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Journal of Clinical Microbiology, August 2002, p. 2755-2759, Vol. 40, No. 8
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.8.2755-2759.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Evaluation of a New Etest for Detecting Metallo-ß-Lactamases in Routine Clinical Testing

Timothy R. Walsh,1* Anne Bolmström,2 Anette Qwärnström,2 and Ana Gales3

University of Bristol, Bristol, United Kingdom,1 AB BIODISK, Solna, Sweden,2 University of Iowa, Iowa City, Iowa3

Received 17 December 2001/ Returned for modification 4 March 2002/ Accepted 7 May 2002

Several Etest (AB BIODISK, Solna, Sweden) gradient formats were developed for detection of metallo-ß-lactamases based on the reduction of imipenem (IP) or ceftazidime (TZ) MICs in the presence of EDTA or 2-mercaptopropionic acid (MPA). The Etest metallo-ß-lactamase (Etest MBL) strips consisted of a double-sided seven-dilution range of IP or TZ (4 to 256 µg/ml) and IP or TZ (1 to 64 µg/ml) overlaid with a constant concentration of EDTA or MPA. The prototype strips were evaluated on several agar media (brain heart infusion agar, Isosensitest agar, nutrient agar, and Mueller-Hinton agar for aerobes and brucella blood agar for anaerobes) with 138 challenge strains: Acinetobacter spp. (n = 9), Aeromonas spp. (n = 8), Chryseobacterium spp. (n = 28), Escherichia coli (n = 1), Klebsiella pneumoniae (n = 4), Pseudomonas aeruginosa (n = 14), Proteus mirabilis (n = 3), Serratia spp. (n = 10), Stenotrophomonas maltophilia (n = 43), Sphingobacterium spp. (n = 3), and Bacteroides fragilis group (n = 15). PCR analysis using specific primers for IMP-1, L1, CcrA, and blaB/C confirmed the presence of the metallo-ß-lactamase genes. Enzyme assays were also performed with IP as an indicator substrate followed by EDTA inhibition profiles. EDTA was found to be a better inhibitor of metallo-ß-lactamases, especially for anaerobes. IP was a better than TZ. Mueller-Hinton agar was the preferred medium, particularly when compared to Isosensitest agar, which frequently produced falsely low MICs for IP. Etest IP plus IP-EDTA with Mueller-Hinton agar had a sensitivity of 94% (79 of 84) and specificity of 95% (124 of 130). The Etest MBL strip appears to be an acceptable diagnostic reagent to detect metallo-ß-lactamase phenotypes in the clinical microbiology laboratory.

* Corresponding author. Mailing address: Department of Pathology and Microbiology, University Walk, University of Bristol, Bristol BS8 1TD, United Kingdom. Phone: 44 117 9287897. Fax: 44 117 9287896. E-mail: t.r.walsh{at}bristol.ac.uk.

Journal of Clinical Microbiology, August 2002, p. 2755-2759, Vol. 40, No. 8
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.8.2755-2759.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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