Application of a Protein Hydrolysate-Based Biostimulant Obtained from Slaughterhouse Sludge on Pepper Crops

Round 1

Reviewer 1 Report

The manuscript is nicely addressing key points in a well-organized approach. I believe that the following concerns deserve additional attention to improve the enthusiasm of readers (enclosed in the word document).

Comments for author File: Comments.pdf

I would like to state that the usage of the English language in some parts of the manuscript is problematic 

Author Response

Reviewer #1:

1. Line no. 56-57 only talks about one example of Ávila-Pozo et al. [5], please state other references as your current research can’t be based or supported by only one reference or only one work (that too by the same author). These and other studies (please refer to this ↓

https://www.sciencedirect.com/science/article/abs/pii/S2352554123000438

https://www.frontiersin.org/articles/10.3389/fsufs.2022.1073010/full

We have included the first reference you suggest. We think that the second reference should not be included as it is a biostimulant from animal manure together with dairy excreta (e.g., feces and urine), dairy products (e.g., milk and ghee), natural resources (e.g., honey), broken seeds or grains and their residues, inedible by-products. Therefore they are not slaughterhouse waste

 

2. It would be good to provide a definition for biostimulants as per European Regulation (EU, 2019/1009) in the introduction. It will make the research work more understandable.

We have included the definition of biostimulants according to the European Regulation (EU, 2019/1009).

 

3. Authors can add one paragraph at the end of the introduction to highlight the knowledge gaps and better justification why the present research is important.

Revised

 

4. Line 69 reference 16 and 17 are not relevant to the statement it supports. Kindly cite a relevant reference in support of the statement.

We don't understand your suggestion. We have read the indicated references again and they mention the nutritional compounds of pepper and its preventive effects on the highlighted diseases.

 

5. For line no 70-71 cite appropriate reference.

We have included the only existing reference on the use of this type of biostimulants in tomato crop

 

6. In the material methods, line no 102-103 please state the pepper crop growth condition like temperature, watering duration etc.

Irrigation has been included in the crop. Regarding the temperature, it is indicated that the conditions of the experiment were in a greenhouse at 25 º C.

 

7. Please state the full form of MAPA abbreviation (line no 130) and ICPS abbreviation (line no. 132)

We have included your suggestion about the ICPS abbreviation. On the other hand, we have observed that we made a mistake when including the reference to the N-Kejldahl determination. We have corrected this reference

 

8. For Table no 3. Plant height, it would be good if some photographic images are provided as the 95- and 140-day treatment showed significant improvement over the controls. These photographs will make the MS interesting for the reader.

We did not take photographs during the experiment and therefore cannot attach them to the manuscript.

9. In the line no. 263-264, the authors are requested to provide other references apart from Ávila-Pozo et al. [5] which also supports their findings.

Revised

 

10.Besides, Conclusion is not well brief in scientific way, many sentences telling the same things/meaning. please improve in precise way. The authors do not provide any perspective about using protein hydrolysate-based biostimulant on pepper crop.

We have rewritten this section including a few more sentences

 

11.Italicize scientific names in text.

Revised

 

12.The current manuscript has lots of faults including mistakes in English grammar and inaccuracy in science.

The manuscript has been reviewed by a native English speaker.

 

 

 

Author Response File: Author Response.docx

Reviewer 2 Report

It deals with biostimulation from protein hydrolysates, for economic purposes and free of fertilizers in the cultivation of peppers, showing important results to disseminate. however, some recommendations must be addressed before continuing with its processing.

etsan observations in the attached file.

Comments for author File: Comments.pdf

Author Response

Reviewer 2

1. Verify and correct the wording (Line 13-14, 23).

Revised

 

2. Could you provide more details about the composition and source of the slaughterhouse sludge? For instance, is it derived from a specific activity or activities at the facility, what type of organic solid waste it consists of, particle size, blood content, etc.?

We have included how slaughterhouse sludge is obtained and the constitution of the organic waste that causes this sludge.

 

3. Line 94-191. Add a reference explaining why these operational conditions were chosen (environmental factors, substrate type, content).

Comment and reference added

 

4. Line 168 – 170. It is not clear where these percentages can be found in the tables.

We have calculated all the percentages indicated in the Results section from the data shown

 

5. Line 197-200. Explain the mechanisms by which this behavior was observed.

Please specify what behavior

 

6. Lines 201-206. Explain and elaborate on the behavior of these results. In other words, why were certain values higher at higher doses, etc.?

The Discussion section explains why the treatment where the biostimulant was applied at a higher dose and directly to the substrate showed better results. It highlights greater root expansion, greater absorption of nutrients, particularly N, which leads to better mineral nutrition, greater concentration of photosynthetic pigments, greater production and quality of the fruit obtained. In the same way, an attempt is made to justify the reasons for these results.

 

7. Lines 22-235. In this section, the state of the art is described, but the specific experimental results are not analyzed and discussed. It is suggested to compare and analyze in more detail with the results obtained in the tables.

It is true that we generalize the discussion according to the results obtained. The reviewer suggests that we compare and specify the discussion based on the different treatments performed. However, we want the reviewer to understand that the results obtained were very homogeneous for all treatments. That is to say, it was seen in all the parameters analyzed that it was the treatment where the biostimulant was applied at a higher dose and via the root system that showed the best results. For this reason the discussion became generalized.

 

8. Based on the literature, it is understood that better results can be achieved with the application of any fertilizer or bio-stimulant. Analyze and discuss the importance of specifically applying the protein hydrolysate-based from slaughterhouse sludge on paper crops. Explain why it might be superior to others, considering the composition and source of this bio-stimulant, not just the dosage and application conditions.

We cannot indicate if this biostimulant is better than others. It is not our intention. With this manuscript we try to use a polluting by-product of the environment (slaughterhouse sludge) in a new biostimulant constituted by enzymatic hydrolysis. In the manuscript we highlight that the importance of the product is in its composition of low molecular weight peptides that are easily assimilated by the plant, and this has a positive impact on said plant.

 

9. At the end of the discussion, add how this bio-stimulant could be beneficial for other crops.

Revised

 

10. In conclusion, briefly include future work or perspectives on the widespread use of this bio-stimulant under real-world conditions, taking into account other environmental factors.

Revised

 

11. Improvement in English wording is required.

The manuscript has been reviewed by a native English speaker.

 

 

 

Reviewer 3 Report

The manuscript “horticulturae-2611436” entitled “Application of a protein hydrolysate-based biostimulant obtained from slaughterhouse sludge on pepper crop” by Ávila-Pozo et al. deals with an interesting subject regarding the response of a protein hydrolysate-based biostimulant obtained from slaughterhouse sludge in pepper plants when applied through root and foliar applications. The results of this study indicated higher values of various parameters, including plant height, number of flowers and fruits, macro and micronutrient content of photosynthetic pigments in the leaves, as well as nutritional content and vitamin C levels in the harvested fruits when the biostimulant was applied at a higher dose 1.4 g/L and through the root application.

 

For publication in “Horticulturae”, the topic and content are appropriate. Scientific content and the manuscript size are appropriate. The subject is very interesting and the authors did a lot of work, and the methodology used is adequate for the objectives of the study. The results are of interest and support the conclusions. When tables are shown, the units are correctly used. Quality of citations are appropriate. In general, the quality of the experiment is well performed and follows rigid scientific logic. However, there are some points that need attention in order for the article to be published:

 

·      The abstract is not written in a way to encourage the reader to read the whole paper. Specifically, the presentation of the key findings of experimental results should be improved and data regarding the mainly measured indicators should be presented.

·      Keywords: Please change some keywords. The title and keywords must not contain the same words.

·      Introduction: In this section, authors should further offer a unique perspective and analysis on the subject that has not been explored in other published material. In addition, they should emphasize a new layer of understanding and insight into the topic, which is valuable for readers.

·      Lines 63 and 97: As a scientific name of plant, Capsicum annuum must be italicized.

·      Materials and Methods: Authors must present in a table the physicochemical properties of the substrate used in their study. 

·      In addition, they must provide information about average lowest and highest recorded temperature, relative humidity and natural daylight cycle in greenhouse during the cultivation period of the research study.

·      Line 98: Please give more details about the plastic pots (height and diameter).

·      Lines 93-153: Authors must provide more information about the methods/materials (e.g., ICP test, UV-visible spectrophotometer and its manufacturer, etc.) they used for the determination of quality parameters in their study.

·       Tables 3-8: Authors must provide F-ratio and p-value for each measurement. In addition, define the treatments of the study in table notes. Furthermore, the correct way to present a mean value is: mean ± SE and letter (e.g., the correct is: “39.2 ± 1.3 a” instead of: “39.2 a ± 1.3”) 

·       The discussion section must be enhanced. The authors should further refer to previous studies with regard to effect of protein hydrolysate-based biostimulants on growth and quality parameters in pepper and other horticultural crops.

·       Finally, I highly recommend to enhance the conclusion section which should provide a good reflection on the results and novelty of this work, as well as your future prospects.

 

 

Thank you for your consideration.

The English language in this manuscript is good. Authors should further check for typesetting errors.

Author Response

Reviewer 3

- The abstract is not written in a way to encourage the reader to read the whole paper. Specifically, the presentation of the key findings of experimental results should be improved and data regarding the mainly measured indicators should be presented.

We have improved the conclusions of the Abstract. On the other hand, the reviewer suggests that data related to the measured indicators be included. In this sense, the authors want the reviewer to take into consideration the word limitation that the abstract has (imposed by the journal) and that would make us exceed the established word limit.

 

- Keywords: Please change some keywords. The title and keywords must not contain the same words.

We have tried to eliminate all the words that coincide with the title of the manuscript except the word "biostimulant", which we thought would be interesting to be part of the keywords.

 

- Introduction: In this section, authors should further offer a unique perspective and analysis on the subject that has not been explored in other published material. In addition, they should emphasize a new layer of understanding and insight into the topic, which is valuable for readers.

We do not understand the reviewer's suggestion. In the Introduction section we highlight that the use of this new biostimulant is unique. There are not many scientific works that use this biostimulant in crops. Specifically, only our research group used it in tomato cultivation (published) and now in green pepper. However, we have highlighted at the end of the section that this study is novel because there is currently no data on the use of this biostimulant obtained from sewage sludge by enzymatic processes in a green pepper crop.

.

- Lines 63 and 97: As a scientific name of plant, Capsicum annuum must be italicized.

Revised

 

- Materials and Methods: Authors must present in a table the physicochemical properties of the substrate used in their study.

Revised

 

- In addition, they must provide information about average lowest and highest recorded temperature, relative humidity and natural daylight cycle in greenhouse during the cultivation period of the research study.

To provide data on the lowest and highest recorded average temperature and relative humidity, we have included along with temperature and humidity the variability data of these parameters during the experiment. Regarding the natural light cycle in the greenhouse, we have indicated that the hours of light correspond to the hours of natural light correspond to the hours of light in the study area during the experimental period.

 

- Line 98: Please give more details about the plastic pots (height and diameter).

We have included the dimensions of the plastic pots

 

- Lines 93-153: Authors must provide more information about the methods/materials (e.g., ICP test, UV-visible spectrophotometer and its manufacturer, etc.) they used for the determination of quality parameters in their study.

Revised

 

- Tables 3-8: Authors must provide F-ratio and p-value for each measurement. In addition, define the treatments of the study in table notes. Furthermore, the correct way to present a mean value is: mean ± SE and letter (e.g., the correct is: “39.2 ± 1.3 a” instead of: “39.2 a ± 1.3”)

We have defined the study treatments in the table notes according to the reviewer's suggestion. On the other hand, we think that it is not necessary to provide F-ratio and p-value data. We think that with the data obtained and separated by homogeneous groups, the significant differences between the treatments can be observed. We hope that this differing opinion that we have with the reviewer is not a reason for rejection

 

- The discussion section must be enhanced. The authors should further refer to previous studies with regard to effect of protein hydrolysate-based biostimulants on growth and quality parameters in pepper and other horticultural crops.

We don't understand your suggestion. The text cites examples of authors who have used biostimulants obtained by enzymatic hydrolysis processes from various organic waste and their effect on plants. However, we have included some more examples.

 

- Finally, I highly recommend to enhance the conclusion section which should provide a good reflection on the results and novelty of this work, as well as your future prospects.

We have rewritten this section including a few more sentences

Round 2

Reviewer 3 Report

The text has been corrected according to my suggestions. Responses to comments are satisfactory. I recommend the manuscript be published in the Horticulturae journal.

The English language in this manuscript is good.