Response of Shoot Growth to Ecological Factors Highlights a Synergistic Relationship Between Yield and Catechin Accumulation in Tea Plant (Camellia sinensis L.)

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Line 81: factors might have not 'factors might has'

Clear the lines 158-160. 

Line 236: this part is very important and should be highlighted in the discussion and conclusion section: 'both high light intensity and moderate substrate water deficit can independently promote shoot growth.'

Line 325: 'The curved surface models with high fitting degree suggested that the growth ability of tea shoots in response to ecological factors affected total catechins content. In conclusion, the growth of tea shoots exerts a synergistic relationship with catechin accumulation in response to variations in ecological factors. '

I have a strong opinion that the theses were shifted here. Namely, in the lines 277 - 279 it is stated that 'Interestingly, the diameter and maximum length of shoots showed a negative correlation with five catechin indicators', also in the lines 296-299 the authors state that 'In summary, leaf number, shoot length, shoot ratio, and shoot diameter were significant factors influencing catechins accumulation. Specifically, the shoot ratio and leaf size significantly affect the total non-esterified catechins (TNEC) and total esterified catechins (TEC) content, respectively.'

The authors need to show full maturity here, otherwise I shall conclude that they are not certain what they are investigating. It is not the growth that influenced catechins accumulation, it is the environment that modifies both the vegetative response and the biochemical response. You did have a hint on it (In conclusion, the growth of tea shoots exerts a synergistic relationship with catechin accumulation in response to variations in ecological factors), but the manuscript is written as if the vegetative growth influenced the adaptive - defensive response.

It needs to be rewritten entirely in the light of the findings - in the young phase (as stated 'bud sprouting and leaf thickening in tea plants have no significant effect on catechin accumulation'), while the maturation (which authors named 'growth') brought the immune and mature response of the plants towards the variable environments. 

Line 436: 'Delving deeper into the regulatory basis for this observed coordination between 
growth and catechin content, the findings reveal that key shoot development-related 
genes...', again I would not name it growth, but the maturation or growth and development to make sure that it is not the green mass- vegetative growth solely, but the whole plant response to the environmental influence. 

Author Response

Comments 1: Line 81: factors might have not 'factors might has'

Response 1: We sincerely apologize for the grammatical error and have revised it to 'factors might have'. In addition, the entire manuscript has been polished by native English speakers.

Comments 2: Clear the lines 158-160.

Response 2: We are extremely grateful for your careful reading. We have deleted lines 158-160 and checked the manuscript once again to avoid the same mistakes.

Comments 3: Line 236: this part is very important and should be highlighted in the discussion and conclusion section: 'both high light intensity and moderate substrate water deficit can independently promote shoot growth.'

Response 3: Thank you for your constructive feedback. (1) To highlight this part in the discussion, we have rewritten Section 4.1 and supplemented the relevant references (lines 372-399). (2) We have highlighted the important part in the conclusion section (lines 447-450).

Comments 4: Line 325: 'The curved surface models with high fitting degree suggested that the growth ability of tea shoots in response to ecological factors affected total catechins content. In conclusion, the growth of tea shoots exerts a synergistic relationship with catechin accumulation in response to variations in ecological factors. ' 

I have a strong opinion that the theses were shifted here. Namely, in the lines 277-279 it is stated that 'Interestingly, the diameter and maximum length of shoots showed a negative correlation with five catechin indicators', also in the lines 296-299 the authors state that 'In summary, leaf number, shoot length, shoot ratio, and shoot diameter were significant factors influencing catechins accumulation. Specifically, the shoot ratio and leaf size significantly affect the total non-esterified catechins (TNEC) and total esterified catechins (TEC) content, respectively.'

The authors need to show full maturity here, otherwise I shall conclude that they are not certain what they are investigating. It is not the growth that influenced catechins accumulation, it is the environment that modifies both the vegetative response and the biochemical response. You did have a hint on it (In conclusion, the growth of tea shoots exerts a synergistic relationship with catechin accumulation in response to variations in ecological factors), but the manuscript is written as if the vegetative growth influenced the adaptive - defensive response.

It needs to be rewritten entirely in the light of the findings - in the young phase (as stated 'bud sprouting and leaf thickening in tea plants have no significant effect on catechin accumulation'), while the maturation (which authors named 'growth') brought the immune and mature response of the plants towards the variable environments.

Response 4: We sincerely apologize for the inappropriate descriptions in the manuscript and appreciate the constructive feedback and the opportunity to revise the manuscript.

(1) Your perspective is well-taken: 'It is not the growth that influenced catechins accumulation, it is the environment that modifies both the vegetative response and the biochemical res' In fact, the results only reveal a coordinated response of shoot growth and catechin accumulation to environmental changes, with no evidence to support a causal relationship between them. Therefore, to clarify the findings, we have carefully revised the descriptions regarding 'growth that influenced catechins accumulation'in Section 3.4, as detailed below.

1. delete lines 284-286 of the original manuscript;
2. revise the content of lines 287-292 of the original manuscript;
3. revise 'significant effects on'to 'significant relationship with' in line 263;
4. delete lines 331-333 of the original manuscript;
5. revise the summary in the lines 277 - 281.

(2) It is very scientific to consider the relationship between growth phases and metabolism from different stages. Inspired by your suggestions, we have added relevant descriptions (lines 263-265). 'These results indicated that the relationship between the morphology and biochemical components of tea plants depended on their growth phases'. In addition, to avoid the analysis without evidence in the results, relevant speculations and literature have been added to the discussion section. (lines 412-419).

Comments 5: Line 436: 'Delving deeper into the regulatory basis for this observed coordination between growth and catechin content, the findings reveal that key shoot development-related genes...', again I would not name it growth, but the maturation or growth and development to make sure that it is not the green mass- vegetative growth solely, but the whole plant response to the environmental influence.

Response 5: Thank you for your suggestion. We have changed "growth" to "growth and development" (line 425).

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Dear Authors,

The manuscript by Xiang et al titled "Response of Shoot Growth to Ecological Factors Highlights a Synergistic Relationship between  Yield and Catechin Accumulation in Tea Plant (Camellia sinensis L.) " study the effects of ecological factors combination on growth and phytochemical accumulation in tea. Below are the comments for manuscript:

1. line 94-95 
Clarify the stable growth mention here (After reaching a stable 94
growth stage..........). Eg, is there any growth scale that can be used for tea growth?
2. Why using only one reference primers for qPCR?
3. Line 156-157 repetitive 160. Remove 156-157, and manage structure of line 160?
4. Concise line 162-204 in 2 paragraphs.
5. Line 83-85 "To validate this hypothesis, experiments were conducted in artificial climate chamber, involving variations in light intensities (L150, L250, L350, L450, and L550 µmol·m⁻²·s⁻¹), relative air humidity (AH40%, AH50%, AH70%, and AH90%), are those paramaters constant in all chambers. In our experience, within the groth chamber there is range of light intensity and relative humidity received by each plant, it all depends on the size of chamber, proximity of light source to plants etc. 

Comments on the Quality of English Language

The quality of english are as follows:

i) Easy to follow

ii) Less grammatical errors 

iii) Need to remove some duplicate sentences

iv) Need to make paper compact by merging paragraphs that talk about similar information.

Author Response

Comments 1: line 94-95 Clarify the stable growth mention here (After reaching a stable 94 growth stage..........). Eg, is there any growth scale that can be used for tea growth?

Response 1: We sincerely appreciate your valuable feedback.

i) In this study, 'the stable growth' means 'tea plants had successfully rooted and developed 2-3 new shoots'. These quantifiable benchmarks ensured that all experimental plants were in a comparable physiological state before the application of different ecological factor treatments. To avoid potential misunderstandings, we have revised this description in the manuscript (lines 98-99) . The transplanted (A) and surviving tea cuttings (B) are shown in the following picture (Please see the attachment).

ii) In addition, the pruning standards before planting have also been supplemented in Section 2.1(lines 93-98). 'The cultivation protocol followed the optimized methods developed by Fujian Zhongke Biology Co., Ltd. Specifically, tea cuttings were pruned to 18-20 cm in height, with excess shoots removed to retain only 4-6 mature leaves on the stem, and fibrous roots were excised. The pruned cuttings were disinfected with a 1% potassium permanganate solution for 1 min before being planted in pots (16 cm in diameter, 18 cm in height).'

Comments 2: Why using only one reference primers for qPCR?

Response 2: Two reference gene primers (GAPDH and β-actin) were used for qPCR, but the CT values of β-actin are higher than those of GAPDH, indicating that GAPDH performed better as a reference gene in the study. The expression levels of the two reference genes across different samples are attached at the end of the text (Please see the attachment).

Comments 3: Line 156-157 repetitive 160. Remove 156-157, and manage structure of line 160?

Response 3: Thank you for your careful reading and constructive feedback. ① We have deleted lines 156-157 of the original manuscript. In addition, the manuscript has been checked once again to avoid the same mistakes. ② We have simplified the structure of Section 3.1 into two coherent paragraphs.

Comments 4: Concise line 162-204 in 2 paragraphs.

Response 4: Thanks for your valuable suggestions. (i) We have simplified lines 162-204. The content of the first paragraph now focuses on the effects of ecological factors on shoot growth, and the second paragraph presents principal component analysis and comprehensive conclusions. (ii) Inspired by your suggestions, we have merged fragmented paragraphs into coherent sections with relevant content in Sections 3.2, 3.3, 3.4, and 3.5.

Comments 5: Line 83-85 "To validate this hypothesis, experiments were conducted in artificial climate chamber, involving variations in light intensities (L150, L250, L350, L450, and L550 µmol·m⁻²·s⁻¹), relative air humidity (AH40%, AH50%, AH70%, and AH90%), are those paramaters constant in all chambers. In our experience, within the growth chamber there is range of light intensity and relative humidity received by each plant, it all depends on the size of chamber, proximity of light source to plants etc.

Response 5: Your observation is correct, and thank you for your kind reminder. We emphasized the range of relative air humidity but overlooked other environmental factors. Therefore, we have added the ranges of environmental factors to the notes of Supplementary Table 2. In addition, we have provided explanations in the Materials and Methods section (lines 110-113).

Comments 6: Comments on the Quality of English Language. The quality of english are as follows:

i) Easy to follow

ii) Less grammatical errors

iii) Need to remove some duplicate sentences

iv) Need to make paper compact by merging paragraphs that talk about similar information.

Response 6: Thank you for your detailed feedback on the English language quality. We have carefully addressed the points raised as follows:

i) delete line 156-157, and carefully check and ensure that there are no duplicate sentences in the manuscript.

ii) merge paragraphs containing similar information. The original multiple paragraphs in Sections 3.1, 3.2, and 3.4.1 were consolidated into two paragraphs per section. Similarly, fragmented paragraphs in Sections 3.3, 3.4.2, 3.5.1, and 3.5.2 were merged into one paragraph per sect

iii) The entire manuscript has been polished by native English speakers.

 

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Thank you for implementing the necessary changes.