Sugar Receding in Aril Benefits the Recalcitrant Seeds of Litchi (Litchi chinensis) and Longan (Dimocarpus longan) to Cope with Dry Spells after Maturation
and Zhaoqi Zhang 1,3,*
Round 1
Reviewer 1 Report
Comments and Suggestions for AuthorsThe work is interesting, but requires additions:
1. Which spectrophotometer was used, which centrifuge....
no specification of the apparatus which is essential for laboratory analyses.
2. No summary or conclusion, please complete this part of the work
3. No full stop at the end of the sentence in line 591
4. please present statistical analyses, which are hardly visible in the work
5. The discussions would also need to be broadened, with more reference to the available literature.
Comments for author File: 
Comments.pdf
Author Response
- Which spectrophotometer was used, which centrifuge....no specification of the apparatus which is essential for laboratory analyses.
Response：Thanks for the comment. We have added some specification for the essential apparatus, such as their lot numbers and company information in the revised manuscript. We also added the parameters of the analyses, such as optical wavelength for the spectrometry, and the temperature and spin speed for the centrifugation.
- No summary or conclusion, please complete this part of the work
Response：We have added the conclusion part in the revised manuscript.
- No full stop at the end of the sentence in line 591
Response：We have added the full stop at the end of that sentence.
- please present statistical analyses, which are hardly visible in the work
Response：Thanks for the comments. You are right that the labels in some of the figures are hardly visible, e. g. Figure 2 D, H, and the labels in figure 4, the lines are fade and the asterisks are too small. Based on these defects, we have checked and improved these labels in all the figures in the revised manuscript.
- The discussions would also need to be broadened, with more reference to the available literature.
Response：Thanks for the comments. The present work aims to study the mechanism of sugar receding on fruit quality. And we found that the sugar receding was involved in the adaptation to environment for the species propagation. It is an interdisciplinary subject of horticultural science and ecology. We consider that too broad topic may make the Horticulturae readers more complicated to understand. Thus, we would like to leave the discussion as it was, hope you can agree with our consideration.
Author Response File: 
Author Response.docx
Reviewer 2 Report
Comments and Suggestions for AuthorsThe article is devoted to present sugar receding in aril benefits the recalcitrant seeds of litchi and longan to cope with dry spells after maturation.
Overall, the article is interesting. However, there are some concerns.
Scientific names should be presented in italics.
In Materials and Methods
Page 3 line 132 : “80℃” or “-80℃” ?
Page 4: “2.6 Seed germination and seedling establishment rate” the author should provide a definition of " Seed germination and seedling establishment"
In Results
Page 5 lines 208-210: “The whole fruit development from the flower bloom to complete fruit drop after maturation last around 110 and 170 days for the litchi and longan respectively” Figure 1 F-I showed 101 and 159 days.
Page 5 line 221-222: “As shown in Figure 1E-F, the aril was initiated at around 52 DAA (May 21th) for the litchi and at 61 DAA (June 18th) for the longan” “Figure 1E-F” should be “Figure 1F-G”
Page 5 line 280-282: “However, the seedling lengths continuously increased to the maximum at 131 DAA, and then a slight drop of the lengths was found at 159 DAA” Seedling length at 131 DAA was significantly decrease compared to 159 DAA.”
Page 8 line 306-307: “more than 90% of the total fruit drop occurs in the early developmental period” Figure 3 A-B shows after 87 and 117 DAA, which is the period after fruit ripening.
Page 9 Figure 3: The order in which the figures are presented should begin with the dropped fruit, germinated seeds, to seedling establishment. (C, E, D, F, H and G)
In Discussion
Page 12 line 451-452: “we found that seeds matured at around 60 DAA for the litchi and at around 75 DAA for the longan, when the arils of the fruits had just started to grow (Figure 1F-G; 2A and E).”
Seed ripening should be 80 and 131 DAA bases on seed weight (Fig. 1 H-I) or 73 and 131 DAA bases on seed vigor (Fig. 2 D-H).
Or the author should provide a definition of "Ripe Seeds" in Materials and Methods
Page 12 line 460-462: “the long duration of seed vigor maintenance on-tree enhance the probability for seed germination and seedling establishment. This special trait evolved by litchi and longan trees can spread the risk over a relatively long period with unpredictable dry weather, which is a good plant regeneration strategy.”
Page 12 line 485-489: “Accordingly, the traits of continuous fruit drop and high seed vigor maintenance on tree of litchis and longans provide multiple germinating times during the season, spread the risk in a relatively long period in unpredictable dry weather, to avoid the high risk of mortality of the seedlings when an excessive single-time fruit drop occasionally encounters a long duration of continuous drought.
However, after fruit ripening the higher drop rate makes seed collection difficult. What is the best way to handle it?
The results of the experiment should be conclused.
Comments for author File: Comments.pdf
Author Response
- Scientific names should be presented in italics.
Responses: Thanks for the comment. We have checked all the scientific names and revised them in italics.
In Materials and Methods
- Page 3 line 132 : “80℃” or “-80℃” ?
Responses: Thanks for the comment. We are sorry that we had made a mistake and “-“ was missing. We have corrected it as “-80℃” in the revised manuscript.
- Page 4: “2.6 Seed germination and seedling establishment rate” the author should provide a definition of " Seed germination and seedling establishment"
Responses: Thanks for the comments. We have added the definition of " Seed germination and seedling establishment" in the revised manuscript. “The seed germination was regarded when the radicle protruded around 2 mm from the seed coat, and the emergence of the first pair of true leaves from the stem above the soil was regarded as the beginning of seedling establishment”.
In Results
- Page 5 lines 208-210: “The whole fruit development from the flower bloom to complete fruit drop after maturation last around 110 and 170 days for the litchi and longan respectively” Figure 1 F-I showed 101 and 159 days.
Responses: Thanks for the comment. We are sorry that the description in the legend was not accurate. By observation, we found the whole period from flower bloom to complete fruit drop last around 110 and 170 days as in the text. However, we collected the last data at 101 and 159 DAA for the litchi and longan respective in the work. After that time point, till 110 and 170 DAA, most of the fruits on tree fell (please seed figure 3 of this work) and the fruit left were not enough for the all the assays for figure 1-2. We have corrected the description in the legend in figure 1 and 2.
- Page 5 line 221-222: “As shown in Figure 1E-F, the aril was initiated at around 52 DAA (May 21th) for the litchi and at 61 DAA (June 18th) for the longan” “Figure 1E-F” should be “Figure 1F-G”
Responses: Thanks for the comment. We have corrected the citing for the figures in the revised text.
- Page 5 line 280-282: “However, the seedling lengths continuously increased to the maximum at 131 DAA, and then a slight drop of the lengths was found at 159 DAA” Seedling length at 131 DAA was significantly decrease compared to 159 DAA.”
Responses: Thanks for the comment. We understand what you mean, and the sentences suggested by you will be better. But you might confuse with the DAA. So, we revised as “The seedling lengths continuously increased to the maximum at 131 DAA, and the length at 159 DAA was significantly decreased compared to 131 DAA”
- Page 8 line 306-307: “more than 90% of the total fruit drop occurs in the early developmental period” Figure 3 A-B shows after 87 and 117 DAA, which is the period after fruit ripening.
Responses: Thanks for the comment. We are sorry that we did not describe clearly. It is known that more than 90% young fruit drop in the early developmental stage. Here we determined the fruit drop rate at the late stages (only after 52 and 61 DAA for the litchi and longan respectively), in another word, we did not count the drop fruits at the early stages (0-52/61 DAA). Thus, we referred the fruit drop rate at the first time point (52/61 DAA) as 0%, and determined the fruit drop rate afterward. When focusing the fruit drop at the late stage (52/61-108/173 DAA), it is true that most of the fruits dropped after 87/131 DAA. We have added some description in the text to avoid the confusion.
- Page 9 Figure 3: The order in which the figures are presented should begin with the dropped fruit, germinated seeds, to seedling establishment. (C, E, D, F, H and G)
Responses: Thanks for the comment. We have changed the order of the images as you suggested.
In Discussion
- Page 12 line 451-452: “we found that seeds matured at around 60 DAA for the litchi and at around 75 DAA for the longan, when the arils of the fruits had just started to grow (Figure 1F-G; 2A and E).”
Seed ripening should be 80 and 131 DAA bases on seed weight (Fig. 1 H-I) or 73 and 131 DAA bases on seed vigor (Fig. 2 D-H).
Or the author should provide a definition of "Ripe Seeds" in Materials and Methods
Response: Thanks for the comment. We have defined “ripe seeds” of litchi and longans in the “Materials and Methods “ of the revised manuscript, in “2.6 Ripe seed definition, seed germination and seedling establishment rate”, as “The seeds are regarded as ripe seeds when they present around 90% seed germination and 80% seedling establishment.” Thus, we have revised the sentence here as “we found that the seeds matured around 2-weeks earlier than the fruits”.
- Page 12 line 460-462: “the long duration of seed vigor maintenance on-tree enhance the probability for seed germination and seedling establishment. This special trait evolved by litchi and longan trees can spread the risk over a relatively long period with unpredictable dry weather, which is a good plant regeneration strategy.”
Page 12 line 485-489: “Accordingly, the traits of continuous fruit drop and high seed vigor maintenance on tree of litchis and longans provide multiple germinating times during the season, spread the risk in a relatively long period in unpredictable dry weather, to avoid the high risk of mortality of the seedlings when an excessive single-time fruit drop occasionally encounters a long duration of continuous drought.
However, after fruit ripening the higher drop rate makes seed collection difficult. What is the best way to handle it?
Response: Thanks for the comment. In this work, we focus on the role of aril on the seed vigor maintenance. Regarding to the seed collection, we consider that the seeds can be collected at any time after seed maturation till complete fruit drop.
- The results of the experiment should be conclused.
Response：We have added the conclusion part in the revised manuscript.
Author Response File: Author Response.docx
Reviewer 3 Report
Comments and Suggestions for AuthorsLine 31 and 117: Italize the species names
Line 71: rephrase: 'or might be terrible'
The introduction section and M&M section were well written.
The results section is partly written as if it were results and discussion, please rewrite it accordingly (like 362-364 and similar).
Although the figure 1 is very beneficial there is to much data for one figure and it should be separated. Part of the content could be transformed into a graphical abstract.
The discussion section is also well-written and appropriately supported by the references.
Overall, this is an excellent study, thus I suggest adding a separate conclusion section to further highlight the obtained results.
Comments for author File: Comments.pdf
Author Response
-
- Line 31 and 117: Italize the species names
Responses: Thanks for the comment. We have italized all the species names in the revised version.
- Line 71: rephrase: 'or might be terrible'
Responses: Thanks for the comment. We rephrased as 'or the seedling might die when a long drought period occurs’ in the revised manuscript.
- The introduction section and M&M section were well written.
Thank you!
- The results section is partly written as if it were results and discussion, please rewrite it accordingly (like 362-364 and similar).
Responses: Thanks for the comment. We have deleted some over-writing in the result part.
- Although the figure 1 is very beneficial there is to much data for one figure and it should be separated. Part of the content could be transformed into a graphical abstract.
Responses: Thanks for the comment. You are right that there quite some data in figure 1. However, we consider that all the data are regarding to one topic. And organizing them together can help readers to understand the weather condition along the fruit growth and development. Thus, we decide to leave the figure 1 as it was.
- The discussion section is also well-written and appropriately supported by the references.
Thank you!
- Overall, this is an excellent study, thus
Thank you!
Author Response File: Author Response.docx
Reviewer 4 Report
Comments and Suggestions for AuthorsThe document presents a subject that does not represent a significant extension of general knowledge, so it does not contribute much in reference to more specific aspects. The tissues surrounding the seed have the function of providing protection and nutrients to the seed; this has been known for a long time.
On the other hand, the evaluation of aril sugars is carried out while the fruits are still on the tree; sugars are not evaluated during the periods when the fruits are separated from the tree and have fallen to the ground, where the aril tissues perform their function of nutrition, protection and moisture supply.
The content of the paper would be more interesting and congruent with the title if the authors would analyze a profile of sugars (by chromatography) and organic acids during the degradation of the fruit regids until the seed is uncovered and germination.
The authors do not present statistical evidence of correlation of variables showing that sugar modification favors seed resistance; correlation analyses are required.
In the document, the information is not presented in a congruent manner in relation to the permanence of the fruit on the tree, first presenting the development of growth and maturation of the fruit on the tree, followed by information on the development of germination and then returning to the topics related to the fruit on the tree. It is advisable to approach the topics of the fruit on the tree together in order to be able to compare them in a better way.
On the other hand, the authors should justify in a scientific manner why the aril and seed samples were stored at 80°C. Explain on a scientific basis the selection of this storage temperature.
Comments for author File: Comments.pdf
Author Response
-
- The document presents a subject that does not represent a significant extension of general knowledge, so it does not contribute much in reference to more specific aspects. The tissues surrounding the seed have the function of providing protection and nutrients to the seed; this has been known for a long time.
Response: Thanks for the comment. It is true that many previous studies have demonstrated that the tissues surrounding the seed have the function of providing protection, while much less studies provided evidence that the tissues can supply nutrients to the seeds. In the present study, we focus on a phenomenon called “sugar receding”, which reduce the fruit quality in the litchi and logan industry. And the mechanism underneath the phenomenon is under debating.
The findings in the present study not only further supply evidence that the surrounding tissues can supply nutrients to the seeds, but also reveal the mechanism of sugar receding that has been puzzled the litchi and longan industry. We found that the sugar receding of aril benefits the recalcitrant seeds of litchi and longan to cope with dry spells after maturation.
- On the other hand, the evaluation of aril sugars is carried out while the fruits are still on the tree; sugars are not evaluated during the periods when the fruits are separated from the tree and have fallen to the ground, where the aril tissues perform their function of nutrition, protection and moisture supply.
Response: Thanks for the comment. In the present study, besides evaluation of aril sugars in the fruits on tree, we did evaluate the sugar contents in the aril of the fruits at 7 days after separated from the trees ( Figure 4 AB, and EF). Further evaluation of the contents in the fallen fruits on the ground will be interesting.
- The content of the paper would be more interesting and congruent with the title if the authors would analyze a profile of sugars (by chromatography) and organic acids during the degradation of the fruit regids until the seed is uncovered and germination.
Response: Thanks for the comment. We agree with you that it will be better if we analyzed a profile of sugars and organic acids in the arils, such as sucrose, glucose and fructose etc.. It is known that the seed germination is dependent on the energy supplied by the various soluble sugars that are oxidized in respiration pathways. Accordingly, we consider that the total soluble sugar contents and TSS can represent the soluble sugar levels for energy supply. For sure, evaluation the sugar and organic acid profiles in the arils until the time point, when the seeds are uncovered and germination on the ground, will provide information regarding the sugar metabolism mechanism. Evaluation of the sugar or organic acid profiles in the whole process in our next work will further support our conclusion.
- The authors do not present statistical evidence of correlation of variables showing that sugar modification favors seed resistance; correlation analyses are required.
Response: Thanks for the comment. You are right that statistical correlation analyses would show the close relation between sugar modification and seed germination or vigor. However, we found that the sugar modification patterns are complicate for litchi and longan aril, which present first accumulation and then decreasing (receding) after fruit maturation. And as we mention in the manuscript, seed mature occurs earlier than fruit (aril) maturation. Furthermore, in the present study, we focus on the effect of sugar receding on seed germination and vigor. Thus, analyses of correlation between the data along the whole fruit development may obtain some confused conclusion. In addition, our conclusion is not only based on the data collected along the fruit development, but also based on the data collected after fruit separation from the trees and blockage the photosynthate supply. In the latter two experiments, the conclusion is evident even when no correlation analyses are presenting. Accordingly, we decide not to carry out the correlation analyses.
- In the document, the information is not presented in a congruent manner in relation to the permanence of the fruit on the tree, first presenting the development of growth and maturation of the fruit on the tree, followed by information on the development of germination and then returning to the topics related to the fruit on the tree. It is advisable to approach the topics of the fruit on the tree together in order to be able to compare them in a better way.
Response: Thanks for the nice suggestion. We also consider it helps to improve the logic of our results. In the revised manuscript, we have moved all the on-tree experiment results together based on your suggestion, for example , we have move the former 3.5 to 3.3.
- On the other hand, the authors should justify in a scientific manner why the aril and seed samples were stored at 80°C. Explain on a scientific basis the selection of this storage temperature.
Responses: Thanks for the comment. We are sorry that we had made a mistake and missed a “-”. We have corrected it as “-80℃” in the revised manuscript.
Author Response File: Author Response.docx
Round 2
Reviewer 4 Report
Comments and Suggestions for AuthorsThe manuscript can now be accepted for publication.
