Ecotoxicological Assessment of Potentially Toxic Elements in Waterworks Sludge Amended Soils Using Bermudagrass Bioassay

Reviewer #1’s comments

My response

Avoid using the confusing term “heavy metals” and replace by trace or toxic metals or trace or toxic elements. (See references: Pourret & Hursthouse, Intern. J. Environ. Res. Public Health, 2019, 16, 4446; Duffus, Pure Appl. Chem., 2002, 793; Hodson, Environ. Pollut., 2004, 129, 341., etc.)

Thank you for your comments.  The term “heavy metals” is replaced by “potentially toxic elements” (Pourret & Hursthouse, 2019).

Section 2.1, line 2: replace profile by concentration. Were those values obtained from a reference (must be given) or by an analytical digestion procedure (must be given), in any case with standard deviation?

Thank you for your comments. The sentence is revised to “the composition of potentially toxic elements … (Table 1) were determined …(Line 83)”

Two sludge samples were used for the determination of the chemical composition. The digestion method and references are provided in Lines 81-85.

Section 2.1, line 3 and following: use past tense to describe methodology (e.g. sludge was oven-dried… etc.).

Thank you for your comments.  T have revised the Materials and Methods to past tense.

Section 2.2, The SI symbol for liter is capital L (mL)

Yes, it is revised to “mL/cm2” (Line 138).

Section 2.3, line 2: Any possible losses of volatile elements during ashing at 480 oC? Many elements listed are not heavy metals (e.g. Al) or even metals (e.g. As). More details must be given on the determination of the studied elements such as Al and As by atomic absorption spectrometry?

The digestion method for the determination of metal contents in plant tissues was developed by Sasmaz et al. (2009) and Yabanli et al. (2014).

Because potentially toxic elements that do not form volatile compounds, loss due to evaporation would be minimal.

Nine potentially toxic elements were determined by flame atomic absorption spectrometry (FAAS)(Ferreira et al., 2018)

Results and Discussion: Unfortunately, several aspects of the discussion and interpretation of results are of limited validity without the presence of a solid statistical analysis showing or not significant differences between the different treatments and because of the absence of quality control on all analyses through the utilization of certified reference material(s). Results are presented with, likely too many significant digits considering the used analytical technique for metal/element determination, and without standard deviations or analytical errors.

Thank you for your comments.  I have performed ANOVA to test the differences between treatments. The results are shown in Figures 1-3.

Reviewer #2’s comments

My response

(Line 3) The title have to included the name of plant studied

Thank you for your comments.  I have included the name of plant in the title. Actually, I have revised the title of the manuscript according to the comments of other reviewers.

(Line 7) the results in the abstract are not enough, So, the authors have to write more details regarding this point

(Line 7) Numerical results have to write especially regarding the important parameters

(Line 7) The authors have to rewrite the abstract part avoiding repetition. When reading the summary part only, it is necessary for the reader to know what this scientific paper contains in brief. Hence, authors must include the experimental treatments that affected the growth of the plant and its chemical analysis, but the authors did not include this in the abstract part

(Line 13) the authors have to write some details about the experimental treatments in brief

I have extensively rewritten the abstract for a better presentation of this study.  Also, I have provided more details about this study in the Abstract. 

However, it is Environments’ requirement that the length of the Abstract should not be longer than 200 words.

I cannot elaborate the experimental design and treatments in the Abstract.

(Line 14) What about the remaining 75%?

Loading rates of waterworks sludge are in terms of sludge weight to the total weight (wt/wt) (Line 120)

(Line 16) Remove this sentence to the end of the abstract part

Yes, removed

(Line 20) Pls write the conclusion in the abstract part in a direct form because it is unclear.

Thank you for your comments.  The Abstract is revised accordingly.

(Line 21) It is preferable do not repeat the words that mentioned in the title, in the keywords 

Thank you for your comments.  I make new keywords that do not repeat in the title

(Line 27) pls delete, no need to these sentences

Thank you for your comments.  I have deleted the sentences.

(Line 39) pls add relevant referance

Yes, a relevant reference is provided.

(Line 58) shorten

Yes, the sentence is shortened.

(Line 70) delete

Yes, the phrase “for 3 months” is deleted.

(Line 102) If you have referance, pls add

Yes, a relevant reference is provided.

(Line 111) correct  

Yes, the phrase “equal to 90g/m2” is deleted because of duplication

(Line 141) add relevant referance

Yes, 2 relevant references are provided.

(Line 141) [34, 35]

Yes, it is revised to “[34, 35]”.

(Line 157) there is nothing regarding the statistical analysis of the current study, this is critical point

(Line 158) the authors have to write the results of the current study in more details supporting these results by numerical results in the entire results section after that discuss these results and support this discussion by the previous literature

Thank you for your comments.  I had performed ANOVA to test the differences between treatments. The results are shown in Figures 1-3.

(Line 159) Figure (1)

Yes, it is revised to “Figure (1)”.

(Line 164) Numerical results have to be added

There are too many numbers to be presented.  For the detailed number, the readers can refer to the Figures.

Yes, will do

(Line 167) the same previous comment

(Line 174) the quality and the resolution of this figure are not good, especially the written data

Because it is a group of diagrams, the size of each diagram is small although the resolution is actually high. To make the figures bigger, I have: 1/ revised the diagram by making bigger font sizes; 2/ rotate the orientation.

(Line 192) Figures (2-5)

Yes, it is revised to “Figures (2-5)”.

(Line 205) the same comment in figure (1)

The same response to the comment on figure (1)

(Line 220) the same

The same response to the comment on figure (1)

(Line 251) OK

Thank you

(Line 282) Ok, but you have to add recent relevant referances regarding the topic of this manuscript (2020-2022)

Thank you for your comments.  I have added a few recent references.  There are 10 recent references (2020-2022) in the reference list.

Reviewer #3’s comments

My response

This paper investigated the heavy metal content of bermudagrass grown under different loading rates and treatments of waterworks sludge. Results of the study may have important application in use of water treatment sludge in land treatment. Authors may wish to consider the following comments in revising of their manuscript

Thank you for your comments. I had revised the MS accordingly.

1.      Please provide details of preparation of mixtures of waterworks sludge and decomposed granite (DG), and waterworks sludge and volcanic soil (VS).

The mixing of waterworks sludge and soils is based on weight (Lines 120-121).

2.      Please present results of control experiments using waterworks sludge only, using DG only and using VS only.

The results of the control group in Experiment 1 are presented in Lines 182-183.

The results of the control group in Experiment 2 are provided in Lines 222-226.

3.      Please provide chemical analysis data of mixture containing waterworks sludge and decomposed granite (DG), and data of mixture containing waterworks sludge and volcanic soil (VS).

Thank you for your comments. I have added a section to detail the chemical data of DG and VS.  The results are listed in table 2.

4.      Please perform mass balance on various heavy metals regarding pot experiments.

Thank you for your comments. However, I am sorry to say that I am not able to calculate the mass balance because the design of the experiment does not serve this purpose.  For example, certain amounts of heavy metals may be leached away through percolation.

5.      In the greenhouse experiments, please provide data of duration of sunlight and intensity of sunlight.

The experiments were conducted in the greenhouse.  Basically, water input and temperature were controlled, but the duration and intensity of sunlight were not.  To control all climatic variables, an incubation chamber is needed.

6.      Please provide government regulations regarding heavy metals related to land application of waterworks sludge.

Hong Kong government have not regulations regarding heavy metals related to land application of waterworks sludge.  That’s why this study was funded by the Hong Kong Government.  They wanted some scientific findings to inform their policy making

7.      Please comment on the limitations of the experimental procedures used in the study.

I have revised the Conclusions and Limitations.  This study represents a preliminary assessment of the ecotoxicological risk of sludge-amended soils.  It tests 9 metals using bermudagrass bioassay. Future studies should include more metals and/or bioassays.