Dear Dr. Ahmad,

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Academic Editor

PLOS One

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5. If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise.

Additional Editor Comments:

Dear Authors,

The reviewers consider the topic relevant and potentially valuable. However, substantial revisions are required before the manuscript can be reconsidered.

Please carefully address the inconsistencies in experimental replication, clearly specifying the number of pots, plants per pot, biological replicates, and technical replicates. All reported units and calculations, particularly those for gibberellic acid, other phytohormones, and cadmium concentrations, must be verified. The surface-sterilization reagent should also be clarified, as sodium hypochlorite rather than sodium chloride is normally used.

The conclusions regarding cadmium uptake should be revised to reflect that the fungal treatment reduced cadmium accumulation under heavy-metal stress relative to the heavy-metal treatment alone, rather than implying a universal reduction in cadmium uptake. Please also strengthen the statistical reporting by providing replicate numbers, measures of variability, significance indicators, and relevant statistical details in all figures and tables.

In addition, the manuscript requires thorough language editing, standardization of abbreviations, treatment labels, units, decimal precision, and figure legends. The Introduction and Discussion should be shortened to remove repetition, clearly state the novelty of the study, and provide greater mechanistic interpretation of phytohormone regulation, antioxidant responses, and possible cadmium immobilization or sequestration. Finally, all references should be carefully audited for relevance, accuracy, publication status, and correct citation placement.

A detailed point-by-point response to all reviewer comments and a clearly marked revised manuscript should be provided with the resubmission.

[Note: HTML markup is below. Please do not edit.]

Reviewers’ comments:

Reviewer’s Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

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2. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

Reviewer #1: This manuscript reports on the isolation of the endophytic fungus Aspergillus fumigatus (strain SOA) from Ziziphus lotus roots and its ability to enhance growth, modulate phytohormone profiles, elevate antioxidant defenses, and reduce cadmium accumulation in okra (Abelmoschus esculentus) under salt and heavy-metal stress conditions. The study addresses a relevant agricultural challenge, employs a reasonably comprehensive experimental design, and presents generally well-supported conclusions. The manuscript is suitable for publication in PLOS ONE following minor revisions as detailed below.

Minor comments:

1. Line 64: “soil salinization is increasing by nearly 10% per year” — this statement requires a citation. Please add the appropriate reference.

2. Lines 279–286 (Section 3.4): Metabolite values are reported with varying decimal precision (e.g., 17.64, 0.0769). Please standardize significant figures consistently throughout the Results.

3. Line 492: “alt” appears to be a typographical error for “Salt.” Please correct.

4. Figure 6 caption refers to measurements over “three weeks” in the text, while figure panels show “1 week,” “2 weeks,” and “7 weeks.” Reconcile caption and figure labels.

5. Figure 11 legend (line 758) references “IBH + salt stress” — “IBH” appears to be a typographical error; likely intended as “HM” or “HM+Salt.” Please correct.

6. Reference 10 (Warrier et al., 2013) is cited in the context of global temperature increase predictions but appears to be a paper on salicylic acid estimation in Eucalyptus. This appears to be a misplaced citation. Please verify and replace with an appropriate climate science reference.

7. Reference 73 (Zhang et al., 2026) is listed with a publication year of 2026. If this is an in-press or advance online publication, please provide the DOI and indicate “in press” or “advance online publication” as appropriate per PLOS ONE style.

8. The abbreviation “FCF” (fungal culture filtrate) used in Section 2.5 (line 178) is not defined at first use. Please define it upon first appearance in the text.

9. Section 2.6 lists eight treatments but the Discussion (line 383) refers to “Aspergillus and Trichoderma species” as a comparative context without a corresponding treatment in this study. This reference is fine but could be slightly misleading; consider adding a brief clarifying phrase that this is a literature comparison.

10. The keywords include “zero hunger,” which, while thematically relevant to food security, is an SDG policy term rather than a scientific keyword. Authors may wish to consider replacing it with a more specific scientific term (e.g., “food security,” “crop productivity,” or “sustainable agriculture”) for better indexing.

Reviewer #2: Title: Endophytic Aspergillus fumigatus Modulates Growth, Oxidative Stress, and Cadmium Accumulation in Okra Under Salinity and Heavy-Metal Stress

The manuscript investigates the role of an endophytic fungal strain, Aspergillus fumigatus SOA, in improving okra growth under salinity and cadmium stress. The authors isolated the fungus from Ziziphus lotus roots and evaluated its plant growth-promoting traits, including hormone production, antioxidant response, secondary metabolites, and cadmium modulation. The study reports that SOA improved plant height, biomass, chlorophyll content, and phytohormonal balance under stress conditions. It also suggests that SOA may help reduce oxidative damage and modulate cadmium uptake in okra. Overall, the work addresses a relevant issue in sustainable agriculture and stress physiology.

The topic is timely because salinity and heavy-metal contamination are serious threats to crop productivity. The use of an endophytic fungus as a biological tool for improving okra tolerance is interesting and practically relevant. The manuscript includes a wide range of physiological, biochemical, hormonal, antioxidant, and elemental analyses, which gives the study good experimental coverage. Molecular identification of the fungal isolate through ITS sequencing strengthens the reliability of the work.

1. The abstract is generally clear, but it would benefit from more precise reporting of the experimental conditions. The authors should briefly mention the salt and cadmium stress levels and include one or two key numerical findings beyond plant height.

2. The introduction contains some repetition about abiotic stress, salinity, and physiological damage. The authors should shorten these repeated parts and focus more directly on the specific gap: the limited evidence on endophytic fungi for okra under combined salt and cadmium stress.

3. The novelty of the work should be stated more clearly at the end of the introduction. It should be clear whether the novelty lies in the fungal strain, the plant model, the combined stress treatment, or the multi-parameter evaluation.

4. Some methodological wording needs correction. For example, phrases such as “misleading morphological features,” “chapel medium,” and “duplicate the osmotic pressure” are unclear and should be revised for scientific accuracy.

5. The fungal isolation section is somewhat repetitive. The authors describe PDA sub-culturing and Czapek broth culturing more than once. This section should be streamlined to improve readability.

6. The surface sterilization protocol should be checked. The manuscript mentions 70% ethanol followed by NaCl 2.0%. Usually, sodium hypochlorite is used for surface sterilization, not sodium chloride. If NaCl was actually used, the authors should justify it; otherwise, this should be corrected.

7. Units should be checked throughout the manuscript. For example, GA is reported as mg/mL in fungal profiling, while other hormones are reported in µg/mL, and plant hormones are later reported as mg/g FW. These unit differences should be verified and made consistent.

8. The discussion is relevant but somewhat repetitive. Several paragraphs restate the results and then compare them with previous studies. The authors should reduce repetition and add a deeper mechanistic explanation of hormone regulation, ROS detoxification, and possible Cd sequestration or immobilization.

9. The figures and legends require careful proofreading. Treatment labels, units, abbreviations, and spelling should be checked throughout. Any inconsistent terms, such as HM, heavy metal, Cd, salt, NaCl, and SOA, should be standardized.

Reviewer #3: 1. Inconsistency in Experimental Replication

The manuscript reports (Lines 242–243)"All experiments were performed in three biological replicates"

However, earlier:

"All treatments had six plants..." (Lines 177–184) and "Each replicated six times" (Line 183)

These statements are contradictory.

Number of pots per treatment

Number of plants per pot

Number of biological replicates

Number of technical replicates

2. Units Need Verification

Several hormone concentrations appear unusually high:

Examples:

GA = 20–28 mg/mL

IAA = 24–31 µg/mL

For fungal culture filtrates, GA concentrations in mg/mL are extremely high and likely incorrect.

Check original calculations and units carefully.

Possible intended units:

µg/mL

ng/mL

This requires verification before publication.

3. Cadmium Uptake Interpretation

In Results:

Control = 9.979 mg/L Cd

SOA = 12.65 mg/L Cd

HM = 13.65 mg/L Cd

HM+SOA = 10.95 mg/L Cd

The conclusion:

"SOA reduced heavy metal toxicity by modulating cadmium uptake"is only partially supported.

SOA alone increased Cd accumulation compared with control.

Recommended wording:

"SOA reduced Cd accumulation under heavy-metal stress conditions compared with HM treatment alone." instead of a broad statement that SOA always reduces Cd uptake.

4. Surface Sterilization Procedure

Line 120 it is mentioned "treated with 70% ethanol for 1 minute, NaCl (2.0%) for 2 minutes"

Normally fungal isolation protocols use “Sodium hypochlorite (NaOCl)” not NaCl.

It should be mentioned clearly that whether NaCl was actually used, or NaOCl.

5. Major Language Errors

Abstract

Line 42

Current:

“with 28.9% increase in plant height”

Correct:

“with a 28.9% increase in plant height”

Line 49

Current:

“enhance plants’ resilience”

Better:

"enhance plant resilience"

Introduction

Line 58

Current:

"Crop productivity was adversely affected"

Should be:

"Crop productivity is adversely affected"

Line 80

Current:

"biomoleculessuch"

Correct:

"biomolecules such"

Line 84

Current:

"up regulated"

Correct:

"upregulated"

Methodology

Line 133

Current:

"The misleading morphological features"

Incorrect wording.

Suggested:

"The morphological characteristics of the isolates"

Line 138

Current:

"To duplicate the osmotic pressure of the fungal cells within the chapel medium"

Problems:

duplicate → simulate

chapel medium → Czapek medium

Suggested:

"To simulate osmotic stress in Czapek medium..."

Line 145

Current:

"To promote the host plant growth"

Better:

"To promote host plant growth"

Line 166

Current:

"Abelmoschus esculentus (L.) genotype seeds"

Incorrect.

Suggested:

"Seeds of Abelmoschus esculentus (L.)"

or

"Seeds of an Abelmoschus esculentus genotype"

Line 203

Current:

"The method determined Gibberellin acid and ABA"

Correct:

"Gibberellic acid and ABA were quantified according to..."

6. Scientific Writing Issues

There are many repetitive sentences in Discussion.

The Discussion repeatedly restates numerical results instead of interpreting them.

Examples:

Lines 397–401

Lines 410–417

Lines 425–429

Lines 444–454

The discussion should focus more on:

Mechanisms, comparison with previous studies, novelty of findings, and less on repeating Results.

Overuse of "The same results have been reported"

Repeated phrases:

"The same results have been documented..."

"The same result has also been reported..."

"The same trends have been reported..."

Use alternatives:

"Similarly,"

"Consistent with previous reports,"

"These findings agree with..."

"Comparable observations have been reported..."

7. Statistical Reporting Issues

The manuscript reports many numerical differences but does not consistently indicate:

p-values

standard errors

significance letters

confidence intervals

Ensure that all figures and tables include:

Mean ± SE (or SD)

Number of replicates (n)

Statistical grouping letters

8. Reference Issues

Some references appear unrelated or weakly connected to the cited statements.

Examples:

Ref. 10 (salicylic acid estimation) is cited in climate-change discussion.

Verify all citations carefully because there may be citation-management errors.

A full reference audit is recommended before resubmission.

Based on the manuscript in its current form, I would suggest a Major Revision before publication.

**********

what does this mean?). If published, this will include your full peer review and any attached files.

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Reviewer #1: Yes:  Naveed Ahmad

Reviewer #2: No

Reviewer #3: No

**********

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Endophytic Aspergillus fumigatus Modulates Growth, Oxidative Stress, and Cadmium Accumulation in Okra Under Salinity and Heavy-Metal Stress

PONE-D-26-29102R1

Dear Dr. Ahmad,

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.

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Kind regards,

Wajid Zaman

Academic Editor

PLOS One

Additional Editor Comments (optional):

Reviewers’ comments:

Reviewer’s Responses to Questions

Comments to the Author

Reviewer #1: All comments have been addressed

Reviewer #2: All comments have been addressed

Reviewer #3: All comments have been addressed

**********

2. Is the manuscript technically sound, and do the data support the conclusions??>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

3. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

4. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

5. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

Reviewer #3: Yes

**********

Reviewer #1: The authors have improved the manuscript thoroughly based on my previous comments and therefore I recommend the acceptance of this work in its current form.

Reviewer #2: fter carefully evaluating the revised manuscript entitled “Endophytic Aspergillus fumigatus Modulates Growth, Oxidative Stress, and Cadmium Accumulation in Okra Under Salinity and Heavy-Metal Stress”, I find that the authors have addressed the reviewers’ comments satisfactorily and have made appropriate revisions that improve the clarity and quality of the manuscript. The study presents valuable findings on the role of an endophytic fungus in enhancing stress tolerance and reducing cadmium accumulation in okra, contributing to the understanding of plant–microbe interactions under combined abiotic stresses. The experimental approach is sound, the data support the conclusions, and the manuscript is now suitable for publication. I recommend acceptance after this first revision, as no major concerns remain. Only minor editorial polishing, if deemed necessary by the editorial office, may be undertaken during the production process.

Reviewer #3: Authors have incorporated all revisions! There are no more quires, which is why I recommend its publication in PLOS ONE.

**********

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:  Naveed Ahmad

Reviewer #2: No

Reviewer #3: No

**********