16S rRNA Gene Sequencing Reveals Bacterial Diversity in Khewra Salt Mine Walls
This article has been Reviewed by the following groups
Listed in
This article is not in any list yet, why not save it to one of your lists.Abstract
The Khewra Salt Mine, also known as the source of Himalayan Pink Salt, is the second-largest salt mine in the world and a significant site for salt export. The mine's salt exhibits both amorphous and crystalline textures, with colors ranging from white to deep red, contributing to its distinctive pink hue. While previous research has extensively explored the mine's soil, plants, and brine samples, little attention has been given to the salt walls, which are a major tourist attraction. This study aimed to investigate the bacterial diversity within the salt walls from four distinct locations, including an under-construction area sampled for the first time. From these four collection points, seven samples were gathered, yielding 19 isolates cultured on 2% NaCl nutrient agar. Bacterial isolates from the under-construction area (US1-1, US1-2, US2-1, and US2-2) and the museum walls (MS1-1 and MS1-2) underwent further biochemical, antibiotic sensitivity, and salt requirement testing, followed by 16S rRNA sequencing. Biochemical assays identified these isolates as gram positive, catalase and oxidase positive rods. All isolates were sensitive to chloramphenicol (30μg/ml). Salt tolerance tests classified them as slightly halophilic, thriving at 2-4% NaCl concentrations. Molecular characterization revealed two strains from the under-construction area, US1-1 and US2-2, as uncultured Bacillus species and Bacillus thuringiensis, respectively, both with 94% sequence similarity. The phylogenetic results show monophyletic associations between these two strains which could be due to their shared salt location and common ancestors. The sequences of US1-2, US2-1, and MS1-1 were 100% identical to Bacillus licheniformis, while MS1-2 matched Staphylococcus saprophyticus with 100% similarity. The results of this study point towards significant diversity of bacteria, slightly halophilic, cocci or bacillus shaped. These halophilic bacterial strains, possessing salt-tolerant genes, present the need for further identification of bacterial colonies in the mine and their evolutionary patterns, which could also help uncover the role these microbes play in the ecosystem of the salt mine. Additionally, they show promise for industrial enzyme production.
Article activity feed
-
-
This manuscript describes the isolation of several strains of halophilic bacteria from the Khewra salt mines in Pakistan, and their subsequent culturing, biochemical characterisation and 16S rRNA sequencing. The results show the presence of Bacillus and Staphylococcus species. This is a study that would be of interest to the field and community. Thank you for addressing the reviewer's comments, however before acceptance into Access Microbiology, the editorial team (Editor Mentor and Mentee) would like to add the following: Please remove accession numbers from the abstract, these should be cited in the Materials and Methods section, not in the abstract. Please also consider moving Tables 8-10 (describing statistical analyses of antibiotic sensitivities) into a supplementary file or integrate into the main text.
-
-
The reviewers have highlighted minor concerns with the work presented. Please ensure that you address their comments.
-
Comments to Author
How is the same salt mine producing 314,845 tons and another time 3,796,634 for different reports in the same year 2021? Lines 90 - 92. This line has no connection with the previous statements. You could introduce the importance of salt and then make such statements. Lines 97 - 101 rephrase these statements to be more active. Lines 107 - 108. This should go to methods. Lines 108 - 112. This should go to methods. 112 - 120 What is the bases for the selection of six isolates from the 19 isolates? Line 165
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have …
Comments to Author
How is the same salt mine producing 314,845 tons and another time 3,796,634 for different reports in the same year 2021? Lines 90 - 92. This line has no connection with the previous statements. You could introduce the importance of salt and then make such statements. Lines 97 - 101 rephrase these statements to be more active. Lines 107 - 108. This should go to methods. Lines 108 - 112. This should go to methods. 112 - 120 What is the bases for the selection of six isolates from the 19 isolates? Line 165
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
-
Comments to Author
Reviewer Comments for Manuscript Number: ACMI-D-24-00130 Comprehensive Analysis of Bacterial Diversity in the Salt Walls of Khewra Salt Mine Using 16S rRNA Gene Sequencing Dear Authors There are several notes regarding your manuscript as follows. The title: I suggest you make a few changes that would be more appropriate to the study. Abstract - The abstract needs some changes and rearrangement of …. Impact Statement: 39 This study has unveiled the diverse bacterial species inhabiting the salt walls of Khewra, focusing 40 particularly on halophiles and their required salt concentrations. By identifying 19 isolates 41 predominantly…………………………………. This paragraph must be at the end of the research before the references .Data Summary: 16S rRNA sequences submitted in GenBank: US1-1 (accession code …
Comments to Author
Reviewer Comments for Manuscript Number: ACMI-D-24-00130 Comprehensive Analysis of Bacterial Diversity in the Salt Walls of Khewra Salt Mine Using 16S rRNA Gene Sequencing Dear Authors There are several notes regarding your manuscript as follows. The title: I suggest you make a few changes that would be more appropriate to the study. Abstract - The abstract needs some changes and rearrangement of …. Impact Statement: 39 This study has unveiled the diverse bacterial species inhabiting the salt walls of Khewra, focusing 40 particularly on halophiles and their required salt concentrations. By identifying 19 isolates 41 predominantly…………………………………. This paragraph must be at the end of the research before the references .Data Summary: 16S rRNA sequences submitted in GenBank: US1-1 (accession code OQ295687), 56 US1-2 (accession code OQ978562), US2-1 (accession code OQ978592), US2-2 (accession code 57 OQ692139), MS1-1 (accession code OQ978561), and MS1-2 (accession code OQ978593). 58 59 Abbreviations: ES- Entrance Salt, CS- Chandni Salt, MS - Museum Salt, US - Under-construction 60 Salt Introduction - Need to write in the passive ….. Salt is an ancient compound used as a preservative in food, chemicals, feedstock, medicine, 65 and detergents. It is derived from brine, soil, and rock salt of dried-up lakes in mines, with 66 varying constituents like magnesium, sodium, potassium, and sulfur that determine its 67 characteristics. Salt intake - Need to write modern studies and write the objects of study in introduction. Please, Add statistical analysis Conclusion 376 5 Conclusion, ….. re-write conclusion with numbered Bibliography/References : add more modern references and relevant to the study field and recent, it is recommended to use software like Mendeley or any modern programs to manage the bibliography list.
Please rate the manuscript for methodological rigour
Good
Please rate the quality of the presentation and structure of the manuscript
Good
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
-