Procalcitonin, C-reactive protein, neutrophil gelatinase-associated lipocalin, resistin and the APTT waveform for the early diagnosis of serious bacterial infection and prediction of outcome in critically ill children

Maryke J. Nielsen; Paul Baines; Rebecca Jennings; Sarah Siner; Ruwanthi Kolamunnage-Dona; Paul Newland; Matthew Peak; Christine Chesters; Graham Jeffers; Colin Downey; Caroline Broughton; Lynsey McColl; Jennifer Preston; Anthony McKeever; Stephane Paulus; Nigel Cunliffe; Enitan D. Carrol

doi:10.1371/journal.pone.0246027

Peer Review History

Original SubmissionFebruary 1, 2020
26 Aug 2020 Decision Letter - José Moreira, Editor PONE-D-20-03003 Procalcitonin, C-Reactive Protein, Neutrophil Gelatinase-Associated Lipocalin, Resistin and the APTT Waveform for the early diagnosis of serious bacterial infection and prediction of outcome in critically ill children. PLOS ONE Dear Dr. Nielsen, Your submission has now been peer reviewed. I agree that the manuscript would benefit from being revised according to the suggestions following and encourage you to do so. Editor Comments to the Authors: Please see the reviewer's comments. Please submit your revised manuscript by Oct 10 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. 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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 [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 ******** 2. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1: Yes Reviewer #2: Yes ****** 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: Yes Reviewer #2: 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 ****** 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 study by Nielsen et al. is a prospective study in the UK evaluating the performance of an array of biomarkers on the diagnosis of SBI upon on admission to the PICU. The models including a combination of PCT, CRP and additional biomarkers indicated an improvement (as compared to CRP alone) in classification of SBI. An array biomarkers to identify SBI in pediatrics are intensely investigated and the literature is considerable in this area. The authors do a nice job on including different biomarkers in their analysis and showing improvement in the discrimination of SBI when using them in combination. Would suggest the following for this article: 1. Can you provide further detail on the diagnosis of SBI by your group specifically the level disagreement within the cases? 2. Are these commonly collected biomarkers for all ICU patients or specific to your study? Further are these POC biomarkers or are they commonly sent out to the laboratory. The external validity may be different if these are biomarkers are not commonly collected or available within real time. 3. I’m unsure whether providing the independent variables associated with prolonged ICU stay and duration of ICU stay is adding much the manuscript. The focus is on specific biomarkers and their combination. Including this analysis looks exploratory at best. Also, it would help the reader if you provide the 95% CIs along with the corresponding ORs, otherwise difficult to assess reliability of your point estimates. 4. It would be of more interest if we start to show the impact on clinical outcomes of these biomarkers and their combination rather than just their performance. Previous studies have evaluated biomarker performance for pediatrics and SBI with similar AUC findings. Reviewer #2: A prospective observation study was conducted to investigate the accuracy of procalcitonin (PCT), neutrophil gelatinase-associated lipocalin (NGAL), resistin, activated partial thromboplastin time (aPTT) waveform and C-reactive protein (CRP) for their discriminative ability in diagnosing serious bacterial infections (SBI) in children. The combination of PCT, resistin, plasma NGAL and CRP resulted in the biggest net reclassification improvement compared to CRP alone. Minor revisions: 1- Line 166: Identify the specific statistical univariate test applied to identify which variable had a statistically significant association with SBI. 2- Line 167: To improve clarity begin the sentence with: “For comparing continuous variables, …” 3- Line 168: To improve clarity begin the sentence with: “For comparing categorical variables, ...” 4- Line 169: Remove “if a count was below 5.” Fisher’s exact tests are appropriate to use when the expected, rather than observed, counts are less than 5. 5- Line 179: Provide a reference for net reclassification improvement. 6- Cite the statistical software used for the analysis. 7- Table 1 and Supplemental Table 5: Note what statistical methods were used to calculate the p-values. Explain that “Difference, (95% CI for location), p-value” pertains to continuous factors only. Possibly only the p-value is necessary. 8- Table 1: Small p-values are expressed as p< 0.001. 9- Line 170: Clarify the following statement. Possibly a more descriptive term can replace “association.” “The primary analysis examined the association between diagnosis of SBI and abnormal biomarkers.” 10- Line 219: Probabilities range from 0 to 1. It’s unclear what a probability of 12.89 represents. 11- P-values never equal zero; express small p-values as < 0.001. 12- Lines 255, 262, etc: Indicate the statistical methods used to identify independent variables. 13- Be sure to fully describe all statistical methods in the statistical methods section of the manuscript. 14- Table 4: Indicate if the models are univariate or multivariate. 15- Line 250: Provide a repeated measures analysis to support the conclusion that only PCT and CRP demonstrated clearly distinct profiles. 16- Supplementary Tables: Indicate the statistical method(s) used to obtain the best fitting models. 17- Add AUC values to the ROC curves. 18- State and justify the study’s target sample size with a pre-study statistical power calculation. The power calculation should include: sample size, alpha level (indicating one or two-sided), and statistical testing method. ****** 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 [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. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step. https://doi.org/10.1371/journal.pone.0246027.r001
Revision 1
2 Dec 2020 Author Response Dear Editor Thank you for the reviewers’ comments on our Manuscript entitled “Procalcitonin, C-Reactive Protein, Neutrophil Gelatinase-Associated Lipocalin, Resistin and the APTT Waveform for the early diagnosis of serious bacterial infection and prediction of outcome in critically ill children”. We have considered the reviewers’ comments and have made the following amendments to the manuscript, figures and supporting information. • Funding Statement: We have amended the funding statement declaring the commercial affiliation of author LMcC and the role of her employer, Select Statistics, in the analysis of the study (Manuscript p2). We can confirm that Select Statistics provided support in the form of salaries for authors [LMcC], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.” • Competing Interests Statement: This has been updated on page 21 of the manuscript about author LMcC. “This does not alter our adherence to PLOS ONE policies on sharing data and materials”. Reviewer 1: 1. Can you provide further detail on the diagnosis of SBI by your group specifically the level disagreement within the cases? Further details have been provided in relation to classification of patients with and without SBI by the multi-disciplinary PICU infection team meetings (Manuscript p12). 2. Are these commonly collected biomarkers for all ICU patients or specific to your study? Further are these POC biomarkers or are they commonly sent out to the laboratory. The external validity may be different if these are biomarkers are not commonly collected or available within real time. We can confirm that CRP was measured in the Alder Hey Hospital’s Clinical Chemistry Laboratory with results available to clinicians typically within hours of the sample being taken. As per standard UK clinical practice this was the only biomarker available to clinicians. All other biomarkers were measured in the laboratory after completion of clinical data collection and follow up. This has been clarified on page 8 and 9 of the manuscript. 3. I’m unsure whether providing the independent variables associated with prolonged ICU stay and duration of ICU stay is adding much the manuscript. The focus is on specific biomarkers and their combination. Including this analysis looks exploratory at best. Also, it would help the reader if you provide the 95% CIs along with the corresponding ORs, otherwise difficult to assess reliability of your point estimates. We have emphasized in the presentation of these results aimed to investigate the roles of the studied biomarkers in comparison to clinical variables as predictors of secondary outcome measures duration of ICU stay, prolonged ICU stay and duration of invasive positive pressure ventilation. 95% CI and significant levels have been added to all ORs specified in the manuscript. 4. It would be of more interest if we start to show the impact on clinical outcomes of these biomarkers and their combination rather than just their performance. Previous studies have evaluated biomarker performance for pediatrics and SBI with similar AUC findings. We note with thanks this comment strongly agree that it important that research can demonstrate the impact on clinical outcomes of these biomarkers. This project was a diagnostic accuracy study in which only CRP, a routinely used biomarker in the U.K., was available to clinicians and was therefore unable to demonstrate impact upon patient outcomes. This study has contributed to informing a multi-centre randomised control trial aiming to determine is the addition of procalcitonin testing to current best practice can safely allow a reduction in duration of antibiotic therapy in hospitalised children with suspected or confirmed bacterial infection compared to current best practice alone. https://www.isrctn.com/ISRCTN11369832?q=&filters=conditionCategory:Infections%20and%20Infestations,ageRange:Child&sort=&offset=7&totalResults=217&page=1&pageSize=10&searchType=basic-search. We have addressed this issue in the discussion (p19). Reviewer 2: 1. Line 166: Identify the specific statistical univariate test applied to identify which variable had a statistically significant association with SBI – We have clarified the statistical tests used for this analysis (p9). 2. Line 167: To improve clarity begin the sentence with: “For comparing continuous variables, …”. We have rephrased the sentences as per the reviewer’s advice. 3. Line 168: To improve clarity begin the sentence with: “For comparing categorical variables, ...” We have rephrased the sentences as per the reviewer’s advice. 4. Line 169: Remove “if a count was below 5.” Fisher’s exact tests are appropriate to use when the expected, rather than observed, counts are less than 5. We have removed “if a count was below 5” as per the reviewer’s recommendation. 5. Line 179: Provide a reference for net reclassification improvement. Reference has been added (p10) 6. Cite the statistical software used for the analysis. Detailed have been added to statistical methods (p10) 7. Table 1 and Supplemental Table 5: Note what statistical methods were used to calculate the p-values. Explain that “Difference, (95% CI for location), p-value” pertains to continuous factors only. Possibly only the p-value is necessary. We concur with the reviewer’s comments and have removed the differences previously stated in Table 1. We have retained the difference results in Supplementary Table 5 since all variables are continuous variables. Statistical methods used to calculate these values have been added to the supporting statistical methods. 8. Table 1: Small p-values are expressed as p< 0.001.- p values have been amended as recommended. 9. Line 170: Clarify the following statement. Possibly a more descriptive term can replace “association.” “The primary analysis examined the association between diagnosis of SBI and abnormal biomarkers.” - This sentence has been replaced by the following “The primary analysis compared the performance of the different study biomarkers and combinations of biomarkers for diagnosis of SBI”. 10. Line 219: Probabilities range from 0 to 1. It’s unclear what a probability of 12.89 represent - We have added in the previously missing % value to the pre-test probability and thank the reviewers for noting this omission. 11. P-values never equal zero; express small p-values as < 0.001. – This has been amended throughout the manuscript. 12. Lines 255, 262, etc: Indicate the statistical methods used to identify independent variables – This is included in supporting statistical methods. 13. Be sure to fully describe all statistical methods in the statistical methods section of the manuscript – We have reviewed statistical methods in both the manuscript and supporting material and believe all methods are explained. 14. Table 4: Indicate if the models are univariate or multivariate. – This has been added to the title of this table. 15. Line 250: Provide a repeated measures analysis to support the conclusion that only PCT and CRP demonstrated clearly distinct profiles. – Details of this analysis and results are now included in the statistical methods (p10) and results section (p17). 16. Supplementary Tables: Indicate the statistical method(s) used to obtain the best fitting models. - This is described in the supplementary statistical methods. 17. Add AUC values to the ROC curves. - AUC values have been added to all ROC curves. 18. State and justify the study’s target sample size with a pre-study statistical power calculation. The power calculation should include: sample size, alpha level (indicating one or two-sided), and statistical testing method.- We have included details of the apriori sample size calculation in the methods section of the manuscript and referred to this in the discussion. We thank the reviewers for their helpful comments, which have helped to improve the manuscript. We hope that all the comments have been satisfactorily addressed and that the manuscript will be deemed suitable for publication. Yours sincerely Dr Maryke Nielsen Corresponding Author Attachments Attachment Submitted filename: Response to Reviewers v 3.docx https://doi.org/10.1371/journal.pone.0246027.r002
13 Jan 2021 Decision Letter - Aleksandar R. Zivkovic, Editor Procalcitonin, C-Reactive Protein, Neutrophil Gelatinase-Associated Lipocalin, Resistin and the APTT Waveform for the early diagnosis of serious bacterial infection and prediction of outcome in critically ill children. PONE-D-20-03003R1 Dear Dr. Nielsen, 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, Aleksandar R. Zivkovic Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0246027.r003
Formally Accepted
25 Jan 2021 Acceptance Letter - Aleksandar R. Zivkovic, Editor PONE-D-20-03003R1 Procalcitonin, C-Reactive Protein, Neutrophil Gelatinase-Associated Lipocalin, Resistin and the APTT Waveform for the early diagnosis of serious bacterial infection and prediction of outcome in critically ill children. Dear Dr. Nielsen: 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 Dr. Aleksandar R. Zivkovic Academic Editor PLOS ONE https://doi.org/10.1371/journal.pone.0246027.r004

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