Effect of Rhizobacteria Application on Nutrient Content, Bioactive Compounds, Antioxidant Activity, Color Properties and Fruit Characteristics of Strawberry Cultivars

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Dear Editor, dear authors,

I have read carefully the paper entitled “Effect of Rhizobacteria Application on Nutrient Content, Bioactive Compounds, Antioxidant Activity, Color Properties and 3 Fruit Characteristics of Strawberry Cultivars”. Article is well written, it is readable, the concept is easy to follow, the scope of work is defined properly and the literature used is contemporary. Statistical analysis significantly contributes to the quality of work

I suggest that the paper should be accepted after a minor correction explained bellow:

Abstract

Line 27

Authors should write the full name of PGPR since it is mentioned for the first time in the text.

Line 29

Authors should change „used as bacteria“ into „used PGPR“

Line 29

What is the unit „cc“?

Keywords

It is not necessary to have „bacteria“ as a keyword.

Material and Methods

Line 131-133

Authors should specify which numbers from the microorganism collections were used?

Line 135

Authors should explain which unit is cc?

Line 140

The authors should explain what the control group is?

Results

Lines 210-211

The authors state twice „fruit weight and fruit length.

Comments for author File: Comments.pdf

Author Response

Responses to Comments of Reviewer 1

General Response:

First, we thank the potential reviewer for her/his valuable comments and suggestions. In this step of revision, we have tried to respond to all comments and addressed all questions. All revised parts have been highlighted in yellow. We hope the revised version of the manuscript gets positive feedback from you and will be acceptable for publication in the Processes journal.

Sincerely.

Comments

Comments 1# I have read carefully the paper entitled “Effect of Rhizobacteria Application on Nutrient Content, Bioactive Compounds, Antioxidant Activity, Color Properties and 3 Fruit Characteristics of Strawberry Cultivars”. Article is well written, it is readable, the concept is easy to follow, the scope of work is defined properly and the literature used is contemporary. Statistical analysis significantly contributes to the quality of work.

I suggest that the paper should be accepted after a minor correction explained bellow:

Response to Comment 1#

We would like to thank Reviewer 1 for her/his valuable contributions.

Abstract

Comments 2# Line 27

Authors should write the full name of PGPR since it is mentioned for the first time in the text.

Response to Comment 2#

The full name of PGPR was written (plant growth promoting rhizobacteria (PGPR))

Comments 3# Line 29

Authors should change „used as bacteria“ into „used PGPR“

Response to Comment 3#

The words "as bacteria" were replaced with " PGPR".

Comments 4# Line 29

What is the unit „cc“?

Response to Comment 4#

Per reviewer suggestion, the standard unit "mL" was used instead of the unit "cc" (cubic centimeters).

Keywords

Comments 5# It is not necessary to have „bacteria“ as a keyword.

Response to Comment 5#

The word "bacteria" was replaced with "Bacterial inoculation".

Material and Methods

Comments 6# Line 131-133

Authors should specify which numbers from the microorganism collections were used?

Response to Comment 6#

The method section revised.

Comments 7# Line 135

Authors should explain which unit is cc?

Response to Comment 7#

Per reviewer suggestion, the standard unit "mL" was used instead of the unit "cc" (cubic centimeters).

Comments 8# Line 140

The authors should explain what the control group is?

Response to Comment 8#

The sentence “The experimental group without bacterial inoculation was considered as the control group.” was added.

Results

Comments 9# Lines 210-211

The authors state twice „fruit weight and fruit length.

Response to Comment 9#

The second words “fruit weight, fruit length” were removed in the sentence.

Reviewer 2 Report

Comments and Suggestions for Authors

Submitted article “Effect of Rhizobacteria Application on Nutrient Content, Bioactive Compounds, Antioxidant Activity, Color Properties and Fruit Characteristics of Strawberry Cultivars” represents original research conducted with an aim to determine the effects of single and combined applications of PGPR bacteria on plant nutrition, biochemical content and fruit characteristics in Albion and Monterey strawberry cultivars. The research is methodologically well established, and the results are statistically validly processed. Also, the achieved results were compared in detail with the relevant results of other authors. The biggest shortcoming of the work is the lack of discussion, at least at the assumption level, about the mechanisms of bacterial influence on the content of macro and microelements, fruit weight, fruit size and biochemical characteristics of strawberry varieties. I am of the opinion that it is absolutely necessary to add data on the origin of the used microorganisms and their identification to the work. It is also necessary to specify accession number under which 16S rDNA sequence is deposited in the GenBank repository or data on the storage of microorganisms in one of the official culture collections. Also, it is necessary to add data on the cultivation conditions for each microorganism, which nutrient broth was used and what were the process parameters - intensity of aeration, mixing, temperature, etc. It is necessary to clarify whether the influence of the total cultivation liquid was tested or whether the biomass of microorganisms was separated and its suspension was prepared for tests. My suggestion is to use SI units throughout the work (ml instead of cc etc.). Without detailed data on microorganisms, the work is not complete, but with the requested additions and corrections, it can be considered for publication.

Author Response

Responses to Comments of Reviewer 2

General Response:

First, we thank the potential reviewer for her/his valuable comments and suggestions. In this step of revision, we have tried to respond to all comments and addressed all questions. All revised parts have been highlighted in turquoise. We hope the revised version of the manuscript gets positive feedback from you and will be acceptable for publication in the Processes journal.

Sincerely.

Comments

Comments 1# Submitted article “Effect of Rhizobacteria Application on Nutrient Content, Bioactive Compounds, Antioxidant Activity, Color Properties and Fruit Characteristics of Strawberry Cultivars” represents original research conducted with an aim to determine the effects of single and combined applications of PGPR bacteria on plant nutrition, biochemical content and fruit characteristics in Albion and Monterey strawberry cultivars. The research is methodologically well established, and the results are statistically validly processed. Also, the achieved results were compared in detail with the relevant results of other authors. The biggest shortcoming of the work is the lack of discussion, at least at the assumption level, about the mechanisms of bacterial influence on the content of macro and microelements, fruit weight, fruit size and biochemical characteristics of strawberry varieties. I am of the opinion that it is absolutely necessary to add data on the origin of the used microorganisms and their identification to the work. It is also necessary to specify accession number under which 16S rDNA sequence is deposited in the GenBank repository or data on the storage of microorganisms in one of the official culture collections. Also, it is necessary to add data on the cultivation conditions for each microorganism, which nutrient broth was used and what were the process parameters - intensity of aeration, mixing, temperature, etc. It is necessary to clarify whether the influence of the total cultivation liquid was tested or whether the biomass of microorganisms was separated and its suspension was prepared for tests. My suggestion is to use SI units throughout the work (ml instead of cc etc.). Without detailed data on microorganisms, the work is not complete, but with the requested additions and corrections, it can be considered for publication.

Response to Comment 1#

The method section has been expanded according to Reviewer 2's suggestion.

“Bacillus subtilis, Bacillus megaterium and Paenibacillus polymyxa were used as bacteria in the areas selected for the experiment. All bacterial strains were obtained from Dr. Metin Turan (The culture collection of the Department of Genetics and Bioengineering, Faculty of Engineering and Architecture at Yeditepe University Istanbul, Türkiye). These bacteria have been reported in the past to have properties that promote plant development and to have the potential to function as biocontrol agents against a broad variety of plant diseases [Turan et al., 2012, Esitken et al., 2006]. Bacteria were grown on nutrient agar by the streak plate inoculation method and kept at 27 °C for 48 hours, and a single colony taken from the cultures that completed their growth at the end of this period was transferred to 250 mL of nutrient broth with 15% glycerol-containing flasks. They had grown aerobically in flasks on a rotating shaker (150 rpm) for 48 hours at 27 °C (Merck KGaA, Darmstadt, Germany). The bacterial suspen-sions were then diluted in sterile distilled water to a final concentration of 108 CFU mL-1. The resultant bacterial suspensions were used for treatment. In the treatment, 10 mL of each bacterial species was applied to the soil where Albion and Monterey cultivars were grown. Bacillus megaterium M3, Bacillus subtilis OSU-142, Paenibacillus polymyxa PGPR bacteria were applied as single treatments. In addition, a mixture of equal amounts of these three bacterial species was applied as a mixed treatment. The study was carried out with a total of four treatments and one control group. The experimental group without bacterial inoculation was considered as the control group.”

Reviewer 3 Report

Comments and Suggestions for Authors

It is necessary to expand the introduction regarding the treatment of different cultivars, as well as the experimental approach, which unfortunately has not been addressed compared to similar publications of interest, nor discussed satisfactorily based on the experimental data obtained.

The project is original precisely because the same type of treatment was carried out on different cultivars. However, it should be noted that the sample population is rather small to proceed with a statistically significant data analysis. Your final dataset turned out to be redundant as the number of variables describing the objects (samples) is much greater than the samples themselves. Additionally, many variables are collinear, which heavily affects the robustness of the PCA model developed. PCA proves to be an effective technique for selecting significant variables and thus proceeding to develop a more robust and consequently more reliable model. Consequently, the PCA you presented is incomplete, as the analysis of the loadings can provide important information that, apparently, has been overlooked. These insights could have been more evident by visualizing the score plot against the principal components.

Compared to what is reported according to your literature review, it can be easily noticed that your data are often not in complete agreement, particularly when, as in your case, the treatment of the two cultivars did not yield similar results but rather, in some cases, opposite ones.

The discussion of the data resulting from the experiments related to the treatment of two cultivars has been neglected, even though both the cluster analysis data tables and the PCA analysis highlight interesting details that distinguish the behavior of the two cultivars with respect to the treatments you considered. The information extracted from the literature is not in complete agreement with the experimental data, especially when considering the two cultivars separately.

In principle, the work is interesting, the study criteria are correct, but the execution and especially the interpretation and discussion of the data prove to be rather lacking and poor in analytical support. Consequently, the bibliographic references should also be consistent with what the experimentation tends to highlight based on the experimental data.

Comments for author File: Comments.pdf

Author Response

Responses to Comments of Reviewer 3

General Response:

First, we thank the potential reviewer for her/his valuable comments and suggestions. In this step of revision, we have tried to respond to all comments and addressed all questions. All revised parts have been highlighted in pink. We hope the revised version of the manuscript gets positive feedback from you and will be acceptable for publication in the Processes journal.

Sincerely.

Comments

Comments 1# It is necessary to expand the introduction regarding the treatment of different cultivars, as well as the experimental approach, which unfortunately has not been addressed compared to similar publications of interest, nor discussed satisfactorily based on the experimental data obtained.

Response to Comment 1#

We thank the referee for their valuable feedback. Based on their feedback, improvements have been made to the manuscript. The experimental approach used has been discussed more comprehensively by comparing it with similar literature and has been related to the experimental data obtained. In this way, discussions regarding both literature and experimental data have been presented in a more satisfactory manner. The text has been updated by making necessary revisions.

Comments 2# The project is original precisely because the same type of treatment was carried out on different cultivars. However, it should be noted that the sample population is rather small to proceed with a statistically significant data analysis. Your final dataset turned out to be redundant as the number of variables describing the objects (samples) is much greater than the samples themselves. Additionally, many variables are collinear, which heavily affects the robustness of the PCA model developed. PCA proves to be an effective technique for selecting significant variables and thus proceeding to develop a more robust and consequently more reliable model. Consequently, the PCA you presented is incomplete, as the analysis of the loadings can provide important information that, apparently, has been overlooked. These insights could have been more evident by visualizing the score plot against the principal components.

Response to Comment 2#

We thank the referee for his feedback. We think that the originality of our study is that the same type of application is carried out on different cultivars. However, we would like to make some explanations about the criticisms regarding the sample size and number of variables.

Regarding the determination that the sample size is small, the cultivars used in our study were selected from a limited population and evaluated considering certain criteria. However, we aimed to address this limitation by optimizing the number of variables and determining important variables with PCA analysis. In this way, we aimed to develop a more powerful model. In addition, we think that our PCA model effectively reveals the distribution and relationships of the existing data. We think that the score graph (Figure 3) presents our findings more clearly and understandably by addressing the loading analyses more comprehensively against the collinearity problem and emphasizing important findings.

Comments 3# Compared to what is reported according to your literature review, it can be easily noticed that your data are often not in complete agreement, particularly when, as in your case, the treatment of the two cultivars did not yield similar results but rather, in some cases, opposite ones.

Response to Comment 3#

The fact that the results we obtained in our study do not fully coincide with the findings in the literature is actually an indication of the original nature of our experiment. The genetic structure and responses to environmental factors of the two different strawberry varieties used in our study (Albion and Monterey) produced different results with the applied PGPR (plant growth promoting rhizobacteria). These different responses between varieties emphasize the role of genetic diversity and environmental interactions. This situation shows that, contrary to the results observed in the literature, different genotypes can give different responses to similar treatments. As stated in the literature, it is known that each strawberry variety can give different responses to environmental conditions, microbial treatments used or nutrient uptake.

The fact that our results differ from the literature indicates that our study has original findings and that strawberry varieties give different responses to PGPR treatments. These differences reveal the importance of conducting more in-depth studies with a wider genotypic variety and under different environmental conditions.

Comments 4#

The discussion of the data resulting from the experiments related to the treatment of two cultivars has been neglected, even though both the cluster analysis data tables and the PCA analysis highlight interesting details that distinguish the behavior of the two cultivars with respect to the treatments you considered. The information extracted from the literature is not in complete agreement with the experimental data, especially when considering the two cultivars separately.

Response to Comment 4#

Thank you for your feedback. With these revisions, we have significantly improved the manuscript and produced a study that is better integrated with the literature, with a more comprehensive interpretation of the analysis results. Your feedback has been very valuable in improving the quality of our study. We tried to reveal the differences on a species basis more clearly by comparing the findings with the studies in the literature.

Comments 5# In principle, the work is interesting, the study criteria are correct, but the execution and especially the interpretation and discussion of the data prove to be rather lacking and poor in analytical support. Consequently, the bibliographic references should also be consistent with what the experimentation tends to highlight based on the experimental data.

Response to Comment 5#

Thank you for your feedback. With these revisions, we have significantly improved the manuscript and produced a study that is better integrated with the literature, with a more comprehensive interpretation of the analysis results.

Reviewer 4 Report

Comments and Suggestions for Authors

Line 211 - There seems to be a repetition of "fruit weight" and "fruit length". Please revise.

I do not know if it can be applied, but if so, can you add the standard variation values to the tables?

This article is very well written, both scientifically and grammatically. 

Comments on the Quality of English Language

Very well written.

Author Response

Responses to Comments of Reviewer 4

General Response:

First, we thank the potential reviewer for her/his valuable comments and suggestions. In this step of revision, we have tried to respond to all comments and addressed all questions. All revised parts have been highlighted in green. We hope the revised version of manuscript gets positive feedback from you and will be acceptable for publication in the Processes journal.

Sincerely.

Comments

Comments 1# Line 211 - There seems to be a repetition of "fruit weight" and "fruit length". Please revise.

Response to Comment 1#

The second words “fruit weight, fruit length” were removed in the sentence.

Comments 2# I do not know if it can be applied, but if so, can you add the standard variation values to the tables?

Response to Comment 2#

Thank you for your suggestion. However, we believe that it is not necessary to add standard deviations or standard errors in order to understand the tables presented in our study more clearly and to prevent readers from having difficulty in a crowded number pile.

Comments 3# This article is very well written, both scientifically and grammatically.

Response to Comment 3#

We would like to thank Reviewer 4 for her/his valuable contributions.