucial insights into the response of underground ecosystems to vegetation changes. It is particularly valuable for understanding how different forest types mediate the ecological effects of bamboo expansion. The authors emphasize the importance of considering the context-dependent patterns in order to challenge assumptions about the ecological impacts of bamboo. 

The article seems correct and logical. However, there are a number of points that require clarification:

Line 231-233. p-value = 0.084. Without further justification, the claim of "significant interaction" appears to be incorrect.

The authors write (222-226): During PF expansion into CF, total soil microbial biomass changed from 10.12 to11.28 nmol g⁻¹, and Bacterial biomass: 6.29–12.17 nmol g⁻¹. But bacterial biomass cannot be greater than total microbial biomass (12.17 > 11.28). Could you explain this ?

Table 3. Complete the legend: during PF expansion into СF here.

Lines 229-231. In your table 4 in column F:B there are p-values ​​both above 0.05 and below 0.05. In this regard, as I see it, this sentence needs to be corrected in the part concerning F:B “Sampling time and forest type significantly interacted to affect total microbial biomass, fungal biomass, and bacterial biomass, but not microbial community composition during PF expansion into CF”.

The same question applies to the part of the results where this sentence (Lines 205-206): “Temporal variation apparently influenced total microbial biomass, bacterial biomass, and fungal biomass”. There is no mention of microbial community composition. Why?

Lines 231-233. You write: “Soil microbial communities varied significantly across sampling periods, whereas the interactive effects between sampling time and forest type were only notable for F:B ratio”. But according to table 4, significant p-values (F:B ratio) indicators are only in the "Forest type" row. How can this be explained?

Section 3.3. Here it is necessary to indicate the correlation coefficients.

L 246. bacterial biomass ? Check this.

Author Response

Comment 1: The topic of the study is significant because it explores the impact of the invasive spread of Moso bamboo on soil microbial communities. This research provides crucial insights into the response of underground ecosystems to vegetation changes. It is particularly valuable for understanding how different forest types mediate the ecological effects of bamboo expansion. The authors emphasize the importance of considering the context-dependent patterns in order to challenge assumptions about the ecological impacts of bamboo. The article seems correct and logical.

Response: Thanks for your recognition of our work. We are willing to make every effort to revise the paper in accordance with your suggestions.

Comment 2: Line 231-233. p-value = 0.084. Without further justification, the claim of "significant interaction" appears to be incorrect.

Response: We have carefully checked the content and deleted the incorrectly worded (redundant) sentences: “Soil microbial communities varied significantly across sampling periods, whereas the interactive effects between sampling time and forest type were only notable for F:B ratio (Table 4). ” in the revised version.

Comment 4: The authors write (222-226): During PF expansion into CF, total soil microbial biomass changed from 10.12 to11.28 nmol g⁻¹, and Bacterial biomass: 6.29–12.17 nmol g⁻¹. But bacterial biomass cannot be greater than total microbial biomass (12.17 > 11.28). Could you explain this ?

Response: Thank you for your suggestion. We have carefully checked the data of soil microbial PLFAs, and corrected the data with descriptive errors. Hopefully the revised version will be more readable. Details please see L207-209, and L227-229. I sincerely apologize for the oversight and appreciate your attention to detail.

Comment 5: Table 3. Complete the legend: during PF expansion into СF here.

Response: Done.

Comment 6: Lines 229-231. In your table 4 in column F:B there are p-values both above 0.05 and below 0.05. In this regard, as I see it, this sentence needs to be corrected in the part concerning F:B “Sampling time and forest type significantly interacted to affect total microbial biomass, fungal biomass, and bacterial biomass, but not microbial community composition during PF expansion into CF”.

Response: Thanks for your suggestion. Here, we have retained our original expression because we aim to emphasize that the interaction between sampling time and forest type has no significant impact on soil microbial community structure (F:B, p = 0.084) (Table 4).

Comment 7: The same question applies to the part of the results where this sentence (Lines 205-206): “Temporal variation apparently influenced total microbial biomass, bacterial biomass, and fungal biomass”. There is no mention of microbial community composition. Why?

Response: In this study, we used the F:B ratio to characterize the microbial community composition. The results in Table 3 indicate that sampling time has no significant effect on F:B (p = 0.295).

Comment 8: Lines 231-233. You write: “Soil microbial communities varied significantly across sampling periods, whereas the interactive effects between sampling time and forest type were only notable for F:B ratio”. But according to table 4, significant p-values (F:B ratio) indicators are only in the "Forest type" row. How can this be explained?

Response: Similar to your COMMENT 2, we have carefully checked the data and deleted the incorrect expression. Thank you again for correcting our work.

Comment 9: Section 3.3. Here it is necessary to indicate the correlation coefficients.

Response: Thank you for your suggestion. We have added the correlation coefficients to Section 3.3 as requested.

Comment 10: L 246. bacterial biomass ? Check this.

Response: Thank you for your reminder. We are carefully checked and confirmed that the expression is correct.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Authors,

Reading carefully the new version of "Contrasting Effects of Moso Bamboo Expansion into Broad-leaved and Coniferous Forests on Soil Microbial Communities" please find my following recommendations:

• L157-163: Please cite the reference methods used
• L165/167: PLease avoid over citation (see ref. 33)
• In my opinion the discussion section should be more seriously treated. I recommend for authors a more deepen/critical/comparative interpretation and discussions of their obtained results related to available literature
• Please provide mechanistic insight in discussion considering the obtained experimental results/available literature
• When talking about limitations of current research the authors might consider also space-for-time inference and compositional data constraints also

Author Response

Response to reviewer’s comments

Comment 1:L157-163: Please cite the reference methods used

Response: thanks for your suggestion. We have supplemented the references related to the measurement methods of indicators.

Comment 2: L165/167: Please avoid over citation (see ref. 33)

Response:thanks for your suggestion. We have adjusted the reference to avoid excessive citation.

Comment 3:In my opinion the discussion section should be more seriously treated. I recommend for authors a more deepen/critical/comparative interpretation and discussions of their obtained results related to available literature; Please provide mechanistic insight in discussion considering the obtained experimental results/available literature

Response:as you suggested, we have optimized the discussion section, and we hope the current version is more logical and readable. Details please see L 307-311 and L336-340 in the revised version.

Comment4: When talking about limitations of current research the authors might consider also space-for-time inference and compositional data constraints also

Response: thanks for your suggestion, we have revised the part of the conclusion that addresses the research limitation as follows: “In addition, given the limitations of the sampling sites in this study and the fact that the space-for-time substitution method used to infer the long-term effects of vegetation succession may fail to fully capture the impacts of nonliner changes over time or historical contingencies (such as extreme climate events), it is suggested that further research should be conducted across a larger spatial scale (e.g., different climate zones) by combining long-term fixed-point observation and including more vegetation types, so as to verify the universality of the “forest type-dependent responses” to triggered by Moso bamboo expansion.”