Hermetia illucens L. Frass in Promoting Soil Fertility in Farming Systems

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

This manuscript investigates the residual fertilization effects of Hermetia illucens L. (Black Soldier Fly) frass (BSFF) on ryegrass productivity and soil fertility across three soil types of contrasting textures and fertility. The topic is timely and relevant to circular agriculture and organic soil amendment studies. The experiment is reasonably designed and the dataset appears substantial; however, the study suffers from several conceptual and technical weaknesses that limit its scientific contribution and reproducibility. The statistical treatment is superficial, the mechanistic interpretations are incomplete, and the discussion lacks integration with current literature on nutrient cycling, soil–plant–microbe interactions, and the chemical behavior of frass-derived organic matter. Figures and tables are numerous but insufficiently synthesized, and the PCA interpretation remains largely descriptive. Overall, the manuscript requires major revision before it can be considered for publication.

1. The introduction mentions “fertility resilience” and “residual effect” but does not clearly define these terms quantitatively or mechanistically.
2. Critical parameters (e.g., initial soil nutrient contents, C/N ratio of BSFF, pH, total organic carbon) are missing.
3. The second-year experiment used soils that had already been treated in the previous year, but potential residual mineral fertilizer effects are not isolated. The study lacks an unfertilized baseline control across both years.
4. The study focuses narrowly on fertility indices but omits the broader ecological functions (carbon sequestration, biodiversity, soil health). It may benefit from incorporating perspectives such as those in "Dual effects of Caragana korshinskii introduction on herbaceous vegetation in Chinese desert areas: short-term degradation and long-term recovery", which discuss short-term versus long-term ecosystem dynamics—a framework that parallels the “residual effect” concept.
5. The PCA discussion merely repeats the clustering patterns without connecting them to soil processes.
6. Since BSFF is described as a slowly releasing fertilizer, kinetic data or modeling (e.g., first-order mineralization) would strengthen the argument.
7. The DHA analysis is interpreted in isolation. The authors should discuss how enzyme activity relates to microbial biomass, SOM changes, and nutrient availability.
8. Frass derived from livestock effluents may introduce trace metals or pathogens. While Cu, Zn, and Mn were measured, there is no discussion of potential accumulation or ecological risks. For context, studies such as “Influence of soil characteristics on the concentrations of Cr, Pb, and Zn in tobacco leaves from Longyan, China, and the associated predictive models”; “Effects of Soil Properties on Pb, Cd, and Cu Contents in Tobacco Leaves of Longyan, China, and Their Prediction Models”provide valuable comparative insights into trace element behavior and should be cited.
9. No discussion connects the residual soil chemistry to the observed yield responses.
10. The conclusion claims improved resilience without quantitative criteria.

Author Response

1. The introduction mentions “fertility resilience” and “residual effect” but does not clearly define these terms quantitatively or mechanistically. R1 - Thank you for the comment and changes were made and hope it is clear now (line 34).
2. Critical parameters (e.g., initial soil nutrient contents, C/N ratio of BSFF, pH, total organic carbon) are missing. R2 - The article referenced in the materials and methods section (Rehan et al., 2024) reports the experimental design, as well as the characterization of the materials used, so it seemed more appropriate not to rewrite everything here and instead refer to the aforementioned article.
3. The second-year experiment used soils that had already been treated in the previous year, but potential residual mineral fertilizer effects are not isolated. The study lacks an unfertilized baseline control across both years. R3 - I appreciate the observation, however, since farmers always use fertilizers, especially nitrogen fertilizers, it doesn't seem right to us to make comparisons that include treatments without any fertilization, besides not achieving any production (we have already done trials with zero fertilization and the lack of plant material prevented us from reaching conclusions other than the obvious one, that plants need nutrients and fertilization is necessary, particularly in soils like sandy soils).
4. The study focuses narrowly on fertility indices but omits the broader ecological functions (carbon sequestration, biodiversity, soil health). It may benefit from incorporating perspectives such as those in "Dual effects of Caragana korshinskii introduction on herbaceous vegetation in Chinese desert areas: short-term degradation and long-term recovery", which discuss short-term versus long-term ecosystem dynamics—a framework that parallels the “residual effect” concept. R4 - I appreciate the suggestion; however, this study does not focus on the aspects mentioned in the article you cited. The introduction of vegetation in arid environments may have different behaviors than the use of organic fertilizers in different soil types, as well as the issue of climate. This study aims to contribute to more comprehensive studies, both under controlled conditions and in the field, under different edaphoclimatic conditions. In fact, some of the evaluations carried out, such as production or enzymatic activity, allow us to infer about the quality of the product tested; however, there were more specific parameters that we were unable to quantify due to various limitations. Nevertheless, the results we were able to obtain seemed interesting and could be a contribution to research in the área.
5. The PCA discussion merely repeats the clustering patterns without connecting them to soil processes. R5 - We thank the reviewer for this valuable observation. We agree that the initial discussion focused primarily on describing the PCA clustering without sufficiently linking it to underlying soil processes. In response, we have revised the PCA discussion to interpret the associations in terms of soil organic matter turnover, nutrient cycling, and microbial activity. We hope that the addition clarifies how the observed treatment groupings reflect soil-specific mechanisms influencing fertility and productivity (line 402).
6. Since BSFF is described as a slowly releasing fertilizer, kinetic data or modeling (e.g., first-order mineralization) would strengthen the argument. R6 - We appreciate the idea; however, the data we obtained is from a preliminar experiment, and we believe more data will be needed for a robust analysis.
7. The DHA analysis is interpreted in isolation. The authors should discuss how enzyme activity relates to microbial biomass, SOM changes, and nutrient availability. R7 - Thank you for the comment and changes were made and hope it is clear now (line 289 and 308).
8. Frass derived from livestock effluents may introduce trace metals or pathogens. While Cu, Zn, and Mn were measured, there is no discussion of potential accumulation or ecological risks. For context, studies such as “Influence of soil characteristics on the concentrations of Cr, Pb, and Zn in tobacco leaves from Longyan, China, and the associated predictive models”; “Effects of Soil Properties on Pb, Cd, and Cu Contents in Tobacco Leaves of Longyan, China, and Their Prediction Models”provide valuable comparative insights into trace element behavior and should be cited. R8 - Thank you for this insightful comment and for highlighting relevant literature. We agree that frass derived from livestock effluents may pose potential risks related to trace metal accumulation and pathogen introduction. In response, we have expanded the Discussion section to address these potential ecological risks and to incorporate the suggested references. This addition strengthens the environmental context of our findings and emphasizes the importance of long-term monitoring of trace metal dynamics in BSFF-amended soils to ensure sustainable agricultural use (line 272).
9. No discussion connects the residual soil chemistry to the observed yield responses. R9 - We thank the reviewer for this observation. We agree that explicitly linking residual soil chemistry to yield responses strengthens the interpretation of our results. In response, we have clarified in the Discussion that higher residual SOM, N, and K in treatments preceded by BSFF (particularly OT and mixed modalities) corresponded with higher fresh weight production, especially in the podzol. The slow-release nature of frass-derived nutrients explains why these treatments maintained higher fertility and contributed to sustained yield, despite the absence of fertilization in the second year. These connections have now been highlighted to demonstrate how residual soil fertility mediates crop performance (line 248).
10. The conclusion claims improved resilience without quantitative criteria. R10 - Thank you for this valuable observation. We acknowledge that the claim of improved resilience was based on indirect indicators rather than explicit quantitative criteria. We have revised the text to clarify this limitation and to avoid overstating the finding, while emphasizing that the observed trends suggest potential resilience benefits that warrant further quantitative study.

Reviewer 2 Report

Comments and Suggestions for Authors

While the study demonstrates the potential benefits of using BSFF as an alternative to traditional mineral fertilizers, several limitations remain:

1. Despite the obvious success of BSFF application on sandy soils, the choice of plant species (Lollium multiflorum) raises doubts regarding the general applicability of the results. Since different plant species respond differently to changes in soil composition, further experiments using other crop species are recommended to confirm the general validity of the findings.
2. The authors note a significant advantage of a mixed fertilizer system; however, it is unclear why the ratio of 75% organic and 25% mineral fertilizers proved optimal. It may be worth exploring a wider range of ratios to determine the best combination for a specific soil and climate.
3. Soil microbiota analysis is based primarily on measurements of enzymatic activity (dehydrogenase). However, the presence of a large number of specific microorganisms involved in nutrient conversion remains unknown. The use of molecular methods to identify microbiota and isolate key groups of bacteria and fungi that play an important role in soil fertility restoration is recommended.
4. The cost and availability of BSFF material were not addressed in the study. The economic feasibility of switching to this type of fertilizer compared to traditional farming methods needs to be assessed.

Regarding minor comments:

1. please indicate the standard error of the data presented in the tables;
2. The "References" section must be formatted in accordance with the journal's requirements.

 

Comments for author File: Comments.pdf

Author Response

1. Despite the obvious success of BSFF application on sandy soils, the choice of plant species (Lollium multiflorum) raises doubts regarding the general applicability of the results. Since different plant species respond differently to changes in soil composition, further experiments using other crop species are recommended to confirm the general validity of the findings. R1 - I appreciate the observation; however, the caveat regarding this issue is made throughout the text. The choice of this crop is due to the fact that it is frequently used as forage, and it also allows for multiple cuts during an annual cycle to assess the production and quality of the plant material.
2. The authors note a significant advantage of a mixed fertilizer system; however, it is unclear why the ratio of 75% organic and 25% mineral fertilizers proved optimal. It may be worth exploring a wider range of ratios to determine the best combination for a specific soil and climate. R2 - Thank you for your observation, and indeed, that is what we have been doing, as we have already published studies testing increasing amounts of frass (6 rates – 25, 50, 75, 100, 125, and 150), as potential fertilizer. In the present study, the aim was to highlight the differences related to soil types, even with different mineral:frass ratios, as there is no ideal ratio for all soil types. Furthermore, this trial was intended as a purely scientific study, evaluating the potential of frass as an organic fertilizer in different soil types, and its residual effect, in the second year.
3. Soil microbiota analysis is based primarily on measurements of enzymatic activity (dehydrogenase). However, the presence of a large number of specific microorganisms involved in nutrient conversion remains unknown. The use of molecular methods to identify microbiota and isolate key groups of bacteria and fungi that play an important role in soil fertility restoration is recommended. R3 - Thank you for your comment. It would indeed be very interesting, however the focus was on functionality, so only the DHA was evaluated.
4. The cost and availability of BSFF material were not addressed in the study. The economic feasibility of switching to this type of fertilizer compared to traditional farming methods needs to be assessed. R4 - Thank you for your valuable feedback. You are correct that the cost and availability of BSFF materials were not included in the current study. Our primary focus was on the agronomic performance of BSFF; however, we acknowledge that a comprehensive economic analysis would provide important insights. We intend to explore this aspect in future work.

Regarding minor comments:

1. please indicate the standard error of the data presented in the tables; Thank you for your comment, we made the changes as suggested.
2. The "References" section must be formatted in accordance with the journal's requirements. Thank you for your comment, we made the changes as suggested.

Reviewer 3 Report

Comments and Suggestions for Authors

The Manuscript ID: sustainability-3972899 evaluates Hermetia illucens L. frass in promoting soil fertility in farming systems. The procedures described and materials utilized in their work properly treat the main question addressed by the research. This was reflected by their preliminary evaluation of the potential residual fertilizing effect of BSFF, obtained by the bio-digestion of an effluent from intensive cattle farming, in a second consecutive year of annual ryegrass cultivation, without any fertilization, in a pot experiment, in three soils with different textures and fertility levels. Interesting results were that fresh mass production of annual ryegrass (Lollium multiflorum L.) was consistently higher in the modalities preceded by exclusively biological fertilization than in the modalities preceded by exclusively chemical fertilization, confirming the fact that there is a slow release of the nutrients contained in the frass. Yet, although there were significant differences between treatments, there was not a linear behavior that could justify the use of frass in opposition of the mineral fertilizer. Across the three tested soil types, it was concluded that soils, OT consistently stood apart, associated with mineral nutrients (K, P, Fe, Zn) and SOM, while MOT(75:25), MOT(50:50), MOT(25:75) and MT clustered together with variables linked to plant growth (FW, DW), N, Cu, and Mn. Thus, the strength of treatment separation varied with soil type: strongest in podzol, intermediate in calcisol, and weakest in fluvisol. This indicates that the effects of treatments depended strongly on soil properties, with mineral-nutrient-driven contrasts most pronounced in podzol. Also, the temporal change in production of annual ryegrass and detected soil properties suggests cumulative improvements in soil biological fertility and nutrient-use efficiency under organic and mixed treatments. This is probably because organic amendments usually need time to build up effects on soil organic matter and microbial communities. It was concluded that growing ryegrass on light soil with low-fertility can produce preferential returns with BSFF compared to more fertile soils, for both green mass production and soil fertility resilience.

The subject is worth publication and the authors did a good job. Yet, further insights might improve the study and specific improvements should be considered:

• At least the contents of the mineral fertilizer used should have been analyzed.
• Compared with other published related material, this paper should give more data for the components of their organic matter used.
• Specific improvements regarding the methodology mainly lie in more requirements to explain the main difference between the three soil types used in addition of course to the aforementioned components of mineral and organic fertilizers utilized.
• Tables and figures should be self-explanatory, e.g. a footnote in table 2 should indicate that: (SOM): The soil organic matter. Likewise, for nd (means: NOT DETECTED?). Also, for N% at Podzol soil, in 1st year of table 2, the same number of 0.04 is twice followed by “ns” but once followed by “nsB”. Please, this needs clarification.
• Scientific names when reported for the first time in the text should be written in full with Authority and systematics; g. Hermetia illucens L. (Diptera: Stratiomyidae)
• The topic is quite relevant to the field as it addresses a specific gap in the field. Yet, in the introductions, the authors would preferably have more focus on the usefulness of Hermetia illucens for recycling organic waste and generating animal feed. This would more highlight the value of their research.
• Some typos, mistakes, and misprinted letters, e.g. “Furthermore, BSFF, preferably as a mixed fertilizer (in a proportion until 75%), was shown to be a promising alternative for Gleyic podzol either in production of ryegrass as in the resilience and promotion of soil fertility.” Instead of “Furthermore, BSFF, preferably as a mixed fertilizer (in a proportion until 75%), shown to be a promising alternative for Gleyic podzol either in production of ryegrass as in the resilience and promotion of soil fertility.”

Therefore, I would suggest accepting it after major revision.

 

Author Response

1. At least the contents of the mineral fertilizer used should have been analyzed. R1 - I appreciate your comment. The article referenced in the materials and methods section (Rehan et al., 2024) reports the experimental design, as well as the characterization of the materials used, so it seemed more appropriate not to rewrite everything here and instead refer to the aforementioned article.
2. Compared with other published related material, this paper should give more data for the components of their organic matter used. R2 - The article referenced in the materials and methods section (Rehan et al., 2024) reports the characterization of the materials used, so it seemed more appropriate not to rewrite everything here and instead refer to the aforementioned article.
3. Specific improvements regarding the methodology mainly lie in more requirements to explain the main difference between the three soil types used in addition of course to the aforementioned components of mineral and organic fertilizers utilized. R3 - Thank you for your comment. The article referenced in the materials and methods section (Rehan et al., 2024) reports the characterization of the materials used, so it seemed more appropriate not to rewrite everything here and instead refer to the aforementioned article.
4. Tables and figures should be self-explanatory, e.g. a footnote in table 2 should indicate that: (SOM): The soil organic matter. Likewise, for nd (means: NOT DETECTED?). Also, for N% at Podzol soil, in 1st year of table 2, the same number of 0.04 is twice followed by “ns” but once followed by “nsB”. Please, this needs clarification. R4 - Thank you for your comment, we made the changes as suggested; regarding the use of lowercase and uppercase letter, as well as “ns”, the explanation is in the table footnote.
5. Scientific names when reported for the first time in the text should be written in full with Authority and systematics; g. Hermetia illucens L. (Diptera: Stratiomyidae). R5 - Thank you for your comment, we made the changes as suggested.
6. The topic is quite relevant to the field as it addresses a specific gap in the field. Yet, in the introductions, the authors would preferably have more focus on the usefulness of Hermetia illucens for recycling organic waste and generating animal feed. This would more highlight the value of their research. R6 - Thank you for your comment, we made the some changes accordingly. However, we have to mention that he purpose of this study was to highlight the value of frass as an organic fertilizer and not the BSF, hence we did not focus the introduction on that subject; however, there is reference to the interest in this dipteran as a biodigester and its potential (line 77 – 90).
7. Some typos, mistakes, and misprinted letters, e.g. “Furthermore, BSFF, preferably as a mixed fertilizer (in a proportion until 75%), was shown to be a promising alternative for Gleyic podzol either in production of ryegrass as in the resilience and promotion of soil fertility.” Instead of “Furthermore, BSFF, preferably as a mixed fertilizer (in a proportion until 75%), shown to be a promising alternative for Gleyic podzol either in production of ryegrass as in the resilience and promotion of soil fertility.” R7 - Thank you for your comment, we made the changes as suggested.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The authors have adequately addressed the main technical issues; however, two minor points still merit attention. Although the environmental contexts differ, the framework proposed in “Dual effects of Caragana korshinskii introduction on herbaceous vegetation in Chinese desert areas: short-term degradation and long-term recovery” is conceptually relevant to your study. It illustrates how short-term declines can evolve into long-term functional recovery—a perspective directly aligned with the “residual effect” you discuss. Citing this work would enrich your theoretical grounding and highlight the temporal dimension of soil fertility responses beyond immediate nutrient changes.

In addition, while you have effectively expanded the discussion on trace-metal risks and included related references, the study “Influence of soil characteristics on the concentrations of Cr, Pb, and Zn in tobacco leaves from Longyan, China, and the associated predictive models” appears to be omitted. Including this citation would further enhance your discussion of metal accumulation mechanisms and comparative modeling across soil conditions.

Author Response

The authors have adequately addressed the main technical issues; however, two minor points still merit attention. Although the environmental contexts differ, the framework proposed in “Dual effects of Caragana korshinskii introduction on herbaceous vegetation in Chinese desert areas: short-term degradation and long-term recovery” is conceptually relevant to your study. It illustrates how short-term declines can evolve into long-term functional recovery—a perspective directly aligned with the “residual effect” you discuss. Citing this work would enrich your theoretical grounding and highlight the temporal dimension of soil fertility responses beyond immediate nutrient changes.

In addition, while you have effectively expanded the discussion on trace-metal risks and included related references, the study “Influence of soil characteristics on the concentrations of Cr, Pb, and Zn in tobacco leaves from Longyan, China, and the associated predictive models” appears to be omitted. Including this citation would further enhance your discussion of metal accumulation mechanisms and comparative modeling across soil conditions.

R. I appreciate the suggestion; however, this study does not focus on the aspects mentioned in the article you cited. The introduction of vegetation in arid environments may have different behaviors than the use of organic fertilizers in different soil types, as well as the issue of climate. This study aims to contribute to more comprehensive studies, both under controlled conditions and in the field, under different edaphoclimatic conditions. In fact, some of the evaluations carried out, such as production or enzymatic activity, allow us to infer about the quality of the product tested; however, there were more specific parameters that we were unable to quantify due to various limitations. Nevertheless, the results we were able to obtain seemed interesting and could be a contribution to research in the area.

Reviewer 3 Report

Comments and Suggestions for Authors

Accepted

Author Response

There is no comment

R. Thank you