Evaluation of a commercial pressure cooker for the preparation of agar media for a diagnostic microbiology laboratory

Joseph E. Rubin; Florence Huby; Roshan P. Madalagama; Shyamali de Alwis; Melinda Wyshynski; Rasika Jinadasa

doi:10.1371/journal.pone.0336988

Peer Review History

Original SubmissionMay 23, 2025
18 Aug 2025 Decision Letter - Tsegaye Alemayehu, Editor Dear Dr.Joseph E. Rubin plosone@plos.org . When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file. A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'. A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'. An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'. If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. 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If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent. If data are owned by a third party, please indicate how others may request data access. 7. If the reviewer comments include a recommendation to cite specific previously published works, please review and evaluate these publications to determine whether they are relevant and should be cited. There is no requirement to cite these works unless the editor has indicated otherwise. [Note: HTML markup is below. Please do not edit.] Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. Is the manuscript technically sound, and do the data support the conclusions? Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? -->?> Reviewer #1: Yes Reviewer #2: No Reviewer #3: I Don't Know ****** 3. Have the authors made all data underlying the findings in their manuscript fully available??> The PLOS Data policy Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ****** 4. Is the manuscript presented in an intelligible fashion and written in standard English??> Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ****** Reviewer #1: Greetings Dear Authors 1- In this manuscript, you wrote 6 times (We)! The rule of manuscript writing is to avoid using (We). So you should delete (We) and use formal scientific words (This study or The current study or The present study). 2- In this manuscript, you wrote 5 times (Our)! The rule of manuscript writing is to avoid using (Our). So you should delete (Our) and use formal scientific words (This study or The current study or The present study). 3- In introduction: you wrote two link of your books, kindly delete them because this considered advertising. 4- In methods, you did not write the date or period of current study for example from May 2024 to December 2024.This is important scientifically. Kind regards Reviewer #2: This is a well-written and clearly structured manuscript that addresses a practical and relevant problem in microbiology laboratories in low-resource settings: the lack of functioning autoclaves for agar media preparation. The study is methodologically sound, with clear descriptions of procedures and an effective comparison between a commercial pressure cooker and a standard autoclave. The inclusion of field testing in a Sri Lankan veterinary diagnostic laboratory, along with Kirby-Bauer susceptibility testing, adds real-world value to the work. However, the novelty of the concept is modest, as pressure cooker sterilization has been reported previously. The main contribution here lies in the practical validation within a specific veterinary diagnostic context and the demonstration of feasibility in a resource-limited field setting. To strengthen the manuscript, I recommend the following: - Clearly position the work as a validation and implementation study, emphasizing what is new in your approach. - Justify the limited range of bacterial species and media tested, and note that results should not be generalized to other organisms or media without further evaluation. - Provide quantitative measurements for observed morphological differences (e.g., colony diameters) and include basic statistical comparisons for zone diameters. - Consider adding a simple cost comparison between an autoclave and the Instant Pot® to highlight economic implications. - Expand the limitations section to note the single model tested, potential nutrient degradation with prolonged heating, small production volumes, and electrical compatibility issues in different regions. Reviewer #3: This article presents a highly practical and beneficial approach, particularly for laboratories that face challenges due to the lack of access to an autoclave. The alternative method proposed could serve as a valuable solution for resource-limited settings. However, further research is recommended, as highlighted by the authors, particularly regarding the use of different brands of pressure cookers in media preparation. Brands that are readily available and suitable for local contexts should be evaluated. In addition, the impact of electrical current capacity on the effectiveness and consistency of results should be examined, as it may influence performance. Lastly, the manuscript formatting should be reviewed to ensure it aligns with the journal’s submission guidelines. ****** what does this mean? ). If published, this will include your full peer review and any attached files. 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Revision 1
30 Sep 2025 Author Response Reviewer #1: 1- In this manuscript, you wrote 6 times (We)! The rule of manuscript writing is to avoid using (We). So you should delete (We) and use formal scientific words (This study or The current study or The present study). • Thank you for identifying this overly casual style, "we" has been removed throughout. 2- In this manuscript, you wrote 5 times (Our)! The rule of manuscript writing is to avoid using (Our). So you should delete (Our) and use formal scientific words (This study or The current study or The present study). • Thank you for identifying this overly casual style, "our" has been removed throughout. 3- In introduction: you wrote two link of your books, kindly delete them because this considered advertising. • Respectfully we disagree with this point. We believe that referencing these manuals provides important context to our project overall. Furthermore, as both documents are 100% freely available open resources which are deposited into our University's library repository, there is no commercial product to advertise. 4- In methods, you did not write the date or period of current study for example from May 2024 to December 2024.This is important scientifically. • We have added a second date to the manuscript. We started this investigation in December 2024 as stated, and then the final work in Sri Lanka was conducted in March, 2025. Reviewer #2: This is a well-written and clearly structured manuscript that addresses a practical and relevant problem in microbiology laboratories in low-resource settings: the lack of functioning autoclaves for agar media preparation. The study is methodologically sound, with clear descriptions of procedures and an effective comparison between a commercial pressure cooker and a standard autoclave. The inclusion of field testing in a Sri Lankan veterinary diagnostic laboratory, along with Kirby-Bauer susceptibility testing, adds real-world value to the work. However, the novelty of the concept is modest, as pressure cooker sterilization has been reported previously. The main contribution here lies in the practical validation within a specific veterinary diagnostic context and the demonstration of feasibility in a resource-limited field setting. To strengthen the manuscript, I recommend the following: - Clearly position the work as a validation and implementation study, emphasizing what is new in your approach. • Thank you for this comment, . We have added additional context to highlight the novelty of using a pressure cooker to prepare media for a diagnostic laboratory (see lines 62-76). We have also added reference to this being a validation study (see line 103) - Justify the limited range of bacterial species and media tested, and note that results should not be generalized to other organisms or media without further evaluation. • Regarding the limited range of bacterial species tested, we agree that this is certainly a limitation. However these organisms do represent the most commonly encountered organisms by veterinary diagnostic laboratories, and the strains which we chose to test were those with zone diameter reference ranges published by the CLSI. We do have some justification on lines 121-126 as to why these species were chosen for our investigation. • Regarding the applicability of our findings, this is indeed a very important point. We were careful to point out where our results are not generalizable in terms of both other organisms and media on lines 284-292. - Provide quantitative measurements for observed morphological differences (e.g., colony diameters) and include basic statistical comparisons for zone diameters. • Thank you for this comment. Both of suggestions are things which had been considered before submitting our initial draft. o With respect to colony size, it is grossly apparent on figures 1 and 2 that there is variation in colony size even one the same media; those colonies growing in the most distant parts of the streak tend to be bigger than those growing close to the densest growth - this is a normal and expected phenotype for a bacterial culture. Consequently, the outcomes of statistical comparisons of colony measurements between media would be highly dependent on which colonies we chose to measure. For this reason we instead chose to use sectioned petri-dishes to directly compare conventionally prepared and pressure cooked media on the same plate; this allows us to see colonies which were grown under exactly the same conditions and for the same length of time side by side without the potentially distorting effects of photography (taking the picture from exactly the same distance, same lighting, holding the plate at the same angle to ensure that lighting is uniform etc.). We believe that the qualitative perspective the reader gains from viewing these images provides a more accurate representation of the differences/similarities observed than would a statistical comparison. o With respect to zone diameters, the Clinical and Laboratory Standards Institute (CLSI) guidelines for the ATCC strain reference ranges which we used are intended to be used as a binary comparison; a control organism is classified as either within or outside the reference interval. Furthermore, it is recognized that a degree of variability between tests of the same organism (no more than 3mm) is expected when evaluating a new lot of media. We have included the reference ranges in table 1, the most important data from this experiment was that our results with both the pressure cooker and autoclaved media fell within the reference ranges and would therefore be considered up to the standards of a diagnostic microbiology laboratory. - Consider adding a simple cost comparison between an autoclave and the Instant Pot® to highlight economic implications. • We have added a notation on lines 81-82 providing a general sense as to the costs of a laboratory autoclave. Because the costs vary widely based on the size of the instrument, manufacturer, distributor and even the country where it is purchased (I understand that import taxes in Sri Lanka can make the same model more expensive to purchase there than it would be in Canada), we have kept this statement general and high level. - Expand the limitations section to note the single model tested, potential nutrient degradation with prolonged heating, small production volumes, and electrical compatibility issues in different regions. • These are also very important points. We have added some language to the manuscript introduction, including reference to previously published paper, which hopefully clarifies why we chose to use this particular model of pressure cooker. • With respect to nutrient degradation, assessing for the chemical changes that occur during pressure cooking vs. autoclaving was beyond the scope of this study. Rather, we acknowledge that these changes occur in our justification for conducting our experiments as we aimed to determine if the media produced would be fit for purpose. • The small production volumes and electrical compatibility issues are both things which we mentioned in the text - whether either of these would be a barrier to using this pressure cooker is very context dependent. Wording was added to highlight the step-down voltage transformers do potentially add an additional cost. Reviewer #3: This article presents a highly practical and beneficial approach, particularly for laboratories that face challenges due to the lack of access to an autoclave. The alternative method proposed could serve as a valuable solution for resource-limited settings. However, further research is recommended, as highlighted by the authors, particularly regarding the use of different brands of pressure cookers in media preparation. Brands that are readily available and suitable for local contexts should be evaluated. In addition, the impact of electrical current capacity on the effectiveness and consistency of results should be examined, as it may influence performance. Lastly, the manuscript formatting should be reviewed to ensure it aligns with the journal’s submission guidelines. • Thank you for these comments, we completely agree that trialing adding different brands of pressure cookers would be an obvious next step. We have added some additional information in the introduction to explain that this particular model was chosen because a previous study demonstrated that it was able to sterilize the Geobacillus spore biological indicator, while other models were not. In the future, we are interested in evaluating appliances which are locally available in Sri Lanka. • With respect to the 230V vs. 110V, this should not have had an impact on the performance of the pressure cooker described here. We were able to source a step-down transformer, and so we were able to use 110V on the ground in Sri Lanka. • We have double checked our formatting and identified some errors in the formatting of our figure legends and references which have now been corrected. Attachments Attachment Submitted filename: R1 Rebuttal letter.docx https://doi.org/10.1371/journal.pone.0336988.r002
4 Nov 2025 Decision Letter - Tsegaye Alemayehu, Editor Evaluation of a commercial pressure cooker for the preparation of agar media for a diagnostic microbiology laboratory PONE-D-25-28065R1 Dear Dr. Rubin, We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements. Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication. An invoice will be generated when your article is formally accepted. Please note, if your institution has a publishing partnership with PLOS and your article meets the relevant criteria, all or part of your publication costs will be covered. Please make sure your user information is up-to-date by logging into Editorial Manager at Editorial Manager® and clicking the ‘Update My Information' link at the top of the page. For questions related to billing, please contact billing support . If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org. Kind regards, Tsegaye Alemayehu, Msc Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: Reviewer's Responses to Questions Comments to the Author Reviewer #1: All comments have been addressed Reviewer #3: All comments have been addressed ******** 2. Is the manuscript technically sound, and do the data support the conclusions??> Reviewer #1: Yes Reviewer #3: Yes ****** 3. Has the statistical analysis been performed appropriately and rigorously? -->?> Reviewer #1: Yes Reviewer #3: I Don't Know ****** 4. Have the authors made all data underlying the findings in their manuscript fully available??> The PLOS Data policy Reviewer #1: Yes Reviewer #3: Yes ****** 5. Is the manuscript presented in an intelligible fashion and written in standard English??> Reviewer #1: Yes Reviewer #3: Yes ****** Reviewer #1: Greetings, Good work. The revisions have significantly improved the manuscript. It is now scientifically stronger and suitable for publication. Kind regards, Reviewer #3: (No Response) ****** what does this mean? ). If published, this will include your full peer review and any attached files. If you choose “no”, your identity will remain anonymous but your review may still be made public. Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy Reviewer #1: No Reviewer #3: No ******** https://doi.org/10.1371/journal.pone.0336988.r003
Formally Accepted
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