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Journal of Clinical Microbiology, January 2000, p. 40-43, Vol. 38, No. 1
0095-1137/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Convenient Test for Screening Metallo-beta -Lactamase-Producing Gram-Negative Bacteria by Using Thiol Compounds

Yoshichika Arakawa,1,* Naohiro Shibata,1 Keigo Shibayama,1 Hiroshi Kurokawa,1 Tetsuya Yagi,1 Hiroshi Fujiwara,1 and Masafumi Goto2

Department of Bacterial and Blood Products, National Institute of Infectious Diseases, Tokyo,1 and Faculty of Pharmaceutical Sciences, Kumamoto University, Kumamoto,2 Japan

Received 28 June 1999/Returned for modification 21 August 1999/Accepted 22 September 1999

A simple disk diffusion test was constructed for detection of IMP-1-type metallo-beta -lactamase-producing gram-negative bacteria. Two Kirby-Bauer disks containing ceftazidime (CAZ) and a filter disk containing a metallo-beta -lactamase inhibitor were used in this test. Several IMP-1 inhibitors such as thiol compounds including 2-mercaptopropionic acid, heavy metal salts, and EDTA were evaluated for this test. Two CAZ disks were placed on a Mueller-Hinton agar plate on which a bacterial suspension was spread according to the method recommended by the National Committee for Clinical Laboratory Standards. The distance between the disks was kept to about 4 to 5 cm, and a filter disk containing a metallo-beta -lactamase inhibitor was placed near one of the CAZ disks within a center-to-center distance of 1.0 to 2.5 cm. For IMP-1-producing strains, the growth-inhibitory zone between the two disks expanded, while no evident change in the shape of the growth-inhibitory zone was observed for CAZ-resistant strains producing serine beta -lactamases such as AmpC or SHV-12. As a result, 2 to 3 µl of undiluted 2-mercaptopropionic acid or mercaptoacetic acid able to block IMP-1 activity gave the most reproducible and clearest results, and CAZ-resistant strains producing AmpC or extended-spectrum beta -lactamases were distinguishable from IMP-1 producers by this test. A similar observation was made with IMP-1-producing clinical isolates such as Serratia marcescens, Klebsiella pneumoniae, Escherichia coli, Enterobacter cloacae, Enterobacter aerogenes, Citrobacter freundii, Proteus vulgaris, Pseudomonas aeruginosa, Pseudomonas putida, Acinetobacter spp., and Alcaligenes xylosoxidans. The specificity and sensitivity of this test were comparable to those of PCR analysis using blaIMP-specific primers. Therefore, this convenient test would be valuable for daily use in clinical laboratories.

* Corresponding author. Mailing address: Department of Bacterial and Blood Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashi-Murayama, Tokyo 208-0011, Japan. Phone: 81-42-561-0771, ext. 500. Fax: 81-42-561-7173. E-mail: yarakawa{at}nih.go.jp.

Journal of Clinical Microbiology, January 2000, p. 40-43, Vol. 38, No. 1
0095-1137/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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