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Journal of Clinical Microbiology, June 2005, p. 2551-2558, Vol. 43, No. 6
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.6.2551-2558.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Practical Methods Using Boronic Acid Compounds for Identification of Class C ß-Lactamase-Producing Klebsiella pneumoniae and Escherichia coli

Tetsuya Yagi, Jun-ichi Wachino, Hiroshi Kurokawa, Satowa Suzuki, Kunikazu Yamane, Yohei Doi, Naohiro Shibata, Haru Kato, Keigo Shibayama, and Yoshichika Arakawa^*

Department of Bacterial Pathogenesis and Infection Control, National Institute of Infectious Diseases, Tokyo, Japan

Received 12 October 2004/ Returned for modification 24 January 2005/ Accepted 24 February 2005

Detection of the resistance mediated by class C ß-lactamases remains a challenging issue, considering that transferable plasmid-mediated class C ß-lactamases are of worldwide concern. Methods for the identification of strains that produce extended-spectrum ß-lactamases (ESBLs) or metallo-ß-lactamases (MBLs) have been developed and applied for routine use in clinical microbiology laboratories, but no practical methods for identification of plasmid-mediated class C producers have been established to date. We therefore developed three simple methods for clinical microbiology laboratories that allow identification of plasmid-mediated class C ß-lactamase-producing bacteria using a boronic acid derivative, 3-aminophenylboronic acid (APB), one of the specific inhibitors of class C ß-lactamases. Detection by the disk potentiation test was based on the enlargement of the growth-inhibitory zone diameter (by greater than or equal to 5 mm) around a Kirby-Bauer disk containing a ceftazidime (CAZ) or a cefotaxime (CTX) disk in combination with APB. In a double-disk synergy test, the discernible expansion of the growth-inhibitory zone around the CAZ or the CTX disk toward a disk containing APB was indicative of class C ß-lactamase production. A greater than or equal to eightfold decrease in the MIC of CAZ or CTX in the presence of APB was the criterion for detection in the microdilution test. By using these methods, Escherichia coli and Klebsiella pneumoniae isolates producing plasmid-mediated class C ß-lactamases, ACT-1, CMY-2, CMY-9, FOX-5, LAT-1, and MOX-1, were successfully distinguished from those producing other classes of ß-lactamases, such as ESBLs and MBLs. These methods will provide useful information needed for targeted antimicrobial therapy and better infection control.

Present address: Division of Respiratory Medicine, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511, Japan.

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