Limitations in predicting reduced susceptibility to third generation cephalosporins in Escherichia coli based on whole genome sequence data

Anna Heydecke; Hong Yin; Eva Tano; Susanne Sütterlin

doi:10.1371/journal.pone.0295233

Peer Review History

Original SubmissionJanuary 23, 2023
15 Sep 2023 Decision Letter - Mohamed O Ahmed, Editor PONE-D-23-01994Limitations in Predicting Reduced Susceptibility to Third Generation Cephalosporins in Escherichia coli Based on Whole Genome Sequence DataPLOS ONE Dear Dr. Sütterlin, 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 30 2023 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. 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Any changes to the reference list should be mentioned in the rebuttal letter that accompanies your revised manuscript. If you need to cite a retracted article, indicate the article’s retracted status in the References list and also include a citation and full reference for the retraction notice. [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: Yes ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: N/A Reviewer #2: Yes ****** 3. 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Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here. Reviewer #1: No Reviewer #2: 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: The manuscript is technically sound and the data backs up the conclusion. However, there is no accession number for the data deposit. The manuscript is generally presented in a standard fashion. However, the writing grammar needs to be touched. Authors explored the limitations in using whole genome sequence data to predict reduced susceptibility to third generation cephalosporins in Escherichia coli. It is of interest and could add benefits to the field. The manuscript is straightforward and the results obtained support the discussion and conclusion. Results are presented and discussed clearly. However, the writing should be checked throughout. Some concerns are raised for manuscript revision. 1. Line 65, I think the use of conventional PCR is more common. Many labs do not have a realtime PCR machine. 2. Line 77, please correct “Escherichia coli” to “E. coli. Please also Please also check throughout. 3. Line 102, a reference for the ESBL screening is needed. Why cepodoxime and cefadroxil were used? For standard ESBL screening, cefotaxime and ceftazidime are mostly used as they are indicators for most ESBLs. 4. Line 207-209, Is there any supporting evidence for “This might be due to low expression……..(=3))”? May be due to mutations in the genes producing truncate proteins? Should “n” be equal to 4? 5. Line 212-213, please mention how many days the isolates were left at room temperature or how many passages to support the loss of bla genes on mobile genetic elements. 6. I agree with the note in Line 303-311. 7. Table 2, In the 2nd row of biology related, the potential explanation is not clear. Is it possible that the genes carried mutations? 8. There are misspellings, typo and grammatical errors throughout. Actually, I am not a native English speaker but I can tell that it is not for smooth reading. The writing should be touched for better reading. For example, -Line 64, “betalactamases” should be corrected to “beta-lactamases”. Please also checkthroughout. - Line 77, please correct “Escherichia coli” to “E. coli and also check throughout. Etc. Reviewer #2: The maniscript of Heydecke and colleagues adress discrepancies betwwen phenotypic and genotypic results applying to resistance to 3GC in E. coli. The paper is well written, the methology sounds well and the results are of interest, even if they are more confirmatory than really new. In particular, the authors clearly explained the reasons for discrepancies, either related to biological issues or to methodologies used for WGS data analysis. General comment: -the number of isolates tested (notably those resistant to 3GC is rather limited) and limits the scope of the results (this may be added in the limitations of the study -Considering that WGS date are available, is there a link between discrepancies of phenotype and genotype and some specific clonal group? Other comments -L76: replace diagnose with detect L212: this implies that phenotypic and genotypic analysis were not done simultaneously, is it correct? L214: beta-lactam Table 1: I think the added value of this table is weak and could be omitted ****** 6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). 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Revision 1
2 Oct 2023 Author Response Response to reviewers PONE-D-23-01994 Limitations in Predicting Reduced Susceptibility to Third Generation Cephalosporins in Escherichia coli Based on Whole Genome Sequence Data PLOS ONE Dear Dr. Ahmed, Please find enclosed our revised manuscript. Replies to each of the reviewers’ questions are given below. We are very grateful for the constructive comments of the reviewers, which have helped us to improve our manuscript. Yours sincerely Susanne Sütterlin Reviewer #1: The manuscript is technically sound and the data backs up the conclusion. However, there is no accession number for the data deposit. The manuscript is generally presented in a standard fashion. However, the writing grammar needs to be touched. Authors explored the limitations in using whole genome sequence data to predict reduced susceptibility to third generation cephalosporins in Escherichia coli. It is of interest and could add benefits to the field. The manuscript is straightforward and the results obtained support the discussion and conclusion. Results are presented and discussed clearly. However, the writing should be checked throughout. Some concerns are raised for manuscript revision. 1. Line 65, I think the use of conventional PCR is more common. Many labs do not have a realtime PCR machine. In Sweden, the use of real-time PCR is standard, while conventional PCR is rarely used in diagnostics anymore. However, the article aims to be of interest to an international audience and thus it seems to be more appropriate to not specify the type of molecular method that can be used. We remove “real-time” and use only PCR. 2. Line 77, please correct “Escherichia coli” to “E. coli. Please also Please also check throughout. We corrected accordingly and checked through the manuscript. 3. Line 102, a reference for the ESBL screening is needed. Why cepodoxime and cefadroxil were used? For standard ESBL screening, cefotaxime and ceftazidime are mostly used as they are indicators for most ESBLs. We followed the recommendations from NordicAST (Nordic committee on antimicrobial susceptibility testing that represents all clinical laboratories in Scandinavia). NordicAST is represented in EUCAST, the European committee. While the use of cefpodoxime and cefadroxil has been removed from the European screening recommendations for ESBL production, the NordicAST kept them in their recommendations. (EUCAST guideline 2013: https://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Resistance_mechanisms/EUCAST_detection_of_resistance_mechanisms_v1.0_20131211.pdf EUCAST guideline 2017: https://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Resistance_mechanisms/EUCAST_detection_of_resistance_mechanisms_170711.pdf NordicAST recommendation 2021 (includes cefpodoxime): https://www.nordicast.org/d/6092?store_referer=true NordicAST breakpoint table v13.0 (includes both cefpodoxime and cefadroxil): https://s3-eu-west-1.amazonaws.com/hl-intranet/files/a00a01520e275b6dfff4b1b7340cc06668471ef8/nordicast_bp_table_se_v_13_0_locked2.xlsx Swedish clinical laboratories use cefadroxil commonly as screening disc in UTI samples, as it can also be used for treatment especially of lower UTI. When the isolate is susceptible to cefadroxil, ESBL production can be ruled out. It is a convenient and economic way of testing ESBL on standard agar plates with space for six antibiotic discs. We would like to suggest to refer to both committee’s websites and use them as references. In order to clarify this, we have rewritten the according part of the manuscript. 4. Line 207-209, Is there any supporting evidence for “This might be due to low expression……..(=3))”? May be due to mutations in the genes producing truncate proteins? Should “n” be equal to 4? Thank you for indicating our mistake, n is equal to 4, not 3, and has been corrected in the manuscript. We chose not to specify potential mechanisms leading to low expression of genes as we did not perform further analysis that can give further evidence for underlying mechanisms. However, we discuss the issue further in the discussion section. 5. Line 212-213, please mention how many days the isolates were left at room temperature or how many passages to support the loss of bla genes on mobile genetic elements. That is a good point! We added information on amount of subculturing and leaving plates at room temperature in the manuscript. 6. I agree with the note in Line 303-311. Ditto. 7. Table 2, In the 2nd row of biology related, the potential explanation is not clear. Is it possible that the genes carried mutations? The isolates were resistant to cephalosporines, however, the synergy test and AmpC-test failed to reveal ESBL-production. Two isolates were AmpC-producers (DHA-1) that were not resistant to cefoxitin, and two carried CTX-9 and CTX-14), however, the synergy test did not indicate inhibition by beta-lactamase inhibitors (probably due to resistance to beta-lactamase inhibitors). 8. There are misspellings, typo and grammatical errors throughout. Actually, I am not a native English speaker but I can tell that it is not for smooth reading. The writing should be touched for better reading. For example, -Line 64, “betalactamases” should be corrected to “beta-lactamases”. Please also checkthroughout. - Line 77, please correct “Escherichia coli” to “E. coli and also check throughout. Etc. The manuscript was sent for language revision to a professional language editor and was checked thorough. Reviewer #2: The maniscript of Heydecke and colleagues adress discrepancies betwwen phenotypic and genotypic results applying to resistance to 3GC in E. coli. The paper is well written, the methology sounds well and the results are of interest, even if they are more confirmatory than really new. In particular, the authors clearly explained the reasons for discrepancies, either related to biological issues or to methodologies used for WGS data analysis. General comment: -the number of isolates tested (notably those resistant to 3GC is rather limited) and limits the scope of the results (this may be added in the limitations of the study We added this limitation to the according section of the manuscript. -Considering that WGS date are available, is there a link between discrepancies of phenotype and genotype and some specific clonal group? We have not analyzed the clonality of the isolates with regard to susceptibility systematically. However, as the isolates derive from different studies where clonality was investigated we did assess the possibility within the revision, but could not find conclusive patterns. Other comments -L76: replace diagnose with detect Changed. L212: this implies that phenotypic and genotypic analysis were not done simultaneously, is it correct? That is correct. As the first reviewer pointed out the same issue, we completed with a clarification accordingly. L214: beta-lactam Changed. Table 1: I think the added value of this table is weak and could be omitted After consideration we agree and omit the table accordingly. Attachments Attachment Submitted filename: Response_to_reviewers.docx https://doi.org/10.1371/journal.pone.0295233.r002
20 Nov 2023 Decision Letter - Mohamed O Ahmed, Editor Limitations in predicting reduced susceptibility to third generation cephalosporins in Escherichia coli based on whole genome sequence data PONE-D-23-01994R1 Dear Dr. Sütterlin, 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, Mohamed O Ahmed, Ph.D Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0295233.r003
Formally Accepted
24 Nov 2023 Acceptance Letter - Mohamed O Ahmed, Editor PONE-D-23-01994R1 Limitations in predicting reduced susceptibility to third generation cephalosporins in Escherichia coli based on whole genome sequence data Dear Dr. Sütterlin: 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 customercare@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 Dr. Mohamed O Ahmed Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0295233.r004

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