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Development and validation of a multiplex polymerase chain reaction assay for detection of the five families of plasmid-encoded colistin resistance.

Authors
  • Jousset, Agnès B1
  • Bernabeu, Sandrine1
  • Bonnin, Remy A2
  • Creton, Elodie3
  • Cotellon, Garance3
  • Sauvadet, Aimie3
  • Naas, Thierry1
  • Dortet, Laurent4
  • 1 Department of Bacteriology-Hygiene, Hôpital de Bicêtre, Assistance Publique - Hôpitaux de Paris, Le Kremlin-Bicêtre, France; Associated French National Reference Centre for Antibiotic Resistance, Le Kremlin-Bicêtre, France; Research Unit EA 7361 'Structure, Dynamic, Function and Expression of Broad-spectrum β-lactamases', Faculty of Medicine, University Paris-Sud, Le Kremlin-Bicêtre, France. , (France)
  • 2 Associated French National Reference Centre for Antibiotic Resistance, Le Kremlin-Bicêtre, France; Research Unit EA 7361 'Structure, Dynamic, Function and Expression of Broad-spectrum β-lactamases', Faculty of Medicine, University Paris-Sud, Le Kremlin-Bicêtre, France. , (France)
  • 3 Associated French National Reference Centre for Antibiotic Resistance, Le Kremlin-Bicêtre, France. , (France)
  • 4 Department of Bacteriology-Hygiene, Hôpital de Bicêtre, Assistance Publique - Hôpitaux de Paris, Le Kremlin-Bicêtre, France; Associated French National Reference Centre for Antibiotic Resistance, Le Kremlin-Bicêtre, France; Research Unit EA 7361 'Structure, Dynamic, Function and Expression of Broad-spectrum β-lactamases', Faculty of Medicine, University Paris-Sud, Le Kremlin-Bicêtre, France. Electronic address: [email protected] , (France)
Type
Published Article
Journal
International journal of antimicrobial agents
Publication Date
Mar 01, 2019
Volume
53
Issue
3
Pages
302–309
Identifiers
DOI: 10.1016/j.ijantimicag.2018.10.022
PMID: 30395987
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Plasmid-mediated colistin resistance is increasingly described worldwide in Enterobacteriaceae from animal and human isolates. Diffusion of these resistance traits among carbapenem-resistant enterobacterial isolates is of particular concern as colistin has become the last resort antibiotic for treating human infections with these organisms. Therefore, being able to monitor the presence of these transferable colistin resistance genes (mcr-1 to mcr-5-variants) is crucial. This paper describes the development of a multiplex polymerase chain reaction (PCR) protocol for detection of all currently known transferable colistin resistance genes in Enterobacteriaceae. Five primer pairs were designed to amplify mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5 gene products in a multiplex PCR. This assay was validated retrospectively on colonies of 50 Escherichia coli, 44 Klebsiella pneumoniae and 12 Salmonella enterica isolates of animal and human origin, all well characterized, and validated prospectively on 450 carbapenem-resistant enterobacterial isolates received by the French National Reference Centre. In addition, 82 Aeromonas spp. and 10 Shewanella spp. known to be the progenitors of mcr-3 and mcr-4 alleles, respectively, were screened. Mcr-multiplex PCR assay displayed 100% specificity, sensitivity, negative predictive value and positive predictive value. The assay was able to detect all variants of the different mcr alleles, and was able to detect chromosomally encoded mcr-4-like variants present in two Shewanella bicestrii JAB-1 and Shewanella woodyi S539. In conclusion, a rapid and robust multiplex PCR assay able to detect all known mcr gene families described in Enterobacteriaceae was developed and validated. This type of test is critical for the epidemiological surveillance of plasmid-encoded resistance, especially in carbapenem-resistant bacteria. Copyright © 2019 Elsevier Ltd. All rights reserved.

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