Artificial Cultivation of Aquatic Plants Promotes Nitrogen Transformation and the Abundance of Key Functional Genes in Agricultural Drainage Ditch Sediments in the Yellow River Irrigation Area in China
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe authors compared the nitrogen transformation rates, microorganism diversity, and key functional genes in agricultural drainage ditch affected by artificial aquatic plants and natural aquatic plants. The topic is very interesting. In my opinion, there are still some minor problems in the manuscript that need further revision.
1. Abstract: “Our results revealed that the denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR) and nitrogen fixation rate (NFR) of rhizosphere sediments exhibited a significant increase in artificially cultivated ditch.” (Line 16-18). According to the data in Figure 3, the denitrification rates in artificially cultivated ditch did not increase significantly. Please have a check, and also check the DNRAR and NFR results.
2. Abstract: “……the richness of the bacterial community and the relative abundances of Bacteroidota, Verrucomicrobiota, Firmicutes, Anaeromyxobacter and Geobacter,” (Line 18-20). Not accurate, please have a check, especially for Verrucomicrobiota, Anaeromyxobacter and Geobacter.
3. Abstract: “due to improvements in environmental conditions.” (Line 21-22). Not very clear, can the environmental conditions be specified?
4. The authors proposed two hypotheses in the introduction (Line 151-165). The conclusions need to be echoed. Are all the nitrogen transformation rates and the bacterial community diversity index greater? These two issues need to be further clarified in the conclusion.
5. “Our study revealed that the DRs of A-Phr and N-Phr were significantly greater than those of A-Typ and N-Typ (Figure 3a),” (Line 460-461). Please discuss the effect of aquatic plant species on denitrification function genes.
Author Response
Comments 1: Abstract: “Our results revealed that the denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR) and nitrogen fixation rate (NFR) of rhizosphere sediments exhibited a significant increase in artificially cultivated ditch.” (Line 16-18). According to the data in Figure 3, the denitrification rates in artificially cultivated ditch did not increase significantly. Please have a check, and also check the DNRAR and NFR results.
Response 1:
Thank you for pointing this out. We agree with this comment. Therefore, we carefully examined the results regarding denitrification rate (DR), dissimilated nitrate reduction rate (DNRAR) and nitrogen fixation rate (NFR) and found that the denitrification rate (DR) and DNRA rate (DNRAR) of A-Phr were significantly higher than those of N-Typ (p<0.05). The nitrogen fixation rate (NFR) of A-Typ was significantly higher than that of N-Phr (p<0.05). (Lines 340-348) In addition, no significant differences were noted between other samples obtained from artificial and natural ditches (Table 1).
Therefore, we have revised the statement “Our results revealed that the denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR) and nitrogen fixation rate (NFR) of rhizosphere sediments exhibited a significant increase in artificially cultivated ditch.” to “Our results revealed that the denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR) and nitrogen fixation rate (NFR) of partial rhizosphere sediments significantly increased in artificially cultivated ditch.” (Lines 16-18)
Table 1 Nitrogen transformation rates in rhizosphere sediments
|
Type of ditch |
DR (umol N kg-1h-1) |
DNRAR (umol N kg-1h-1) |
NFR (umol N kg-1h-1) |
|
A-Phr |
3.06±0.067a |
2.14±0.12a |
0.83±0.086ab |
|
A-Typ |
2.50±0.16b |
1.63±0.19b |
0.93±0.19a |
|
N-Phr |
3.08±0.089a |
1.93±0.18ab |
0.64±0.05b |
|
N-Typ |
2.60±0.16b |
1.65±0.26b |
0.73±0.099ab |
Comments 2: Abstract: “……the richness of the bacterial community and the relative abundances of Bacteroidota, Verrucomicrobiota, Firmicutes, Anaeromyxobacter and Geobacter,” (Line 18-20). Not accurate, please have a check, especially for Verrucomicrobiota, Anaeromyxobacter and Geobacter.
Response 2:
We agree with the reviewer. After careful examination of relevant data, it was clear that the Chao1 and ACE richness indices of A-Phr and A-Typ bacterial communities were significantly higher than those of N-Phr and N-Typ (p<0.05). (Lines 353-355) The relative abundances of Bacteroidota, Firmicutes and Geobacter in rhizosphere sediments of A-Phr and A-Typ were significantly higher than those of N-Phr and N-Typ (p<0.05). However, the relative abundance of Verrucomicrobiota in A-Phr rhizosphere sediments was significantly higher than that of N-Phr and N-Typ (p<0.05). In addition, the relative abundance of Anaeromyxobacter in A-Phr and A-Typ rhizosphere sediments was significantly higher than that of N-Typ (p<0.05). (Lines 395-407) (Table 2)
Therefore, we modified the following 3 points:
(1) “Compared with those in natural ditches, the richness of the bacterial community and the relative abundances of Bacteroidota, Verrucomicrobiota, Firmicutes, Anaeromyxobacter and Geobacter,…”
was changed to
“Compared with those in natural ditches, the richness of the bacterial community and the relative abundances of Bacteroidota, Firmicutes and Geobacter…” (Lines 18-20)
(2) “Their relative abundances in the rhizosphere sediments of the artificially cultivated ditch exceeded those of the natural ditches.”
was changed to
“The relative abundances of partial rhizosphere sediment samples in the artificially cultivated ditch significantly exceeded those of the natural ditches.” (Lines 580-582)
(3) “These results confirmed a significantly greater abundance of the nitrogen cycle bacterial phyla Bacteroidota, Verrucomicrobiota, and Firmicutes in both A-Phr and A-Typ than in N-Phr and N-Typ.”
was changed to
“These results confirmed a significantly greater abundance of the nitrogen cycle bacterial phyla Bacteroidota and Firmicutes in both A-Phr and A-Typ than in N-Phr and N-Typ.” (Lines 596-598)
Table 2 Relative abundance at the phylum and the genus level of bacteria (%)
|
Type of ditch |
Bacteroidota |
Verrucomicrobiota |
Firmicutes |
Anaeromyxobacter |
Geobacter |
|
A-Phr |
25.13±1.66a |
9.62±1.17a |
6.69±1.02a |
3.66±0.15a |
2.01±0.09a |
|
A-Typ |
27.82±2.43a |
5.59±0.03c |
7.01±0.08a |
0.95±0.33b |
0.80±0.20b |
|
N-Phr |
16.68±4.90b |
5.33±0.48c |
1.05±0.42c |
3.19±0.54a |
0.14±0.04c |
|
N-Typ |
18.98±2.34b |
7.13±0.94b |
3.67±1.17b |
0.26±0.08c |
0.26±0.05c |
Comments 3: Abstract: “due to improvements in environmental conditions.” (Line 21-22). Not very clear, can the environmental conditions be specified?
Response 3:
Thank you for pointing this out. We agree with this comment. According to Figures 5a, 5b and 5c in the paper, we amended "…due to improvements in environmental conditions." to "… due to improvements in the nitrogen and carbon environments." (Lines 21-22)
Comments 4: The authors proposed two hypotheses in the introduction (Line 151-165). The conclusions need to be echoed. Are all the nitrogen transformation rates and the bacterial community diversity index greater? These two issues need to be further clarified in the conclusion.
Response 4:
We agree with the reviewer. After discussing this issue, we have made the following two modifications.
(1) “A-Phr exhibited a high DR and DNRAR, while A-Typ demonstrated a high rate of nitrogen fixation. The richness indices of the A-Phr and A-Typ bacterial communities, as well as the relative abundances of bacterial phyla (Bacteroidota, Firmicutes) and bacterial genera (Geobacter, Sulfuritalea) involved in nitrogen cycling, were all significantly greater than those of the natural ditches.”
was changed to
“Compared with those in rhizosphere sediments in natural ditches, the nitrogen transformation rates in partial rhizosphere sediments in artificially cultivated ditches are greater. Specifically, the DR and DNRAR of A-Phr were significantly elevated compared to those of N-Typ, and the NFR of A-Typ was notably higher than that of N-Phr. However, no significant differences between other samples from artificially cultivated ditches and natural ditches were observed. The Chao1 and ACE richness indices of bacterial communities in A-Phr and A-Typ were markedly greater than those in N-Phr and N-Typ, whereas the Shannon diversity index showed no significant difference. Furthermore, the relative abundances of Bacteroidota and Firmicutes involved in the nitrogen cycle within A-Phr and A-Typ, as well as Geobacter and Phitalea, were significantly higher than those found in N-Phr and N-Typ. The relative abundance of denitrifying (napA, norB) and nitrate-dissimilating (nrfC, nrfA) genes in A-Phr was notably higher than that observed in both N-Phr and N-Typ, and the relative abundance of nitrogen-fixing genes (nifD, nifK, nifH) in A-Typ was significantly greater than that found in both N-Phr and N-Typ.” (Lines 870-883)
(2) “…leading to an increase in the abundance of functional genes related to denitrification (napA, norB), dissimilatory nitrate reduction (nrfC, nrfA), and nitrogen fixation (nifD, nifK, nifH) …”
was changed to
“These improvements promote an increase in the abundance of partial functional genes related to denitrification, dissimilatory nitrate reduction, and nitrogen fixation.” (Lines 890-892)
Comments 5: “Our study revealed that the DRs of A-Phr and N-Phr were significantly greater than those of A-Typ and N-Typ (Figure 3a),” (Line 460-461). Please discuss the effect of aquatic plant species on denitrification function genes.
Response 5:
Thank you for pointing this out. We agree with this comment. Therefore, we modify the text as follows.
“(1) The microenvironment of the rhizosphere varies among different plant species, leading to differences in the potential denitrification rates and abundance of denitrification functional genes [13]. The cultivation of plants in drainage ditches has been shown to increase the DR [149]. Due to the superior competitive advantage of a single plant community in terms of nitrogen uptake, it diminishes the available nitrate for denitrification. As a result, a multiplant community may not necessarily lead to an increase in the DR [150]. Therefore, the difference in denitrification rates between A-Phr and N-Phr was not statistically significant (Figure 3a). However, the root system of P. australis can secrete more oxygen and sugar exudates than that of T. orientalis [26,89], resulting in higher denitrification rates and abundances of denitrification functional genes in A-Phr and N-Phr than in T. orientalis (Figure 3a, Figure 6a).”
was changed to
“(1) The microenvironment of the rhizosphere varies among different plant species, leading to differences in the potential denitrification rates and abundance of denitrification functional genes (nirK, nirS and nosZ) [13]. Compared to shrub and tree soils, herbaceous soils exhibit a higher diversity of species along with greater carbon and nitrogen content, resulting in elevated abundances of nirS and nirK genes [149]. Significant differences in the abundance of denitrification functional genes nirS and nirK were observed across various aquatic plant species, which showed a notable positive correlation with rhizosphere NH4+-N and NO3--N levels [150]. In this study, the denitrification functional genes napA, nirK, nosZ, and norB in A-Phr were found to be significantly more abundant than those in A-Typ; similarly, N-Phr exhibited significantly higher levels of nirK, nosZ, norB, and nirS compared to N-Typ (Figure 6a). These variations are primarily associated with carbon and nitrogen content as well as dissolved oxygen levels within the rhizosphere environment (Table S4). This phenomenon may be attributed to P. australis roots secreting more oxygen and sugar exudates than those from T. orientalis [26,91], thereby modifying environmental conditions within rhizosphere sediments to create a more conducive habitat for denitrifying bacteria. The cultivation of plants in drainage ditches has been shown to enhance the DR [151]. Due to the superior competitive advantage of a single plant community in terms of nitrogen uptake, it diminishes the available nitrate for denitrification. As a result, a multiplant community may not necessarily lead to an increase in the DR [150]. Therefore, the difference in denitrification rates between A-Phr and N-Phr was not statistically significant (Figure 3a), and the denitrification rate and denitrification functional genes in A-Phr and N-Phr were significantly higher than those of A-Typ and N-Typ (Figure 3a, Figure 6a).” (Line 701-723)
Author Response File: 
Author Response.doc
Reviewer 2 Report
Comments and Suggestions for AuthorsThe article titled “Artificial cultivation of aquatic plants promotes nitrogen transformation and key functional genes in agricultural drainage ditch sediments in the Yellow River irrigation area, China” investigates the nitrogen transformation rates in rhizosphere sediments of artificial planting and natural farmland drainage ditches and their influencing mechanisms. It also shows the diversity and structural composition of bacterial communities and key functional genes of the nitrogen cycle. The study outputs would assist researchers in using aquatic plants to reduce nitrogen pollution in water bodies. Although the study objectives are mentioned, some points should be considered before final acceptance:
1- In Line 33, it’s mentioned “economic development…”; however, no any estimation about the economic performance was mentioned through the paper!!
2- The authors should mention why they have estimated denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR), and nitrogen fixation rate (NFR)
3- Add more details about the materials of the diversity and composition of the bacterial community
44 - The captions of some tables and figures such as “Table 1. Composition of aquatic plant species in agricultural drainage ditches” should be mentioned in details
55 - What is the sensitivity/statistical analysis for the results in Figure 4; what are the symbols “A”, “B”, “AB”, etc.
66 - What is the advantage of the proposed principal component analysis (PCA) or RDA in Figure 5 compared with the other analyses in the literature
77 - The suggested mechanism for enhancing the relative abundance of nitrogen transformation functional genes in rhizosphere sediments should be illustrated
8- What about the real application of the proposed environmental and microbial factors on the nitrogen conversion rate approach
9- The expected drawbacks and limitations of the study should be mentioned
10- What will happen for the presence of extreme environmental conditions (e.g., toxicants) in the rhizosphere soil
Comments on the Quality of English Language
Moderate editing of English language required.
Author Response
Comments 1: In Line 33, it’s mentioned “economic development…”; however, no any estimation about the economic performance was mentioned through the paper!!
Response 1:
Thank you for pointing this out. We agree with this comment. Therefore, we have added the following text.
“Despite the initial investment required for artificial planting of aquatic vegetation in drainage ditches, including costs for procurement, plantation and ongoing management, this measure has the potential to improve the environment of farmland. By returning aquatic plant compost to the field and reducing overall fertilizer input, ecological circular agriculture can be promoted. Furthermore, the landscaping of aquatic plants can stimulate local economic development through the advancement of rural tourism and by increasing the added value of ecological agricultural products.” (Lines 858-865)
Comments 2: The authors should mention why they have estimated denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR), and nitrogen fixation rate (NFR)
Response 2: We agree with the reviewer. We have made the following changes.
“The Yellow River irrigation area in Ningxia is located in the arid region of the middle temperate zone in Northwest China and is characterized by unique climate, soil and hydrological conditions. This region experiences ample sunshine, significant temperature fluctuations, drought, minimal precipitation, and high evaporation rates. Rivers and lakes in this region exhibit elevated salinity levels, while the ecological attributes of aquatic plants are distinguished by their distinct growth patterns and nutrient absorption mechanisms.”
was changed to
“The Yellow River irrigation area in Ningxia is located in the arid region of the middle temperate zone in Northwest China and is characterized by unique climate, soil, and hydrological environment conditions and aquatic plants with distinctive ecological features. Owing to ample sunshine, significant temperature fluctuations, drought, minimal precipitation, and high evaporation rates, the rivers and ditches in this region generally exhibit elevated salinity levels. According to previous studies [44,45] and the preliminary findings of our research group, nitrification and ammonia oxidation are inhibited in anaerobic sediment environments with salt accumulation and ammonium deficiency. Consequently, our focus lies on investigating the DR, DNRAR, and NFR.” (Lines 137-145)
Comments 3: Add more details about the materials of the diversity and composition of the bacterial community
Response 3:
Thank you for pointing this out. We agree with this comment. Therefore, we have added the following text.
“The full-length bacterial 16S rRNA gene was amplified using primers 27F_(16S-F) (5'-AGRGTTTGATYNTGGCTCAG-3') and 1492R_(16S-R) (5'-TASGGHTACCTTGTTASGA CTT-3') [52] by a PCR instrument (Veriti96well 9902, ABI, USA). After amplification, the PCR products were recovered through 2% agarose gel electrophoresis, purified, detected, and quantified. Subsequently, library construction was performed using the SMRTbell Template Prep Kit. Following the standard protocol of Biomarker Technologies (Beijing, China), sequencing of the samples was conducted on the PacBio Sequel II platform. Usearch software was used to cluster reads at a similarity level of 97.0% and obtain OTUs. Sample alpha diversity indices were evaluated using QIIME2 2020.6 software [53]. Taxonomic annotation was performed using the Silva 138 database (Release 138, https://www.arb-silva.de/documentation/release-138/) [54].”
was changed to
“The DNA concentration was detected by adding 1X dsDNA HS Working Solution (Yisheng Biotechnology (Shanghai) Co., LTD.) using an enzyme-labelling apparatus (Synergy HTX, Gene Company Limited). The full-length bacterial 16S rRNA gene was amplified using primers 27F_(16S-F) (5'-AGRGTTTGATYNTGGCTCAG-3') and 1492R_(16S-R) (5'-TASGGHTACCTTGTTASGA CTT-3') [54] with a PCR instrument (Veriti 96-well 9902, ABI, USA). After amplification, the PCR products were analyzed by electrophoresis with a 2% agarose gel, quantified with a LabChip, and then mixed. Damaged repair and terminal repair along with attachment of the mixed products were performed using the SMRTbell Template Prep Kit. AMpure PB magnetic beads were used to purify and recover the library. The final library passed the concentration requirement (Qubit). A polymerase kit was used to bind the library to the primer and polymerase, and the final reaction products were purified using cleanup beads. Sequencing was performed using the PacBio Sequel II platform. CCS sequences were extracted from the original data, and lima (v1.7.0) software was used to perform barcode identification, length filtering, and chimaera removal from the CCS sequences to obtain effective CCS sequences. The effective CCS sequences were classified into OTUs/ASVs (later uniformly called based on features) by clustering/de-noising. Using Silva 138 (Release 138, https://www.arb-silva.de/documentation/release-138/) [55] as the reference database, the feature sequences were annotated for taxonomy using a naive Bayesian classifier combined with comparison methods. The Alpha diversity index of samples was evaluated using QIIME2 2020.6 software [56].” (Lines 289-309)
Comments 4: The captions of some tables and figures such as “Table 1. Composition of aquatic plant species in agricultural drainage ditches” should be mentioned in details
Response 4: We agree with the reviewer. We have added the following text to the footnote of Table 1.
“The dominant species in the artificial planting ditch are Phragmites australis, Typha orientalis, Nymphaea tetragona, and occasionally Echinochloa crusgalli. The dominant species in natural ditch 1 is Phragmites australis. Associated species include Scirpus triqueter and Echinochloa crusgalli, and Lemna minor is occasionally observed. The dominant species in natural ditch 2 is Typha orientalis. Associated species include Phragmites australis and Scirpus triqueter, and Lemna minor is occasionally observed.” (Lines 204-209)
Comments 5: What is the sensitivity/statistical analysis for the results in Figure 4; what are the symbols “A”, “B”, “AB”, etc.
Response 5:
Thank you for pointing this out. We agree with this comment. Therefore, we have added “The different capital letters indicate significant differences among the different rhizosphere sediment samples (p<0.05) according to one-way ANOVA and LSD.” to the legends of Figure 3. (Lines 344-346)
We have added “The different capital/lowercase letters indicate significant differences among the different rhizosphere sediment samples (p<0.05) according to one-way ANOVA and LSD.” to the legend of Figure 4. (Lines 363-365)
Comments 6: What is the advantage of the proposed principal component analysis (PCA) or RDA in Figure 5 compared with the other analyses in the literature
Response 6: Thank you for pointing this out. By integrating principal component analysis with regression analysis, redundancy analysis (RDA) is employed to investigate the relationship between nitrogen conversion rates and various microbial and environmental factors in Figure 5. This approach facilitates a deeper understanding of how environmental variables influence the diversity and structural composition of bacterial communities. Additionally, it generates principal coordinate diagrams that visually represent both bacterial communities and their associated environmental variables, thereby simplifying the interpretation of complex ecological data. Ultimately, this method aids in identifying specific environmental and microbial factors correlated with nitrogen conversion rates.
Comments 7: The suggested mechanism for enhancing the relative abundance of nitrogen transformation functional genes in rhizosphere sediments should be illustrated
Comments 8: What about the real application of the proposed environmental and microbial factors on the nitrogen conversion rate approach
Responses 7 and 8:
We agree with the reviewer. Based on the combined information from comments 7 and 8, we added the following text to the Discussion.
“It is recommended to desilt gutters at regular intervals [183] and plant saline-tolerant plants with strong nitrogen absorption capacity [13]. These methods can enhance the rhizosphere effect; improve DO and ORP and carbon and nitrogen levels in rhizosphere sediments; and promote the expression of nitrogen conversion functional genes. As a result, these practices can increase the rate of nitrogen conversion. In addition, the application of microbial inoculants is also an effective measure to improve the nitrogen conversion rate of sediments. It is important to further study how different microbial inoculants change the microbial communities in sediments, thereby influencing the nitrogen conversion rate.” (Lines 845-853)
Comments 9: The expected drawbacks and limitations of the study should be mentioned
Response 9:
Thank you for pointing this out. We agree with this comment. Therefore, we add the following text.
“Given that samples collected for this study were obtained at a single time point, they only represent sediment nitrogen conversion rates and bacterial community structures during that specific period, thus presenting certain limitations. Subsequently, we should further study the differences and influencing factors of nitrogen conversion rates in rhizosphere sediments in different seasons.” (Lines 841-845)
Comments 10: What will happen for the presence of extreme environmental conditions (e.g., toxicants) in the rhizosphere soil
Response 10: We agree with the reviewer. We add the following information to the manuscript.
“The issue of farmland salinization within the Yellow River irrigation area is becoming increasingly severe. An accumulation of salts in drainage ditch sediments can diminish permeability, water retention capacity, and nutrient content, consequently inhibiting plant growth while adversely impacting microbial composition, diversity, and functionality [184]. Future research should focus on screening salt-tolerant plant growth-promoting rhizobia.” (Lines 853-858)
Author Response File: Author Response.doc
Round 2
Reviewer 2 Report
Comments and Suggestions for AuthorsThe authors responses are satisfactory
Comments on the Quality of English Language
Minor editing of English language required.
Author Response
Comments 1: - In the abstract, quickly describe your aim and experimental setting and plants. I suggest to add a sentence in between line 15-16 like: “... is a crucial strategy for mitigating regional water eutrophication. We here compare key processes of nitrogen transformations occurring in the rhizophere of sediments of a ditch artificially planted with a mix of species (Phragmites australis, Typha orientalis, Nymphaea tetragon) with the rhizosphere of ditch occupied by naturally occurring aquatic vegetation, dominated either by P. australis or T. orientalis. Our results revealed that the denitrification….”
Response 1:
Thank you for pointing this out. We agree with this comment. Therefore, we have added the following text.
“We here compare key processes of nitrogen transformations occurring in the rhizophere of sediments of a ditch artificially planted with a mix of species (Phragmites australis, Typha orientalis, Nymphaea tetragon) with the rhizosphere of ditch occupied by naturally occurring aquatic vegetation, dominated either by P. australis or T. orientalis.” (Lines 16-19)
Comments 2: - In the abstract, lines16-17, but also later lines 580, 871..: you write “… and nitrogen fixation rate (NFR) of partial rhizosphere sediments significantly increased…”. For me, “partial rhizosphere sediments” is not understandable. From fig.3 you show that N-processes are species dependent (as in fig 5 d,e). So may be you can rephrase as “Our results revealed a species effect with an increased denitrification rate (DR) and dissimilatory nitrate reduction to ammonium rate (DNRAR) in the cultivated ditch for P. australis, compared to the naturally occurring T. orientalis vegetation. Nitrogen fixation rate (NFR) increased in the artificial setting with T. orientalis in comparison to natural P. australis vegetation.”
Response 2:
We agree with the reviewer. Therefore, we modified the following 2 points:
(1) “Our results revealed that the denitrification rate (DR), dissimilatory nitrate reduction to ammonium rate (DNRAR) and nitrogen fixation rate (NFR) of partial rhizosphere sediments significantly increased in artificially cultivated ditch. ”
was changed to
“Our results revealed a species effect with an increased denitrification rate (DR) and dissimilatory nitrate reduction to ammonium rate (DNRAR) in the cultivated ditch for P. australis, compared to the naturally occurring T. orientalis vegetation. Nitrogen fixation rate (NFR) increased in the artificial setting with T. orientalis in comparison to natural P. australis vegetation.” (Lines 19-23)
(2) “Compared with those in rhizosphere sediments in natural ditches, the nitrogen transformation rates in partial rhizosphere sediments in artificially cultivated ditches are greater. Specifically, the DR and DNRAR of A-Phr were significantly elevated compared to those of N-Typ, and the NFR of A-Typ was notably higher than that of N-Phr. However, no significant differences between other samples from artificially cultivated ditches and natural ditches were observed.”
was changed to
“A species effect was observed, characterized by an increased DR and DNRAR in the cultivated ditch of P. australis compared to the naturally occurring vegetation of T. orientalis. Additionally, the NFR was found to be elevated in the artificial setting with T. orientalis relative to natural P. australis vegetation.” (Lines 873-876)
Comments 3: - Abstract L18-22, this sentence may be simplified to “…the relative abundances of Bacteroidota, Firmicutes and Geobacter were significantly greater in the rhizosphere of artificially cultivated ditch due a greater availability in nitrogen and organic carbon.”
Response 3:
Thank you for pointing this out. We agree with this comment. Therefore, we modify the text as follows.
“Compared with those in natural ditches, the richness of the bacterial community and the relative abundances of Bacteroidota, Firmicutes and Geobacter, which are involved primarily in the nitrogen cycle, in the rhizosphere sediments of artificially cultivated ditch were significantly greater due to improvements in the nitrogen and carbon environments.”
was changed to
“The richness of the bacterial community and the relative abundances of Bacteroidota, Firmicutes and Geobacter were significantly greater in the rhizosphere of artificially cultivated ditch due a greater availability in nitrogen and organic carbon.” (Lines 23-25)
Comments 4: - In conclusion, L890: “… in the abundance of partial functional gene…”: ?? meaning? May be to rephrase as “.. an increase in the abundance of functional genes, depending on species, related to denitrification…”
Response 4:
We agree with the reviewer. After discussing this issue, we have made the following modifications.
“These improvements promote an increase in the abundance of partial functional genes related to denitrification, dissimilatory nitrate reduction, and nitrogen fixation.”
was changed to
“These improvements promote an increase in the abundance of functional genes related to denitrification, dissimilatory nitrate reduction, and nitrogen fixation, contingent upon species variation.” (Lines 891-893)
Author Response File: Author Response.doc
