Phenotypic Detection of Carbapenem-Susceptible Metallo-β-Lactamase-Producing Gram-Negative Bacilli in the Clinical Laboratory

Abstract

Rapid detection of metallo-␤-lactamase (MBL)-producing gram-negative pathogens is critical to prevent their widespread dissemination. Thus far, no standardized phenotypic method is available, and previously reported techniques have poor sensitivity for detecting carbapenem-susceptible MBL-carrying isolates, an increasingly described phenomenon. We developed a phenotypic detection method using both a double-disk synergy test and a combined-disk test with imipenem and 292 g EDTA on one agar plate. Genotypic confirmation was used for validation. Of the 134 clinical isolates, 84 were confirmed to carry an MBL. Of these, 51 (61%) were susceptible to at least one carbapenem, and 22 (26%) were isolated from blood. The phenotypic method correctly differentiated all MBL-producing isolates (sensitivity, 100%). Fifty-one of the 52 MBLnegative isolates were correctly differentiated (specificity, 98%). This study reports the validation of a simple and accurate MBL detection method that can be easily incorporated into the daily routine of a clinical laboratory. Early detection of MBL-carrying organisms, including those with susceptibility to carbapenems, is of paramount clinical importance, as it allows rapid initiation of strict infection control practices as well as therapeutic guidance for confirmed infection.

The molecular basis of β-lactamase production in Gram-negative bacteria from Saudi Arabia

“…VIM was also predominant among 75 randomly selected A. baumannii isolates (65 % of which were carbapenem resistant), accounting for 96 % of the isolates. This confirms results of previous studies that showed that the number of isolates carrying a gene for the VIM-type enzyme is much higher than the number of isolates phenotypically resistant to carbapenems (Franklin et al, 2006;Peleg et al, 2005).…”

Prevalence of IncI1-Iγ and IncFIA-FIB type plasmids in extended-spectrum β-lactamase-producing Klebsiella pneumoniae strains isolated from the NICU of a North Indian hospital

“…An increase in the diameter of the inhibition zone by ¢4 mm around the imipenem+EDTA, meropenem+EDTA and ertapenem+EDTA as compared to that of the imipenem, meropenem and ertapenem alone indicated the presence of MBL (Franklin et al, 2006).…”

An evaluation of four different phenotypic techniques for detection of metallo-β-lactamase producing <i> Pseudomonas aeruginosa</i>

“…[3] Currently, no standardised method for MBL detection has been proposed and despite PCR being highly accurate and reliable, its accessibility is often limited to reference laboratories. [4] Several non molecular techniques have been studied, all taking advantage of the enzyme's zinc dependence by using chelating agents, such as EDTA or 2 mercaptopropionic acid, to inhibit its activity. [4] Our institute, a tertiary care centre in India, has a very high prevalence of nosocomial infections due to P. aeruginosa.…”

“…[4] Several non molecular techniques have been studied, all taking advantage of the enzyme's zinc dependence by using chelating agents, such as EDTA or 2 mercaptopropionic acid, to inhibit its activity. [4] Our institute, a tertiary care centre in India, has a very high prevalence of nosocomial infections due to P. aeruginosa. [5,6] We have also found a very high prevalence of multidrug resistant (MDR) and ESBL positive gram negative bacteria in ICUs and other wards of our hospitals.…”