Biological assay for the detection of metallo-beta-lactamases in Bacteroides fragilis

A biological assay was developed for the detection of carbapenemases, particularly metallo-beta-lactamases, in Bacteroides fragilis. The isolates tested possessed the gene (cfiA) responsible for metallo-beta-lactamase production, and showed reduced susceptibility to imipenem. Carbapenemase activity was investigated spectrophotometrically and by a biological assay in which sonicates of bacterial cells were mixed with imipenem in wells cut into Isosensitest agar inoculated with an Escherichia coli indicator organism. After incubation, zones of inhibition were measured. Reductions in zone size compared to a beta-lactamase-negative control, indicating carbapenemase production, were observed with all strains that exhibited hydrolysis of imipenem when measured spectrophotometrically, and with one isolate in which activity was not detected by spectrophotometry. Inclusion of EDTA in the well mixtures abolished the reduction in zone size, indicating the presence of metallo-beta-lactamase. This simple method can detect weak carbapenemase activity that may be overlooked by spectrophotometry.