Bacillus subtilis Simultaneously Detoxified Aflatoxin B1 and Zearalenone

Round 1

Reviewer 1 Report

1. The abstract seems a bit confusing as far as the results are concerned, it's tough to follow. You say that Bacillus subtilis had strong mycotoxin-degrading activity, and you previously stated up to 40%. Later it says - The heat-inactivated extracellular fraction of B. subtilis ZJ-2019-1 showed 79.85%. It isn't easy to see under what conditions, to translate the abstract a bit.

2. Badly chosen keywords same as in the title. The visibility of your work will be reduced. Because search engines mostly search at the level of titles, keywords, and abstracts. Your keywords and title are the same!

3. B. subtilis ZJ-2019 you did not introduce the abbreviation in the abstract.

4. Maybe "strengthen the goal" a little more. Point out the differences from your previous work, and what is new, when I read the methodology, I too many references to your previous work.

5. Row 107-108 phosphate buffer what pH value?

6. Row 110 which pressure, in which vessel, autoclave?

7. Somewhere you specify sterile PBS, somewhere you don't, please make it uniform, I don't see any logic for it to be different.

8. In line 130, refer again to the HPLC method reference. Somehow I miss the validation parameters and conditions for performing the analysis, but then at least there is a reference everywhere. The same as Chapter 3.5. Suppose any modification has been made which is, conditions, validation parameters. Thank you.

9. Wherever you state the pressure under which pressure, the same phosphate buffer as the pH buffer.

10. Row 148, how did you decide on this (for 3 h at 58℃)?

11. Have you thought maybe about some potential side effects of metals and their removal?

12. We have concluded, but not with a clear primal solution, which conditions provide the best results. I think you have to point that out.

Minor editing of English language required

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 2 Report

The manuscript entitled “Bacillus subtilis simultaneously detoxified aflatoxin B1 and 2 zearalenone” presents a significant method for decontaminating mycotoxins which represent a major threat to human dietary and livestock feed.  The authors used HPLC for detecting and quantifying mycotoxins in experimental trials, and they used appropriate stat. analysis methods. The authors also defined the nature of the active ingredient within the supernatant of B. subtilis by applying EDTA, SDS, and proteinase K. The English language is slightly clear and proper data presentation method was used. However, I have a few comments on the hypothesis, methods, and conclusions.

1.     The manuscript does not clearly explain the work scheme, which causes a confusion for readers. Please introduce the experimental plan first in the abstract before describing your results. You need to mention that you tested both the bacterial culture and the heat-treated supernatant.

2.     Please use proper scientific words throughout the manuscript.

a.     Bacterial solution: bacterial culture or broth.

b.     PBS has saline, so it is not just phosphate buffer.

3.     The whole manuscript is unclear and needs to be rewritten by a professional. It

4.     The experimental description needs to be written by a microbiologist or other closely related fields that deal with microorganisms.

5.     Lines 134: please explain how this experiment can help you define the active ingredient and its mode of action.

6.     Line 170: what does 20 mL of bacteria mean? Did you use media? What are the optimal growth conditions?

7.     I think you should use black and white in your histograms as you don’t have multiple parameters within the same panel.

8.     I believe that boiling is close to autoclaving and the boiled cells/supernatant treatments can be used in the supplementary section.

9.     After heat treatment, you did not study whether it was enough to inactivate the bacteria or not. Therefore, you should call it heat-treated instead of heat-inactivated because you do not know whether it becomes inactive or not.

10.  Line 190: what is the residual rate? Please introduce this term in the materials and methods section and explain how you made the calculation.

11.  Figure 3 panel A: I wonder why 22^oC was suppressive while the temperatures before and after enhanced degradation rate.

12.  Figure 4: how do you explain that heat treatment enhanced the ability of bacterial supernatant to detoxify/degrade the aflatoxins?

13.  Experimental data are missing control treatments such as Fig1A & B, Fig 2C & D, Fig 3C & D, Fig 5A, D, E & F. Please revisit all histograms.

14.   What is the difference between Fig 2 and Fig 5 panels A, B, D & E. Similarly, Fig 3 & Fig 6.

15.  Have you studied the stability of mycotoxins under different media, pH, incubation temp, and chemicals like EDTA & SDS?

16.  Please look at my comments on the attached PDF file.

Comments for author File: Comments.pdf

The sentences need to be rephrased by a professional writer and microbiologist. The sentences need to be arranged in a meaningful manner within the same paragraph. 

For example, you introduce your hypothesis, you clearly explain your experimental plan, you briefly mention what methods were used, and you provide a conclusion based on the data you saw. Please avoid redundancy. 

Author Response

Please see the attachment.

Author Response File: Author Response.docx

Reviewer 3 Report

1-    Substitute the keywords present in the title (aflatoxin B1 & zearalenone).

2-    The HPLC detection method for aflatoxin B1 & zearalenone should be added to the methodology.

3-    Many parts of the results should be transferred to the discussion particularly that explain the results.

Author Response

请参阅附件。

Author Response File: Author Response.docx

Round 2

Reviewer 1 Report

Accept in present form