Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea?

Masoumeh Khalvandi; Mohammadreza Amerian; Hematollah Pirdashti; Sara Keramati

doi:10.1371/journal.pone.0254076

Peer Review History

Original SubmissionFebruary 5, 2021
24 Feb 2021 Decision Letter - Mayank Gururani, Editor PONE-D-21-02576 Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? PLOS ONE Dear Dr. Khalvandi, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by 10th March 2021. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. 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(Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: Critical comments General comments This is a well written article explaining the results of a well conducted experiment on synergistic influence of AM and Pirimorphospora sp. on salinity stress in Peppermint. The article has impressive findings, acceptable for publication in the esteemed Plos-1 journal. However, the language need minor corrections. It seems that the authors have some self-contradictory arguments in the discussion. The authors may do minor corrections giving due credit to the specific comments given below. Specific comments Page 1: Introduction: line 1-2: “which its demand has grown dramatically in recent decades” – may be corrected as: ‘the demand of which has grown dramatically in recent decades’ Line 3: “It is evident that peppermint essential oil” – may be modified as: ‘it is evident that the essential oil’ Line 5-6: “as well as the reactive oxygen species (ROS) scavenging activity because of its valuable constituents” – ‘as well as scavenging of reactive oxygen species (ROS)’ 2nd Para – line 1: - avoid ‘Nowadays’; the sentence may begin as ‘The sustainability …’ Line 5: no need of repeating – ‘reactive oxygen species (ROS)’, better put the short form only – ROS because the same is already defined above Line 7: “stresses; it can show’ – ‘stresses, because the same can show’ Page 3: 2nd para – 2nd line: not ‘sever’ – but ‘severe’ Page 4: para 3- line 2: not ‘fungi symbiosis’ – ‘fungal symbiosis’ Discussion: Page 4-5: line 1-4: “In the present study, the root colonization with P. indica and AMF were remarkably reduced by increasing salinity (6 to 9 dSm−1). Similar observations were reported in other plants (Wu et al., 2010; Hadian-Deljou, et al, 2020; khalvandi et al, 2019). The reason that salinity decreased fungal colonization might be attributed to the adverse 5 effects of salinity on photosynthesis, along with a reduction in carbon supply” The above statements seem contradictory to the claim below on page 5, para 5 – line 1-4: “Our findings showed that P. indica, AMF and their co-inoculation symbiosis mitigated the inhibitory effect of salinity on the photosynthetic capability of peppermint” Last para – line 1-2: “The endophytic fungus can mitigate the toxic influence of salinity on chloroplast and chlorophyll by releasing cytokinin-like substances which maintain the relative stability of the organelle” – how can the fungi mitigate the toxic influence, when the fungi themselves are negatively affected by salinity? “Nevertheless, endophytic fungi symbiosis mitigated the inhibitory effect of salt stress on the mineral uptake. It has been well documented that endophytic fungi can prevent the toxicity of Na+ in aerial parts of plants through accumulation of Na+ ions in fungal cytosol, hyphae wall and vesicles” – more explanations are required for this argument, especially when the fungi are harmfully affected by salt stress • Overall, the researchers have observed evidence for synergistic effect Pirimorphospora and AM in alleviating salt stress in peppermint; however, their arguments need more explanations to avoid self-contradictory appearance as they found salt stress also negatively affecting the plant metabolism as well as fungal colonization. Although, they provide evidences of previous positive findings of synergistic influence of AM and Pirimorphospora in certain plants, they need to report findings showing no such influences as well ( for example: DOI: 10.1080/01904160903435409 ) Reviewer #2: The study applied single P. indica, AMF and co-inoculation of these two endophytic fungi to investigate the potential of enhancing the ability of peppermint to alleviate the negative effect of the salinity stress. Therefore, they are sure interesting and meaningful. However, there are some issues to be addressed before the manuscript is ready for publication. Some language mistakes need to be revised and re-checked. Below are the few comments that can provide the glimpse of flaws in the manuscript. Q 1 [Introduction] Piriformospora indica, also named as Sebacinales indica, the author needs to add it. Q 2 [Materials and Methods] Give more specific info on inoculum: e.g. mycelia mass or chlamydospores per ml of P. indica and AMF. And provide the method or reference of fungi inoculation and detection. Q 3 [Results] This study has many figure, but author doesn’t or good explain the result of the figure. Q 4 [Discussion] The conclusions drawn by the authors that P. indica, AMF and their co-inoculation symbiosis mitigated the inhibitory effect of salinity on the photosynthetic capability of peppermint is not really supported by those parameters. The authors mention that higher photosynthetic capability attributed to the better physiological status. Why the author not measure the ROS scavengers related enzyme activity to support the point? Kindly explain. Q 5 The Fig. 2 was not clear, please replace it and provide the scale or microscope magnification. The microscopic observation pictures of the control plant roots are also need provided. “chlamidospore” probably “chlamydospore”? Please check it. Q 6 Fig. 3, add the abbreviation of Pi. Give the test sample number of root colonization of each group. Q 7 All the column figures in this study have no error bars, please check/add it. Q 8 Fig. 5 (a-c) different with other column figure like Fig. 4, please check/replace it. Q 9 Fig. 6, “AMF* Pi” or “Pi AMF”? “PI” or “Pi”? Same with Fig. 7, 8, 10, 12 and 13, keep one unified format. please check/replace it. Q 10 I suggest the authors provide an intuitive picture of each treatment group sample peppermint to show the result of this study. Here two excellent publications on symbiosis of P. indica and Mentha piperita should be read in this study to give some reference. Dolatabadi, H.K., Goltapeh, E.M., Moieni, A., Varma, A., 2012. Evaluation of different densities of auxin and endophytic fungi (Piriformospora indica and Sebacina vermifera) on Mentha piperita and Thymus vulgaris growth. Afr J Biotechnol, 11: 1644–1650. Dolatabadi, H.K., Goltapeh, E.M., Safari, M., Golafaie, T.P., 2017. Potential effect of Piriformospora indica on plant growth and essential oil yield in Mentha piperita. Plant Pathol Quar, 7: 96–104. Overall, the MS needs to be comprehensively revised both in respects of writing and analysis/interpretation of results. Reviewer #3: Introduction: 1st Para- line 1:- ‘Which its’ can be written as ‘and its’ 2nd Para- line 1:- Yield is negatively affected (Kindly add references) 3rd Para- 2nd last line:- ‘Quercus ilex’ should be italics Material and Methods Growth Conditions:- 1st para- line 3:- ‘Three replications’; always use minimum five replications Line 5:- Kindly mention the spore counts of P indica and AMF Line 6:- Kindly specify the ratio of Caspian sea water and distilled water Physiological parameters: Line 3-5:- Kindly elaborate the methods used for analysis Results: 1st Para- 2nd last line:- Kindly confirm the synergistic association between P indica & AMF through plating techniques. ******* 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). 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Revision 1
25 Mar 2021 Author Response Prof. Mayank Gururani Editor-in-Chief PLOS ONE Journal Dear Prof., Mayank Gururani Attached please find our revised manuscript entitled: Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? (No.: PONE-D-21-02576) for publication in PLOS ONE Journal. Thank you for giving us this opportunity to revise and resubmit our manuscript. We appreciate your comments and the suggestions made by referees to improve the manuscript. We responded to all of the reviewers' suggestions. We hope that after these enhancements the manuscript can now be accepted for publication; however, we are certainly willing to consider further changes if necessary. We look forward to hearing from you. Yours faithfully, Masoumeh Khalvandi In behalf authors Response to Review Comments (Manuscript Number: PONE-D-21-02576) Response to Reviewer #1 Comments Reviewer point #1: Page 1: Introduction: line 1-2: “which its demand has grown dramatically in recent decades” – may be corrected as: ‘the demand of which has grown dramatically in recent decades’ Author response #1: the correction has been made in the revised manuscript. Reviewer point #2: Line 3: “It is evident that peppermint essential oil” – may be modified as: ‘it is evident that the essential oil’ Author response #2: “It is evident that peppermint essential oil” was modified to ‘it is evident that the essential oil’ Reviewer point #3: Line 5-6: “as well as the reactive oxygen species (ROS) scavenging activity because of its valuable constituents” – ‘as well as scavenging of reactive oxygen species (ROS)’ Author response #3: This has been corrected in the revised manuscript. Reviewer point #4: 2nd Para – line 1: - avoid ‘Nowadays’; the sentence may begin as ‘The sustainability …’ Author response #4: (Nowadays) was removed in the introduction. Reviewer point #5: Line 5: no need of repeating – ‘reactive oxygen species (ROS)’, better put the short form only – ROS because the same is already defined above Author response #5: done Reviewer point #6: Line 7: “stresses; it can show’ – ‘stresses, because the same can show’ Author response #6: “stresses; it can show’ was modified to ‘stresses, because the same can show’ Reviewer point #7: Page 3: 2nd para – 2nd line: not ‘sever’ – but ‘severe’ Author response #7: ‘sever’ was changed to ‘severe’ Reviewer point #8: Page 4: para 3- line 2: not ‘fungi symbiosis’ – ‘fungal symbiosis’ Author response #8: the correction has been made in the revised manuscript. ‘fungi symbiosis’ was modified to ‘fungal symbiosis’ Reviewer point #9: Discussion: Page 4-5: line 1-4: “In the present study, the root colonization with P. indica and AMF were remarkably reduced by increasing salinity (6 to 9 dSm−1). Similar observations were reported in other plants (Wu et al., 2010; Hadian-Deljou, et al, 2020; khalvandi et al, 2019). The reason that salinity decreased fungal colonization might be attributed to the adverse 5 effects of salinity on photosynthesis, along with a reduction in carbon supply” The above statements seem contradictory to the claim below on page 5, para 5 – line 1-4: “Our findings showed that P. indica, AMF and their co-inoculation symbiosis mitigated the inhibitory effect of salinity on the photosynthetic capability of peppermint” Reviewer point #10: Last para – line 1-2: “The endophytic fungus can mitigate the toxic influence of salinity on chloroplast and chlorophyll by releasing cytokinin-like substances which maintain the relative stability of the organelle” – how can the fungi mitigate the toxic influence, when the fungi themselves are negatively affected by salinity? “Nevertheless, endophytic fungi symbiosis mitigated the inhibitory effect of salt stress on the mineral uptake. It has been well documented that endophytic fungi can prevent the toxicity of Na+ in aerial parts of plants through accumulation of Na+ ions in fungal cytosol, hyphae wall and vesicles” – more explanations are required for this argument, especially when the fungi are harmfully affected by salt stress Author response #9 & 10: In this experiment, symbiotic relationship with fungi under no stress and mild stress conditions had a positive significant effect on the examined traits. On the other hand, we observed negative effects of high salinity levels on colonization and physiological parameters of peppermint plants. Although, fungi themselves are negatively affected by salinity, but the results showed that compared to the control plants, symbiotic relationship with these fungi improved the photosynthetic apparatus function. Based on the reports of some researchers, these changes in the presence of fungi and under severe salinity stress can be due to: The participating AMF induces expression of genes involved in N+ extrusion to the soil solution, K+ acquisition (by phloem loading and unloading), and release into the xylem, therefore maintaining a favorable Na: K ratio. Colonization by AMF differentially affects the expression of the plasma membrane and tonoplast aquaporins (PIPs and TIPs), which consequently improves the water status of the plant. The formation of AM (arbuscular mycorrhiza) surges the capacity of the plant to mend photosystem-II (PSII) and boosts the quantum efficiency of PSII under salt stress conditions by mounting the transcript levels of chloroplast genes encoding antenna proteins involved in the transfer of excitation energy. Furthermore, AM-induced interplay of phytohormones, including strigolactones, abscisic acid, gibberellic acid, salicylic acid, and jasmonic acid have also been associated with the salt tolerance mechanism (Heikham Evelin et al, 2019). Reviewer point #11: Overall, the researchers have observed evidence for synergistic effect Pirimorphospora and AM in alleviating salt stress in peppermint; however, their arguments need more explanations to avoid self-contradictory appearance as they found salt stress also negatively affecting the plant metabolism as well as fungal colonization. Although, they provide evidences of previous positive findings of synergistic influence of AM and Pirimorphospora in certain plants, they need to report findings showing no such influences as well ( for example: DOI: 10.1080/01904160903435409 ) Author response #11: The correction was made in Conclusion section. Response to Reviewer #2 Comments Reviewer point #1: [Introduction] Piriformospora indica, also named as Sebacinales indica, the author needs to add it. Author response #1: "Sebacinales indica" was added to the text Reviewer point #2: [Materials and Methods] Give more specific info on inoculum: e.g. mycelia mass or chlamydospores per ml of P. indica and AMF. And provide the method or reference of fungi inoculation and detection. Author response #2: The methods of fungi inoculation and amount of AMF and P. indica spores were added to the Materials and Methods. Reviewer point #3: [Results] This study has many figure, but author doesn’t or good explain the result of the figure. Author response #3: Thank you for pointing this out. The reviewer is correct, we checked all figures and we found in Fv/fm some mistake was made because of careless writing. This has been corrected in the revised manuscript. Also, Analysis of variance was added to the Results. Figures 7, 10, and 12 were referred to the text. Reviewer point #4: [Discussion] The conclusions drawn by the authors that P. indica, AMF and their co-inoculation symbiosis mitigated the inhibitory effect of salinity on the photosynthetic capability of peppermint is not really supported by those parameters. The authors mention that higher photosynthetic capability attributed to the better physiological status. Why the author not measure the ROS scavengers related enzyme activity to support the point? Kindly explain. Author response #4: Thank you for your suggestion. We studied the antioxidant enzymes, such as catalase and superoxide dismutase, and we have considered it for another article. But if necessary, we can also represent them. Reviewer point #5: The Fig. 2 was not clear, please replace it and provide the scale or microscope magnification. The microscopic observation pictures of the control plant roots are also need provided. “chlamidospore” probably “chlamydospore”? Please check it. Author response #5: the scale or microscope magnification was (10- 40X). the scale was added to the pictures. We separated the pictures to make the pictures clearer. “chlamidospore” was modified to “chlamydospore Reviewer point #6: Fig. 3, add the abbreviation of Pi. Give the test sample number of root colonization of each group. Author response #6: The correction was done in text. Reviewer point #7: All the column figures in this study have no error bars, please check/add it. Author response #7: Error bars was added to the all figures. Reviewer point #8: Fig. 5 (a-c) different with other column figure like Fig. 4, please check/replace it. Author response #8: Thank you for pointing this out. The reviewer is correct, this mistake was made because of using the wrong way of lettering Means. We learned from one of our colleagues that there is a more accurate way to Lettering means. then, we did the Lettering method through the SAS program itself, and all figures have been corrected. Reviewer point #9: Fig. 6, “AMF* Pi” or “Pi AMF”? “PI” or “Pi”? Same with Fig. 7, 8, 10, 12 and 13, keep one unified format. please check/replace it. Author response #9: The reviewer is correct; the correction has been made in the revised manuscript. Reviewer point #10: I suggest the authors provide an intuitive picture of each treatment group sample peppermint to show the result of this study. Author response #10: Thank you for your valuable suggestion. This research has already been done and we only have some pictures from the time of planting that we present. Reviewer point #11: Here two excellent publications on symbiosis of P. indica and Mentha piperita should be read in this study to give some reference. Dolatabadi, H.K., Goltapeh, E.M., Moieni, A., Varma, A., 2012. Evaluation of different densities of auxin and endophytic fungi (Piriformospora indica and Sebacina vermifera) on Mentha piperita and Thymus vulgaris growth. Afr J Biotechnol, 11: 1644–1650. Dolatabadi, H.K., Goltapeh, E.M., Safari, M., Golafaie, T.P., 2017. Potential effect of Piriformospora indica on plant growth and essential oil yield in Mentha piperita. Plant Pathol Quar, 7: 96–104. Author response #11: These references were added to the revised manuscript. Response to Reviewer #3 Comments Reviewer point #1: 1st Para- line 1:- ‘Which its’ can be written as ‘and its’ Author response #1: ‘Which its’ was modified to ‘and its’ Reviewer point #2: 2nd Para- line 1:- Yield is negatively affected (Kindly add references) Author response #2: The appropriate reference was added to "Yield is negatively affected" Reviewer point #3: 3rd Para- 2nd last line:- ‘Quercus ilex’ should be italics Author response #3: The correction was done in text. Reviewer point #4: 1st para- line 3:- ‘Three replications’; always use minimum five replications Author response #4: Thanks to the referee’s suggestion, definitely conducting the experiment with five replications would be more accurate. However, we found authoritative articles that tested their experiment with three repetitions, such as: 1. Bharti, N., Pandey, S. S., Barnawal, D., Patel, V. K., & Kalra, A. (2016). Plant growth promoting rhizobacteria Dietzia natronolimnaea modulates the expression of stress responsive genes providing protection of wheat from salinity stress. Scientific reports, 6(1), 1-16. 2. Gururani, M. A., Venkatesh, J., Ghosh, R., Strasser, R. J., Ponpandian, L. N., & Bae, H. (2018). Chlorophyll-a fluorescence evaluation of PEG-induced osmotic stress on PSII activity in Arabidopsis plants expressing SIP1. Plant Biosystems-An International Journal Dealing with all Aspects of Plant Biology, 152(5), 945-952. 3. Li, X., Zhao, C., Zhang, T., Wang, G., Amombo, E., Xie, Y., & Fu, J. (2021). Exogenous Aspergillus aculeatus Enhances Drought and Heat Tolerance of Perennial Ryegrass. Frontiers in Microbiology, 12, 307. 4. Varghese, N., Alyammahi, O., Nasreddine, S., Alhassani, A., & Gururani, M. A. (2019). Melatonin positively influences the photosynthetic machinery and antioxidant system of Avena sativa during salinity stress. Plants, 8(12), 610. 5. Yamane, K., Oi, T., Enomoto, S., Nakao, T., Arai, S., Miyake, H., & Taniguchi, M. (2018). Three‐dimensional ultrastructure of chloroplast pockets formed under salinity stress. Plant, cell & environment, 41(3), 563-575. 6. Ji, J., Yue, J., Xie, T., Chen, W., Du, C., Chang, E., ... & Shi, S. (2018). Roles of γ-aminobutyric acid on salinity-responsive genes at transcriptomic level in poplar: involving in abscisic acid and ethylene-signalling pathways. Planta, 248(3), 675-690. Reviewer point #5: Line 5:- Kindly mention the spore counts of P indica and AMF Author response #5: 10 g of Arbuscular mycorrhizal inoculum (with a density of 120 active spores per gram) and 10 ml of Piriformospora indica suspension (1× 109) were added to the pots. Reviewer point #6: Line 6:- Kindly specify the ratio of Caspian sea water and distilled water Author response #6: In order to reach each level of salinity stress: determined amount of the Caspian Sea water and distilled water were mixed and then the desired salinity was determined by EC meter. Reviewer point #7: Physiological parameters: Line 3-5:- Kindly elaborate the methods used for analysis Author response #7: The methods used for analysis were added to the Materials and Methods section. Chlorophyll fluorescence parameters were measured in the last fully developed leaf by using pulse amplitude modulated fluorometer (PAM-2500, Walz, Germany). First, leaves were placed in darkness for 30 minutes using specific leaf clamps. The samples were exposed to low-intensity light [< 0.1 μmol (photon) m−2 s−1, red light]. Then, a saturating light pulse [> 8,000 μmol (photon) m−2 s−1, white light) was turned on for 1 s (one pulse). The minimum fluorescence (Fo) and maximum fluorescence (Fm) were determined in dark-adapted leaves. The variable fluorescence (Fv) and maximum quantum photosystem II efficiency (Fv/Fm) were evaluated based on Equations 1 and 2. and the effective photochemical quantum efficiency II [ФPSII], electron transfer rate [ETR, μmol(electron) m–2 s–1), non-photochemical quenching (NPQ), the quantum yield of regulated energy dissipation (YNPQ), and Quantum yield of non-regulated energy dissipation in PSII (Y (NO)) were calculated by using Equations 3 and 6 (Khanghahi et al, 2012). 1) Fv= Fm- Fo 2) Fv/Fm= (Fm- Fo)/ Fm 3) ФPSII= (Fm´ − F)/Fm´ 4) NPQ = (Fm/Fm') – 1 5) Y(NPQ) = (F/Fm') – (F/Fm) 6) Y(NO) = F/Fm F= steady-state fluorescenc Fm'= maximum fluorescence measured in light-exposed leaf samples. Reviewer point #8: Results: 1st Para- 2nd last line:- Kindly confirm the synergistic association between P indica & AMF through plating techniques. Author response #8: Thank you for your valuable suggestion, we definitely investigate the synergistic association between P indica & AMF through plating techniques in our future research; however, this research has already been conducted and done; thus, we can't add a new experiment. Attachments Attachment Submitted filename: Response to Reviewers.docx* https://doi.org/10.1371/journal.pone.0254076.r002
20 Apr 2021 Decision Letter - Mayank Gururani, Editor PONE-D-21-02576R1 Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? PLOS ONE Dear Dr. Khalvandi, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by 5th May 2021. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. 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If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) Reviewer #3: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #3: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: (No Response) Reviewer #3: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #3: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: No Reviewer #3: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: General comments: The article is improved a lot after the revision, but the authors were not thorough in a critical review of the entire text. Some minor corrections in the abstract, introduction and materials & methods part of the article are pointed out below. But the discussion part needs thorough revision. The authors discuss many physiological effects in plants concerning salt stress based on previous literature but not specific to their findings. Discussion in a research paper is meant to discuss the results and not the previous literature. Some contradictions are also in their argument. They must thoroughly revise the discussion. Let them go through the specific comments pointed out below while reviewing the discussion. If they limit the discussion to their actual findings, the entire discussion can be summarized to 1/3rd of what is presented. Specific comments 1. Abstract: First sentence of the abstracts seems complex and needs to be simplified 2. Introduction – 2nd paragraph – 3rd line: ‘Photosynthesis process’ – please delete ‘process’ 3. Introduction – 4th paragraph – Line No.12: ‘which are all pharmaceutical useful’ – please modify as ‘pharmaceutically useful’ 4. Material and Methods: 1st paragraph – 2nd line: ‘This research was carried out in a factorial experiment in a….’ – may be modified as: ‘A factorial experiment was carried out in a....’ 5. Discussion – 9th paragraph: ‘Several reports have reported a direct correlation between nutrient imbalance and the simultaneous decline in PSII function in salt-stressed plants’ - the sentence may be modified as ‘Several reports suggest a direct correlation between nutrient imbalance and the simultaneous decline in PSII function in salt-stressed plants’ – which are these reports? Please refer to the reports; the reference of Yang et al. (2021) is not sufficient to substantiate the argument of ‘several reports’; otherwise, modify the sentence. 9th paragraph – line 7: ‘fungi symbiosis’ – may be modified as fungal symbiosis 9th paragraph: lines 6-7: ‘Nevertheless, endophytic fungi symbiosis mitigated the inhibitory effect of salt stress on the mineral uptake’ – please mention the evidence which the authors have presented as results in their study to substantiate this argument; otherwise delete this speculative argument. (the first sentence of this paragraph is ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake because of the high rate of Na+ absorption’, is contradictory to this argument. Line 11: ‘osmotic stress)’ – delete the bracket 10th paragraph: lines 5-7: ‘In agreement with the present results, Xu et al. (2016) reported that the positive effect of AMF on Fv/Fm increasing in maize seedlings can be attributed to the absorption of mineral nutrients, activating mediated genes and sink stimulation’ – how can it be in agreement with, especially when the authors argue in the 9th paragraph that ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake because of the high rate of Na+ absorption’ ? - please note the contradiction and revise I think the 11th and 12th paragraph are unnecessary as these do not discuss any specific findings of the researchers. Overall, the discussion need to be thoroughly revised Reviewer #3: The manuscript describe a technically sound piece of scientific research. Authors have adequately addressed the comments raised in a previous round of review and this manuscript is now acceptable for publication ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: Yes: Joseph George Ray Reviewer #3: Yes:** Md Nafe Aziz [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0254076.r003
Revision 2
5 May 2021 Author Response Prof. Mayank Gururani Editor-in-Chief PLOS ONE Journal Dear Prof., Mayank Gururani Attached please find our revised manuscript entitled: Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? (No.: PONE-D-21-02576) for publication in PLOS ONE Journal. Thank you for giving us this opportunity to revise and resubmit our manuscript. We appreciate your comments and the suggestions made by referees to improve the manuscript. We responded to all of the reviewers' suggestions. We hope that after these enhancements the manuscript can now be accepted for publication; however, we are certainly willing to consider further changes if necessary. We look forward to hearing from you. Yours faithfully, Masoumeh Khalvandi In behalf authors Response to Review Comments (Manuscript Number: PONE-D-21-02576) Response to Reviewer #1 Comments Reviewer point #1: If they limit the discussion to their actual findings, the entire discussion can be summarized to 1/3rd of what is presented. Author response #1: The length of the discussion part has been reduced. And these sentences were removed in the discussion part. • The reason that salinity decreased fungal colonization might be attributed to the adverse effects of salinity on photosynthesis, along with a reduction in carbon supply from plant to endophytic fungus. This can also be supported by the fact that salinity induced changes in morphological features of fungal hyphae. Besides, • These results are similar to what has been recorded in wheat (lyas et al, 2020), and maize (Wang et al, 2020). • inhibition of QA re-oxidation, and degradation of D1 and D2 proteins in the PSII which • Salinity stress probably saturates the electron transport chain that leads to accumulation of protons, followed by an increase in the amount of NPQ. The increase in NPQ, which is actually an indicator of heat loss in the processes of the excessive energy removal, shows the high capacity of the xanthophyll cycle (Nilkens et al., 2010; Murchie and Lawson, 2013). • Higher values of photochemical activity of PSII in plants which have a symbiotic relationship with fungi are mainly associated with: the increased density of photosynthetic units, the improved reaction center activity, enhancing the plants’ ability to use light energy, and facilitating the electron transport to NADP (Xu et al, 2016; Moreira et al. 2015; Zhang et al, 2018; Shahabivand et al, 2017; Ghorbani et al, 2018; Shahabivand et al, 2012). Furthermore, microbial inoculation can alleviate the adverse impacts of salinity on the photochemical activity of PSII through expression of the genes encoding D1 and D2 proteins, improving water absorption, modulating membrane damage, and improving chlorophyll content (Chen et al, 2017). • AMF has been reported to regulate root permeability by doubling the expression of the LSPLP1 gene, the strategy which leads to a higher tolerance to osmotic stress) (Heikham Evelin et al, 2019). • In addition to what is mentioned above, salinity suppresses the biosynthesis of essential oil. The reason for the reduction in the synthesis of essential oils can be related to the low availability of assimilates as a result of reduced photosynthetic activity and inactivation of the PSII reaction center (Zuo et al, 2017). However, even when photosynthesis assimilation approaches zero (due to stomatal restriction, damage to chlorophyll, and chloroplast membrane system under severe stress conditions) monoterpenes biosynthesis still occurs due to alternative carbon sources allocation (Lavoir et al, 2009). Naturally, monoterpenes are biosynthesized through MEP/ DOXP pathway and are fueled by a constant supply of substrates, high amounts of acetyl-CoA, and ATP and NADPH+H cofactors (Niinemets et al., 2002, Behn et al, 2010). This can be the reason for the tight link between methyl erythritol-4-osphate (MEP) and photosynthesis, as the Calvin cycle in chloroplasts is the main carbon sources for terpenoid biosynthesis and the MEP pathway is a place for reducing excess electron flow in the photosynthetic electron transfer chain (Valifard et al, 2018). • It is believed that the monoterpene pattern is mostly determined by NADPH+H+ -dependent reductive interconversions, like a reduction of menthone to menthol (Behn et al, 2010). These modifications in the biosynthesis of monoterpenes can be linked to alterations in the photosynthetic electron transfer chain, the production of photosynthetic NDPH2 (Maffei and Codignola, 1990), and oxidation of monoterpenes by ROS to generate oxygenated forms under environmental stress (Zuo et al, 2017). These references were also removed in the revised manuscript. 1. Agron. 10, 160–162.Wu, M., Wei, Q., Xu, L., Li, H., Oelmüller, R., & Zhang, W. (2018). Piriformospora indica enhances phosphorus absorption by stimulating acid phosphatase activities and organic acid accumulation in Brassica napus. Plant and Soil, 432(1-2), 333-344. 2. Behn, H., Albert, A., Marx, F., Noga, G., & Ulbrich, A. (2010). Ultraviolet-B and photosynthetically active radiation interactively affect yield and pattern of monoterpenes in leaves of peppermint (Mentha× piperita L.). Journal of agricultural and food chemistry, 58(12), 7361-7367. 3. Flexas, J., Escalona, J. M., & Medrano, H. (1999). Water stress induces different levels of photosynthesis and electron transport rate regulation in grapevines. Plant, Cell & Environment, 22(1), 39-48. 4. lyas, N., Mazhar, R., Yasmin, H., Khan, W., Iqbal, S., Enshasy, H. E., & Dailin, D. J. (2020). Rhizobacteria isolated from saline soil induce systemic tolerance in wheat (Triticum aestivum L.) against salinity stress. Agronomy, 10(7), 989. 5. Lavoir, A. V., Staudt, M., Schnitzler, J. P., Landais, D., Massol, F., Rocheteau, A., ... & Rambal, S. (2009). Drought reduced monoterpene emissions from the evergreen Mediterranean oak Quercus ilex: results from a throughfall displacement experiment. Biogeosciences, 6(7), 1167-1180. 6. Maffei, M., & Codignola, A. (1990). Photosynthesis, photorespiration and herbicide effect on terpene production in peppermint (Mentha piperita L.). Journal of Essential Oil Research, 2(6), 275-286. 7. Sebastian, A., & Prasad, M. N. V. (2019). Photosynthetic light reactions in Oryza sativa L. under Cd stress: Influence of iron, calcium, and zinc supplements. The EuroBiotech Journal. 3(4): 175-181. 8. Zuo, Z., Wang, B., Ying, B., Zhou, L., & Zhang, R. (2017). Monoterpene emissions contribute to thermotolerance in Cinnamomum camphora. Trees, 31(6), 1759-1771. Reviewer point #2: Abstract: First sentence of the abstracts seems complex and needs to be simplified Author response #2: The correction was done in Abstract. Reviewer point #3: Introduction – 2nd paragraph – 3rd line: ‘Photosynthesis process’ – please delete ‘process’ Author response #3: process was deleted in ‘Photosynthesis process’ Reviewer point #4: Introduction – 4th paragraph – Line No.12: ‘which are all pharmaceutical useful’ – please modify as ‘pharmaceutically useful’ Author response #4: ‘which are all pharmaceutical useful’ was modified to ‘pharmaceutically useful’ Reviewer point #5: Material and Methods: 1st paragraph – 2nd line: ‘This research was carried out in a factorial experiment in a….’ – may be modified as: ‘A factorial experiment was carried out in a....’ Author response #5: ‘This research was carried out in a factorial experiment in a….’ was modified to ‘A factorial experiment was carried out in a....’ Reviewer point #6: Discussion – 9th paragraph: ‘Several reports have reported a direct correlation between nutrient imbalance and the simultaneous decline in PSII function in salt-stressed plants’ - the sentence may be modified as ‘Several reports suggest a direct correlation between nutrient imbalance and the simultaneous decline in PSII function in salt-stressed plants’ Author response #6: The correction has been made in the revised manuscript. Reviewer point #7: which are these reports? Please refer to the reports; the reference of Yang et al. (2021) is not sufficient to substantiate the argument of ‘several reports’; otherwise, modify the sentence. Author response #7: Thank you for pointing this out. The following articles were added to the references. • Loudari, A., Benadis, C., Naciri, R., Soulaimani, A., Zeroual, Y., Gharous, M. E., Kalaji, H M., & Oukarroum, A. (2020). Salt stress affects mineral nutrition in shoots and roots and chlorophyll a fluorescence of tomato plants grown in hydroponic culture. Journal of Plant Interactions, 15(1), 398-405. • Qu, C., Liu, C., Gong, X., Li, C., Hong, M., Wang, L., & Hong, F. (2012). Impairment of maize seedling photosynthesis caused by a combination of potassium deficiency and salt stress. Environmental and Experimental Botany, 75, 134-141 Reviewer point #8: 9th paragraph – line 7: ‘fungi symbiosis’ – may be modified as fungal symbiosis Author response #8: ‘fungi symbiosis’ was modified to ‘fungal symbiosis’ Reviewer point #9: 9th paragraph: lines 6-7: ‘Nevertheless, endophytic fungi symbiosis mitigated the inhibitory effect of salt stress on the mineral uptake’ – please mention the evidence which the authors have presented as results in their study to substantiate this argument; otherwise delete this speculative argument. (the first sentence of this paragraph is ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake because of the high rate of Na+ absorption’, is contradictory to this argument. Author response #9: ‘Nevertheless, endophytic fungi symbiosis mitigated the inhibitory effect of salt stress on the mineral uptake’ was modified to ‘Nevertheless, in inoculated plant endophytic fungal symbiosis increased P and K+ content (Figure. 6a) and decreased Na+ uptake in peppermint leaves (Figure. 4c).’ In order to avoid ambiguity and contradiction, ‘in non-inoculated plant’ and ‘in inoculated plant’ were added to the text of this paragraph. Reviewer point #10: Line 11: ‘osmotic stress)’ – delete the bracket Author response #10: done Reviewer point #11: 10th paragraph: lines 5-7: ‘In agreement with the present results, Xu et al. (2016) reported that the positive effect of AMF on Fv/Fm increasing in maize seedlings can be attributed to the absorption of mineral nutrients, activating mediated genes and sink stimulation’ – how can it be in agreement with, especially when the authors argue in the 9th paragraph that ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake because of the high rate of Na+ absorption’ ? - please note the contradiction and revise Author response #11: In order to avoid ambiguity and contradiction, ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake because of the high rate of Na+ absorption’ was modified to ‘The results of this study showed that severe salinity stress remarkably reduced P and K+ uptake in non-inoculated plant because of the high rate of Na+ absorption.’ Thank you for your valuable suggestion. After reviewing articles in various journals, we found that they also reported such contradictions, and in all of them, although the rate of colonization and formation of fungal hyphae and spores decreased under salinity stress, its presence in the plant roots was able to reduce the negative effects of salinity stress on the plant and improves the amount of some plant parameters. • Klinsukon, C., Lumyong, S., Kuyper, T. W., & Boonlue, S. (2021). Colonization by arbuscular mycorrhizal fungi improves salinity tolerance of eucalyptus (Eucalyptus camaldulensis) seedlings. Scientific Reports, 11(1), 1-10. • Zai, X. M., Fan, J. J., Hao, Z. P., Liu, X. M., & Zhang, W. X. (2021). Effect of co-inoculation with arbuscular mycorrhizal fungi and phosphate solubilizing fungi on nutrient uptake and photosynthesis of beach palm under salt stress environment. Scientific Reports, 11(1), 1-11. • Parvin, S., Van Geel, M., Yeasmin, T., Verbruggen, E., & Honnay, O. (2020). Effects of single and multiple species inocula of arbuscular mycorrhizal fungi on the salinity tolerance of a Bangladeshi rice (Oryza sativa L.) cultivar. Mycorrhiza, 30(4), 431-444. • Kaya, C., Ashraf, M., Sonmez, O., Aydemir, S., Tuna, A. L., & Cullu, M. A. (2009). The influence of arbuscular mycorrhizal colonisation on key growth parameters and fruit yield of pepper plants grown at high salinity. Scientia horticulturae, 121(1), 1-6. • Jahromi, F., Aroca, R., Porcel, R., & Ruiz-Lozano, J. M. (2008). Influence of salinity on the in vitro development of Glomus intraradices and on the in vivo physiological and molecular responses of mycorrhizal lettuce plants. Microbial Ecology, 55(1), 45. Reviewer point #12: I think the 11th and 12th paragraph are unnecessary as these do not discuss any specific findings of the researchers. Author response #12: These 11th and 12th paragraphs was delated in the revised manuscript. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0254076.r004
21 May 2021 Decision Letter - Mayank Gururani, Editor PONE-D-21-02576R2 Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? PLOS ONE Dear Dr. Khalvandi, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by 5 June 2021. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. Please include the following items when submitting your revised manuscript: A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter. If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: http://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols. We look forward to receiving your revised manuscript. Kind regards, Mayank Gururani Academic Editor PLOS ONE Journal Requirements: Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: (No Response) Reviewer #4: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #4: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A Reviewer #4: Yes ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #4: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: No Reviewer #4: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: I appreciate the improvements done in the submitted revision text, but feel that the authors need to summarize the discussion further. The lengthy and imprecise debate can only tarnish the brilliance of findings in any good research. Therefore, I request the authors to revise the ‘discussion’ to clarify the exact relevance of their findings. Please use the comments given in the ‘discussion’ part of the revised 'manuscript with track changes' as a model for the revision. Please avoid excessive references to previous findings; limit the citations to a maximum of 1-2 most relevant authorities in each case. Please also thoroughly check the entire text before submission of the revision for spelling and grammar; summarize or rephrase sentences to improve clarity wherever necessary. Finally, check the references thoroughly after deletion of unnecessary citations in the text during revision of the text. Reviewer #4: The authors have adequately addressed the points raised by the previous reviewers and the manuscript can be accepted for publication. However, the article will look more authentic if you could mention the source of both the Piriformospora indica and Arbuscular mycorrhizal fungi used in this work. If available, mention the culture depository details as well. ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: Yes:** Joseph George Ray Reviewer #4: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. Attachments Attachment Submitted filename: PONE-D-21-02576_R2 - with corrections.pdf https://doi.org/10.1371/journal.pone.0254076.r005
Revision 3
10 Jun 2021 Author Response Prof. Mayank Gururani Editor-in-Chief PLOS ONE Journal Dear Prof., Mayank Gururani Attached please find our revised manuscript entitled: Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? (No.: PONE-D-21-02576) for publication in PLOS ONE Journal. Thank you for giving us this opportunity to revise and resubmit our manuscript. We appreciate your comments and the suggestions made by referees to improve the manuscript. We responded to all of the reviewers' suggestions. We hope that after these enhancements the manuscript can now be accepted for publication; however, we are certainly willing to consider further changes if necessary. We look forward to hearing from you. Yours faithfully, Masoumeh Khalvandi In behalf authors Response to Review Comments (Manuscript Number: PONE-D-21-02576) Response to Reviewer #1 Comments Reviewer point #1: The lengthy and imprecise debate can only tarnish the brilliance of findings in any good research. Therefore, I request the authors to revise the ‘discussion’ to clarify the exact relevance of their findings. Please use the comments given in the ‘discussion’ part of the revised 'manuscript with track changes' as a model for the revision. Please avoid excessive references to previous findings; limit the citations to a maximum of 1-2 most relevant authorities in each case. Please also thoroughly check the entire text before submission of the revision for spelling and grammar; summarize or rephrase sentences to improve clarity wherever necessary. Finally, check the references thoroughly after deletion of unnecessary citations in the text during revision of the text. Author response #1: All requested corrections were made in the revised manuscript. The comments made in the ‘discussion’ part of the 'revised manuscript with track changes' were used as a model for revision. citations were reduced to a maximum of 1-2 most relevant authorities in each case. unnecessary references were deleted. The text was grammatically reviewed and corrected. The following references have been deleted: 1. Calatayud, A., Roca, D., Martínez, P.F., (2006). Spatial-temporal variations in rose leaves under water stress conditions studied by chlorophyll ﬂuorescence imaging. Plant Physiol. Biochem. 44, 564e573 2. Coban, O., Göktürk Baydar, N., (2016). Brassinosteroid effects on some physical and biochemical properties and secondary metabolite accumulation in peppermint (Mentha piperita L.) under salt stress. Ind Crops Prod. 86, 251-258. 3. Gupta, R., Singh, A., Srivastava, M., Singh, V., Gupta, M. M., & Pandey, R. (2017). Microbial modulation of bacoside A biosynthetic pathway and systemic defense mechanism in Bacopa monnieri under Meloidogyne incognita stress. Scientific reports, 7(1), 1-11. 4. Hasan, M., Ma, F., Prodhan, Z., Li, F., Shen, H., Chen, Y., & Wang, X. (2018). Molecular and physio-biochemical characterization of cotton species for assessing drought stress tolerance. International journal of molecular sciences, 19(9), 2636. 5. Khatri, K., & Rathore, M. S. (2019). Photosystem photochemistry, prompt and delayed fluorescence, photosynthetic responses and electron flow in tobacco under drought and salt stress. Photosynthetica, 57(1), 61-74. 6. Kromdijk, J., Głowacka, K., Leonelli, L., Gabilly, S.T., Iwai, M., Niyogi, K.K., Long, S.P. (2016). Improving photosynthesis and crop productivity by accelerating recovery from photoprotection. Science 354, 857–861. 7. Kumar, G. (2020). Unit 7 Topic: Arbuscular Mycorrhizal Colonization In Plant Roots. 8. Li, D., Liu, H., Qiao, Y., Wang, Y., Cai, Z., Dong, B., Shi, C., Liu, Y., Li, X., and Liu, M. (2013). Effects of elevated CO2 on the growth, seed yield, and water use efficiency of soybean (Glycine max (L.) Merr.) under drought stress. Agricultural Water Management, 129, 105-112. 9. Li, Y., Song, H., Zhou, L., Xu, Z., & Zhou, G. (2019). Tracking chlorophyll fluorescence as an indicator of drought and rewatering across the entire leaf lifespan in a maize field. Agricultural Water Management, 211, 190-201. 10. Loomis, W.D., Corteau, R., 1972. Essential oil biosynthesis. Recent Adv Phytochem. 6, 147-185. 11. McMillen, B.G., Juniper, S., Abbott, L.K., (1998). Inhibition of hyphal growth of a Vesicular arbuscular mycorrhizal fungus in soil containing sodium chloride limits the spread of infection from spores. Soil Biol. Biochem. 30, 1639–1646. 12. Moreira, B. C., Junior, P. P., Jordao, T. C., da Silva, M. D. C. S., Stürmer, S. L., Salomão, L. C. C., ... & Kasuya, M. C. M. (2016). Effect of inoculation of symbiotic fungi on the growth and antioxidant enzymes’ activities in the presence of Fusarium subglutinans f. sp. ananas in pineapple plantlets. Acta Physiologiae Plantarum, 38(10), 1-14. 13. Niinemets, Ü., Hauff, K., Bertin, N., Tenhunen, J. D., Steinbrecher, R., & Seufert, G. (2002). Monoterpene emissions in relation to foliar photosynthetic and structural variables in Mediterranean evergreen Quercus species. New Phytologist, 153(2), 243-256. 14. Nilkens, M., Kress, E., Lambrev, P., Miloslavina, Y., Müller, M., Holzwarth, A.R., Jahns, P. (2010). Identification of a slowly inducible zeaxanthin-dependent component of nonphotochemical quenching of chlorophyll fluorescence generated under steady-state conditions in Arabidopsis. 15. Ormeño, E., Olivier, R., Mévy, J. P., Baldy, V., & Fernandez, C. (2009). Compost may affect volatile and semi-volatile plant emissions through nitrogen supply and chlorophyll fluorescence. Chemosphere, 77(1), 94-104. 16. Rai, M., Acharya, D., Singh, A., Varma, A., (2001). Positive growth responses of the medicinal plants Spilanthes calva and Withania somnifera to inoculation by Piriformospora indica in a field trial. Mycorrhiza. 11, 123-128. 17. Rapparini, F., Llusià, J., & Peñuelas, J. (2008). Effect of arbuscular mycorrhizal (AM) colonization on terpene emission and content of Artemisia annua L. Plant Biology, 10(1), 108-122. 18. Shahabivand, S., Parvaneh, A., & Aliloo, A. A. (2017). Root endophytic fungus Piriformospora indica affected growth, cadmium partitioning and chlorophyll fluorescence of sunflower under cadmium toxicity. Ecotoxicology and environmental safety, 145, 496-502. 19. Sun, C., Johnson, J. M., Cai, D., Sherameti, I., Oelmüller, R., & Lou, B. (2010). Piriformospora indica confers drought tolerance in Chinese cabbage leaves by stimulating antioxidant enzymes, the expression of drought-related genes and the plastid-localized CAS protein. Journal of plant physiology, 167(12), 1009-1017. 20. Tarraf, W., Ruta, C., Cillis, F.D., Tagarelli, A., Tedone, L., Mastro, G.D., (2015). Effects of mycorrhiza on growth and essential oil production in selected aromatic plants. J. 21. Vickers, C. E., Gershenzon, J., Lerdau, M. T., & Loreto, F. (2009). A unified mechanism of action for volatile isoprenoids in plant abiotic stress. Nature chemical biology, 5(5), 283. 22. Tsai, M.L., Wu, C.T., Lin, T.F., Lin, W.C., Huang, Y.C., Yang, C.H., (2013). Chemical composition and biological properties of essential oils of two mint species. Trop. J. Pharm. Res. 12, 577–582. Reviewer #4: The authors have adequately addressed the points raised by the previous reviewers and the manuscript can be accepted for publication. However, the article will look more authentic if you could mention the source of both the Piriformospora indica and Arbuscular mycorrhizal fungi used in this work. If available, mention the culture depository details as well. Author response #1: The source of both the Piriformospora indica and Arbuscular mycorrhizal fungi used in this work was added in text. We obtained both he Piriformospora indica and Arbuscular mycorrhizal fungi from Sari Agricultural Sciences and Natural Resources University. But the main source of these two fungi was as follows: Mycorrhiza fungi inoculum (consisted of spores in a sand and mycorrhizal roots mixture) was prepared from Turan Biotechnology Company, Shahrood, Iran. The Piriformospora indica culture was kindly gifted by Prof. Karl-Heinz Kogel, Institute of Phytopathology and Applied Zoology, University of Giessen, Germany. P. indica was cultured in liquid Kafer’s medium at 24◦C for 10 days (Yaghoubian et al., 2019). Journal Requirements: Please review your reference list to ensure that it is complete and correct. If you have cited papers that have been retracted, please include the rationale for doing so in the manuscript text, or remove these references and replace them with relevant current references. Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. Author response #1: reference list has been corrected in the revised manuscript. "Arnon, D. I. (1949). Copper enzymes in isolated chloroplasts. Polyphenol oxidase in Beta vulgaris. Plant physiology, 24(1), 1" was replaced with "Hameed, A., Akram, N. A., Saleem, M. H., Ashraf, M., Ahmed, S., Ali, S., ... & Alyemeni, M. N. (2021). Seed treatment with α-tocopherol regulates growth and key physio-biochemical attributes in carrot (Daucus carota l.) plants under water limited regimes. Agronomy, 11(3), 469". "Hemming, D., (2013). Plant sciences reviews 2012. CABI. 280 pp. (online ISSN 1749-8848)" was replaced with " Sun, Z., Wang, H., Wang, J., Zhou, L., Yang, P., (2014). Chemical composition and anti-inflammatory, cytotoxic and antioxidant activities of essential oil from Leaves of Mentha piperita grown in china. PLoS One. 12, e114767." "Bertamini, M., Faralli, M., Varotto, C., Grando, M. S., & Cappellin, L. (2021). Leaf Monoterpene Emission Limits Photosynthetic Downregulation under Heat Stress in Field-Grown Grapevine. Plants, 10(1), 181" was corrected. Attachments Attachment Submitted filename: Response to Reviewers.docx https://doi.org/10.1371/journal.pone.0254076.r006
21 Jun 2021 Decision Letter - Mayank Gururani, Editor Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? PONE-D-21-02576R3 Dear Dr. Khalvandi, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Mayank Gururani Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A ****** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes ****** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: I think the authors have addressed all the comments satisfactorily and corrected the language to the extent possible from their side. ****** 7. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review?** For information about this choice, including consent withdrawal, please see our Privacy Policy. Reviewer #1: No https://doi.org/10.1371/journal.pone.0254076.r007
Formally Accepted
1 Jul 2021 Acceptance Letter - Mayank Gururani, Editor PONE-D-21-02576R3 Does co-inoculation of mycorrhiza and Piriformospora indica fungi enhance the efficiency of Chlorophyll fluorescence and essential oil composition in peppermint under irrigation with saline water from the Caspian Sea? Dear Dr. Khalvandi: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Dr. Mayank Gururani Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0254076.r008

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