Reconciling the potentially irreconcilable? Genotypic and Phenotypic Amoxicillin-Clavulanate Resistance in Escherichia coli

Timothy J. Davies*, Nicole Stoesser, Anna E. Sheppard, Manal Abuoun, Philip Fowler, Jeremy Swann, T. Phuong Quan, David Griffiths, Alison Vaughan, Marcus Morgan, Hang T.T. Phan, Katie J. Jeffery, Monique Andersson, Matthew Ellington, Oskar Ekelund, Neil Woodford, Amy J. Mathers, Robert A. Bonomo, Derrick W. Crook, Tim E.A. PetoMuna F. Anjum, A. Sarah Walker

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

Resistance to amoxicillin-clavulanate, a widely used beta-lactam/ beta-lactamase inhibitor combination antibiotic, is rising globally, and yet susceptibility testing remains challenging. To test whether whole-genome sequencing (WGS) could provide a more reliable assessment of susceptibility than traditional methods, we predicted resistance from WGS for 976 Escherichia coli bloodstream infection isolates from Oxfordshire, United Kingdom, comparing against phenotypes from the BD Phoenix (calibrated against EUCAST guidelines). A total of 339/976 (35%) isolates were amoxicillin-clavulanate resistant. Predictions based solely on beta-lactamase presence/absence performed poorly (sensitivity, 23% [78/339]) but improved when genetic features associated with penicillinase hyperproduction (e.g., promoter mutations and copy number estimates) were considered (sensitivity, 82% [277/339]; P < 0.0001). Most discrepancies occurred in isolates with MICs within ±1 doubling dilution of the breakpoint. We investigated two potential causes: the phenotypic reference and the binary resistant/susceptible classification. We performed reference standard, replicated phenotyping in a random stratified subsample of 261/976 (27%) isolates using agar dilution, following both EUCAST and CLSI guidelines, which use different clavulanate concentrations. As well as disagreeing with each other, neither agar dilution phenotype aligned perfectly with genetic features. A random-effects model investigating associations between genetic features and MICs showed that some genetic features had small, variable and additive effects, resulting in variable resistance classification. Using model fixed-effects to predict MICs for the non-agar dilution isolates, predicted MICs were in essential agreement (±1 doubling dilution) with observed (BD Phoenix) MICs for 691/715 (97%) isolates. This suggests amoxicillin-clavulanate resistance in E. coli is quantitative, rather than qualitative,

Original languageEnglish
Article numbere02026-19
JournalAntimicrobial Agents and Chemotherapy
Volume64
Issue number6
DOIs
Publication statusPublished - Jun 2020

Bibliographical note

Funding Information:
This study uses data provided by patients and collected by the NHS as part of their care and support. We thank the people of Oxfordshire who contributed to the Infections in Oxfordshire Research Database. The Research Database Team included R. Alstead, C. Bunch, D. C. W. Crook, J. Davies, J. Finney, J. Gearing (community), L. O?Connor, T. E. A. Peto (PI), T. P. Quan, J. Robinson (community), B. Shine, A. S. Walker, D. Waller, and D. Wyllie. The Patient and Public Panel included G. Blower, C. Mancey, P. McLoughlin, and B. Nichols. We also thank the HPRU Steering Group (N. French, C. Marwick, J. Coia, and M. Sharland). The study was funded by the National Institute for Health Research Health Protection Research Unit (NIHR HPRU) in Healthcare Associated Infections and Antimicrobial Resistance at Oxford University in partnership with Public Health England (PHE; grant HPRU-2012-10041). D.W.C., T.E.A.P., and A.S.W. are supported by the NIHR Oxford Biomedical Research Centre. The report presents independent research funded by the National Institute for Health Research. The views expressed in this publication are those of the authors and not necessarily those of the NHS, the National Institute for Health Research, the Department of Health or Public Health England. NS is funded by a PHE/University of Oxford Clinical Lectureship. D.W.C., T.E.A.P., and A.S.W. are NIHR Senior Investigators. T.J.D., N.S., M.J.E., N.W., D.W.C., T.E.A.P., M.F.A., and A.S.W. designed the study. K.J., M.A. (OUH), M.M., and T.P.Q. obtained the automated susceptibility phenotypes from archived BD phoenix records. T.J.D. and M.A. (APHA) performed agar dilution on samples. D.G. and A.V. sequenced isolates. T.J.D., H.P., and J.S. ran resistance genotype prediction on samples. T.J.D., A.S., N.S., O.E., R.B., and A.M. interpreted the genetic results and established rules regarding the relationship with phenotype. T.J.D. and A.S.W. fitted random-effects models to the data. T.J.D., N.S., A.S., P.F., A.S.W., and M.F.A. prepared the first draft. All authors commented on the data and its interpretation, revised the content critically, and approved the final version. The authors have no competing interests to declare. However, for N.W. and M.J.E., PHE?s AMRHAI Reference Unit has received financial support for conference attendance, lectures, research projects or contracted evaluations from numerous sources, including Accelerate Diagnostics; Achaogen, Inc.; Allecra Therapeutics; Amplex; AstraZeneca UK, Ltd.; AusDiagnostics; Basilea Pharmaceutica; Becton, Dickinson Diagnostics; bioM?rieux; Bio-Rad Laboratories; The BSAC; Cepheid; Check-Points B.V.; Cubist Pharmaceuticals; Department of Health; Enigma Diagnostics; European Centre for Disease Prevention and Control; Food Standards Agency; GlaxoSmithKline Services, Ltd.; Helperby Therapeutics; Henry Stewart Talks; IHMA, Ltd.; Innovate UK; Kalidex Pharmaceuticals; Melinta Therapeutics; Merck Sharpe & Dohme Corp.; Meiji Seika Pharma Co., Ltd.; Mobidiag; Momentum Biosciences, Ltd.; Neem Biotech; NIHR; Nordic Pharma, Ltd.; Norgine Pharmaceuticals; Rempex Pharmaceuticals, Ltd.; Roche, Rokitan, Ltd.; Smith & Nephew UK, Ltd.; Shionogi & Co., Ltd.; Trius Therapeutics; VenatoRx Pharmaceuticals; Wockhardt, Ltd.; and the World Health Organization.

Funding Information:
The study was funded by the National Institute for Health Research Health Protection Research Unit (NIHR HPRU) in Healthcare Associated Infections and Antimicrobial Resistance at Oxford University in partnership with Public Health England (PHE; grant HPRU-2012-10041). D.W.C., T.E.A.P., and A.S.W. are supported by the NIHR Oxford Biomedical Research Centre. The report presents independent research funded by the National Institute for Health Research. The views expressed in this publication are those of the authors and not necessarily those of the NHS, the National Institute for Health Research, the Department of Health or Public Health England. NS is funded by a PHE/University of Oxford Clinical Lectureship. D.W.C., T.E.A.P., and A.S.W. are NIHR Senior Investigators.

Publisher Copyright:
Copyright © 2020 Davies et al.

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