Initial Medium Optimization of Nigrospora oryzae JL-4 and Its Biocontrol Potential on Solanum rostratum

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

In this work, Zheng et al. optimized the fermentation medium to improve the growth of Nigrospora Oryzae JL-4. The authors present the motivation that this strain, which was previously isolated and described by the authors, has the potential as a biocontrol agent against Solanum Sortratum. The development of an effective and scalable fungal fermentation process is essential for the practical application of biological weed control. Here, the authors screened the best carbon and nitrogen sources and later utilized an L9(34) orthogonal experimental design to identify the optimal C:N ratio for N. oryzae growth. The robust experimental design included the identification of optimal fermentation medium components, ratios, inoculum amount, fermentation volume, and process variables such as ideal temperature and pH. Additionally, the study assessed the safety of the fungal broth on six varied crops. It found no adverse effects on Zea mays, S. melongena, Festuca arundinacea, Bromus inermis, and Medicago sativa. However, there was a moderate susceptibility observed in Triticum aestivum and S. lycopersicum, which presents a notable result.

Given this is among the first findings that suggest the potential of N. oryzae for development as microbial herbicides targeting S. rostratum, it is noteworthy and compelling. However, throughout the text, the sentence structure could be revised to enhance clarity and readability. The controls have not been appropriately selected. In certain sections, the experimental design appears robust, yet the explanations do not adequately convey the significance of the findings.

Major comments:

1.      The term 'fermentation medium optimization' might be too broad for a study conducted solely in shake flasks. Given that shake flask experiments are primarily useful for preliminary screenings and early-stage process development, they don't encapsulate the complexities of full-scale fermentation. It would be more precise to refer to this as 'shake flask culture optimization' or 'initial medium optimization. 

2.      While the current study provides valuable insights under controlled conditions, the efficacy and safety of the fermentation broth would be more comprehensively accessed through field trials. These trials would consider environmental influences and assess potential ecological impacts, interactions with other species, and performance variations due to weather and soil conditions. 

3.      The Author should provide more context of orthogonal experimental design, and why this design was preferred over other DOEs.  Were all three levels explored for each factor? It seems only three experiments were conducted based on the table, whereas typically L9 (3⁴) indicates nine experiments. 

4.      The Results and Discussion section requires substantial revision. The paper fails to address catabolite repression or diauxic growth patterns in the context of mixed carbon supplementation. Additionally, the lack of controls, mentions of the base medium, and unclear descriptions of the carbon sources and other physiological parameters during nitrogen source testing detract from the clarity and reliability of the findings. 

5.      Captions should also be rewritten. 

Minor comments:

1.      Submerged cultivation was conducted using a rotary shaker at 120 rpm and 30 °C for 6 days in 150 mL conical flasks containing 60 mL of nutrient medium inoculated with 3 mL of seed liquid: All this comes out to 5% inoculum and

2.      The authors should also include NH4NO3 as a nitrogen source for the best comparison with previous studies.

3.      The discussion section could be elaborated further.

Text:

1.      For instance, abstract, Line number 11. The sentence is not clear and lacks important context. The sentence states 25% twice without specifying what these percentages refer to. It's not clear what the fermentation volume is.

2.      Page 1: Line number 25 needs proper formatting. The theme fonts and text do not match the rest of the manuscript.

3.      Page 2: Lines 42-44 - It's not immediately clear whether the fungi were isolated from diseased leaves or if the leaves were isolated with 5 fungal species on them.

 

4.      Page 7: Line numbers: 243 typographical error. 

Comments on the Quality of English Language

The text necessitates substantial revision.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding corrections highlighted in track changes in the re-submitted files.

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript examines the use of the fungus Nigrospora oryzae JL-4 as a biocontrol for the Chinese invasive weed Solanum rostratum. The primary objective of this work was to optimize the fermentation growth conditions of N. oryzae JL-4 to maximize its biological control effect on S. rostratum. It was determined that a pH 5.0 medium containing a liquid medium containing a specific ratio of lactose, glycerol, yeast and ammonium sulfate was most effective. It was also found that growth for 4 days at 20 degrees C produced a broth that exhibited the highest level of biocontrol against S. rostratum. The manuscript was well organized and clearly written. The findings of the manuscript could help in the biocontrol of invasive weeds. The manuscript does require minor revision. With respect to the yeast utilized in the medium, was the yeast live or lyophilized? Was the yeast a powder? Could yeast extract have been utilized as an alternative? With respect to the References, the citations appear to be inconsistent. Some of the journal titles are fully written out (references 1, 2 and 4) while others are abbreviated.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding corrections highlighted in track changes in the re-submitted files.

Author Response File: Author Response.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

 The authors have incorporated major changes; however, extensive English language editing is still required before publication. 

Comments on the Quality of English Language

Despite the adequately addressed technical content, significant improvements to the manuscript's language are required to ensure clarity and effective communication of the author's ideas.

Author Response

Thank you very much for taking the time to review this manuscript. Please find the detailed responses below and the corresponding corrections highlighted in track changes in the re-submitted files.

Author Response File: Author Response.pdf