The Scientific World Journal Volume 2012, Article ID 654939, 7 pages doi:10.1100/2012/654939 The cientificWorldJOURNAL Research Article Phenotypic Detection of Metallo-β-Lactamase in Imipenem-Resistant Pseudomonas aeruginosa Yalda Khosravi, 1 Mun Fai Loke, 2 Eng Guan Chua, 1 Sun Tee Tay, 1 and Jamuna Vadivelu 1 1 Department of Medical Microbiology, University of Malaya, 50603 Kuala Lumpur, Malaysia 2 Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia Correspondence should be addressed to Jamuna Vadivelu, jamun@ummc.edu.my Received 19 March 2012; Accepted 12 April 2012 Academic Editors: I. F. N. Hung, Z. Markiewicz, and J. Sanchez Copyright © 2012 Yalda Khosravi et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Carbapenems are the primary choice of treatment for severe Pseudomonas aeruginosa infection. However, the emergence of carbapenem resistance due to the production of metallo-β-lactamases (MBLs) is of global concern. In this study, 90 imipenem- (IPM- or IP-) resistant P. aeruginosa (IRPA) isolates, including 32 previously tested positive and genotyped for MBL genes by PCR, were subjected to double-disk synergy test (DDST), combined disk test (CDT), and imipenem/imipenem-inhibitor (IP/IPI) E-test to evaluate their MBLs detection capability. All three methods were shown to have a sensitivity of 100%. However, DDST was the most specific of the three (96.6%), followed by IP/IPI E-test interpreted based on the single criteria of IP/IPI ≥ 8 as positive (62.1%), and CDT was the least specific (43.1%). Based on the data from this evaluation, we propose that only IRPA with IP MIC > 16 μg/mL and IP/IPI ≥ 8 by IP/IPI E-test should be taken as positive for MBL activity. With the new dual interpretation criteria, the MBL IP/IPI E-test was shown to achieve 100% sensitivity as well as specificity for the IRPA in this study. Therefore, the IP/IPI E-test is a viable alternative phenotypic assay to detect MBL production in IRPA in our population in circumstances where PCR detection is not a feasible option. 1. Introduction Carbapenems, including imipenem (IPM or IP) and mero- penem, are the most potent antibacterial agents used for the treatment of infections initiated by multidrug-resistant gram-negative bacilli [1]. However, acquired resistance to carbapenems has been increasingly reported globally, which can be attributed to the evolution of divergent β-lactamases in numerous gram-negative bacteria (including Pseudomo- nas aeruginosa). P. aeruginosa is an important nosocomial pathogen that is intrinsically resistant to multiple antibiotics. Amongst the various β-lactamases that have been identified to date, the genetically mobile metallo-β-lactamases (MBLs) are the most versatile ones as they are able to hydrolyse all β- lactams except monobactams [2]. Genes encoding for MBL were shown to be carried on large transferable plasmids or were associated with transposons, allowing horizontal trans- fer of these MBL genes among different bacterial genera and species [3]. To date, five types of acquired MBL genes (IMP, VIM, SPM, GIM, and SIM) have been identified based on their divergent protein molecular structures [4–6]. While IMP and VIM variants have been reported worldwide, members of SPM, GIM, and SIM are restricted to certain geographical regions [7, 8]. Although PCR-based genotyping remains as the golden standard for MBL detection and classification, its use is mainly restricted to research purposes. As genotyping infor- mation is necessary, diagnostic centers and laboratories still rely mostly on culture-based phenotypic test as a means for rapid detection of MBL activity. So far, many variations of phenotypic assays for MBLs detection have been reported, and these assays are not standardized. Early detection of MBL-producing organisms is critical as it allows for the prompt use of appropriate antibiotics to effectively control infection. It has been well documented that the activity of MBLs is dependent on zinc or cadmium [4, 9–13]. Several screening methods incorporating the use of metal chelating agents, such as ethylenediaminetetraacetic acid (EDTA) and thiol-based compounds like 2-mercaptopropionic acid (2- MPA), which are capable of blocking MBL activity, have been developed to detect MBL-producing organisms [14–18]. A double-disk synergy test (DDST) using a IPM or ceftazidime