Root-System Architectures of Two Cuban Rice Cultivars with Salt Stress at Early Development Stages

Round 1

Reviewer 1 Report

In this manuscript, the author studied the root-system architectures of two Cuban rice cultivars with salt stress at early development stages. The author analyzed root architecture traits of two rice cultivars (INCA LP-5 and Perla de Cuba) at early plant development in response to salt, phosphorus (P) limitation, and a brassinosteroid. Seeds were placed in I) paper rolls for seven days and II) mini-rhizotrons for 21 days without or with salt (50 mM NaCl), without or with 24-epibrassinolide (10-6 M) pre-treatment, and with two levels of P (10 or 1 ppm). The root system of LP-5 was larger in size and extent, while the roots of Perla were growing denser. Salt affected mainly the size- and extent-related root characteristics and explained about 70% of the variance. The effect of P was more pronounced without salt treatment. In Perla, P supply reduced the salt effect on root growth. The brassinosteroid had hardly any effect on the development of the plants in both experiments. Due to the high dependence on experimental factors, root length and related traits can be recommended for selecting young rice cultivars regarding salt stress and P deprivation.

The manuscript is very well written, and it covers all the experimental aspects. However, for the betterment of this manuscript, I have few suggestions for the authors mentioned below:

Change at

L19 The brassinoisteroid had hardly any effect to The brassinosteroid had hardly any effect.

L90 material with an optimal sowing in to material with optimal sowing in.

At L99, it is pretreatment, whereas at L14, L187, L191, L203, L276, L421, it is pre-treatment, which is true? In this manuscript, pre-treatment words came seven times, whereas pretreatment words came three times. Choose the same word for the whole manuscript.

L124 with low concentration of P to with a low concentration of P.

L130 shoot fresh weight was measured and a root image acquired to shoot fresh weight were measured and a root image was acquired.

L163 radarcharts to radar charts?

L164 was used with z-scored data over all treatments and both cultivars to was used with z-scored data overall treatments, and both cultivars.

L169 Although, the shoot weight to Although the shoot weight.

L211 whereas in Perla the P concentration to whereas in Perla, the P concentration.

Change from salt treated to salt-treated in all-over manuscript.

L213 Sentence looks incomplete.

L239 showed a similar behavior to showed similar behavior.

L244 showed more divers reactions to showed more diverse reactions.

L350 varies with clear differences to vary with clear differences

L433 widely in dependence of growing conditions to widely in dependence on growing conditions.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Reviewer 2 Report

Changes in root architecture are believed to play important role in adaption of plants to their environment. The reviewed article reports on the results of comparison of the effects of salt stress, P deficit and treatment with brassinosteroid (BR) applied to rice cultivars separately and in combination. The article convincingly demonstrates applicability of root image analysis using software packages for the study of plant responses related to their salt tolerance. This report is likely to be useful in breeding for plant resistance and therefore interesting for readers. Still I have some recommendations for improvement of the article.

I have some general recommendations:

1. Significance of the difference in distinct characteristics between the cultivars is not indicated either in the tables of figures, where significance of the effects of factors is shown for each cultivar separately. Authors should extend ANOVA they used including LSD or Duncan test. This allows proving significance of the difference between all the means, which are usually marked with letters, different letters indicating significantly different means.
2. There are a lot of abbreviations in the text, which are deciphered in the Table 1. However, I think that it is not convenient for readers to return to the table each time they would need to remember that the abbreviations mean. I advice to decipher them each time in the figure and table legends. Illustrations should be self-sufficient.
3. It is unclear, which criteria are considered to be important in terms of resistance. In some places it seems that authors consider maintenance of shoot weight as criteria of plants resistance, but if this is so, it should be stated clearer
4. Authors should read and cite reports differentiating osmotic and toxic effects of salinity (articles of Munns and Tester may be useful). The reported delay in toxic effects of ions may be useful for explaining less detrimental effects of salt on young seedlings compared to elder plants detected in the present research.
5. It is frequently unclear, which illustrations support certain statements. Reference to illustrations should not be omitted (especially in the Discussion).

 

My further detailed comments are as follows

 

1. The title of the article should be changed. In its present form it does not reflect the study of the effects of P deficit and BR treatments, which were significant parts of this research.
2. In the abstract, the study of the effects of phosphorus and BR treatments alongside with the salt stress is not justified. Some phrases relevant in this respect may be extracted from the introduction. If authors exceed the recommended size of the abstract, they may delete some details of obtained results.
3. Formulation of the aim of the present research should be improved. In the present form it just describes what was done. At least the word “comparison” of the effects of… on rice cultivars should be added.
4. Lines 47-48. “Salt stress often inhibits root growth more 48 than shoot growth, which reduces the root-to-shoot ratio” – some references are needed here.
5. Lines 89-91. It would be relevant to summarize that used cultivar differed in the optimal time for sowing.
6. Line 110. “Every treatment combination (cultivar × salt × BR) was replicated five times and placed in the same plastic bag” – the number of replicates and how many bags were used is unclear.
7. Line 169. “LP-5 than in Perla (average 63 mg and 55 mg, respectively, pCV < 0.05)”. It is unclear what pCV means and which approach was used for verifying significance of the difference between cultivars. Marking means with letters according to the results of LSD or Duncan test (see above) would make this comparison easier without the need of providing any comments in the brackets or figure legends.
8. Line 172. “Perla reacted with slightly higher shoot weights and slightly lower root weights” – although no statistic analysis of significance of the difference is provided, these responses do not look significant. I think it would be better to describe these results as showing capacity of the plants to maintain shoot and root growth under conditions of salt stress at the level of the control (unstressed) plants.
9. Lines 174-175. “Root-to-shoot ratio was slightly, but not significantly lower when plants were treated with salt” – The effects was obviously statistically insignificant and should not be described in this way.

• Line 176, 207-208, 297. “The RL was higher in LP-5 than in Perla”. “In both cultivars, the P concentration was reduced at P depletion”. “LP-5 developed higher biomasses and a larger root system during the experiments” – I think it important to mention that these effects were detected in the absence of salt stress. In the presence of NaCl the difference between cultivars seems insignificant.
• Lines 209-211. “In LP-5, the BR pre-treated group without salt and with P had about twice the P concentration (5.2 mg g-1) than the non-treated control group (2.3 mg g-1), whereas in Perla the P concentration with BR (1.9 mg g-1) was about half of the non-treated without salt and with P (3.4 mg g-1).” – cannot this be described simpler? I propose “BR increased P concentration in LP-5 and decreased it in Perla”.
• Variability of P concentration in the plants should be somehow explained. Otherwise it produces an impression of inaccurate measurement.

13. Line 245. “Here, TOO, the influence of NaCl and P on the variance of the traits was in general 245 high in LP-5” – I do not understand what “too” is doing here. I failed to find any discussion of variability of the data above.

• Line 246. “In Perla, BR treatment significantly influenced maximum and median number of roots” – I failed to find the data on medium number of roots (MeNR) in table 3.

15. Lines 341-342. “Considering the lower reduction of shoot biomass due to salt stress for Perla than LP-5, under this experimental conditions a lower root-to-shoot ratio seems to be beneficial” – The sentence sounds as if Perla had lower root-to-shoot mass ratio than LP-5. But it seems to me that the difference was not statistically significant (see my comments above about necessity of statistical analysis of significance of the difference between cultivars and presentation of its results as letter mark of means).

• Line 378-379 “Under salt stress, the NS increased for Perla, which was not found for LP-5” – but increased net root area should increase Na uptake by the plants thereby reducing salt resistance, which seems opposite to what authors think about Perla.

I hope my comments won’t discourage the authors. I actually like the article and just wanted to help authors to improve it. I hope my comments will be useful.

Author Response

Please see the attachment.

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Authors followed all my comments and revised the text in accordance. I am quite satisifed. I only advise authors to use Duncan test in the future. It allows comparison between two cultivars and different number of treatments. It helps to mark significantly different means with different letters, which allows comparison between all variants. If you use Statistica program, it shows how to apply Duncan test

This manuscript is a resubmission of an earlier submission. The following is a list of the peer review reports and author responses from that submission.