Peer Review History
Original SubmissionMay 20, 2020 |
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PONE-D-20-14041 Discovery of beta-lactamase CMY-10 inhibitors for combination therapy against multi-drug resistant Enterobacteriaceae PLOS ONE Dear Dr. Syed Sikander Azam, Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process. Please submit your revised manuscript by Oct 26 2020 11:59PM. If you will need more time than this to complete your revisions, please reply to this message or contact the journal office at 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. Please include the following items when submitting your revised manuscript:
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Please know it is PLOS ONE policy for corresponding authors to declare, on behalf of all authors, all potential competing interests for the purposes of transparency. PLOS defines a competing interest as anything that interferes with, or could reasonably be perceived as interfering with, the full and objective presentation, peer review, editorial decision-making, or publication of research or non-research articles submitted to one of the journals. Competing interests can be financial or non-financial, professional, or personal. Competing interests can arise in relationship to an organization or another person. Please follow this link to our website for more details on competing interests: http://journals.plos.org/plosone/s/competing-interests 3. Please include a copy of Table 6 which you refer to in your text on page 25 [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? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Partly Reviewer #3: Partly ********** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: N/A ********** 3. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: No Reviewer #2: Yes Reviewer #3: Yes ********** 4. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ********** 5. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: This is an interesting paper highlighting an innovative approach at antimicrobial resistance of Enterobacteriaceae spp. The paper is well written and detailed. However, I have some questions about article: Authors should explain a potential action mechanism of compound 11. Can author clarify the correlation between lab tests and potential clinical applications? Reviewer #2: In the present study, the Authors describe an experimental model to discover beta-lactamase inhibitors, focusing on the characterization of the antimicrobial properties of the identified compounds. The work is very interesting but in my opinion some aspects need to be improved to make the study more clear and comprehensible to the reader. Introduction In the nomenclature of bacterial species of Enterobacter it is sufficient to write the Genus in full for the first time, then the dotted Genus can be used (for example E. agglomerans); In the last paragraph of the introduction, write in full "site identification by ligand competitive saturation" and put the acronym SILCS in brackets, as it is mentioned for the first time in this section. Results When a bacterial species is named for the first time, the name must be written in full (Escherichia coli); To make the comprehension clearer for a non-expert reader, Authors should write in full and clarify the meaning of the acronyms "VS", "MC", "LGFE" and "MLRF"; Experimental analysis It would be clearer if the Authors already specified in the section Results that the Beta-lactamase activity assay was performed spectrophotometrically using a chromogenic substrate such as Nitrocefin; The legend of Figure 4 should be described more clearly, it is not clear; Add the sentence "in vitro" in the title "Susceptibility testing"; In vitro susceptibility tests were performed on clinical isolates of Enterobacter spp. the authors should have used ATCC reference strains of the same Genus. This aspect needs to be clarified, the sensitivity towards the molecule could be different between bacterial strains of different genera; The first time the acronym MIC is used, the meaning must be written in full; It would be interesting and would been strengthened the data if the in vitro susceptibility tests were also carried out in broth dilution in order to verify a bacteriostatic or bactericidal effect of the compounds; In the SEM analysis paragraph the authors should clarify the observations, the paragraph is not clear, should be rewritten. Furthermore, the resolution of Figure 6 is very poor, it is not possible to appreciate the “extensive structural damage” reported. In the legend of Figure 6, the nomenclature of the bacterial species must be corrected, the species must be always written in lower case; Also in the paragraph "Activity of compounds similar to compound 11" the authors should clarify that the reference strains used do not belong to the same genus as the clinical isolates, and the test should be carried out with an Enterobacter reference strain. In my opinion is wrong to use reference bacterial strains belonging to different genera, indeed the behavior of the compounds selected between ATCC strains and clinical isolates is not comparable (see compound 20). From the main text it would seem that it has not been verified whether the analyzed bacterial strains are β-lactamase CMY-10 producers. This should be clearly demonstrated for example with a molecular assay; The resolution of Figures 4, 5, 6 and 7 is too poor, these are not clear. In Figure 5 the written are not legible; In my opinion, to make the tables (Table 2; S4; S6 and S7) more streamlined, the Authors could directly report the average value without reporting the values of the individual batches, and Figure 8 should be moved as the first figure, to make the experimental design clearer. Discussion In the discussion paragraph the Authors indicate "disc diffusion assay", but in the Results section the Authors indicate "epsilometric test". However, the diameters of the inhibition zones are commented in the section Results. What kind of test was done? This aspect is confusing and should be clarified; In my opinion, the results of the agar diffusion tests should be commented in the Results section, making the discussion more streamlined. Material and Methods The bacterial suspension must be indicated with “CFU/ml” and not “CFU/spot”. Replace the sentence; E-test strips containing the tested compounds were produced by the authors? This aspect should be clarified and the specific MIC values obtained should be reported in the Results; In the Synergistic assays the molecules were only tested in a 1: 1 ratio? Different ratio should also be checked. Table S3, which reports a partial antibiogram of clinical strains, should be completed and improved, so it is not clear. Reviewer #3: The article “Discovery of beta-lactamase CMY-10 inhibitors for combination therapy against multidrug resistant Enterobacteriaceae” by Parvaiz and coworkers deals with the identification of a novel β-lactamase inhibitors. The study is of interest because is crucial to support investigation to counter the antimicrobial resistance development in pathogens. However, the study is overall preliminary and lacks some crucial experimental work to drive proper conclusion. The title mentions beta-lactamase CMY-10 but the paper lacks an experimental demonstration that substance 11 specifically inhibits this enzyme. How do the authors explain this? ABSTRACT The abstract needs to be rewritten to reflect the aim, methods, results, and conclusions. For example, the sentence “Of these, one compound shows promising activity in β-lactamase activity assay, susceptibility testing against ATCC strains and MDR clinical isolates, with synergistic assay indicating its potential as a β-lactam enhancer and β-lactamase inhibitor” is not clear: what are the strains analyzed? what do the authors mean by “susceptibility testing”? Please rephrase. P. 8, Line 14: Please change “11 compounds” with “Eleven compounds”. P. 8, Line 18-20: “Structural similarity search against the active compound yielded 28 more compounds, the majority of which also showed β-lactamase inhibition potential and antibacterial activity”. This sentence is not clear, please rephrase. AUTHOR SUMMARY P. 9, Line 10-12: , please rephrase the sentence “Compounds screened as potential inhibitors of CMY-10 in the current study have the potential to be used in combination therapy as non-β-lactam-based β-lactamase inhibitors against MDR clinical isolates that have been found resistant against last-line antibiotics”, because the same sentence is already written in the abstract. Some sentences are repetitions of what previously said. INTRODUCTION Please, explain briefly the classification of �-lactamases and the reason for the choice of identify new inhibitors only against class C �-lactamase. RESULTS P.12: IMP should be spelled out P.13: The names of the bacterial species in the all text must be written in italics (E. coli) P.14, line 20: The experimental strategy used in β-lactamase activity assay is only briefly explained in the text. Clarification on this point may help. Which strains were used in this assay? P.15, line 1: what is the Control? P.15, line 2: The order of Supplementary Table is wrong…. Table S2 appears first in the text while Table S1 is at the end of the paper in the materials and methods section P.15, line 8-10: “Epsilometer test (E-test) was performed to quantify antimicrobial susceptibility of clinical isolates against advanced generation of macrolide and third and fourth generation antibiotics”: Please indicate antibiotics or refer to the supplementary table P.15, line 12: Table S3 is not clear and incomplete, the header is wrong, some of the data refer to Epsiolmeter test results (the bottom), the other to the diffusion test…. Please explain. Why are imipenem and meropenem antibiotics not also tested? P.16, line 1-2: Why do the authors test the substances against ATCC reference strains of S. aureus and E. coli? why don't they test the substances against a reference ATCC strain of Enterobacter? Please, perform these experiments P16, line 3: “three multi-drug resistant clinical isolates”: only 3 isolates of Enterobacter spp were tested. Please, increase the number of clinical isolates tested. P.16, line 6: Table 2: indicate the method used in the antibacterial activity assays. What are the 3 batches? three replicated experiments? if yes, please report only the average with DS in the table (this also applies to the other tables in supplementary). What is the MIC of compound 11? It is preferable to determine MIC of this compound by using the broth microdilution method. Please, perform these experiments. P.17, line 1: the synergy assay should be performed using the microbroth checkerboard method. The authors should perform this experiment for at least compound 11 and discuss the results. P.17, line 2: Why have you chosen this antimicrobial (cefixime)? Please explain. P.17, line 12: Fig. 6: the resolution of the figure is too low and this does not allow to verify the data. What is the control of experiment? the untreated strains? the images of the strains treated only with cefixime are missing. P.18, line 3: “Results obtained from the β- lactamase assay suggest that”…. Please indicate the total number of compounds similar to compound 11 found (28) P.18, line 4: “having structural similarity”: is the chemical structure of the compounds shown in Table S1? if yes, please refer to the above table. DISCUSSION The Discussion section is a bit long and it would also benefit from some condensation. MATERIAL AND METHODS P.24, line 20: In my opinion, the sentence “The complete protocol used in this study is shown in Fig 8“ should be moved to the beginning of the results, in this way the experimental procedure is immediately clear to the reader of the manuscript. P.29, line 4: The method described for the β-lactamase activity assay is not clear. The authors should describe in detail the method used. Which sample is used in the assay? bacteria? which bacterial strains? what are the conditions used to grow bacteria? P.29, line 15: describe punctually the protocol P.29, line 16: did the authors characterize (at the genomic level) the clinical strains of Enterobacter used? do these strains contain the gene that encodes for CMY-10? why did the authors not test Enterobacter ATCC reference strains? Please, perform these experiments. P.29, line 22: Change the sentence: “The test was run for 24 hrs at 37 °C” with the sentence: “The plates were incubated for 24 hrs at 37 °C” P.31, line 4: Table S1?????? The order of Supplementary Table is wrong. ********** 6. PLOS authors have the option to publish the peer review history of their article (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 #2: No Reviewer #3: No [NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.] While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. 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Revision 1 |
Discovery of beta-lactamase CMY-10 inhibitors for combination therapy against multi-drug resistant Enterobacteriaceae PONE-D-20-14041R1 Dear Dr. Syed Sikander Azam, 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 for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org. 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, Massimiliano Galdiero, M.D., Ph.D. Academic Editor PLOS ONE Additional Editor Comments (optional): Reviewers' comments: Reviewer's Responses to Questions Comments to the Author 1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the “Comments to the Author” section, enter your conflict of interest statement in the “Confidential to Editor” section, and submit your "Accept" recommendation. Reviewer #1: All comments have been addressed Reviewer #2: 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? The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ********** 3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: N/A ********** 4. Have the authors made all data underlying the findings in their manuscript fully available? The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ********** 5. Is the manuscript presented in an intelligible fashion and written in standard English? PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: Yes Reviewer #2: Yes Reviewer #3: Yes ********** 6. Review Comments to the Author Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters) Reviewer #1: THIS PAPER IS CLEARLY REVISIONED FOLLOWING REVIEWER' SUGGESTIONS. I BELIEVE THAT AFTER THIS REVISION THE PAPER IS SUITABLE FOR PUBBLICATION. Reviewer #2: Authors should pay attention to the nomenclature of bacteria throughout the text including the tables (see Table 2). The bacterial species must always be written in lowercase italics. Furthermore, the units of measurement reported throughout the text must be checked (see Table 4 and related text). Finally, in the Molecular assay, as in the electrophoresis there is no control it would be useful to insert in the text a reference on the high molecular weight plasmid. Reviewer #3: (No Response) ********** 7. PLOS authors have the option to publish the peer review history of their article (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 #2: No Reviewer #3: No |
Formally Accepted |
PONE-D-20-14041R1 Discovery of beta-lactamase CMY-10 inhibitors for combination therapy against multi-drug resistant Enterobacteriaceae Dear Dr. Azam: I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. 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. If we can help with anything else, please email us at plosone@plos.org. Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staff on behalf of Prof. Massimiliano Galdiero Academic Editor PLOS ONE |
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