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Proteomic assay for rapid characterisation of Staphylococcus aureus antimicrobial resistance mechanisms directly from blood cultures

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European Journal of Clinical Microbiology & Infectious Diseases Aims and scope Submit manuscript

Abstract

Purpose

Staphylococcus aureus is one of the most common pathogens causing bloodstream infection. A rapid characterisation of resistance to methicillin and, occasionally, to aminoglycosides for particular indications, is therefore crucial to quickly adapt the treatment and improve the clinical outcomes of septic patients. Among analytical technologies, targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) has emerged as a promising tool to detect resistance mechanisms in clinical samples.

Methods

A rapid proteomic method was developed to detect and quantify the most clinically relevant antimicrobial resistance effectors in S. aureus in the context of sepsis: PBP2a, PBP2c, APH(3’)-III, ANT(4’)-I, and AAC(6’)-APH(2’’), directly from positive blood cultures and in less than 70 min including a 30-min cefoxitin-induction step. The method was tested on spiked blood culture bottles inoculated with 124 S.aureus, accounting for the known genomic diversity of SCCmec types and the genetic background of the strains.

Results

This method provided 99% agreement for PBP2a (n = 98/99 strains) detection. Agreement was 100% for PBP2c (n = 5/5), APH(3’)-III (n = 16/16), and ANT(4’)-I (n = 20/20), and 94% for AAC(6')-APH(2'') (n = 16/17). Across the entire strain collection, 100% negative agreement was reported for each of the 5 resistance proteins. Additionally, relative quantification of ANT(4’)-I expression allowed to discriminate kanamycin-susceptible and -resistant strains, in all strains harbouring the ant(4’)-Ia gene.

Conclusion

The LC-MS/MS method presented herein demonstrates its ability to provide a reliable determination of S. aureus resistance mechanisms, directly from positive blood cultures and in a short turnaround time, as required in clinical laboratories.

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Data availability

MRM raw data and transition lists are available via PASSEL with the accession number PASS05841 (https://db.systemsbiology.net/sbeams/cgi/PeptideAtlas/PASS_View). The 124 S.aureus genomes are available on ENA (accession numbers in Supplementary Table 1).

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Acknowledgements

We would like to thank anyone involved in the IdBIORIV project, whether at the Institut des Sciences Analytiques, at the Infectious Agents Institute, at the French National Reference Centre for Staphylococci or at the Centre International de Recherche en Infectiologie. We thank Patricia Martins-Simoes and Sylvere Bastien for valuable technical help. We thank Sciex company for providing the technical support and the instruments used in the present work. We also thank Shanez Haouari from the Hospices Civils de Lyon (France) for her help in manuscript preparation.

Funding

This study and the IdBIORIV project were supported financially by the French country through the Agence Nationale de la Recherche and the « investissement d’avenir» program (ANR-18-RHUS-0013).

Author information

Author notes

  1. Jérôme Lemoine and François Vandenesch have equal contribution.

Authors and Affiliations

  1. Institut des Sciences Analytiques, Université de Lyon, Université Claude Bernard Lyon 1, Centre National de la Recherche Scientifique (CNRS), UMR 5280, Villeurbanne, France

    Francis Deforet, Romain Carrière, Pierre L.’Aour Dufour & Jérôme Lemoine

  2. Hospices Civils de Lyon, Institut des Agents Infectieux, Lyon, France

    Pierre L.’Aour Dufour, Roxane Prat, Chloé Desbiolles, Noémie Cottin, Alicia Reuzeau, Olivier Dauwalder, Tiphaine Cecchini & François Vandenesch

  3. Centre National de Référence des Staphylocoques, Institut des agents infectieux, Hospices Civils de Lyon, Lyon, France

    Céline Dupieux-Chabert, Anne Tristan & François Vandenesch

  4. CIRI, Centre International de Recherche en Infectiologie, Université de Lyon, Inserm, U1111, Université Claude Bernard Lyon 1, Centre National de la Recherche Scientifique (CNRS), UMR5308, École Normale Supérieure (ENS) de Lyon, Lyon, France

    Anne Tristan & François Vandenesch

Authors
  1. Francis Deforet
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  2. Romain Carrière
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  3. Pierre L.’Aour Dufour
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  4. Roxane Prat
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  5. Chloé Desbiolles
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  6. Noémie Cottin
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  7. Alicia Reuzeau
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  8. Olivier Dauwalder
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  9. Céline Dupieux-Chabert
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  10. Anne Tristan
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  11. Tiphaine Cecchini
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  12. Jérôme Lemoine
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  13. François Vandenesch
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Contributions

FD, RC, PLD, CDC, AT, TC, OD, JL and FV contributed to the study conception and design. Material preparation, data collection and analysis were performed by FD, RC, PLD, RP, CD, NC and AR with the support of TC and OD. The first draft of the manuscript was written by FD, FV and JL and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to François Vandenesch.

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Competing interest

The authors declare the following conflicts of interest: F.D, J.L and F.V have filled a patent regarding the method to improve the sensitivity of MRSA detection; R.C., J.L and F.V are co-founders of Weezion company. All other authors declare no conflict of interest.

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Deforet, F., Carrière, R., Dufour, P.L. et al. Proteomic assay for rapid characterisation of Staphylococcus aureus antimicrobial resistance mechanisms directly from blood cultures. Eur J Clin Microbiol Infect Dis (2024). https://doi.org/10.1007/s10096-024-04811-0

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