Effect of Plant Growth Promoting Rhizobacteria on the Development and Biochemical Composition of Cucumber under Different Substrate Moisture Levels

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

The manuscript is not well written, has many errors such as line 77 where they stated (Strains were re-inoculated). The seeds itself are inoculated not the strains. Also, lines 79:81 the design is factorial in RCBD. There is no mention of the multiple comparison method which was used to compare among treatments. There are no tables at all in the manuscript as well as significance letters are badly arranged. Where is the information about the interaction?.

Comments on the Quality of English Language

English Language should be improved.

Author Response

Replay Reviewer 1

1. The manuscript is not well written, has many errors such as line 77 where they stated (Strains were re-inoculated). The seeds itself are inoculated not the strains.

Answer

Manuscript was revised, mistakes were corrected, i e. line 77

Change

“Strains were re-inoculated at 15 days after sowing (DAS), applying 15 mL of each rhizobacterium on the stem base at a concentration of 1 x 10⁸ CFU mL^-1”

By

“Plants were re-inoculated at 15 days after sowing (DAS), applying 15 mL of each rhizobacterium on the stem base at a concentration of 1 x 10⁸ CFU mL^-1”

 

2. Also, lines 79:81 the design is factorial in RCBD. There is no mention of the multiple comparison method which was used to compare among treatments.

Answer

In lines 125-127 the comparison method was mentioned, a section was added explaining the ANAVA.

Change

“The data obtained were subjected to normality Shapiro and Wilk’s test. The variables that showed normality in their data were subjected to a one-way analysis of variance (ANOVA); separation of measures was performed with Tukey’s test (p < 0.05).”

By

“2.6. Statistical Analyses

The data obtained were subjected to normality Shapiro and Wilk’s test. The variables that showed normality in their data were subjected to a one-way analysis of variance (ANOVA); separation of measures was performed with Tukey’s test (p < 0.05).”

 

3. There are no tables at all in the manuscript as well as significance letters are badly arranged.

Answer

Tables were not considered necessary in the submitted manuscript, important letters were corrected.

 

4. Where is the information about the interaction?.

Interactions between bacteria were not analyzed; this work was aimed on the individual behavior of each microorganism. The study of bacterial consortia is not ruled out in future research.

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript addressed how plant-associated bacteria ehance growth and antioxidant activity in cucumbers. The manuscript is generally good, however, the following points need to be considered before acceptance.

1- The title can be improved to accurately express the manuscript (Boosting Growth and Antioxidant Defenses in Cucumbers with Plant-Friendly Bacteria).

2-Abstract: Specific data about plant growth promotion should be included. 

Introduction: The motivation/novelty of the work should clearly addressed. Introduction should be substantiated with latest relevant work.

3- The rationale of selecting the bacterial strains (Pseudomonas paralactis

(KBendo6p7), Sinorhizobium meliloti (KBecto9p6) and Acinetobacter radioresistens) should be discussed throughly with citation of relevent studies.

4- I wonder why didn't the authors try the bacterial strains as a consortium.

5- Figures including the positive impact on the plant growth parameters should be added in the results section. 

6-   section (2.4. Nitrogen and Phosphorus Content in Plant and Statistical Analyses) should be splitted into two (2.4 Nitrogen and Phosphorus Content in Plant ) and 2.5 Statistical Analyses

7- S. meliloti has a  typical role in nitrogen fixation for legumes raises the question of whether this strain possesses specific adaptations for cucumbers. It would be valuable to discuss if the observed benefits are unique to this interaction or if they represent a more generalizable effect of S. meliloti on plant growth.

8-  Please elaborate more on why PGPR-moisture interaction wasn't significant for P uptake.

9- Please highlight the potential of increased antioxidants to scavenge free radicals and protect cells under stress.

10. The scientific names in the references sould be italic (e.g., reference 1, 4, 19, 33). Please check all the references. 

 

Comments on the Quality of English Language

Minor editing of English language required

Author Response

Replay Reviewer 2

1. The title can be improved to accurately express the manuscript (Boosting Growth and Antioxidant Defenses in Cucumbers with Plant-Friendly Bacteria).

Authors consider the title appropriate and reflects the work carried out.

2. Abstract: Specific data about plant growth promotion should be included. 

Answer

Specific data about plant growth promotion were included.

Change

“Abstract: Cucumis sativus L. plants are subjected to stress during production, affecting their growth, development, and fruit quality due to abiotic factors. Thus, the aim of this research is to evaluate Pseudomonas paralactis, Sinorhizobium meliloti and Acinetobacter radioresistens plant growth-promoting rhizobacterial (PGPR) effect on C. sativus plants under three substrate moisture levels (100, 75, and 50%). A randomized complete block design (RCBD) was used with A x B arrangement: (1) factor A (inoculant) with four levels; (2) PGPR and control (without bacterium); (3) factor B (substrate moisture content) with three levels. Height, root length, and fresh weight increased by inoculating PGPR. The phenol, flavonoid, and antioxidant capacity contents were statistically different (p < 0.05) in plants with PGPR while statistical differences were observed in flavonoid and antioxidant capacity contents (p < 0.05). Nevertheless, PGPR interaction and substrate moisture content only showed statistical differences in nitrogen assimilation, therefore, they may be considered as alternative to obtain vigorous C. sativus plants.”

By

“Abstract: Cucumis sativus L. plants are subjected to stress during production, affecting their growth, development, and fruit quality due to abiotic factors. Thus, the aim of this research is to evaluate Pseudomonas paralactis, Sinorhizobium meliloti and Acinetobacter radioresistens plant growth-promoting rhizobacterial (PGPR) effect on C. sativus plants under three substrate moisture levels (100, 75, and 50%). A randomized complete block design (RCBD) was used with A x B arrangement: (1) factor A (inoculant) with four levels; (2) PGPR and control (without bacterium); (3) factor B (substrate moisture content) with three levels. Plant height, root length, and fresh weight increased by inoculating PGPR (121%, 135% and 134% respectively); likewise, these variables increased with higher moisture content (177%, 204% and 234% respectively), while the effect of the interactions of the PGPR and the moisture content in the substrate showed statistical differences in plant height increasing of 197% and root length 267%. On the other hand, the content of phenols, flavonoids and antioxidant capacity was statistically different (p≤ 0.05) in inoculated plants with PGPR increasing by 117%, 126% and 150% respectively, likewise; In the moisture content of the substrate, statistical differences were observed, increasing the flavonoid content (114%) and antioxidant capacity (116%). Regarding the assimilation of nitrogen, it was higher by 274% and phosphorus by 124% with the PGPR inoculation, the moisture content increased the nitrogen content un plant (257%) and phosphorus content in plant (135%), with significant differences (p≤ 0.05); However, the interaction of PGPR and moisture content only presented statistical differences in nitrogen assimilation. PGPR can be considered as an alternative to obtain vigorous cucumber seedlings.”

 

 

Introduction: The motivation/novelty of the work should clearly addressed. Introduction should be substantiated with latest relevant work.

Change citations

4. Li, J.; Wang, X.; Lin, X.; Yan, G.; Liu, L.; Zheng, H.; Zhao, B.; Tang, J.; Guo, Y.D. Alginate-derived oligosaccharides promote water stress tolerance in cucumber (Cucumis sativus L.). Plant Physiology and Biochemistry 2018, 130, 80-88.
5. Kaur, G., Asthir, B. Molecular responses to drought stress in plants. Biol Plant 2017, 61, 201–209. https://doi.org/10.1007/s10535-016-0700-9.
6. Sarker, U.; Oba, S. Drought stress effects on growth, ROS markers, compatible solutes, phenolics, flavonoids, and antioxidant activity in Amaranthus tricolor. Applied biochemistry and biotechnology 2018, 186, 999-1016. https://doi.org/10.1007/s12010-018-2784-5.

 

By

 

4. Roy, S., Kapoor, R., Mathur, P. Revisiting Changes in Growth, Physiology and Stress Responses of Plants under the Effect of Enhanced CO2 and Temperature. Plant and Cell Physiology 2024, 65, 4-19. https://doi.org/10.1093/pcp/pcad121
5. Considine, M. J., Foyer, C. H. Oxygen and reactive oxygen species-dependent regulation of plant growth and development. Plant Physiology 2021, 186, 79-92. https://doi.org/10.1093/plphys/kiaa077
6. Suman, S., Bagal, D., Jain, D., Singh, R., Singh, I. K., Singh, A. Biotic stresses on plants: reactive oxygen species generation and antioxidant mechanism. In Frontiers in plant-soil interaction 2021, 381-411. https://doi.org/10.1016/B978-0-323-90943-3.00014-6
7. The rationale of selecting the bacterial strains (Pseudomonas paralactis (KBendo6p7), Sinorhizobium meliloti (KBecto9p6) and Acinetobacter radioresistens) should be discussed throughly with citation of relevent studies.

 

 

4. I wonder why didn't the authors try the bacterial strains as a consortium.

Answer

Interactions between bacteria were not analyzed; this work was aimed on the individual behavior of each microorganism. The study of bacterial consortia is not ruled out in future research.

 

5. Figures including the positive impact on the plant growth parameters should be added in the results section. 

Answer

The results show the increase in the variables expressed in percentages due to the treatments studied

 

 

6. Section (2.4. Nitrogen and Phosphorus Content in Plant and Statistical Analyses) should be splitted into two (2.4 Nitrogen and Phosphorus Content in Plant ) and 2.5 Statistical Analyses

Answer

Sections suggested was added.

Change

“2.4. Nitrogen and Phosphorus Content in Plant and Statistical Analyses

Nitrogen content in plants was determined by the method proposed by Bremmer and Keeney [28], whereas phosphorus content was assessed by Olsen and Sommers’ [29] method. The data obtained were subjected to normality Shapiro and Wilk’s test. The variables that showed normality in their data were subjected to a one-way analysis of variance (ANOVA); separation of measures was performed with Tukey’s test (p < 0.05).”

 

By

“2.4. Nitrogen Content in Plant

Nitrogen content in plants was determined by the method proposed by Bremmer and Keeney [28].

 

2.5. Phosphorus Content in Plant

Phosphorus content was assessed by Olsen and Sommers’ [29] method.

 

2.6. Statistical Analyses

The data obtained were subjected to normality Shapiro and Wilk’s test. The variables that showed normality in their data were subjected to a one-way analysis of variance (ANOVA); separation of measures was performed with Tukey’s test (p < 0.05).”

7. meliloti has a  typical role in nitrogen fixation for legumes raises the question of whether this strain possesses specific adaptations for cucumbers. It would be valuable to discuss if the observed benefits are unique to this interaction or if they represent a more generalizable effect of S. meliloti on plant growth.

Answer

1. meliloti is a free living microorganism. It effects with other crops was studied. i.e.

Galleguillos, C., Aguirre, C., Barea, J. M., & Azcón, R. (2000). Growth promoting effect of two Sinorhizobium meliloti strains (a wild type and its genetically modified derivative) on a non-legume plant species in specific interaction with two arbuscular mycorrhizal fungi. Plant Science, 159(1), 57-63.

Gou, J. Y., Suo, S. Z., Shao, K. Z., Zhao, Q., Yao, D., Li, H. P., ... & Rensing, C. (2020). Biofertilizers with beneficial rhizobacteria improved plant growth and yield in chili (Capsicum annuum L.). World Journal of Microbiology and Biotechnology, 36, 1-12.

Msimbira, L. A., Naamala, J., Antar, M., Subramanian, S., & Smith, D. L. (2022). Effect of microbial cell-free supernatants extracted from a range of pH levels on corn (Zea mays L.) and tomato (Solanum lycopersicum L.) seed germination and seedling growth. Frontiers in Sustainable Food Systems, 6, 789335.

Orozco-Mosqueda, M. D. C., Flores, A., Rojas-Sánchez, B., Urtis-Flores, C. A., Morales-Cedeño, L. R., Valencia-Marin, M. F., ... & Santoyo, G. (2021). Plant growth-promoting bacteria as bioinoculants: attributes and challenges for sustainable crop improvement. Agronomy, 11(6), 1167.

 

8. Please elaborate more on why PGPR-moisture interaction wasn't significant for P uptake.

Answer

In line 303-309 the topic is addressed.

9. Please highlight the potential of increased antioxidants to scavenge free radicals and protect cells under stress.

 

In lines 317-325 the topic is addressed

“Research on stress effect due to drought evidence that the presence of reactive oxygen species (ROS), such as superoxide, hydrogen peroxide, hydroxyl free radicals, including free oxygen, react with proteins, lipids, and DNA, altering the normal functioning of the cells. In this sense, chloroplasts, mitochondria, and peroxisomal signal are the first front of cellular defense facing oxidation, in studies on non-enzymatic antioxidants, the results have evidenced the increment of total phenol, flavonoid, and antioxidant capacity as defense mechanisms facing stress by drought, mainly by reducing hydrogen peroxide [6]”

10. The scientific names in the references sould be italic (e.g., reference 1, 4, 19, 33). Please check all the references.

Answer

format of scientific names in all references was changed.

 

Reviewer 3 Report

Comments and Suggestions for Authors

In general, I like this paper, but there are several quations og presentation that can be much improved.  While reading I made the folloing notes:

Fig 1;  the substrate moisture should be given for a) because as shown in  b),  this has a larger effect than PGPR.  In particular, the control in 1 a) is about 8 cm, like the 75% moisture in 1 b).  And in para. 3.1, line 131 : plant height 177%, percent of what? Presumably of the 50% moisture column.  All this is not clear.

Simialr criticisms apply to line 156.  Perhaps clarified by adding moisture (50%?)

Fig 5 a) again this needs to have the moisture %age added.

Similarly for Fig 6 a) and probably for Fig 7a) and 8a)and 9a) and 11a)

Fig 10 shows the most striking effect of PGPR even at 50% moisture;  commpare to the other results of hight etc, this seems to jsutify more comment.

There are a number of msitkes in English presentation, such as
line 306, where the sentence has no meaning.

The only really striking results that confirms the Conclusion is Fig 10.  I did not find the other Figs convincing, mainly because of lack of data in the a) parts of the figures mentioned.

Comments on the Quality of English Language

The langauge is slightly uneasy in places. I picked out one meaningless sentence, but there are others that need to be checked.

Author Response

Replay Reviewer 3

Fig 1;  the substrate moisture should be given for a) because as shown in  b),  this has a larger effect than PGPR.  In particular, the control in 1 a) is about 8 cm, like the 75% moisture in 1 b).  And in para. 3.1, line 131 : plant height 177%, percent of what? Presumably of the 50% moisture column.  All this is not clear.

Simialr criticisms apply to line 156.  Perhaps clarified by adding moisture (50%?)

Fig 5 a) again this needs to have the moisture %age added.

Similarly for Fig 6 a) and probably for Fig 7a) and 8a)and 9a) and 11a)

Fig 10 shows the most striking effect of PGPR even at 50% moisture;  commpare to the other results of hight etc, this seems to jsutify more comment.

There are a number of msitkes in English presentation, such as line 306, where the sentence has no meaning.

The only really striking results that confirms the Conclusion is Fig 10.  I did not find the other Figs convincing, mainly because of lack of data in the a) parts of the figures mentioned

 

Answer

The increased values ​​in the evaluated variables (and their corresponding figure) were corrected by adding the highlighted percentages for each treatment. Corrected substrate moisture factor names to differentiate.

The manuscript underwent an English language review to correct errors.