Synergistic Removal of β-Hexachlorocyclohexane from Water via Microorganism–Plant Technology and Analysis of Bacterial Community Characteristics
Round 1
Reviewer 1 Report
Recommendation: Minor revision
This is a review of the manuscript entitled: ‘Synergistic removal of β-HCH from water by microorganism and plant and analysis of bacterial community characteristics’.
This is an interesting manuscript exploring the use of β-15 HCH degrading bacteria——Ochrobactrum (B, F) and Pseudomonas (J) combined with Canna for microbial, plant and microbial-plant repair hydroponics experiments”. Overall, the research writing is pretty decent making it easily understandable for the audience. This work would be more interesting to Water Journal if some revision have been made.
To me the author present many results but lack in some of the discussion part on each experiment, and the results still required connectivity between experiments. I also suggest the authors state more of their novelty of this contribution.
1. The abstract had merely results. I recommend putting more of the new finding (with discussion) in the abstract. This would definitely make this article stand out.
2. The introduction could have been more attractive in term of stating the new finding and stated while this quite complicated combination would be more interesting than other known effective methods.
3. At the end, however, I'd like the authors to reflect on the value of manuscript by telling readers how all this research material could be used in practice and applied to the conventional industrial wastewater treatment plant. Without some application, the value of this work will be lost.
Author Response
Comment 1: The abstract had merely results. I recommend putting more of the new finding (with discussion) in the abstract. This would definitely make this article stand out.
Response: Thanks for your decision and constructive comments on my manuscript, we have carefully revised the abstract.
Comment 2: The introduction could have been more attractive in term of stating the new finding and stated while this quite complicated combination would be more interesting than other known effective methods.
Response: Thanks for your suggestions. We added more content to enrich the introduction in line 52-115.
Comment 3: At the end, however, I'd like the authors to reflect on the value of manuscript by telling readers how all this research material could be used in practice and applied to the conventional industrial wastewater treatment plant. Without some application, the value of this work will be lost.
Response: Thanks for your suggestions. We added the use of bacterial suspension in practical engineering and the prospect of the future in line 270-277 and 502-507. We added some account: “In practical engineering, the strains used in this study can be made into biological agents and introduced into polluted water. However, Ochrobactrum may contain pathogens that are harmful to humans and animals. Therefore, it is important to remember that access to sites with polluted water in which this bacterial suspension has been used should be restricted for 6 days. Previous experiments have confirmed that these three strains will stop growing after 6 days and then lyse and die; and the high-efficiency bacterial degrading agents for target pollutants are potentially available natural biological agents, simple to operate, and have incredibly high environmental and economic benefits in the large-scale removal of organic pollutants in sewage. Thus, they represent a promising alternative for treating β-HCH-contaminated water in combination with plants.”
Reviewer 2 Report
The paper is very important, but is necessary:
1) add recent references in the introduction;
2) improve english in the manuscript;
3) add more chromatographic information.
The paper is very important, but is necessary:
1) add recent references in the introduction;
2) improve english in the manuscript;
3) add more chromatographic information.
Author Response
Comment 1: add recent references in the introduction.
Response: Thanks for your suggestions. We added to the literatures in the introduction.
Comment 2: improve english in the manuscript.
Response: Thanks for your suggestion. We have tried our best to polish the language in the revised manuscript.
Comment 3: add more chromatographic information.
Response: Thanks for your suggestion. We have supplemented the chromatogram in the supplementary in figure A2 and A3.
Reviewer 3 Report
The manuscript, entitled "Synergistic removal of β-HCH from water by microorganisms and plant and analysis of bacterial community characteristics" is based on a novel idea but author needs careful revision of manuscript for better understanding. So, I recommend the "major revision" of this manuscript after handling the suggestions listed below.
- Line 2-3; Title: Title needs revision it should be eye catching and more comprehensive. It is suggested to use word "and" only once and avoid abbreviations in the title.
- Line 32-33; Key words: Arrange key words alphabetically and add few more.
- Line 35-74; Revise introduction and add major sources of β-HCH pollution and role of specific microbes in its degradation. Also mention the threshold level of β-HCH pollution for plants and water reservoirs. Ochrobactrum is also a human pathogen, kindly mention inoculation of this bacteria is safe.
- Line 98-105; Materials and methods: as we know bioremediation/phytoremediation is a slow process so do you think a 7 day experiment is enough for this study.
- Line 110-116; Materials and methods: kindly mention how many canna plantlets were used for each treatment and also mention the age of canna plantlets used for this experiment. Also mention the experimental design used for this study.
- Line 120-154; Materials and methods: Provide appropriate references for different adopted protocols for analysis.
- Line 162; Results and Discussion: Simply revise this "B and J(p<0.05)" as "B and J (p<0.05)".
- Line 210; Results and Discussion: Check out this reference style "Shahid et al., (2019)[29]."
- Lines 345–362; Conclusions: Add briefly the future recommendations at the end of this section and the benefits of this research for the general public.
- Line 379-467. References: Kindly ensure the citation of all the references in the text with the correct sequence and numbers provided in the reference list.
Carefully revise the whole manuscript for English language grammar check.
Author Response
Comment 1: Line 2-3; Title: Title needs revision it should be eye catching and more comprehensive. It is suggested to use word "and" only once and avoid abbreviations in the title.
Response: Thanks for your suggestion. We have revised the title as “Synergistic removal of β-hexachlorocyclohexane from water via microorganism–plant technology and analysis of bacterial community characteristics.”
Comment 2: Line 32-33; Key words: Arrange key words alphabetically and add few more.
Response: Thanks for your suggestion. We have reordered and supplemented the keywords.
Comment 3:Line 35-74; Revise introduction and add major sources of β-HCH pollution and role of specific microbes in its degradation. Also mention the threshold level of β-HCH pollution for plants and water reservoirs. Ochrobactrum is also a human pathogen, kindly mention inoculation of this bacteria is safe.
Response: Thank you for your rigorous and professional suggestion.
We added major sources of β-HCH pollution and the threshold level of β-HCH pollution for plants and water reservoirs in line 51-69: “In 1991, it was found that γ-HCH in technical hexachlorocyclohexane was the main insecticide component, and so the production and use of HCHs in China gradually decreased. However, due to their characteristics of lipophilicity, high toxicity, persistence, and limited biodegradability, many isomers of hexachlorocyclohexane persist at the production site and migrate through environmental carriers, having harmful effects on human health and the ecological environment. The chlorine atom in HCHs makes these organochlorine compounds highly stable in the environment, and, of these, β-HCH is the most stable and difficult to degrade. At present, HCHs are still commonly detected in surface water, groundwater, rivers, and oceans, with a detection rate of almost 100% , about 1.1~14.8 ng·L−1. Studies have shown that HCHs concentration of 4.72 ~ 11.19 ng·g−1 have also been detected in aquatic plants.”
We describe the degradation of specific microorganisms in line 76-96 : “Microbe–plant joint remediation technology overcomes the limitations of traditional treatment processes and makes the remediation process more environmentally and economically beneficial. However, the combination of indigenous microorganisms and plants often has a limited ability to degrade specific pollutants, so the injection of exogenous high-efficiency degrading strains can improve the problem of pesticide contamination. This green and sustainable remediation technology can accelerate natural degradation and reduce secondary pollution. Specific contaminant-degrading bacteria are crucial for successful remediation, and studies have shown that this method is possible …This may be attributed to the fact that exogenous high-efficiency degrading bacteria degrade toxic pesticides into simple inorganic substances, and the dehalogenation products formed by microorganisms after pesticide degradation are absorbed by plants and further oxidized by enzymes involved in degradation to improve the removal efficiency of pollutants in the system.”
We gave a reminder of the application of Ochrobactrum in lines 271-278. We added some caveats: “In practical engineering, the strains used in this study can be made into biological agents and introduced into polluted waters. However, Ochrobactrum may contain pathogens that are harmful to humans and animals. Therefore, it is important to remember that access to sites with polluted water in which this bacterial suspension has been used should be restricted for 6 days. Previous experiments have confirmed that these three strains will stop growing after 6 days and then lyse and die.”
Comment 4:Line 98-105; Materials and methods: as we know bioremediation/phytoremediation is a slow process so do you think a 7 days experiment is enough for this study.
Response: Thank you for your rigorous and professional suggestion. According to our previous study, the three degraded strains used all stopped proliferating and began to lysis and die after 6 days, so the experiment period was set at 7 days.
Comment 5: Line 110-116; Materials and methods: kindly mention how many canna plantlets were used for each treatment and also mention the age of canna plantlets used for this experiment. Also mention the experimental design used for this study.
Response: Thanks for your suggestion. We added to the content in line 119-121 and 160-161: “ Each phytoremediation and microbe–plant remediation unit included two Canna samples that were about 60~70cm tall. The experimental design in line 169-171 was as follows: control, microbial remediation, plant remediation, and microbe–plant remediation phases, and all experiments were repeated three times, with an experimental period of 7 days.
Comment 6: Line 120-154; Materials and methods: Provide appropriate references for different adopted protocols for analysis.
Response: Thanks for your suggestion. We have supplemented the materials and methods section in line 155-201.
Comment 7: Line 162; Results and Discussion: Simply revise this "B and J(p<0.05)" as "B and J (p<0.05)".
Response: Thanks for your suggestion. We carefully examined the full text of the experimental group naming and modification.
Comment 8: Line 210; Results and Discussion: Check out this reference style "Shahid et al., (2019)[29]."
Response: Thanks for your suggestion. We carefully examined and modified the citation patterns of the full text.
Comment 9: Lines 345–362; Conclusions: Add briefly the future recommendations at the end of this section and the benefits of this research for the general public.
Response: Thanks for your suggestion. We briefly added the future recommendations at the end of this section and the benefits of this research for the general public in line 500-505, we added some account: “ The high-efficiency bacterial degrading agents for target pollutants are potentially available natural biological agents, simple to operate, and have incredibly high environmental and economic benefits in the large-scale removal of organic pollutants in sewage. Thus, they represent a promising alternative for treating β-HCH-contaminated water in combination with plants.”
Comment 10: Line 379-467. References: Kindly ensure the citation of all the references in the text with the correct sequence and numbers provided in the reference list.
Response: Thanks for your suggestion. We carefully checked the order and number of all the references.
Reviewer 4 Report
In the MS entitled, Synergistic removal of β-HCH from water by microorganism 2 and plant and analysis of bacterial community characteristics, authors have subjected β-HCH contaminated water to 3 different kinds of bacteria alone and in association with Canna plant. There results revealed that the combination of Canna and bacteria have more bioremediation ability. The work is interesting but language and data representation is not up to the mark.
Please see the following comments for further improvement of the MS.
· Title must be revised. Avoid repetition of same words.
· In abstract- ass 1-2 lines of background and research gap.
· L76 2.1. Main reagent must be Main reagents.
· L84 Mention in brief how β-HCH-degrading bacteria were screened?
· In figure 1- Highlight the bacterial strain used for the study.
· Mention the dimensions of vases used for the experiment.
· A supplementary figure showing experimental set up will be useful for the readers.
· In 2.3.2. Phytoremediation- mention age/growth stage of Canna plantlets.
· The methods are not s ufficiently described. Revise the materials and methods section thoroughly.
· In results, Bacterial strain type- J, showed higher degradation (60%) of β-HCH at 100 μg·L−1 as compared to 10 μg·L−1 (30%). Justify the results.
· Discussion must be improved. At several places only results are written.
· Avoid writing results in the conclusion.
· If possible, carry out phytotoxicity analysis of bioremediated water after harvesting of Canna plants and studied bacteria.
Extensive Language editing is required.
Author Response
Comment 1: Title must be revised. Avoid repetition of same words.
Response: Thanks for your suggestion. We have revised the title as “Synergistic removal of β-hexachlorocyclohexane from water via microorganism–plant technology and analysis of bacterial community characteristics.”
Comment 2: In abstract- ass 1-2 lines of background and research gap.
Response: Thanks for your suggestion. We have supplemented the abstract: “In recent years, β-Hexachlorocyclohexane(β-HCH) has been detected frequently in water, seriously threatening human health and ecological balance.”
Comment 3: L76 2.1. Main reagent must be Main reagents.
Response: Thanks for your suggestion. We have modified the reagent part of the experiment in line 117-131.
Comment 4: L84 Mention in brief how β-HCH-degrading bacteria were screened?
Response: Thanks for your suggestion. We've added screening methods for strains in line 133-138, we added some account: “For the strain screening method, a mixture of mud and water from the rhizosphere of Canna in a β-HCH-contaminated area was cultured in 100 mL of enrichment medium for 48h, diluted 10-6 times, and coated on beef extract peptone solid medium. After being cultured at 37℃ for 24h, a single colony was selected to verify the degradation ability. Finally, three strains of β-HCH-degrading bacteria were identified: Och1, Och2, and Pse1.”
Comment 5: In figure 1- Highlight the bacterial strain used for the study.
Response: Thanks for your suggestion. We've highlighted the names of the strains in color.
Comment 6: Mention the dimensions of vases used for the experiment.
Response: Thanks for your suggestion. We increased the size of the vessel is 10cm*10cm*30cm in line 160.
Comment 7: A supplementary figure showing experimental set up will be useful for the readers.
Response: Thanks for your suggestion. We have supplemented the experimental procedure diagram in the supplementary in figure A1.
Comment 8: In 2.3.2. Phytoremediation- mention age/growth stage of Canna plantlets.
Response: Thanks for your suggestion. We have described the dimensions of Canna in line 161-162 and 182-184, we added some account: “Each phytoremediation and microbe–plant remediation unit included two Canna samples that were about 60~70cm tall and Canna seedlings with similar growth potential were collected, washed and pre-cultured in deionized water without β-HCH.
Comment 9: The methods are not s ufficiently described. Revise the materials and methods section thoroughly.
Response: Thanks for your suggestion. We have modified the materials and methods section.
Comment 10: In results, Bacterial strain type-J, showed higher degradation (60%) of β-HCH at 100 μg·L−1 as compared to 10 μg·L−1 (30%). Justify the results.
Response: Thanks for your suggestion. We added discussion and contrast in line 251-271, we added some account: “ The results showed that under 10 μg·L−1 β-HCH stress, the β-HCH degradation rate for Och1 was 33.49%, which was 3.18 and 1.18 times that of Och2 and Pse1, respectively. Compared with Och2, the degradation rate of β-HCH was significantly increased by Och1 and Pse1(p<0.05); these results indicated that Och1 and Pse1 could effectively degrade β-HCH at low concentrations. Under 100 μg·L−1 β-HCH stress, Och1 and Och2 showed poor tolerance to β-HCH con-centration, and the degradation rates were only 5.72% and 0.62%. However, Pse1 showed strong degradation ability, and the removal rate of β-HCH reached 60.02%, which indicated that the degrading bacteria were susceptible to the external environment. The tolerance and degradation ability of β-HCH was different among different species of bacteria. It was worth noting that different concentrations of β-HCH had a more significant effect on different strains. When the concentration of β-HCH increased from 10μg·L−1 to 100μg·L−1, the degradation rate of β-HCH was significantly decreased by Och1 and Och2, which may be because the proliferation of Och1 and Och2 was inhibited, and the cell viability was reduced at this concentration. The advantage of microbial remediation is that it can consume organic pollutants in the environment as a carbon source to maintain its metabolic growth, completely removing them from the water by mineralizing them into harmless end products.”
Comment 11: iscussion must be improved. At several places only results are written.
Response: Thanks for your suggestion. We have revised the discussion section and added some comparison with literature to the chapters 3.2.
Comment 12: Avoid writing results in the conclusion.
Response: Thanks for your suggestion. We revised the conclusion in line 488-506.
Comment 13: If possible, carry out phytotoxicity analysis of bioremediated water after harvesting of Canna plants and studied bacteria.
Response: Thanks for your suggestion. Your suggestion will make our results more in-depth, but we cannot continue the experiment at present due to work reasons. We will conduct detailed research in this direction in the follow-up experiment.
Reviewer 5 Report
Check for typos (space, dot, capitol letter etc..) and for less common used abbreviation (instead r/min is more common rpm).
The name Canna refer only to the genus. It is needed also indicate the species
Could be better not use the internal laboratory clone name (B, F and J) but some abbreviation, like Och 1, Och 2, Pse 1 or something like this to facilitate the interpretation of data and the difference between the two Och species. Also other abbreviation, like CK, K and M aren’t so intutitive. Provide more information regarding these three strains and why you chose these (I wasn't able to find the paper that you cited) and create a specific paragraph in the result and discussion part in with also include the 16S rDNA sequence analysis.
In the material and method part, in some case, the procedures aren’t clear and also some details (centrifuge speed, temperature, time, etc..) are missing (see 122-128, 130-141, 145, 150-154)
The figures captions lack of information like the statistic used (Fig. 2 and 3) and the statistic appear not congruent with the data (Fig. 2). The Fig. 4 could be enriched with the statistical analysis and could be more correct use the same X-Axis name like in Fig.3.
65 what do you mean for inorganic?
82 do you mean HPLC grade?
102 It isn’t clear why you add methanol. It isn’t also clear that the final concentration used in the experiment of β-HCH is 10 μg/L or 100 μg/L. I suggest rearranging a little the phrase living the indication that β-HCH stock was dissolved in acetone.
115 What kinds of water do you use?
116 It isn’t clear the CK water composition
119 B, F and J are the selected clone of the bacteria used, but it isn’t clear why you used M to refer to the presence of Canna.
168 not clear sentences
242 use translocations instead transshipment
256 accumulations instead enrichment
270 accumulate instead enrich.
328-341 degraded or degrading bacteria? Only two increasing percentage reported but 3 growing conditions are cited.
342 not clear legend, try to separate better the Fig. 5A and 5B
In the “Analysis of Species composition of bacterial community at genus level” it isn’t clear and explained why you don’t find in the BM, FM and JM samples (water or root) a relevant presence of the inoculated genus Ochrobactrum and Pseudomonas. If this is the correct result, it is very difficult conclude that these selected strains are able to degrade the β-HCH and also that they can differentially influence the accumulation of β-HCH in Canna. It is more reasonable think that they can influence the bacterium community before disappear in favor of other genus like Methylophilus and other, that are more involved in the detoxification from β-HCH and to influence Canna to accumulate and translocate this contaminant.
The level of English is acceptable, but it is needed to improve. There are some errors that can bring to misunderstand the content (see 328). In some case appear that it is accepted the translated word from the translation system without a revision
Author Response
Comment 1: Check for typos (space, dot, capitol letter etc..) and for less common used abbreviation (instead r/min is more common rpm).
Response: Thanks for your suggestion. We are deeply sorry for our carelessness, and we have checked and modified the whole text according to your suggestions.
Comment 2: The name Canna refer only to the genus. It is needed also indicate the species
Response: Thanks for your suggestion. We have a more detailed explanation of the Canna information in line 179 (Canna indica L.).
Comment 3: Could be better not use the internal laboratory clone name (B, F and J) but some abbreviation, like Och 1, Och 2, Pse 1 or something like this to facilitate the interpretation of data and the difference between the two Och species. Also other abbreviation, like CK, K and M aren’t so intutitive. Provide more information regarding these three strains and why you chose these (I wasn't able to find the paper that you cited) and create a specific paragraph in the result and discussion part in with also include the 16S rDNA sequence analysis.
Response: Thanks for your suggestion.(1) We have modified the name of the experimental group according to your suggestion. (2) More detailed studies on the three strains, such as strain identification, degradation performance, and influencing conditions, have been published in our article in a Chinese journal.(named Screening and identification of bacteria β-HCH rhizosphere degradation in constructed wetland plants) (3) We added the content of strain identification in the results and discussion 3.1 section and the sequence information of strains in the appendix.
Comment 4: In the material and method part, in some case, the procedures aren’t clear and also some details (centrifuge speed, temperature, time, etc..) are missing (see 122-128, 130-141, 145, 150-154)
Response: Thanks for your suggestion. We have carefully revised the materials and methods section.
Comment 5: The figures captions lack of information like the statistic used (Fig. 2 and 3) and the statistic appear not congruent with the data (Fig. 2). The Fig. 4 could be enriched with the statistical analysis and could be more correct use the same X-Axis name like in Fig.3.
Response: Thanks for your suggestion. We have explained the statistical method in line 235-238, and modified and supplemented the picture information. we added some account: “Values are expressed as mean ± standard deviation (SD). Different lowercase letters (a, b) indicate significant (p < 0.05) differences in concentration of β-HCH among conditions (like colors should be compared with each other), and *, p < 0.05; **, p < 0.01; -, p > 0.05. Like colors are compared to each other.”
Comment 6: 65 what do you mean for inorganic?
Response: Thanks for your suggestion. The inorganic culture medium are composed of KH2PO4 2 g，NH4Cl 1 g，K2HPO4 7.5 g，NaCl 0.5 g，MgCl 0.1 g, (RO)-water 1L.
Comment 7: 82 do you mean HPLC grade?
Response: Thanks for your suggestion. Yes.
Comment 8: 102 It isn’t clear why you add methanol. It isn’t also clear that the final concentration used in the experiment of β-HCH is 10 μg/L or 100 μg/L. I suggest rearranging a little the phrase living the indication that β-HCH stock was dissolved in acetone.
Response: Thanks for your suggestion. (1) Methanol was added to the experimental facility as an additional carbon source. (2) In our experiment, β-HCH was set to two stress concentrations of 10 μg/L and 100 μg/L, respectively. (3) The β-HCH standard solution was dissolved in acetone and then added to the experimental water. The acetone would evaporate quickly.
Comment 9: 115 What kinds of water do you use?
Response: Thanks for your suggestion. The water in the experimental device was all deionized water, which we have explained in the materials and methods section.
Comment 10: 116 It isn’t clear the CK water composition
Response: Thanks for your suggestion. I'm sorry for causing your misunderstanding. CK was represented for deionized water in pre-culture Canna, which does not contain β-HCH.
Comment 11: 119 B, F and J are the selected clone of the bacteria used, but it isn’t clear why you used M to refer to the presence of Canna.
Response: Thanks for your suggestion. M comes from the Chinese word for Canna.
Comment 12: 168 not clear sentences
Response: Thanks for your suggestion. We have revised this part.
Comment 13: 242 use translocations instead transshipment
Response: Thanks for your suggestions. In line 362, “transshipment” has changed to “translocations”.
Comment 14: 256 accumulations instead enrichment
Response: Thanks for your suggestion. In line 373, “enrichment” has changed to “accumulations”.
Comment 15: 270 accumulate instead enrich.
Response: Thanks for your suggestion. In line 392, “enrich” has changed to “accumulate”.
Comment 16: 328-341 degraded or degrading bacteria? Only two increasing percentage reported but 3 growing conditions are cited.
Response: Thanks for your suggestion. I am very sorry that we did not make it clear. We compared it according to the two stress concentrations of β-HCH and have revised the description of this part in line 458-463. We change the content to : “Compared with the treatments that were not inoculated with degrading bacteria under 10 μg·L−1 and 100 μg·L−1 β-HCH stress, the total proportion of Methylophilus in Canna-degrading bacteria treatments increased by 78.59% and 48.04%, respectively. The total proportion of Paracoccus and Methylobacillus increased by 3.28% and 1.92%, 0.48%, and 4.34%, respectively. The total proportion of Methylophilus in the Canna root samples of the Canna-degrading bacteria treatments increased by 40.88% and 36.61%, respectively.
Comment 17: 342 not clear legend, try to separate better the Fig. 5a and 5b.
Response: Thanks for your suggestions. We do apologize for the size of the legend and images; Figure 5 have been revised to make them clearer.
Comment 18: In the “Analysis of Species composition of bacterial community at genus level” it isn’t clear and explained why you don’t find in the BM, FM and JM samples (water or root) a relevant presence of the inoculated genus Ochrobactrum and Pseudomonas. If this is the correct result, it is very difficult conclude that these selected strains are able to degrade the β-HCH and also that they can differentially influence the accumulation of β-HCH in Canna. It is more reasonable think that they can influence the bacterium community before disappear in favor of other genus like Methylophilus and other, that are more involved in the detoxification from β-HCH and to influence Canna to accumulate and translocate this contaminant.
Response: Thank you for your rigorous and professional suggestion. We have modified the content according to your suggestion in line 471-476, we modify the content as: “Although no degraded bacteria were directly found in the experimental water and the Canna rhizosphere; according to the experimental results, this may be due to the degradation bacteria promoting the growth of biodegradable β-HCH bacteria, such as Methylophilus, which were more involved in the detoxification of β-HCH before their disappearance.”
Comment 19: The level of English is acceptable, but it is needed to improve. There are some errors that can bring to misunderstand the content (see 328). In some case appear that it is accepted the translated word from the translation system without a revision
Response: Thanks for your suggestion. We have tried our best to polish the language in the revised manuscript.
Round 2
Reviewer 3 Report
Acepted in currunt form
Minor english editing required
Reviewer 4 Report
The authors have significantly improved the manuscript. It can be accepted for publication.
Minor editing of the English language is required.
Reviewer 5 Report
Dear authors,
all the comments that I suggested were taken in consideration. Now the work is more clear and easy to read.
