Effects of Azorhizobium caulinodans and Piriformospora indica Co-Inoculation on Growth and Fruit Quality of Tomato (Solanum lycopersicum L.) under Salt Stress
Round 1
Reviewer 1 Report
Overall, the English language of the paper is not according to standards. It needs drastic revision by some expert. Moreover, there is use of very long sentences which the readers will not like. There is a need to break the long sentences into small sentences. The other minor comments are as follow:
Line 20: most menaces- This sentence need revision
Line 21-22: Delete the second sentence in abstract
Line 25-38. Break the long sentence into small sentences
Line 25: Add the authority along with scientific name
Line 40: Improved
Line 47-50: just write one sentence about the area of salinity in world
Line 122-123: Importance of tomato is in agronomy or its importance should be related to health?
Line 123-124: Just provide one citation and do not put full link
Line 139: Instead of professor name, better to mention name of laboratory. You can write professor name in acknowledgment.
Line 156: Replace the word sowed by sown
Line 180-183: English language needs improvement
Line 201: Do not start sentence with a digit. Same comment for line 244 and remaining parts of paper
Line 271: Give more details about statistical analysis
Line 291-294: revise the sentences
Author Response
Answers to Reviewer 1
Overall, the English language of the paper is not according to standards. It needs drastic revision by some expert. Moreover, there is use of very long sentences which the readers will not like. There is a need to break the long sentences into small sentences. The other minor comments are as follow:
Line 20: most menaces- This sentence need revision
Done!
Line 21-22: Delete the second sentence in abstract
Done!
Line 25-38. Break the long sentence into small sentences
Done!
Line 25: Add the authority along with scientific name
The full Latin name of A. caulinodans was introduced completely in Line 23.
Line 40: Improved
Done!
Line 47-50: just write one sentence about the area of salinity in world
Done!
Line 122-123: Importance of tomato is in agronomy or its importance should be related to health?
We deleted the sentence.
Line 123-124: Just provide one citation and do not put full link
The full link was deleted and the citation was listed in References
Line 139: Instead of professor name, better to mention name of laboratory. You can write professor name in acknowledgment.
Done!
Line 156: Replace the word sowed by sown
Done!
Line 180-183: English language needs improvement
Done!
Line 201: Do not start sentence with a digit. Same comment for line 244 and remaining parts of paper
Done!
Line 271: Give more details about statistical analysis
Done!
Line 291-294: revise the sentences
Done!
Reviewer 2 Report
In this manuscript, Xu and colleagues show the effect of co-inoculation of Azorhizobium caulinodans, a PGPB, and Piriformospora indica, a PGPF, to tomato plants, in control or salt stress conditions. The use of microorganisms to improve stress resistance in plants is very interesting, and some good results are already available in the literature. The use of microorganisms in agriculture is a promising option for a greener agriculture. However, although nice results are obtained by authors when inoculating the microorganisms, the manuscript must be improved in order to be published, in particular aspects concerning figures. Some of my comments are:
Major comments:
- The treatment with both microbial treatments (Azca + Piin) under control conditions is missing.
- Infection of tomato stems and roots by A. caoulinodans in Fig. 1A and 1B is not clear, I do not know what the arrows are pointing. Improve these images to clearly show bacteria cells. In addition, survival of microbes after salt treatment should be documented, pictures showed in the manuscript are taken before starting salt treatments.
- Shoot height and base diameter in Fig. 2 should be represented in different figures, and results corresponding to 1 and 2 weeks in the same. In this figure, letters indicating statistical differences in base diameter after two weeks of treatments are missing.
- The style should be the same for all figures, fig. 7B, 8B, 9 and 10 are different from the rest of the figures.
Minor comments:
- In materials and methods, the recipe for arabinose gluconate medium should be expressed in molar units, not in mg/l
- In MM, line 157: what is the composition of the sterilized culture medium bought from Jinrun Biotechnological and used to grow the plants?
- Line 158: 14 h light + 8 h darkness makes 22h, it should be 24h.
- Line 160: “After germination, the seedlings were selected, 2 seedlings per pot”. This sentence is confusing, reformulate.
- Line 161: What do you mean by “These seedlings were watered according to the moisture of culture medium”?
- What is the soil volume used in pots? Depending on this volume, 100 ml of sterile water to avoid salt accumulation could be enough or not.
- For Micrography, the camera used should be indicated in MM
- Line 228: Tlie should say The
- Line 207: were extracted
- Line 209: change “chlorophyll concentration” for “photosynthetic pigments concentration”
- Line 237: yeast-GR and DTNB should be fully written the first time they appear in the text.
- Line 293, 296, and throughout the text. Delete p<0.05, this should be indicated in figure legends, not in the text.
- English should be improved, there are many grammatical mistakes
- Letters A, B, C, etc in figures are not indicated.
- 3 is just a picture, it does not show differences in growth or tomato maturity. A scale bar or reference should be indicated in figures, as pictures are clearly not taken from the same distance.
- Line 354: Dabrowski et al., 2016
- In Fig. 5, separate soluble sugars from soluble proteins, make fig. 5A and B with data.
Author Response
Answers to Reviewer 2
In this manuscript, Xu and colleagues show the effect of co-inoculation of Azorhizobium caulinodans, a PGPB, and Piriformospora indica, a PGPF, to tomato plants, in control or salt stress conditions. The use of microorganisms to improve stress resistance in plants is very interesting, and some good results are already available in the literature. The use of microorganisms in agriculture is a promising option for a greener agriculture. However, although nice results are obtained by authors when inoculating the microorganisms, the manuscript must be improved in order to be published, in particular aspects concerning figures. Some of my comments are:
At first we thank the reviewer for his good opinions
Major comments:
The treatment with both microbial treatments (Azca + Piin) under control conditions is missing.
Based on our early observation, Azca and Piin can live well together, thus when we designed the whole experiment, we omitted the treatment (Azca+Piin) under control in order to reduce the number of treatments.
Infection of tomato stems and roots by A. caoulinodans in Fig. 1A and 1B is not clear, I do not know what the arrows are pointing. Improve these images to clearly show bacteria cells. In addition, survival of microbes after salt treatment should be documented, pictures showed in the manuscript are taken before starting salt treatments.
The inoculation of the two microbes was followed by salt stress treatment. Because A. caulinodans colonized in stems of tomato plants, salt stress treatment did not affect its colonization in stems.
In addition, all the pictures only showed the two microbes colonized in roots or stems, suggesting they certainly infected roots and stems. According to other research articles related to A. caulinodans and P. indica, we only confirmed their colonization in roots and stems.
Shoot height and base diameter in Fig. 2 should be represented in different figures, and results corresponding to 1 and 2 weeks in the same. In this figure, letters indicating statistical differences in base diameter after two weeks of treatments are missing.
The reviewer suggested another way to show results. Fig. 2 showed the results according to different harvest periods. We revised Fig. 2.
The style should be the same for all figures, fig. 7B, 8B, 9 and 10 are different from the rest of the figures.
All the figures clearly showed the experimental results, and statistical results were also showed. Thus they were not revised.
Minor comments:
In materials and methods, the recipe for arabinose gluconate medium should be expressed in molar units, not in mg/l
The arabionose gluconate medium was prepared according to a usual method, suggesting concentrations of all components in the medium in 1 L of the medium.
In MM, line 157: what is the composition of the sterilized culture medium bought from Jinrun Biotechnological and used to grow the plants?
The medium composition was: Perlite: peat: coconut shell powder=1:2:1
We have added the composition in the text.
Line 158: 14 h light + 8 h darkness makes 22h, it should be 24h.
So sorry for the mistake. We have revised it: 16 h light + 8 h darkness.
Line 160: “After germination, the seedlings were selected, 2 seedlings per pot”. This sentence is confusing, reformulate.
Done!
Line 161: What do you mean by “These seedlings were watered according to the moisture of culture medium”?
Because the culture medium possessed good water-holding capacity, these tomato seedlings were watered every week in their early growth period. As these tomato seedlings grew bigger and bigger, they consumed more and more water, thus water was provided according to moisture of the culture medium.
What is the soil volume used in pots? Depending on this volume, 100 ml of sterile water to avoid salt accumulation could be enough or not.
We did not know the soil volume and only weighed 1.5 kg of culture medium for one pot. In the revised text, we added the explanation.
According to MS degree treatise of Mr. Xu ZW (Yangtze University, 2021), we revised the treatment of salt stress, seen in the text.
Because the culture medium were often kept wet and 100 ml of sterile water was provided for avoidance of salt accumulation. Maybe 100 ml is not enough, but we saw part of water outflowed from the bottom holes of the pots.
For Micrography, the camera used should be indicated in MM
Done!
Line 228: Tlie should say The
Done!
Line 207: were extracted
Done!
Line 209: change “chlorophyll concentration” for “photosynthetic pigments concentration”
Done!
Line 237: yeast-GR and DTNB should be fully written the first time they appear in the text.
Done!
Line 293, 296, and throughout the text. Delete p<0.05, this should be indicated in figure legends, not in the text.
We have deleted p<0.05 and p>0.05 in the text and revised the captions of all the figures, as shown in the revised manuscript.
English should be improved, there are many grammatical mistakes
We have read the manuscript few times and got rid of these mistakes as possible as we could.
Letters A, B, C, etc in figures are not indicated.
In few of the figures, there were three or four small figures in them, letters A, B, C etc indicated small figures in the figures, respectively. Results presented in these small figures were mentioned, thus it is very convenient to find these results presented in the small figures with A, B, C, etc.
3 is just a picture, it does not show differences in growth or tomato maturity. A scale bar or reference should be indicated in figures, as pictures are clearly not taken from the same distance.
Fig. 3 presented the growth status of tomato plants under different treatments, and the results presented in Fig. 3 were only used to show growth status, acting in concert with these results shown in Fig. 4.
By the way, when we took the photos, we did not put a scale aside the pots, because the sizes of these pots were fixed and specific.
Line 354: Dabrowski et al., 2016
The spelling was correct!
In Fig. 5, separate soluble sugars from soluble proteins, make fig. 5A and B with data.
Fig. 5 is a simple histogram with two y axes, presenting changes in concentrations of soluble sugars and proteins. We did think that the figure should not be divided into two small figures. But we changed the colors of statistical results, as shown in the text.
Reviewer 3 Report
First of all, You need to pay attention to spaces between words and there are plenty of examples in the paper (e.g. Lines: 101,143,144,608,615,623,666,669,682,710).
Lines 212-215: Authors must provide a more detailed description of applied methods. There is no information about extraction. The listed references describe only the procedure for measurement on UV-VIS spectrophotometer.
Lines 217-230: Authors must provide information on how they calculated enzyme activity presented in Fig 6 from a direct reading of absorbance. There is no information about the extraction.
Line 259: Please provide more details on the applied method.
Line 263-265: Vanillin assay is a method for determining flavan-3-ols (catechin is flavan-3-ol) and not for the determination of anthocyanins as mentioned in the paper. Authors must change this.
Author Response
Answers to Reviewer 3
First of all, You need to pay attention to spaces between words and there are plenty of examples in the paper (e.g. Lines: 101,143,144,608,615,623,666,669,682,710).
We examined the spaces above mentioned. If the manuscript is accepted for publishing, there will not be these questions in its new version
Lines 212-215: Authors must provide a more detailed description of applied methods. There is no information about extraction. The listed references describe only the procedure for measurement on UV-VIS spectrophotometer.
Analysis methods for concentrations of soluble sugars and proteins are usual and simple ones, thus it is not necessary to describe their detailed processes
Lines 217-230: Authors must provide information on how they calculated enzyme activity presented in Fig 6 from a direct reading of absorbance. There is no information about the extraction.
We added the information, as shown in the text.
Line 259: Please provide more details on the applied method.
The determination of MDA concentrations is too easy and simple, and we thought that it was necessary to describe the whole process.
Line 263-265: Vanillin assay is a method for determining flavan-3-ols (catechin is flavan-3-ol) and not for the determination of anthocyanins as mentioned in the paper. Authors must change this.
The method was used to determine the concentrations of total proanthocynins in tomato fruits, not for anthocyanins.
We revised the mistake in the text.
Round 2
Reviewer 1 Report
Authors have incorporated all comments
Author Response
At first we thank the reviewer for his encouragement. We improved all the text in English language.
Author Response File: 
Author Response.docx
Reviewer 2 Report
Althought not all my concerns have been satisfied, authors have made an effort to make most of them or to explain why they consider not to do so. Therefore, I recommend this manuscript for publication in Horticulturae
Author Response
At first we thank the reviewer for his encouragement. We improved the two parts: results and conclusion, as shown in the text.
Author Response File: Author Response.docx
Reviewer 3 Report
In the sub section „ Assay of enzymes“ authors must provide an equation for the conversion of measured Absorbance (A) at the given wavelength to enzyme activity expressed in U/(g min) or U/g.
Author Response
We improved the experimental methods and added the three equations to express the activities of the enzymes, as show in the text.
Author Response File: Author Response.docx
