Thermithiobacillus plumbiphilus AAFK—Arsenic-Resistant Bacteria Isolated from Arsenopyrite Material
Rajender Boddula
Round 1
Reviewer 1 Report
Comments and Suggestions for Authors1. Is the methodology for isolating Thermithiobacillus plumbiphilus AAFK from arsenopyrite material clearly described, and is it scientifically robust?
2. Are the genetic mechanisms for arsenic resistance, as identified in the genome of T. plumbiphilus AAFK, thoroughly analyzed and well-supported by experimental evidence?
3. Does the study sufficiently compare the arsenic resistance system in T. plumbiphilus AAFK with those in acidophilic bacteria like A. ferrooxidans?
4. Is the formation of biofilm by T. plumbiphilus AAFK adequately characterized in terms of structure, composition, and function?
5. Are the roles of biofilm in adhesion, mineral interactions, and quorum sensing convincingly explained with supporting experimental data?
6. Does the paper adequately address the potential of T. plumbiphilus AAFK as an auxiliary bacterial culture in arsenopyrite bioleaching?
7. Are the proposed arsenic reduction mechanisms involving polysaccharides and sugars sufficiently detailed and supported by experimental evidence?
8. Is the presence of genes like ACR3, phosphate transporters, and arsenate reductase in T. plumbiphilus AAFK properly linked to the observed arsenic resistance?
9. Are the environmental and biotechnological implications of utilizing T. plumbiphilus AAFK in arsenic-rich environments adequately discussed?
Author Response
Reviewer 1
- Is the methodology for isolating Thermithiobacillus plumbiphilus AAFK from arsenopyrite material clearly described, and is it scientifically robust?
- Yes, the methodology is described clearly and supplemented in connection with specific comments of other reviewers.
- Are the genetic mechanisms for arsenic resistance, as identified in the genome of T. plumbiphilus AAFK, thoroughly analyzed and well-supported by experimental evidence?
- Yes, the genetic mechanisms for arsenic resistance are identified in the genome of T. plumbiphilus and shown at first time both for the species and genus. The As-resistance is confirmed in experiments with the strain.
- Does the study sufficiently compare the arsenic resistance system in T. plumbiphilus AAFK with those in acidophilic bacteria like A. ferrooxidans?
- The paper contains references which confirm the mentioned similarity.
- Is the formation of biofilm by T. plumbiphilus AAFK adequately characterized in terms of structure, composition, and function?
- It is the first description of the biofilm with Thermithiobacillus. It contains detailed description of its structure and composition. Functions of the biofilm are suggested in accordance with known publications and open for discussion, they will be the object of a special further research.
- Are the roles of biofilm in adhesion, mineral interactions, and quorum sensing convincingly explained with supporting experimental data?
- The aim of this paper is to describe the strain T. plumbiphilus and its primary and essential characteristics. The role of biofilms is a separate large theme beyond the scope of this article. The presented study provides specific new data on biofilm formation by Thermithiobacillus. Questions for broad discussion are raised. Functions of the biofilm are suggested and open for discussion.
- Does the paper adequately address the potential of T. plumbiphilus AAFK as an auxiliary bacterial culture in arsenopyrite bioleaching?
- Yes, thank you.
- Are the proposed arsenic reduction mechanisms involving polysaccharides and sugars sufficiently detailed and supported by experimental evidence?
- As far as this topic is beyond the scope of this article, the text is shortened and limited.
- Is the presence of genes like ACR3, phosphate transporters, and arsenate reductase in T. plumbiphilus AAFK properly linked to the observed arsenic resistance?
- Yes, thank you.
- Are the environmental and biotechnological implications of utilizing T. plumbiphilus AAFK in arsenic-rich environments adequately discussed?
- The paper dedicated to new information about the T. plumbiphilus properties. In this paper, the possible industrial implications of the strain are limited with some suggestions.
Reviewer 2 Report
Comments and Suggestions for AuthorsThe manuscript entitled ‘Thermithiobacillus plumbiphilus AAFK-arsenic-resistant bacteria isolated from arsenopyrite material’ was study on the new isolated strain (later identified as T. plumbiphillus AAFK) paying particular attention to its high resistance to arsenic. The research objectives included: testing the effect of different arsenic compounds on bacterial oxidative activity, analyzing the genome and genes responsible for arsenic resistance, and studying the resulting biofilm and its possible role. The work is meaningful. But the quality of the manuscript should be improved.
1. In line 67, what is the meaning of Gen?
2. From lines 121 to 122, please offer more details of the medium.
3. Some short paragraph that only included one sentence, with an example from lines 147 to 148. I suggested the author rewrite to increase the readability.
4. From lines 206 to 214, too many words. I suggested the author show these results by table of figures.
5. In figure 1, please improve the quality of the figure. In addition, many bacteria on genus level with very low abundance. It is very hard to see in the figure. I suggested the author remove them or show them in another formation.
6. The quality of figures 4 and 5 should be improved. For example, the words in the figures were too small to read.
7. Why does the author show the two identifications result in figures 2 and 3?
8. In figure 6, I don’t think the author concluded the cell structure was solid. It would be better to put some reference or element analysis here.
9. In figure 8, it is very hard to read the words in the figure. The cited reference also needs to be listed.
10. In the conclusion, the author should highlight their new findings. In addition, I don’t agree the author to put reference in this part.
Comments on the Quality of English Languageno comments
Author Response
Reviewer 2:
- In line 67, what is the meaning of Gen?
- Thank you for the thorough reading. The mistyped words “Gen 16S rRNA” are replaced with correct “Bacterial 16S ribosomal RNA (rRNA) genes”.
- From lines 121 to 122, please offer more details of the medium.
- Thank you again. The renovated text is detailed: “Bacterial thiosulfate oxidation in the mentioned liquid medium ATCC 290”.
- Some short paragraph that only included one sentence, with an example from lines 147 to 148. I suggested the author rewrite to increase the readability.
- Three paragraphs describing microscopy are merged into a modified new one.
- From lines 206 to 214, too many words. I suggested the author show these results by table of figures.
- Thank you. The text is shortened and the paragraphs are united.
- In figure 1, please improve the quality of the figure. In addition, many bacteria on genus level with very low abundance. It is very hard to see in the figure. I suggested the author remove them or show them in another formation.
- The main information is changed to be clearer and presented as a text. Data on genera/groups present in minor amounts do not allow us to assess their role, but may also be useful for a general understanding of the microbiota.
- The quality of figures 4 and 5 should be improved. For example, the words in the figures were too small to read.
- Thank you for your recommendations. The figures are improved to be more readable.
- Why does the author show the two identifications result in figures 2 and 3?
- The text paragraphs related to the figures are combined into one and supplemented. The subsection heading is also changed accordingly. The principal difference in these figures is that 1 is identification by 16S RNA, while 2 is by genome: until now, data on the genome of T. plumbiphilus were absent. Figure 1 is only sufficient for identification, since genome analysis (Figure 2) confirms the validity of the presence of thermophilic and mesophilic species within one genus.
- In figure 6, I don’t think the author concluded the cell structure was solid. It would be better to put some reference or element analysis here.
- Thank you, the text is replaced with a more accurate description.
- In figure 8, it is very hard to read the words in the figure. The cited reference also needs to be listed.
- Thank you for your recommendations. The figure and legend are changed.
- In the conclusion, the author should highlight their new findings. In addition, I don’t agree the author to put reference in this part.
- Thank you, the text is changed and the references are removed.
Reviewer 3 Report
Comments and Suggestions for Authors1. Line 30: Should be 'mainly' not 'nainly'.
2. Line 49: Correct the grammatical structure of this sentence: `The aim of the study was study`
3. Line 113: Was optimal pH estimated or known before culturing? If pH 7 was used for culturing, why are authors saying, optimal pH was estimated?
4. Line 121: How do authors know 2g/L is the optimum concentration of each these As-compounds? Cite references or provide data to support this claim.
5. Line 122: Which method/technique was used to measure sulfate? If mentioned elsewhere in the paper, provide the section where its described.
6. Line 124: Provide reference for Bradford method.
7. Line 144: Instead of 'by' write 'previously'.
8. Line 217: What is `HB2-32-21 `?
9. Line 230: Label Y-axis in figure 1.
Comments on the Quality of English LanguageEnglish needs to be rechecked. Some grammatical errors and repetitive sentences.
Author Response
Reviewer 3:
- Line 30: Should be 'mainly' not 'nainly'.
- Thank you. The mistyped word is corrected.
- Line 49: Correct the grammatical structure of this sentence: `The aim of the study was study`.
- Thank you again. The new version is: “The aim of the presented research was study on…”.
- Line 113: Was optimal pH estimated or known before culturing? If pH 7 was used for culturing, why are authors saying, optimal pH was estimated?
- The text is changed to be clearer for readers: “Culture of autotrophic bacteria able of thiosulfate oxidation was isolated in the medium DSMZ 71 (https://www.dsmz.de/microorganisms/medium/pdf/DSMZ_Medium71.pdf. Accessed October 05, 2024) with pH 4.5. Here and below this isolated strain is designated as AAFK. As far as the selected pH 4.5 allowed detection of various acidophilic, moderately acidophilic and neutrophilic thiobacilli, the optimal pH value for the isolated strain AAFK was estimated in additional culturing experiments. The later culturing was carried out in the ATCC medium 290 “S6 Medium for Thiobacilli” (https://www.atcc.org/-/media/product-assets/documents/microbial-media-formulations/2/9/0/atcc-medium-290.pdf. Accessed October 05, 2024) with the found optimal pH 7.0.”
- 4. Line 121: How do authors know 2g/L is the optimum concentration of each these As-compounds? Cite references or provide data to support this claim.
- The references and comments are added.
- Line 122: Which method/technique was used to measure sulfate? If mentioned elsewhere in the paper, provide the section where its described.
- The sentence is supplemented with the reference: “(section 2.7. “Chemical analyses” below)”.
- Line 124: Provide reference for Bradford method.
- The reference is added: Kruger, N.J. The Bradford method for protein quantitation. In: Walker, J.M. (eds) The Protein Protocols Handbook. Springer Protocols Handbooks. Humana Press, Totowa, NJ. 2009. https://doi.org/10.1007/978-1-59745-198-7_4.
- Line 144: Instead of 'by' write 'previously'.
- Thank you, the sentence is modified to: “as described by Kanehisa with coauthors previously [11]”.
- Line 217: What is `HB2-32-21 `?
- Explanation is added in the text: “…the group HB2-32-21 which combines representatives of Alteromonadaceae, a family of Pseudomonadota”.
- Line 230: Label Y-axis in figure 1.
- The ordinate axis presents percentage of the detected bacterial taxa.
Round 2
Reviewer 2 Report
Comments and Suggestions for Authorsno comments
