Effects of Long-Term Fertilization on Soil Physical and Chemical Properties of “Dangshansuli” Pear Orchard

Round 1

Reviewer 1 Report (Previous Reviewer 1)

Comments and Suggestions for Authors

I find the manuscript significantly improved now thanks to suggestions from previous round of reviews. 

Author Response

Please check the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report (Previous Reviewer 2)

Comments and Suggestions for Authors

Dear author,
I find the changes made to the manuscript satisfactory.

Author Response

Please see the attached document.

Author Response File: Author Response.pdf

Reviewer 3 Report (New Reviewer)

Comments and Suggestions for Authors

In this study, the authors evaluate the impacts of NPK fertilization on soil properties in a pear orchard after 30 years of application. By analyzing the physicochemical properties of the soil, mineral and heavy metal contents across five soil depths, and the allelopathic effects, the authors concluded that mineral fertilization contributes to significant changes in soil properties.

Overall, the manuscript is logically organized. The language is quite fluent, but some sentences include awkward expressions. Many grammatical or typographical errors also occur. Thus, it would be necessary to check the language throughout. Moreover, there are several points that need to be clarified to improve the manuscript. Please see the attached file.

Comments for author File: Comments.pdf

Author Response

Please see the attached document.

Author Response File: Author Response.pdf

Round 2

Reviewer 3 Report (New Reviewer)

Comments and Suggestions for Authors

I have carefully reviewed the authors’ responses and the revised version of the manuscript, and I am very satisfied. The Authors have successfully addressed my concerns in this resubmitted version of the manuscript. As such, I consider the manuscript suitable for publication.

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Thee paper is easy to be read and understand but there are several week point.

The authors used terms like "soil structure" and "physical" although no such characteristics were presented in the paper.

It is not clear were soil samples taken from pear orchard or genotypes collection? Commercial orchard and genotypes collection are not bee managed in the same way.

The final conclusion about usage of organic fertilizers is not supported by the results. The results show increase in soil fertility, but no linkage to usage of organic fertilizers is shown.

There are some individual comments and corrections inserted in the file.

Comments for author File: Comments.pdf

Author Response

Response to reviewers

Reviewer 1:

(1) The authors used terms like "soil structure" and "physical" although no such characteristics were presented in the paper.

Response: Thank you for your kind reminder. We have deleted this description in the revised manuscript.

• It is not clear were soil samples taken from pear orchard or genotypes collection? Commercial orchard and genotypes collection are not bee managed in the same way.

Response: Thank you for your kind reminder. The samples were taken from “Dangshan” pear orchard. We added this information in material and methods. Soil sampling was collected in May 2020. The treatment group consisted of a “Dangshansuli” pear orchard that was subjected to Long-term fertilization (Fertilization was primarily based on a ternary NPK compound fertilizer, supplemented with a small amount of nitrogen fertilizer) for 30 years, while an unfertilized area served as the control. To study the effect of fertilization, all other management operations—including irrigation, pruning, pest and disease control, and weed management—were kept the same in both the fertilized and unfertilized pear orchards.

(3)The final conclusion about usage of organic fertilizers is not supported by the results. The results show increase in soil fertility, but no linkage to usage of organic fertilizers is shown.

There are some individual comments and corrections inserted in the file.

Response: Thank you for your kind reminder and valuable suggestions. According to your suggestion, we rewrote the conclusion as “This study clarifies the multi-faceted effects of 30-year chemical-based fertilization on soil in a “Dangshansuli” pear orchard. The co-occurrence of topsoil acidification and organic matter accumulation highlights the complex interactions among nutrient inputs, root-driven carbon cycling, and soil chemistry in perennial cropping systems. Although fertilization increased soil nitrogen and phosphorus, it also caused plant-available iron deficiency and elevated copper to near-phytotoxic levels, along with gradual accumulation of other heavy metals within regulatory limits. Furthermore, long-term fertilization shifted the soil’s biochemical composition, introducing allelochemicals such as octadecane, which inhibited pear seedling growth. Together, these results indicate that conventional mineral fertilization, while maintaining productivity, promotes nutrient imbalance and latent biological stress in soil. Thus, we recommend adopting integrated nutrient management—reducing mineral NPK inputs while consistently applying quality organic amendments to alleviate acidification, improve micronutrient supply, and strengthen soil biological resilience in “Dangshan” pear orchard.” 

Reviewer 2 Report

Comments and Suggestions for Authors

I want to commend the authors for the relevance of the chosen research topic. However, I would like to highlight some points that need to be reviewed and/or improved.

1 - Introduction
The paragraphs should be anchored by a greater number of references and more recent citations.

 

2 -  Materials and Methods - I suggest that the authors add information about the type of soil found in the study areas.

3 - L. 149: Fig. A – organin matter (could it be "organic matter"?

4 - L. 169: Fig. A - I recommend that the letters corresponding to the deviation bars in Fig. A be revised. In Fig. C - correction to the figure title - "Totai-N"

5 - L. 206: Fig. d - some letters corresponding to the deviation bars appear to be incorrect.

6 - Figures - I suggest that the lettering of the different figures be standardized (e.g., the letters for the figures in L. 169 are in uppercase and those in L. 206 are in lowercase).

7 - Regarding the figures, in general, I believe that a "Heatmap" type figure, or another that correlates the different levels of nutrients and contaminants, would make the understanding of the results clearer.

8 - Conclusion - I suggest that this topic be rewritten and that the answer to the hypothesis mentioned in the introduction be included.

9 - References - More than 50% das referências were published more than 5 or 10 years ago. I suggest that the authors corroborate the manuscript, especially the discussion, with more references and from more recent dates.

10 - Supplementary - I suggest that in the tables showing the results obtained in gas chromatography, this analysis be mentioned in the caption.

Author Response

1 - Introduction
The paragraphs should be anchored by a greater number of references and more recent citations.

Response: Thank you for your valuable suggestions. In the revised Introduction, we updated the citations of the references.

2 - Materials and Methods - I suggest that the authors add information about the type of soil found in the study areas.

Response: Thank you for your kind reminder. We supplied the type of soil as “The treatment group consisted of a “Dangshansuli” pear orchard that was subjected to Long-term fertilization (Fertilization was primarily based on a ternary NPK compound fertilizer, supplemented with a small amount of nitrogen fertilizer) for 30 years, while an unfertilized area served as the control)”.

3 - L. 149: Fig. A – organin matter (could it be "organic matter"?

Response: We are really sorry for this. In the Revised Figure 2， we have modified ‘organin’ to ‘organic’.

4 - L. 169: Fig. A - I recommend that the letters corresponding to the deviation bars in Fig. A be revised. In Fig. C - correction to the figure title - "Totai-N"

Response: Thank you for your kind reminder. We have reanalyzed the difference significance with modifications, and we modified ‘Totai-N’ to ‘Total-N’.

 

5 - L. 206: Fig. d - some letters corresponding to the deviation bars appear to be incorrect.

Response: Thank you for your kind reminder. We have reanalyzed the difference significance with modifications.

6 - Figures - I suggest that the lettering of the different figures be standardized (e.g., the letters for the figures in L. 169 are in uppercase and those in L. 206 are in lowercase).

Response: Thank you for your kind reminder. We have modified the lowercase in Figure 6 to uppercase.

7 - Regarding the figures, in general, I believe that a "Heatmap" type figure, or another that correlates the different levels of nutrients and contaminants, would make the understanding of the results clearer.

Response: We appreciate your suggestion to include a heatmap to illustrate correlations between different nutrient and contaminant levels. After careful consideration, we decided to maintain the current figures to allow readers to better observe the vertical distribution patterns of each parameter across soil layers. Given the multidimensional nature of our dataset, a single heatmap might obscure some important layer-specific trends. However, we have expanded the Discussion to explicitly describe key correlations between major nutrients and heavy metals. Thank you again for your suggestion.

8 - Conclusion - I suggest that this topic be rewritten and that the answer to the hypothesis mentioned in the introduction be included.

Response: Thank you for your valuable suggestion. According to your suggestions, we rewrote the conclusion as “This study clarifies the multi-faceted effects of 30-year chemical-based fertilization on soil in a “Dangshansuli” pear orchard. The co-occurrence of topsoil acidification and organic matter accumulation highlights the complex interactions among nutrient inputs, root-driven carbon cycling, and soil chemistry in perennial cropping systems. Although fertilization increased soil nitrogen and phosphorus, it also caused plant-available iron deficiency and elevated copper to near-phytotoxic levels, along with gradual accumulation of other heavy metals within regulatory limits. Furthermore, long-term fertilization shifted the soil’s biochemical composition, introducing allelochemicals such as octadecane, which inhibited pear seedling growth. Together, these results indicate that conventional mineral fertilization, while maintaining productivity, promotes nutrient imbalance and latent biological stress in soil. Thus, we recommend adopting integrated nutrient management—reducing mineral NPK inputs while consistently applying quality organic amendments to alleviate acidification, improve micronutrient supply, and strengthen soil biological resilience in “Dangshan” pear orchard.”

9 - References - More than 50% das referências were published more than 5 or 10 years ago. I suggest that the authors corroborate the manuscript, especially the discussion, with more references and from more recent dates.

Response: Thank you for your kind reminder. We have updated the literature citations in the revised manuscript.

10 - Supplementary - I suggest that in the tables showing the results obtained in gas chromatography, this analysis be mentioned in the caption.

Response: We appreciate your comment. In response, the raw data obtained from the gas chromatography–mass spectrometry (GC-MS) analysis have been incorporated within the supplementary material (see the original data in supplementary material).

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

The present manuscript addresses a descriptive study aimed at identifying the implications of fertilization on the physical and chemical properties of a pear orchard. The results indicate an increase in organic matter and a decrease in soil pH, which contrasts with widely documented global trends associated with long-term inorganic fertilization. Indeed, international organizations such as the UN have highlighted the need to identify alternatives to inorganic fertilizers, given their well-known negative impacts on soil fertility and ecosystem integrity.

However, the current study lacks a clear and precise description of the management history of the orchard during the 30-year period evaluated.

 

Below you will find several suggestions to strengthen and clarify your manuscript.

 

General observations

 

Authors 1 and 2 are marked with a #, but the manuscript does not indicate what this symbol refers to.

 

Likewise, the last two authors are marked with *, but no explanation is provided.

 

Abstract

 

This section should not contain subsections. Please remove the “Conclusion” heading and integrate that sentence into the paragraph.

 

Introduction

 

The introduction requires strengthening. What observations have been reported regarding allelopathy in other crops? What has been documented about long-term fertilization in similar systems? How does this study contribute to preventing similar issues in other agroecosystems?

 

Please be more explicit regarding what the study aims to address and how this objective was approached. Alternatively, clearly state that this is a descriptive article reporting the long-term effects of fertilization in a pear production system.

 

It is necessary to expand the information about the study site, including the type of fertilizer used (organic or inorganic) and the management practices applied during the 30-year period. These details are essential to justify the long-term implications observed.

 

Results

 

The Results section should be concise and strictly descriptive. This means avoiding explanations, justifications, or interpretations of why the findings occurred. All explanatory content should be reserved for the Discussion.

 

The figure labels are inconsistent. The legend states that the pH graph corresponds to panel (A), but in the image this graph appears as (B), and vice versa. The text matches the graphical order, so the error lies in the legend.

 

Please standardize how figures are labeled. Figures 2 through 5 use uppercase letters for subpanels, but Figure 6 uses lowercase letters in parentheses. In the text, the figure is referenced using uppercase letters. Consistency is required.

 

For Figure 7, consider using arrows, circles, or other markers to highlight differences, as the visual differences among plants are not readily apparent at first glance.

 

It appears that the results of section 2.5 (“Seeds germination assay”) were not described. Even if the table was moved to the supplementary material, the main text must present a subsection summarizing the principal findings of this assay.

 

Supplementary Material

 

Supplementary tables are still part of your scientific manuscript, and therefore must maintain the same level of rigor. Each table should be self-contained and self-explanatory. Please include a note at the end defining what the values represent (e.g., means), indicate the sample size (n), and specify whether any statistical analysis was performed and which one.

 

Discussion

 

It is essential to describe in detail how the soil was managed and what type of fertilization was applied throughout the 30-year period. Without this information, the results cannot be adequately contextualized, as they diverge from well-documented global trends. Inorganic fertilization typically contributes to severe soil degradation; if this were not the case, soil loss would not be a major global concern and the search for alternatives to inorganic agrochemicals would not be a priority.

 

The discussion must justify and explain—mechanistically—how the fertilization regime and long-term management could have contributed to an increase in organic matter and a lower pH compared to the control soil. This requires a deeper interpretation based on soil biochemistry and long-term nutrient dynamics.

 

Lines 251–254: The manuscript does not evaluate the aspect mentioned by the authors in these lines, and therefore it is not appropriate to discuss it. The Discussion should align strictly with the Materials and Methods and with the study’s objectives, providing clear mechanistic explanations of the observed results.

 

Overall, the discussion is currently insufficient and lacks structure.

 

Suggested structure for improving the Discussion:

 

1. a) Begin with a broad interpretation of your main findings
2. b) Compare your results with previously published literature
3. c) Provide a mechanistic explanation—rooted in soil science and plant physiology—of why these results occurred
4. d) Emphasize the novelty or importance of your study
5. e) Acknowledge limitations
6. f) Present implications and future research perspectives

 

Conclusion

The conclusion contradicts widely observed global trends regarding the long-term effects of inorganic fertilization on soil. Therefore, it is crucial to be extremely precise in explaining the particular conditions of this case that justify the atypical results.

Author Response

The present manuscript addresses a descriptive study aimed at identifying the implications of fertilization on the physical and chemical properties of a pear orchard. The results indicate an increase in organic matter and a decrease in soil pH, which contrasts with widely documented global trends associated with long-term inorganic fertilization. Indeed, international organizations such as the UN have highlighted the need to identify alternatives to inorganic fertilizers, given their well-known negative impacts on soil fertility and ecosystem integrity.

However, the current study lacks a clear and precise description of the management history of the orchard during the 30-year period evaluated.

Response: Thank you very much for your valuable suggestions.

Below you will find several suggestions to strengthen and clarify your manuscript.

• General observations

Authors 1 and 2 are marked with a #, but the manuscript does not indicate what this symbol refers to, Likewise, the last two authors are marked with *, but no explanation is provided.

Response: Thank you for your kind reminder. We have supplied the note of ‘#’ and ‘*’.

• Abstract

This section should not contain subsections. Please remove the “Conclusion” heading and integrate that sentence into the paragraph.

Response: Thank you for your kind reminder. We have removed the “Conclusion” heading and integrate that sentence into the paragraph

• Introduction

The introduction requires strengthening. What observations have been reported regarding allelopathy in other crops? What has been documented about long-term fertilization in similar systems? How does this study contribute to preventing similar issues in other agroecosystems?

Response: Thank you very much for your valuable suggestions. According to your questions, we have rephrased this sentence, the modified content is as follows:“In recent years, the application of livestock and poultry manure has gained attention as an environmentally sustainable alternative, and many fruit growers increasingly prefer manure over commercial fertilizers [9,10]. However, long-term manure application may also pose risks, such as elevating concentrations of toxic heavy metals (e.g., Hg, Cu, Zn, and Cr) and promoting the accumulation of allelochemicals in soil [11-15]. Allelopathy, the phenomenon by which plants release biochemicals that influence the growth and development of themselves or neighboring plants, is increasingly recognized as a contributing factor to replanting problems and soil sickness in perennial cropping systems [16,17]. For instance, in black walnut and banana orchards, phenolic acids and certain terpenoids released from root exudates or decomposing residues have been shown to inhibit seedling growth and reduce root vitality [18,19]. These allelopathic effects are largely attributed to secondary metabolites such as alkaloids, phenolic compounds, fatty acids, alkanes, and terpenoids [16,17]. Therefore, analyzing the diversity of allelochemicals and their biological activity is crucial for understanding soil–plant feedbacks in intensively managed orchards.”

 

Please be more explicit regarding what the study aims to address and how this objective was approached. Alternatively, clearly state that this is a descriptive article reporting the long-term effects of fertilization in a pear production system.

Response: Thank you very much for your valuable suggestions. According to your suggestions, we modified this as “Pears are one of the most important economic fruit crops worldwide. According to the Food and Agriculture Organization of the United Nations [20], China accounts for 67.3% of the worldwide pear cultivation area. The “Dangshansuli” pear, a local cultivar from Dangshan County, Anhui Province, has a cultivation history of more than 200 years and is renowned for its juiciness, pleasant flavor, and sweet aroma. Currently, it represents one-quarter of China’s total pear production [21]. However, the widespread and often excessive use of fertilizers in these has led to marked change in soil physicochemical properties, triggering issues such as soil acidification, nutrient imbalance, and potential heavy metal enrichment [22]. While the effects of longterm fertilization have been studied in other fruit systems such as apples [23] and highbush blueberries [24], the specific impacts on soil chemistry, metal accumulation, and allelochemical profiles in “Dangshansuli” pear orchards remain poorly documented.

This study aims to systematically assess how longterm fertilization affects soil chemical properties across different soil layers in a “Dangshansuli” pear production system. Specifically, we compared mineral elements, heavy metals, and organic compound profiles between soils that had received longterm fertilization and an unfertilized control area maintained for over 30 years. We hypothesized that prolonged chemical fertilizer application would result in soil acidification and increased accumulation of heavy metals. By characterizing these changes, this work seeks to provide a scientific basis for developing improved fertilization strategies that can mitigate adverse effects on soil health. The findings are expected to offer insights applicable not only to “Dangshansuli” pear orchards but also to other intensively managed fruitgrowing systems facing similar sustainability challenges.”

 

It is necessary to expand the information about the study site, including the type of fertilizer used (organic or inorganic) and the management practices applied during the 30-year period. These details are essential to justify the long-term implications observed.

Response: We are really sorry for the misunderstanding description. In the revised manuscript, we have provided this information in the material and methods as below:“Soil sampling was collected in May 2020. The treatment group consisted of a “Dangshansuli” pear orchard that was subjected to Long-term fertilization (Fertilization was primarily based on a ternary NPK compound fertilizer, supplemented with a small amount of nitrogen fertilizer) for 30 years, while an unfertilized area served as the control. To study the effect of fertilization, all other management operations—including irrigation, pruning, pest and disease control, and weed management—were kept the same in both the fertilized and unfertilized pear orchards. ”

Results

 

The Results section should be concise and strictly descriptive. This means avoiding explanations, justifications, or interpretations of why the findings occurred. All explanatory content should be reserved for the Discussion.

Response: Thank you very much for your valuable suggestions. According to your suggestions, we have revised the results section to be strictly descriptive.

The figure labels are inconsistent. The legend states that the pH graph corresponds to panel (A), but in the image this graph appears as (B), and vice versa. The text matches the graphical order, so the error lies in the legend.

Response: Thank you for your kind reminder. We have modified the quotation in the manuscript.

Please standardize how figures are labeled. Figures 2 through 5 use uppercase letters for subpanels, but Figure 6 uses lowercase letters in parentheses. In the text, the figure is referenced using uppercase letters. Consistency is required.

Response: Thank you for your kind reminder. We have changed lowercase letters to uppercase letters for consistently.

For Figure 7, consider using arrows, circles, or other markers to highlight differences, as the visual differences among plants are not readily apparent at first glance.

Response: Thank you for your valuable suggestion. We used arrows to mark the difference of different treatment.

It appears that the results of section 2.5 (“Seeds germination assay”) were not described. Even if the table was moved to the supplementary material, the main text must present a subsection summarizing the principal findings of this assay.

Response: Thank you for your valuable suggestion. We supplied this description in the results as “Meanwhile, the seed germination assay revealed that at a concentration of 1.0 g·mL⁻¹, seeds treated with the fertilized soil extract exhibited a significantly higher germination rate compared to those treated with the unfertilized soil extract (Table S3).”

 

Supplementary Material

 

Supplementary tables are still part of your scientific manuscript, and therefore must maintain the same level of rigor. Each table should be self-contained and self-explanatory. Please include a note at the end defining what the values represent (e.g., means), indicate the sample size (n), and specify whether any statistical analysis was performed and which one.

Response: Thank you for your valuable suggestion. We have supplied the note at the end of Table S3 as “Note: Control: distilled water control; S_0.25: soil extract from fertilized plots at 0.25 g·mL^-1; S_0.5: soil extract from fertilized plots at 0.50 g·mL^-1; S1: soil extract from fertilized plots at 1.00 g·mL^-1; W_0.25: soil extract from unfertilized plots at 0.25 g·mL^-1; W_0.5: soil extract from unfertilized plots at 0.50 g·mL^-1; W₁: soil extract from unfertilized plots at 1.00 g·mL^-1. Different lowercase letters above the error bars (n = 5) indicate significant differences at p < 0.05, as determined by Duncan’s multiple range test.”

 

Discussion

 

It is essential to describe in detail how the soil was managed and what type of fertilization was applied throughout the 30-year period. Without this information, the results cannot be adequately contextualized, as they diverge from well-documented global trends. Inorganic fertilization typically contributes to severe soil degradation; if this were not the case, soil loss would not be a major global concern and the search for alternatives to inorganic agrochemicals would not be a priority.

Response: Thank you for your valuable suggestion. We described fertilized and unfertilized soil management in detail in the material and methods. The supplied content was as “Soil sampling was collected in May 2020. The treatment group consisted of a “Dangshansuli” pear orchard that was subjected to Long-term fertilization (Fertilization was primarily based on a ternary NPK compound fertilizer, supplemented with a small amount of nitrogen fertilizer) for 30 years, while an unfertilized area served as the control. To study the effect of fertilization, all other management operations—including irrigation, pruning, pest and disease control, and weed management—were kept the same in both the fertilized and unfertilized pear orchards. ”

 

The discussion must justify and explain—mechanistically—how the fertilization regime and long-term management could have contributed to an increase in organic matter and a lower pH compared to the control soil. This requires a deeper interpretation based on soil biochemistry and long-term nutrient dynamics.

Response: Thank you for your valuable suggestion. According to your suggestions, we modified the discussion in deeply.

Lines 251–254: The manuscript does not evaluate the aspect mentioned by the authors in these lines, and therefore it is not appropriate to discuss it. The Discussion should align strictly with the Materials and Methods and with the study’s objectives, providing clear mechanistic explanations of the observed results.

Response: Thank you for your valuable suggestion. According to your suggestions, To avoid ambiguity, we deleted the sentence.

 

Overall, the discussion is currently insufficient and lacks structure.

 

Suggested structure for improving the Discussion:

 

1. a) Begin with a broad interpretation of your main findings
2. b) Compare your results with previously published literature
3. c) Provide a mechanistic explanation—rooted in soil science and plant physiology—of why these results occurred
4. d) Emphasize the novelty or importance of your study
5. e) Acknowledge limitations
6. f) Present implications and future research perspectives

Response: Thank you for your valuable suggestion. According to your suggestions, we rewrote the discussion as below:

4.1 Soil Physicochemical Characteristics Under Long-term Fertilization

The influence of fertilization on soil physicochemical properties is highly dependent on climatic and site-specific conditions [25,26]. In this study, systematic assessment of a 30-year NPK fertilization regime in a “Dangshansuli” pear orchard revealed distinct alterations in soil chemistry. Notably, long-term fertilization resulted in elevated soil organic matter (SOM) content alongside reduced pH in the 0–20 cm layer. This pattern aligns with observations in other perennial orchard systems, where sustained productivity can enhance belowground carbon inputs through increased root turnover and litter deposition [27–29]. The significant increases in total nitrogen, phosphorus, alkaline hydrolyzable nitrogen, and available phosphorus within the 0–40 cm depth are direct outcomes of cumulative nutrient applications. In contrast, the more moderate increase in potassium and the reduction in exchangeable calcium and magnesium—particularly in the surface layer—are mechanistically explained by ion-exchange dynamics. Prolonged input of NH₄⁺ and K⁺ from fertilizers promotes nitrification (releasing H⁺) and the competitive displacement of Ca²⁺ and Mg²⁺ from soil colloids by K⁺ and H⁺, thereby lowering base saturation and buffering capacity [27,30]. This study highlights that in perennial fruit systems, the feedback between fertilization, enhanced root-derived carbon inputs, and nutrient cycling can partially offset the typical decline in SOM associated with inorganic fertilizers, even as soil acidification advances.

4.2 Influence of Soil Trace Elements Under Long-term Fertilization

The availability of trace elements was strongly mediated by the altered soil chemical environment, particularly the decrease in pH. Significantly higher concentrations of available Fe, Zn, Cu, Mn, and B were found in the fertilized 0–40 cm soil layer, consistent with the general increase in metal solubility under acidic conditions. Specifically, the available Cu content exceeded twice the optimal level for plant growth, which can be attributed to combined effects of enhanced mobility at lower pH and historical use of copper-based fungicides, a common practice in orchard management [27,31]. Paradoxically, despite sufficient total Fe content, available Fe remained critically low (11 mg·kg⁻¹ in the 20–40 cm layer), explaining the widespread iron chlorosis observed in the pear leaves. This deficiency likely results from the interplay between moderate acidity and high calcium content, which favors the oxidation and precipitation of iron into insoluble forms, thereby limiting its phytoavailability—a typical physiological constraint in calcareous soils undergoing acidification.

4.3 Soil Heavy Metals Under Long-term Fertilization

Our findings confirm that long-term fertilization, even with predominantly mineral sources, can lead to the accumulation of several heavy metals in the soil profile [14,28]. Significant enrichment of Cu and Hg was detected in the surface layer (0–20 cm), while Ni, Cr, As, and Mn levels were elevated throughout the sampled depth, with Ni showing particularly notable enrichment. Importantly, all measured concentrations remained below the limits established by China's soil environmental quality standards, indicating no immediate contamination risk. This gradual accumulation likely originates from trace impurities in mineral fertilizers and atmospheric deposition over decades. While short-term applications of organic amendments such as pig manure compost have been reported to improve soil fertility without sharply increasing metal loads [8,14,32], our long-term context demonstrates a progressive buildup. Since pear roots are predominantly concentrated in the 0–40 cm layer, this accumulation warrants careful attention. To mitigate further metal loading and improve overall soil physicochemical health, a strategic shift toward integrated fertilization—reducing mineral inputs while incorporating high-quality, well-composted organic amendments—is strongly recommended for Dangshan pear orchards.

4.4 Influence of Soil Chemical Compounds Composition Under Long-term Fertilization

Long-term fertilization fundamentally altered the soil's organic chemical profile. A greater diversity and total content of organic compounds were detected in the fertilized 0–60 cm soil (Table S1). Furthermore, soil extracts from fertilized plots enhanced seed germination (Table S3), indicating a complex mixture of both stimulatory and inhibitory substances. Notably, several compounds–including alkanes (e.g., 2,6,10-trimethylpentadecane, octadecane) and organic acids (e.g., benzoic acid, dodecanoic acid)–were detected exclusively in the fertilized soil (Table S2). Their presence suggests an origin linked to the fertilization regime, either as direct components, metabolic byproducts, or via fertilizer-induced shifts in microbial activity. Among these, octadecane and benzoic acid are established allelochemicals known to inhibit the growth of various plants and algae at elevated concentrations [33-35]. In our bioassay, octadecane exhibited clear phytotoxicity toward tissue-cultured “Shanli” pear seedlings, inducing leaf tip blackening and curling at 0.18 mmol·L-1 (Figure 7). This finding is particularly significant, as it suggests that long-term fertilization may not only modify nutrient and heavy metal profiles but also introduce or promote the accumulation of specific bioactive compounds. Such compounds could contribute to soil fatigue or replanting problems—a relatively underexplored consequence of continuous orchard management.

Conclusion

The conclusion contradicts widely observed global trends regarding the long-term effects of inorganic fertilization on soil. Therefore, it is crucial to be extremely precise in explaining the particular conditions of this case that justify the atypical results.

Response: Thank you for your valuable suggestion. According to your suggestions, we rewrote the conclusion as “This study clarifies the multi-faceted effects of 30-year chemical-based fertilization on soil in a “Dangshansuli” pear orchard. The co-occurrence of topsoil acidification and organic matter accumulation highlights the complex interactions among nutrient inputs, root-driven carbon cycling, and soil chemistry in perennial cropping systems. Although fertilization increased soil nitrogen and phosphorus, it also caused plant-available iron deficiency and elevated copper to near-phytotoxic levels, along with gradual accumulation of other heavy metals within regulatory limits. Furthermore, long-term fertilization shifted the soil’s biochemical composition, introducing allelochemicals such as octadecane, which inhibited pear seedling growth. Together, these results indicate that conventional mineral fertilization, while maintaining productivity, promotes nutrient imbalance and latent biological stress in soil. Thus, we recommend adopting integrated nutrient management—reducing mineral NPK inputs while consistently applying quality organic amendments to alleviate acidification, improve micronutrient supply, and strengthen soil biological resilience in “Dangshan” pear orchard.”

 

 

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Thank you for your explanations and changes in the manuscript that significantly improved the quality.

Author Response

1. Title: Effects of Long-term Fertilization on Soil Chemical Properties of “Dangshansuli” Pear Orchard.

Response: We appreciate your acknowledgment.

2. As many as 12 authors signed for the research presented in the manuscript! The scope and quality of the research presented casts doubt on the contribution of so many signed authors.

Response: Thank you for raising this valid concern regarding the number of authors. We agree that all listed authors must have made substantial contributions. In direct response to your comment, we have critically re-evaluated the authorship list and have revised the manuscript to include only the 8 core contributors. The specific contributions of each author are now detailed in the ‘Author contributions’ section. We believe this revised list accurately reflects the actual intellectual and practical work undertaken for this study.

3. Abstract, line 15-16: Indicate the type of soil on which this research was conducted.

Response: Thank you for your valuable suggestions. This experiment was conducted on a typical “sandy soil” collected from the field. We have supplied the type of the soil (sandy soil) in the revised manuscript in line 15. The revised content was description as:“In May 2020, a total of 30 soil samples were collected from a long-term fertilization and an unfertilized sandy control plot that had been maintained for 30 years. ”

4. Key words: - These words are already found in the title of the manuscript: "Dangshansuli”; long-term fertilization; - "soil physicochemical" not studied (Why is this keyword even here?) - other keywords should be added, such as: allelochemical properties, heavy metals, type of soil.
5. Response: Thank you for your kind reminder. We agree that the original list of key words was not suitable. We have revised it as follows: The type of soil; mineral elements; chemical compounds; allelochemical properties.
6. Introduction Line 34-37: The amounts of nitrogen fertilizers applied to apple plantations in China are highly debatable. The cited works did not explain why 5 times the amount of nitrogen fertilizers was applied.

Response: Thank you for this comment. To address the concern and improve clarity, we have revised the text in the manuscript. The relevant sentence was as below: “This pattern of inefficient resource use is widespread in orchard management; where growers of crops such as pears and apples often apply excessive nitrogen fertilizer in pursuit of higher yields [3,4].”

Reference:

Dong CX., Jiang HB., Zhao JW., Xu YC. Current fertilization in pear orchards in China. Soils.  2012, 44, 754–761.

Wang N.; Wolf J.; Zhang FS. Towards sustainable intensification of apple production in China — Yield gaps and nutrient use efficiency in apple farming systems. J Integr Agric. 2016, 15 (4):716-725. https://doi.org/10.1016/S2095-3119(15)61099-1.

7. This manuscript also does not explain the reasons for the irrational consumption of nitrogen. Then, the efficiency of using nitrogen from applied fertilizer in the most optimal conditions does not exceed 50% (proven by using nitrogen fertilizers labeled with the isotope 15N). With an application of 400-600 kg of nitrogen, nitrogen use efficiency must be >15%. It is completely pointless to investigate the multi-year "waste" of nitrogen fertilizer from: 400-600 kg hm^-2!

Response: Thank you for your critical comment regarding nitrogen use efficiency. We have revised the introduction to more precisely frame the issue of fertilizer use in orchard systems. As you rightly highlighted, it is crucial to clarify the nature of the problem. According to your suggestions, we removed the previous description and incorporated the dual issues of over-application and nutrient imbalance. The revised text was as below: “This pattern of inefficient resource use is widespread in orchard management; where growers of crops such as pears and apples often apply excessive nitrogen fertilizer in pursuit of higher yields [3,4]. Compounding this issue, fertilization in many orchards relies heavily on mineral inputs with limited organic supplementation [5,6]. This combination of over-application and nutrient imbalance leads to a significant resource wastage and contributes to soil degradation and environmental pollution [7,8].”

Reference:

Gao X., Zhao X., Wu P., Yang M.,Ye M., Tian L., Zou Y., Wu Y., Zhang F., Siddique KHM. The economic–environmental trade-off of growing apple trees in the drylands of China: a conceptual framework for sustainable intensification. J Clean Prod. 2021, 296:126497. https://doi.org/10.1016/j.jclepro.2021.126497.

Guo S.; Xiong W.; Xu H.; Hang X.; Liu H.; Xun W.; Li R.; Shen Q. Continuous application of different fertilizers induces distinct bulk and rhizosphere soil protist communities. Eur J Soil Biol. 2018, 88: 8-14. https://doi.org/10.1016/j.ejsobi.2018.05.007.

Srivastav AL., Patel N., Rani L., Kumar P., Dutt I., Maddodi B., Chaudhary VK. Sustainable options for fertilizer management in agriculture to prevent water contamination: a review. Environ. Dev. Sustain. 2023, 1-25. https://doi.org/10.1007/s10668-023-03117-z.

Ning CC.; Gao PD.; Wang BQ.; Lin WP.; Jiang NH.; Cai KZ. Impacts of chemical fertilizerreduction and organic amendments supplementation on soil nutrient, enzyme activity andheavy metal content. J Integr Agric. 2017, 16 (8): 1819-1831. https://doi.org/10.1016/s2095-3119(16)61476-4.

 

8. Materials and Methods Line 82-87: - Indicate the type of soil on which the research was conducted (FAO or USDA soil classification). - What are the local specificities of pear cultivation and support this with an appropriate reference. - Why is pear grown in the specified region (for food, for industrial processing, or something else)? Figure 1: Incomplete figure caption (figures and tables should be self-explanatory). For example: Soil profile under a “Dangshansuli” pear orchard. Schematic diagram of the soil sampling location. Line 105-124 and 140-146: Not all determination methods are described, nor are appropriate literature sources for the methods used listed.

Response: Thank you for your valuable comments. We have carefully revised the manuscript according to your suggestions. The specific responses and modifications are as follows:

• Materials and Methods Line 82-87

Response: We have added the soil classification and local cultivation context as suggested. The revised text was “Dangshan provides optimal conditions for pear cultivation, characterized by its flat terrain, loose and well-drained sandy soil, distinct seasons, and a climate with concurrent rainfall and warm periods, as well as abundant sunshine [25]. The pear trees in both the long-term fertilized and unfertilized orchards were cultivated for fruit production. From each orchard, three “Dangshansuli” pear trees (Pyrus bretschneideri Rehd.) with uniform growth vigor all selected, all grafted onto Pyrus betulifolia Bunge rootstock.”

• Figure 1: Incomplete figure caption (figures and tables should be self-explanatory). For example: Soil profile under a “Dangshansuli” pear orchard. Schematic diagram of the soil sampling location.

Response: We have revised the caption of Figure 1 to make it self-explanatory as recommended. The Revised caption was “Soil profile under a “Dangshansuli” pear orchard. Schematic diagram of the soil sampling location.”

8.3 Line 105-124 and 140-146: Not all determination methods are described, nor are appropriate literature sources for the methods used listed.

Response: We have supplemented the description of analytical methods and added corresponding references for each determination. The supplement methods of soil physical and chemical properties determination was shown as below:

“10 g of air-dried soil were placed into a 200 mL conical flask, mixed with 50 mL of ultrapure water, and stirred for 2 min using a magnetic stirrer. The flask was then sealed with sealing film and allowed to stand for 30 min before measurement. After calibration with standard solutions, the electrode was inserted into the supernatant, and the pH value was read directly from the meter and recorded.

1 g of soil was placed into a test tube, and organic carbon in the soil was oxidized using the sulfuric acid–potassium dichromate method. The mixture was heated in an oil bath, and the remaining potassium dichromate was titrated with ferrous sulfate. The organic matter content was calculated based on the amount of potassium dichromate consumed [26].

The minerals and trace elements were determined using wet analysis and ICP OES analytical methods [27]. Heavy metals (Cu, Zn, Co, Ni, Cr, Pb) were extracted with HCl and determined by ICPOES, as was measured by hydride generationatomic fluorescence spectrometry, and Hg by coldvapor atomic fluorescence spectrometry [28].”

 

9. Results The manuscript contains 7 figures and 0 tables. It would be more appropriate to present the data from Figures 4 and 5 in a tables.

Response: Thank you for your valuable suggestions. According to your suggestion, we have modified Figure 4 and 5 to Table 1 and Table 2 respectively. The updated tables have been incorporated into the main text.

 

10. Discussion The discussion of the results obtained is very modest (unsatisfactory). Pear is grown all over the world and these results should be compared with similar research in Europe, America and Asia.

Response: Thank you very much for your valuable suggestions. According to your suggestions, we modified this as below:

4.1 Soil Physicochemical Characteristics Under Long-term Fertilization

The influence of fertilization on soil physicochemical properties is highly dependent on climatic and site-specific conditions [31,32]. In this study, systematic assessment of a 30-year NPK fertilization regime in a “Dangshansuli” pear orchard revealed distinct alterations in soil chemistry. Notably, long-term fertilization resulted in elevated soil organic matter (SOM) content alongside reduced pH in the 0–20 cm layer. This pattern aligns with observations in other perennial orchard systems, where sustained productivity can enhance belowground carbon inputs through increased root turnover and litter deposition [33–35]. Similar SOM accumulation under mineral fertilization has been reported in apple orchards in the United States [36]. The significant increases in total nitrogen, phosphorus, alkaline hydrolyzable nitrogen, and available phosphorus within the 0–40 cm depth are direct outcomes of cumulative nutrient applications. In contrast, the more moderate increase in potassium and the reduction in exchangeable calcium and magnesium—particularly in the surface layer—are mechanistically explained by ion-exchange dynamics. Prolonged input of NH₄⁺ and K⁺ from fertilizers promotes nitrification (releasing H⁺) and the competitive displacement of Ca²⁺ and Mg²⁺ from soil colloids by K⁺ and H⁺, thereby lowering base saturation and buffering capacity [33,37]. This acidification pathway is widely recognized in intensively fertilized orchards worldwide. Over-application of inorganic fertilizers is a major driver of soil acidification, a concern increasingly reported across fruit-growing regions in Asia and Europe [38,39]. In response, studies have shown that organic amendments—such as organic fertilizer, bioorganic fertilizer, and humic acid—can mitigate acidification, enhance microbial activity, and improve crop yield, offering a sustainable strategy for maintaining soil health in perennial fruit systems [40,41].

4.2 Influence of Soil Trace Elements Under Long-term Fertilization

The availability of trace elements was strongly mediated by soil acidification. Significantly higher concentrations of available Fe, Zn, Cu, Mn, and B in the fertilized 0–40 cm reflect increased metal solubility at lower pH—a trend consistent with studies in acidic orchard soils in Spain [42]. Notably, the available Cu content exceeded twice the optimal level for plant growth, which can be attributed to combined effects of enhanced mobility at lower pH and historical use of copper-based fungicides, a common practice in orchard management [33,43]. Paradoxically, despite sufficient total Fe content, available Fe remained critically low (11 mg·kg⁻¹ in the 20–40 cm layer), explaining the widespread iron chlorosis observed in the pear leaves. The contrast between Cu accumulation and Fe limitation underscores the complex bioavailability shifts induced by longterm fertilization, with implications for nutrient management in “Dangshan” peargrowing regions.

4.3 Soil Heavy Metals Under Long-term Fertilization

Our findings confirm that long-term fertilization, even with predominantly mineral sources, can lead to the accumulation of several heavy metals in the soil profile [14,34]. Significant enrichment of Cu and Hg was detected in the surface layer (0–20 cm), while Ni, Cr, As, and Mn levels were elevated throughout the sampled depth, with Ni showing particularly notable enrichment. Importantly, all measured concentrations remained below the limits established by China's soil environmental quality standards, indicating no immediate contamination risk. This gradual accumulation likely originates from trace impurities in mineral fertilizers and atmospheric deposition over decades. While short-term applications of organic amendments such as pig manure compost have been reported to improve soil fertility without sharply increasing metal loads [8,14,44], however, our long-term data demonstrates a cumulative effects. Compared to other orchard soils where application of chicken and swine manure have limited metal accumulation [45]. Our findings demonstrate that the “Dangshan” region requires an adjustment to its current fertilization strategy. It is recommended to transition toward an integrated approach—specifically, reducing mineral fertilizer inputs while incorporating high-quality composted organic amendments. This shift would help mitigate further heavy metal accumulation and enhance soil health, aligning with sustainable orchard management practices widely adopted in the most fruit orchards [40,41].

4.4 Influence of Soil Chemical Compounds Composition Under Long-term Fertilization

Long-term fertilization fundamentally altered the soil's organic chemical profile. The greater diversity and total content of organic compounds in fertilized 0–60 cm soil (Table S1), along with enhanced seed germination in extracts (Table S3), indicate a complex mixture of both stimulatory and inhibitory substances. Notably, several compounds–including alkanes (e.g., 2,6,10-trimethylpentadecane, octadecane) and organic acids (e.g., benzoic acid, dodecanoic acid)–were detected exclusively in the fertilized soil (Table S2). Their presence suggests an origin linked to the fertilization, either as direct components, metabolic byproducts, or via fertilizer-induced shifts in microbial activity. Among these, octadecane and benzoic acid are established allelochemicals known to inhibit the growth of various plants and algae at elevated concentrations [46-48]. In our bioassay, octadecane induced phytotoxicity in tissue-cultured “Shanli” pear seedlings at 0.18 mmol·L^-1 (Figure 7). This finding is particularly significant, as it suggests that long-term fertilization may not only modify nutrient and heavy metal profiles but also introduce or promote the accumulation of specific bioactive compounds that could inhibit plant growth—a phenomenon reported in fruit crop orchards [18]. The global relevance of this mechanism warrants further crossregional investigation.

11. Conclusion The conclusion is general and does not rely on the results of the research. The conclusion does not state any numerical values or correlations!

Response: Thank you for your valuable suggestions. According to your suggestion, we have revised the conclusion as: “This study elucidates the complex long-term effects of continuous mineral fertilization in sandy soil orchard ecosystems. The findings demonstrate that such fertilization practices simultaneously promote soil acidification and alter organic matter dynamics, highlighting the intricate relationship. Fertilization was shown to shift soil nutrient profiles, potentially causing micronutrient imbalances and facilitating the gradual accumulation of certain metals. Additionally, the introduction of allelopathic substances into the soil was identified as a significant secondary effect. Collectively, these results indicate that reliance on conventional mineral fertilizers can aggravate soil degradation, disrupt nutrient equilibrium, and promote the buildup of bioactive compounds. Therefore, transitioning to integrated nutrient management strategies by reducing synthetic inputs and incorporating systematic organic amendments is recommended to mitigate these adverse effects and improve soil health in intensively managed orchard production systems.”

12. References Very modest and inadequate selection of cited references. The general impression is that scientists for whom agriculture (fruit growing) is the primary scientific discipline did not participate in the writing of this manuscript.

Response: Thank you for raising this crucial point. We fully agree with your assessment and acknowledge the significant limitation posed by the narrow selection of references in the manuscript. In response to the your comments, we have comprehensively updated and modified the reference list to incorporate the latest research in horticultural science. The revised references are as follows:

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