Round 1

Reviewer 1 Report

Comments and Suggestions for Authors This article is an update or further development of work presented in ref 13. The process of decontamination of soil with bioaugmentation with known decomposing organisms is a valid approach to soil contamination. Overall, it is a good research paper presenting interesting results, valid for publication. However, the authors need to ascertain that the article stands on its own, and does not require the reader to read other work first, to be able to follow the work done here and the results obtained.   In particular: The authors do not explain how the relative abundances of different taxa correspond to activity of various mentioned enzymes. Are for instance the taxa mentioned in Fig 3A known for their cathechol 1,2-dioxygenase activity, and other taxa are not included? "To better evaluate the metabolic potential of the bacterial ecology during the process 325 of TPH depletion, the contribution of the different bacterial taxa to the abundance of func-326 tional features of interest was analyzed, by evaluating the contribution of specific enzy-327 matic commission (EC) numbers. Results obtained are shown in Figures 3-5." - this is very unclear. I do not understand what was done here. Did the authors measure activities? Quantified DNA?   While I perfectly understand testing toxicity towards plants, I do not know what is the rationale to test if a soil is toxic to human intestinal tract?    p4, Table 1: kilogram is kg, not Kg 167: methods should be described without referencing another article. A very brief description of a method should be added. 181: there is a space inside the primer sequence 213: glutamine, not glutammine 214: line break unneccesary 260-261: "This section may be divided by subheadings. It 260 should provide a concise and precise description of the experimental results, their inter-261 pretation, as well as the experimental conclusions that can be drawn." ??? 265: Val-ues   The Fig 8 legend does not fit with the other legends, it is a part of the figure itself. 

Author Response

Dear Reviewer, thanks for your efforts in our behalf that improved the quality of the manuscript

We explained all the reason at the base of our approach on the predctive functional metagenomic and we modified all the parts you suggested.

Kind Regards

Dear Reviewer, thanks for your efforts in our behalf that improved the quality of the manuscript

We explained all the reason at the base of our approach on the predictive functional metagenomic and we modified all the parts you suggested.

Kind Regards

Simona Di Gregorio

 

This article is an update or further development of work presented in ref 13. The process of decontamination of soil with bioaugmentation with known decomposing organisms is a valid approach to soil contamination. Overall, it is a good research paper presenting interesting results, valid for publication. However, the authors need to ascertain that the article stands on its own, and does not require the reader to read other work first, to be able to follow the work done here and the results obtained. 

 In particular: The authors do not explain how the relative abundances of different taxa correspond to activity of various mentioned enzymes.

The PICRUSt2 method consists of phylogenetic placement, hidden-state prediction and sample-wise gene and pathway abundance tabulation. ASV sequences and abundances are taken as input, and gene family and pathway abundances are output. All necessary reference tree and trait databases for the default workflow are included in the PICRUSt2 implementation.

Very practically, it means that several functions are inferred by matching the closest reference organism (which annotated genome is present in PICRUSt2 reference database) by 16S sequence homology: this implies the assumption that closely related taxa have a closely related genome and can express closely related genes, proportionally to the number of copies present. Of course, as it is mentioned in the article, we are talking about inference, that can be achieved directly from metabarcoding analysis.

No direct measurements of enzymatic activities were performed: to do this and to be sure to address effective activities without using a culturomic approach (that indeed gives a partial and growth medium dependent portrait of the community), we should have done a proteomic study, an heterologous expression of retrieved protein sequences for target genes, and related ad hoc high throughput enzymatic assays. All this stuff goes beyond the scope of this article.

Are for instance the taxa mentioned in Fig 3A known for their cathechol 1,2-dioxygenase activity, and other taxa are not included?

PICRUSt2 inference gives a suggestion of which taxa detected in the microbial community has the potential to express the reported gene. The effective capability of retrieved taxa is then confirmed by literature. Other taxa renown to express e.g. cathecholi 1,2- oxygenase activity not included can:

• Be below detection threshold for 16S metabarcoding method
• Have a cumulative contribution in all samples to cathechol 1,2-oxygenase below 0.1% (arbitrarily considered irrelevant)
• Have a non-significative variation across samples, or a variation below 10^0.3 (arbitrarily considered irrelevant)

Of course, in terms of reference to literature, few words can be spent on unclassified taxa, if not giving a reference to the general metabolic capabilities of the members of their lowest identified taxonomic rank: PICRUSt2, anyway, can still give a non-trivial indication of what these unclassified taxa could potentially do.

 

To better evaluate the metabolic potential of the bacterial ecology during the process of TPH depletion, the contribution of the different bacterial taxa to the abundance of functional features of interest was analyzed, by evaluating the contribution of specific enzymatic commission (EC) numbers. Results obtained are shown in Figures 3-5." - this is very unclear. I do not understand what was done here. Did the authors measure activities? Quantified DNA? 

Referring to EC inference, PICRUSt2 can produce two different outputs: the stratified output is a data table that contains the inferred gene counts (any gene that is present in every reference genome, so a huge amount of data) for each ASV per sample, obtained as described before. The unstratified output is the sum of each ASV contribution to each EC in each sample. In practice the phrase you report means that we took the stratified contribution table output and filtered it for the ECs of our interest. We did not perform qPCR/enzymatic activities on specific genes.

To avoid misunderstanding, any reference to activity has been substituted with “feature”

While I perfectly understand testing toxicity towards plants, I do not know what is the rationale to test if a soil is toxic to human intestinal tract?   

• Soil components can be ingested indirectly as mentioned in the article, (e.g. eating vegetables not properly clean) but also directly by airborne particles of soil. In risk assessment the evaluation of toxicity effect on intestinal human tract is relevant, as you can see, for example, in Yan L, Franco AM, Elio P. Health risk assessment via ingestion and inhalation of soil PTE of an urban area. Chemosphere. 2021 Oct;281:130964. doi: 10.1016/j.chemosphere.2021.130964. Epub 2021 May 20. PMID: 34289622. More specifically, for sensitive individuals that quite commonly are in contact with soil matrix, like children, you can find useful this reference Moya, J., Phillips, L. A review of soil and dust ingestion studies for children. J Expo Sci Environ Epidemiol24, 545–554 (2014). https://doi.org/10.1038/jes.2014.17

p4, Table 1: kilogram is kg, not Kg

done

167: methods should be described without referencing another article.

it’s usual to do this when bioinformatic tools, reference to already described methods, or standard procedures present in norms (e.g. UNI EN ISO 16703) are employed

A very brief description of a method should be added.

done

181: there is a space inside the primer sequence

removed

213: glutamine, not glutammine

corrected

214: line break unneccesary

corrected

260-261: "This section may be divided by subheadings. It 260 should provide a concise and precise description of the experimental results, their inter-261 pretation, as well as the experimental conclusions that can be drawn." ???

It was a typo from an internal revision row. removed

265: Val-ues  

corrected

The Fig 8 legend does not fit with the other legends, it is a part of the figure itself.

Modified

Simona Di Gregorio

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

Very detailed data but it was challenging to decipher the small writing even when "zooming" in. Reading a hard copy can be challenging.

 

Comments on the Quality of English Language

In the introduction and discussion, please read over for grammatical errors.

For example; Introduction ( maybe concise for clear reading?)

Line 42 add but not toxic

line 45: define grabbing read the citation # 2 as listed but did not find a clear description.

line 50: Two sentences?

line 54: ones their?

Author Response

Dear Reviewer, thanks for your efforts in our behalf that improved the quality of the manuscript

we modified all the parts you suggested.

Kind Regards

Simona Di Gregorio

In the introduction and discussion, please read over for grammatical errors.

For example; Introduction ( maybe concise for clear reading?)

The Introduction and discussion were extensively revised (in yellow)

Line 42 add but not toxic

Done

Line 45: define grabbing read the citation # 2 as listed but did not find a clear description.

We meant soil exploitation for urban activities, we changed the citation that better explain the concept

Line 50: Two sentences?

This part has been extensively revised

Line 54: ones their?

This part has been extensively revised

 

Author Response File: Author Response.pdf

Reviewer 3 Report

Comments and Suggestions for Authors

In this study, the authors examined the Bio-based decontamination and detoxification of total petroleum hydrocarbon contaminated dredged sediments in order to produce constructed technosols in the frame of the circular economy…..

 

The authors have investigated an impact of hydrocarburoclastic ascomycetes, Lambertella sp. MUT 5852, was bioaugmented to dredged sediments co-composting with a lignocellulosic matrix on the depletion of total petroleum hydrocarbons. However, in the opinion of the reviewer, various anomalies are detected in the text which should be corrected before agreeing to submit to the Journal "water".

 

Scientific comments

1-      In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

 

2-      Heading 2.7 should be moved down to the next page.

 

3- The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

In this study, the authors examined the Bio-based decontamination and detoxification of total petroleum hydrocarbon contaminated dredged sediments in order to produce constructed technosols in the frame of the circular economy…..

 

The authors have investigated an impact of hydrocarburoclastic ascomycetes, Lambertella sp. MUT 5852, was bioaugmented to dredged sediments co-composting with a lignocellulosic matrix on the depletion of total petroleum hydrocarbons. However, in the opinion of the reviewer, various anomalies are detected in the text which should be corrected before agreeing to submit to the Journal "water".

 

Scientific comments

1-      In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

 

2-      Heading 2.7 should be moved down to the next page.

 

3- The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

In this study, the authors examined the Bio-based decontamination and detoxification of total petroleum hydrocarbon contaminated dredged sediments in order to produce constructed technosols in the frame of the circular economy…..

 

The authors have investigated an impact of hydrocarburoclastic ascomycetes, Lambertella sp. MUT 5852, was bioaugmented to dredged sediments co-composting with a lignocellulosic matrix on the depletion of total petroleum hydrocarbons. However, in the opinion of the reviewer, various anomalies are detected in the text which should be corrected before agreeing to submit to the Journal "water".

 

Scientific comments

1-      In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

 

2-      Heading 2.7 should be moved down to the next page.

 

3- The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

In this study, the authors examined the Bio-based decontamination and detoxification of total petroleum hydrocarbon contaminated dredged sediments in order to produce constructed technosols in the frame of the circular economy…..

 

The authors have investigated an impact of hydrocarburoclastic ascomycetes, Lambertella sp. MUT 5852, was bioaugmented to dredged sediments co-composting with a lignocellulosic matrix on the depletion of total petroleum hydrocarbons. However, in the opinion of the reviewer, various anomalies are detected in the text which should be corrected before agreeing to submit to the Journal "water".

 

Scientific comments

1-      In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

 

2-      Heading 2.7 should be moved down to the next page.

 

3- The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

In this study, the authors examined the Bio-based decontamination and detoxification of total petroleum hydrocarbon contaminated dredged sediments in order to produce constructed technosols in the frame of the circular economy…..

 

The authors have investigated an impact of hydrocarburoclastic ascomycetes, Lambertella sp. MUT 5852, was bioaugmented to dredged sediments co-composting with a lignocellulosic matrix on the depletion of total petroleum hydrocarbons. However, in the opinion of the reviewer, various anomalies are detected in the text which should be corrected before agreeing to submit to the Journal "water".

Scientific comments

1-      In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

2-      Heading 2.7 should be moved down to the next page.

   3- The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

Author Response

Dear Reviewer, thanks for your efforts in our behalf that improved the quality of the manuscript

we modified all the parts you suggested.

Kind Regards

Simona Di Gregorio

In Table 1, the authors should harmonize the units of measurement and write "dredged sediments (mg/Kg)" as follows.

done

 Heading 2.7 should be moved down to the next page.

 done

The images in figures 2, 3, 4 and 5 are blurred and need to be improved!

Sorry but they all have at least 600dpi resolution: at least from my PC, they all look defined. We zoomed them to the maximum wideness possible and resulted to be not blurred

Author Response File: Author Response.pdf