A sensitive and specific phenotypic assay for detection of metallo-β-lactamases and KPC in Klebsiella pneumoniae with the use of meropenem disks supplemented with aminophenylboronic acid, dipicolinic acid and cloxacillin

C. G. Giske*, L. Gezelius, Samuelsen, Marina Warner, A. Sundsfjord, Neil Woodford

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

158 Citations (Scopus)

Abstract

Enterobacteriaceae producing carbapenemases, such as KPC or metallo-β-lactamases (MBLs), have emerged on several continents. Phenotypic tests are urgently needed for their rapid and accurate detection. A novel carbapenemase detection test, comprising a meropenem disk, and meropenem disks supplemented with 730μg of EDTA, 1000μg of dipicolinic acid (DPA), 600μg of aminophenylboronic acid (APBA), or 750μg of cloxacillin, was evaluated against Klebsiella pneumoniae isolates with KPC (n=34), VIM (n=21), IMP (n=4) or OXA-48 (n=9) carbapenemases, and carbapenem-resistant Enterobacteriaceae with porin loss in combination with an extended-spectrum β-lactamase (ESBL) (n=9) or AmpC hyperproduction (n=5). Commercially available diagnostics tablets from Rosco containing meropenem and the same inhibitors as described above (except EDTA) were also evaluated. An increased meropenem inhibition zone was sought in the presence of each added β-lactamase inhibitor. APBA had excellent sensitivity for detecting K. pneumoniae with KPC enzymes. Isolates with combined AmpC hyperproduction and porin loss were also positive in the APBA test but, unlike KPC producers, showed cloxacillin synergy. Both DPA and EDTA had excellent sensitivity for detection of MBL-producing K. pneumoniae. However, EDTA showed poor specificity, with positive results noted for 1/9 ESBL-producing isolates, for 4/34 KPC-producing isolates, and for 4/9 OXA-48-producing isolates, whereas all of these were negative when DPA was used. The in-house test distinguished accurately between several different mechanisms mediating reduced susceptibility to carbapenems in Enterobacteriaceae. The commercial combination tablets from Rosco performed similarly to the in-house test, with the exception of one false-positive MBL result and one false-positive KPC result among the OXA-48 producers.

Original languageEnglish
Pages (from-to)552-556
Number of pages5
JournalClinical Microbiology and Infection
Volume17
Issue number4
DOIs
Publication statusPublished - Apr 2011

Bibliographical note

Funding Information:
C. G. Giske has received conference support from Calixa Therapeutics Inc. N. Woodford has received grants and conference support from major pharmaceutical companies, including all those that manufacture and supply carbapenems internationally (AstraZeneca, MSD, Johnson and Johnson/Janssen-Cilag). The other authors have no conflicts of interest to declare.

Funding Information:
We thank J. B. Patel, CDC, for providing clinical isolates of K. pneumoniae producing KPC β-lactamases. We would also like to thank A. Vatopoulos, Department of Microbiology, National School of Public Health, Athens, Greece, for providing clinical isolates of K. pneumoniae producing VIM-1 β-lactamase. Ø. Samuelsen is supported by a grant from the Northern Norway Regional Health Authority Medical Research Programme.

Keywords

  • IMP
  • KPC
  • OXA-48
  • VIM

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