Examining the Diversity of Rhizosphere Soil Bacterial Communities and Screening of Growth-Promoting Bacteria from the Rhizosphere Soil of Haloxylon ammodendron in Xinjiang
María Lorena Castrillo
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe manuscript is ok and easy to read.
Author Response
We appreciate for Reviewers’ warm work earnestly, and hope that the correction will meet with approval.
Once again, thank you very much for your comments and suggestions
Reviewer 2 Report
Comments and Suggestions for AuthorsThe manuscript. 'Study on Diversity of Rhizosphere Soil Microbial Community and Screening of Growth promoting Bacteria from Rhizosphere Soil of Haloxylon ammodendron in Xinjiang' is an interesting work focusing on microbial diversity in rhizosphere. Few points need to be considered before the acceptance for publication.
1. The material and method section needs to be revised to elaborate the methodology. For example - The authors have not explained DNA extraction methods before DNA sequencing.
2. References are missing in the methodology. Authors should consider adding references to the methods adopted.
3. The referencing style is not the same throughout the manuscript. For eg. In paragraph 3.1 reference style is different for two references quoted.
4. Certain topographical errors need to be taken care of. For eg. Line 246
Author Response
Please see the attachment.
Author Response File: 
Author Response.pdf
Reviewer 3 Report
Comments and Suggestions for AuthorsOverall, the article provides valuable insights into the diversity of rhizosphere soil microbial communities associated with Haloxylon ammodendron and the screening of growth-promoting bacteria. However, there are several areas where modifications could enhance the quality and clarity of the manuscript:
Abstract:
The abstract provides a concise overview of the study; however, it could be improved by including specific findings or conclusions. For example, highlighting the key results obtained from high-throughput sequencing and growth-promoting bacteria screening.
Introduction:
The introduction effectively outlines the importance of studying rhizosphere microbial communities for plant health and growth, but it could be more focused. Consider streamlining the introduction to clearly state the research objectives and gap in knowledge that the study aims to address.
Materials and Methods:
Provide more detailed descriptions of the methods used, including specific reagents, equipment, and procedures.
Clarify the rationale behind the selection of specific methods, such as why particular media were chosen for bacterial screening assays.
Results and Analysis:
Organize the results section logically, following the sequence of methods presented in the materials and methods section.
Provide more context for the results presented in tables and figures, including interpretation and implications.
Clarify any abbreviations or acronyms used in tables and figures to improve readability.
Discussion:
The discussion should focus on interpreting the results in the context of existing literature and research objectives.
Provide a clear synthesis of the findings and their implications for understanding the role of rhizosphere microbial communities in plant growth and soil health.
Consider discussing the limitations of the study and avenues for future research to address remaining questions or uncertainties.
Comments on the Quality of English LanguageEnsure consistency in terminology and formatting throughout the manuscript.
Proofread the manuscript for grammar, spelling, and punctuation errors to improve readability and professionalism.
Author Response
Please see the attachment.
Author Response File: Author Response.pdf
Reviewer 4 Report
Comments and Suggestions for AuthorsGeneral comments
The authors analyzed the bacterial community in the rhizosphere of Haloxylon ammodendron using next-generation sequencing and isolated several bacteria. Several characteristics of the isolated strains were then investigated, including their phosphate solubility capacity.
Although the study contains information on the rhizosphere bacteria of Haloxylon ammodendron, the purpose of the study and the novelty of the results obtained are not clear. The experimental methods and interpretation of the results are inadequate in some areas. In addition, the manuscript contains numerous formatting errors. Hence, It must be said that this manuscript is incomplete.
Major comments
1. There are papers about bacteria in the rhizosphere of Haloxylon as cited by the authors. An overview of those papers and a description of the remaining issues are needed. Based on them, the objectives of this study should be established and new findings from this study should be discussed.
2. It is stated that 10 samples were collected from the rhizosphere of 2-year and 5-year grown Haloxylon(Line 82-84), but the results (Line158-212) show only one data set for 2-year and 5-year. Nevertheless, it discusses "significant differences" (Line301). On the other hand, Fig. 5 has more than 10 plots for 2-year and 5-year, respectively. It also does not state how the endophytic (Line131 and 223) and rhizosphere bacteria were separated. It is not stated how the 14 isolates were selected. In addition, 14 isolates are insufficient for discussion. It is unclear what kind of experiments and analyses were conducted, and it is difficult to determine whether the experiments, analyses, and discussions were properly conducted.
3. Many formatting errors are present in the manuscript. For example, unnecessary spaces (Lines 15 and 26), missing spaces (Lines 35 and 101, etc.), italics not converted (Lines 79 and 83, etc.), missing manufacturer's description (Lines 118-120), missing text (Lines 123-124), missing figure (Lines 250-251), and uncommon article structure (Lines 157 and 258).
4. Cited references are not listed in many areas where citations are required. For example, medium composition (Line104-107), Primers (Line117), screening methods (Line129-156). The reference for Chenhua Li et al. (Line64-65) is not present in the literature list.
5. Line259 and 282-283: Actinomycetes is one of the taxonomic group of bacteria. Plus, Fungi do not appear to have been investigated in this study.
Author Response
Please see the attachment.
Author Response File: Author Response.pdf
Round 2
Reviewer 2 Report
Comments and Suggestions for AuthorsThe authors have revised the manuscript nicely.
Author Response
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Response to Reviewer two Comments |
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1. Summary |
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
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2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
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Does the introduction provide sufficient background and include all relevant references? |
Yes/Can be improved/Must be improved/Not applicable |
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Are all the cited references relevant to the research? |
Yes/Can be improved/Must be improved/Not applicable |
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Is the research design appropriate? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the methods adequately described? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the results clearly presented? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the conclusions supported by the results? |
Yes/Can be improved/Must be improved/Not applicable |
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[我们真诚地感谢审稿人的热情工作，并希望更正能得到批准。 再次感谢您的意见和建议] |
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Author Response File: 
Author Response.docx
Reviewer 3 Report
Comments and Suggestions for AuthorsI am pleased to report that the authors have diligently addressed all the concerns and queries I raised during the review process. Their revisions demonstrate a thorough understanding of the feedback provided and have significantly strengthened the manuscript.
Based on my assessment, I believe the manuscript is now suitable for publication pending any final editorial checks. I commend the authors for their responsiveness and commitment to improving the quality of their work.
Author Response
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Response to Reviewer three Comments |
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1. Summary |
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
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2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
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Does the introduction provide sufficient background and include all relevant references? |
Yes/Can be improved/Must be improved/Not applicable |
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Are all the cited references relevant to the research? |
Yes/Can be improved/Must be improved/Not applicable |
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Is the research design appropriate? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the methods adequately described? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the results clearly presented? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the conclusions supported by the results? |
Yes/Can be improved/Must be improved/Not applicable |
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[We appreciate for Reviewers’ warm work earnestly, and hope that the correction will meet with approval. 再次感谢您的意见和建议] |
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Author Response File: Author Response.docx
Reviewer 4 Report
Comments and Suggestions for AuthorsThe authors have revised the manuscript in response to the reviewers' comments. However, the manuscript is not appropriate as a scientific paper because of the following points.
1. No experiments, analyses, or discussions were conducted that lead to the conclusions (L50-L52) described in the manuscript
2. The purpose and novelty of this study are still unclear.
3. Since it is submitted as a Brief Report, redundant information should be put in the body of the text or moved to the SI. For example, Table1, 2, 3, and 4, Figure1.
4. Figures 1, 2, 3 are too small to grasp the content.
5. It is not clear whether the difference at the phylum level between year 2 and year 5 (L256-261) is a significant difference or within variation. Nevertheless, it is discussed as if there was a difference (L434-437).
6. The characteristics possessed by some bacteria are described in an expanded interpretation of the characteristics of that taxon. Not all bacteria of the phylum Proteobacteria have nitrogen-fixing ability. (L437-439)
7. The description is unclear or inappropriate.
L244, The word "gate" could be a mistake for "phylum."
L347-349, The sentence is incomplete.
L376, It is unclear what "the azotobacter medium" indicates, as it appears for the first time.
L410, It is unclear to which bacteria the "antagonistic bacteria" refers in the results.
L418-419, It is unclear what " Cistanciola deserticola” indicates, as it appears for the first time.
8. The content of the discussion chapter is not appropriate for the discussion of this study. The contents are not described in the results (L445-447, L449-450, L468-470). The contents are contradictory to the contents described in the results (L326-330 and L455-457). Only references to the cited literature are introduced, but their relevance to this study is not described (L482-501).
Sorry if I have offended you with my many negative comments.
Author Response
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Response to Reviewer Four Comments |
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1. Summary |
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Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding revisions/corrections highlighted/in track changes in the re-submitted files
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2. Questions for General Evaluation |
Reviewer’s Evaluation |
Response and Revisions |
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Does the introduction provide sufficient background and include all relevant references? |
Yes/Can be improved/Must be improved/Not applicable |
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Are all the cited references relevant to the research? |
Yes/Can be improved/Must be improved/Not applicable |
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Is the research design appropriate? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the methods adequately described? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the results clearly presented? |
Yes/Can be improved/Must be improved/Not applicable |
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Are the conclusions supported by the results? |
Yes/Can be improved/Must be improved/Not applicable |
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3. Point-by-point response to Comments and Suggestions for Authors |
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Comments 1: [No experiments, analyses, or discussions were conducted that lead to the conclusions (L50-L52) described in the manuscript] |
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Response 1: [ Discussion Soil microorganisms are the key drivers of plant productivity in the terrestrial ecosystem. Therefore, we have studied and analyzed theCistanche deserticola-Haloxylon ammodendron bacterial community structure of rhizosphere soil bacteria and the screening of growth-promoting bacteria. This study is of great significance for the influence of rhizosphere bacteria on soil nutrient release, improvement of cistanche yield and quality, and access to nutrient elements [32]. ] |
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Comments 2: [he purpose and novelty of this study are still unclear.] |
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Response 2: [Rhizosphere soil bacterial microbial community structure is very important for plant growth and development, and it interacts and promotes each other with plant roots. However, the composition characteristics of bacterial microbial community in rhizosphere soil of 2-year and 5-year Haloxylon ammodendron have not been clearly defined. The purpose of this study was to determine the microbial in the rhizosphere soil bacteria were analyzed by isolating, purifying and identifying the dominant microorganisms combined with high-throughput sequencing technology in order to identify the characteristics of microbial diversity in the rhizosphere soil bacteria of 2-year and 5-year Haloxylon ammodendron in Turpan, Xinjiang.]
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Comments 3:[Since it is submitted as a Brief Report, redundant information should be put in the body of the text or moved to the SI. For example, Table1, 2, 3, and 4, Figure1.] |
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Response 3: [Tables 1, 2, 3 and 4 have been placed in S1] |
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Comments 4:[Figures 1, 2, 3 are too small to grasp the content.] |
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Response 4: [Figures 1, 2, and 3 have been enlarged] |
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Comments 5: [It is not clear whether the difference at the phylum level between year 2 and year 5 (L256-261) is a significant difference or within variation. Nevertheless, it is discussed as if there was a difference (L434-437).] |
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Response 5: [The results of high-throughput amplification sequencing showed that the rhizosphere soil bacteria belonged to 45 phyla, 109 classes, 288 orders, 451 families, 826 genera and 404 species. The rhizosphere bacteria belonging to 56 bacterial phyla, 148 classes, 369 orders, 601 families, 1062 genera and 671 species distributed in the rhizosphere soil in 5 years (Table 5). The dominant bacterial phyla are Firmicutes, Proteobacteria, Actinobacteriota, Bacteroidota, Crenarchaeota and Patesc ibacteria, Chloroflexi, Gemmatimonadota, Myxococcota, Acidobacteriota, etc., The relative abundance of bacteria in rhizosphere soil was 31.59%, 36.24%, 3.67%, 5.45%, 0.81%, 3.01%, 4.21%, 4.12%, 1.41% and 2.26%, respectively. The relative abundance of bacteria in rhizosphere soil in 5 years was 27.73%, 33.36%, 13.61%, 7.16%, 2.57%, 2.01%, 3.51%, 1.47%, 1.33%, 1.69%, respectively. There was no significant difference between 2 and 5 years at door level.The sum of relative abundance of Firmicutes, Proteobacteria, Actinobacteriota and Bacteroidota is over 70%. Firmicutes and Proteobacteria decreased with the increase of tree age, while Bacteroidota decreased with the increase of tree age. Actinobacteriota, Bacteroidota and Crenarchaeota increased with the increase of tree age (Figure 2). Discussion It was found that the relative abundance of Firmicutes, Proteobacteria and Patescibacteria in two years was higher than that in five years. The relative abundance of 5-year old Actinobacteriota, Bacteroidota and Crenarchaeota was higher than that of 2-year old Halocardia. However, the relative abundance difference is between 1%-10%, which is not significant. ] |
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Comments 6: [It is not clear whether the difference at the phylum level between year 2 and year 5 (L256-261) is a significant difference or within variation. Nevertheless, it is discussed as if there was a difference (L434-437).] |
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Response 6: [Studies have shown that Proteobacteria are widely distributed, Rhizobium sp, Shinella zoogloeoides, Pseudomonas fluorescens, Pseudomonas extremaustralis, Sphingomonas paucimobilis, Stenotrophomonas sp, have strong nitrogen fixation ability, and have great variable morphological and physiological characteristics, which gives them a great competitive advantage in the ecological niche [36]. ] |
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Comments 7: [The description is unclear or inappropriate. L244, The word "gate" could be a mistake for "phylum." L347-349, The sentence is incomplete. L376, It is unclear what "the azotobacter medium" indicates, as it appears for the first time. L410, It is unclear to which bacteria the "antagonistic bacteria" refers in the results. L418-419, It is unclear what " Cistanciola deserticola” indicates, as it appears for the first time.] |
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Response 7: [ (1)Comparative analysis of Phylum level (2)Among them, sg21 and Bacillus sp reached 100%, sg1 and Bacillus sp reached 100%, and sg9F and Streptomyces levis reached 100%. sg16 and Phyllobacterium phragmitis have formed a cluster with 99.85% similarity, and ss4 has formed a cluster with Sinorhizobium meliloti with 99.75% similarity (Figure 6). (3)Ashby medium (4)Therefore, we studied and analyzed the bacterial microbial community structure in the rhizosphere soil as well as the screening of growth promotion bacteria. (5)Cistanche deserticola] |
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Comments 8: [The content of the discussion chapter is not appropriate for the discussion of this study. The contents are not described in the results (L445-447, L449-450, L468-470). The contents are contradictory to the contents described in the results (L326-330 and L455-457). Only references to the cited literature are introduced, but their relevance to this study is not described (L482-501).] |
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Response 8: [ (1) Discussion Soil microorganisms are the key drivers of plant productivity in the terrestrial ecosystem. Therefore, we have studied and analyzed theCistanche deserticola-Haloxylon ammodendron bacterial community structure of rhizosphere soil bacteria and the screening of growth-promoting bacteria. This study is of great significance for the influence of rhizosphere bacteria on soil nutrient release, improvement of cistanche yield and quality, and access to nutrient elements [32]. Therefore, we studied and analyzed the bacterial microbial community structure in the rhizosphere soil as well as the screening of growth promotion bacteria. The number of bacteria in plant rhizosphere microorganisms is the largest, and the types of soil microorganisms will also change when the external environment changes [33,34]. In this study, the variation of bacterial microbial diversity in rhizosphere soil of cistanticola deserticola in Gaochang District of Turpan City, Xinjiang Province was studied for the first time by using high-throughput amplification sequencing. The dominant rhizosphere soil bacteria in 2 and 5 years were Firmicutes, Proteobacteria, Actinobacteriota and Bacteroidota, and the total relative abundance of these four groups was over 70%. This is consistent with the research results of Wang Anlin, who showed that Proteobacteria, actinomycetes, cyanobacteria and chloromycetes accounted for 76.05% of the relative abundance of soil bacteria in the forest, belonging to the dominant bacteria in soil [35]. It was found that the relative abundance of Firmicutes, Proteobacteria and Patescibacteria in two years was higher than that in five years. The relative abundance of 5-year old Actinobacteriota, Bacteroidota and Crenarchaeota is higher than that of 2-year old Halocarpacta, but the relative abundance difference is between 1%-10%, and the difference is not significant. Studies have shown that Proteobacteria is widely distributed. Among them, Rhizobium sp., Shinella zoogloeoides, Pseudomonas fluorescens and Pseudomonas psychrophile extremaustralis, Sphingomonas paucimobilis, Stenotrophomonas sp) and others have strong nitrogen fixation ability and great morphological and physiological characteristics, which makes them have great competitive advantages in the ecological niche [36]. In addition, the relative abundance of actinomycetes in the rhizosphere soil of Haloxylon is significantly higher, because actinomycetes are Gram-positive bacteria, which not only have strong drought resistance, but also participate in the complex decomposition process of organic matter, providing favorable conditions for the growth and development of plants [37]. Chloroflexi is a phototrophic microorganism that is mainly involved in microbial photosynthesis [38]. In this study, The relative abundance of Pseudomonas, Bacillus and Lactobacillus in the 2-year old species was higher than that in the 5-year old species. The 5-year-old Acinetobacter, Nitrosopumilaceae and Ralstonia genera, in which Pseudomonas and Bacillus are basophilic bacteria. The living environment of basophilic microorganisms is alkaline soil, carbonate lake, high pH value environment[39].Studies have shown that the number of microorganisms in rhizosphere soil of Haloxylon ammodendron in different growth years are different to varying degrees, which is related to the species and content of root exudates of Haloxylon ammodendron growing under different environmental conditions [40]. In this study, there were differences between the rhizosphere bacterial communities of 2-year old and 5-year old Haloxylon haloxylon. There were 10724 unique OTUs in the rhizosphere soil of 5-year old Haloxylon ammodendron, and 3329 OTUs in the rhizosphere soil of different tree ages, accounting for 16.98% of the total number of OTUS. It can be seen that the microbial species in the rhizosphere soil of 5-year old Haloxylon ammodendron were significantly higher than those in the rhizosphere soil of 2-year old Haloxylon ammodendron. Different years of Haloxylon ammodendron can also change the underground rhizosphere flora, affecting the scale of soil rhizosphere effect and special rhizosphere microorganisms. This study also concluded that haloxylon of different years affected the rhizosphere effect and changed the number of microorganisms [41]. The number of inorganic phosphorus bacteria, iron-bearing bacteria and azotobacter increased in the rhizosphere soil of Haloxylon ammodendron, while the number of potassium solubilizing bacteria decreased in the rhizosphere soil. Organophosphorus bacteria decreased in rhizosphere soil. The number of inorganic phosphorus bacteria, iron-bearing bacteria and azotobacter increased in the rhizosphere soil of Haloxylon ammodendron, while the number of potassium solubilizing bacteria decreased in the rhizosphere soil. Organophosphorus bacteria decreased in rhizosphere soil. The inorganic phosphorus bacteria in this study belong to Phyllobacterium phragmitis, Bacillus, and Sinorhizobium meliloti respectively. There have been many studies on the growth promoting effect of Bacillus on plants [42,43]. The isolated bacillus sg16 has the dual characteristics of phosphorus solubilizing and nitrogen fixation. The synthesis and release of low molecular weight organic acids by Bacillus can promote the dissolution of inorganic phosphorus in soil. The hydroxyl and carboxyl groups in organic acids chelate phosphates to bind cations and ultimately convert phosphates into soluble forms [44]. In this study, the microbial community structure diversity of the rhizosphere soil bacteria in the 2-year and 5-year rhizosphere soil in Turpan, Xinjiang was analyzed. Through the isolation, purification and identification of the dominant microorganisms in the roots of Haloxylon ammodendron, and combined with high-throughput sequencing technology, the main microbial groups and diversity of the rhizosphere soil of Haloxylon ammodendron were analyzed. There are still shortcomings in the process of this experiment. This study studied the microbial community structure diversity of rhizosphere soil bacteria in 2 and 5 years, but did not study the bacterial diversity in different growth periods, different ecological niches and more different years of Haloxylon ammodendron. The IAA production ability of the isolated strains was not tested, and the IAA production ability can be tested later, and the bacterial diversity of different growth periods, different ecological niches and more different years can be studied. (2) Composition of rhizosphere soil bacterial community of Haloxylon ammodendron, The dominant bacterial genera are Pseudomonas, Bacillus, Acinetobacter, Nitrosopumilaceae, Lactobacillus and Ralstonia and Marinobacter, Viridibacillus, Pseudarthrobacter, Achromobacter, etc. The relative abundance of bacteria in rhizosphere soil of Haloxylon ammodendron in 2 years was 9.32%, 11.68%, 2.71%, 0.24%, 2.71%, 1.34%, 0.94%, 1.64%, 0.04%, 1.70%, respectively. The relative abundance of bacteria in the rhizosphere soil in 5 years was 1.16%, 10.18%, 2.74%, 1.71%, 1.47%, 1.51%, 1.26%, 1.61%, 2.61%, 2.49%, respectively (Figure 3).The relative abundance of Pseudomonas, Bacillus and Lactobacillus in the 2-year old species was higher than that in the 5-year old species. Acinetobacter, Nitrosopumilaceae , Ralstonia were higher in 5 years than in 2 years. (3) L449-450、L468-470 has been deleted, L326-330 and L455-457 have been modified It was found that the relative abundance of Firmicutes, Proteobacteria and Patescibacteria in two years was higher than that in five years. The relative abundance of 5-year old Actinobacteriota, Bacteroidota and Crenarchaeota was higher than that of 2-year old Halocardia. However, the relative abundance difference is between 1%-10%, (3) L482-501has been deleted] |
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[We appreciate for Reviewers’ warm work earnestly, and hope that the correction will meet with approval. Once again, thank you very much for your comments and suggestions] |
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