Session: Poster Session: β-Lactamase Detection in the Clinical Laboratory
Saturday, October 25, 2008: 12:00 AM
Room: Hall C
Background: NXL104 is a non-β-lactam covalent inhibitor of a broad spectrum of serine β-lactamases of classes A and C, including ESBLs (Extended Spectrum β-Lactamases) and KPC carbapenemases. The KPCs are of greater concern, conferring resistance to all β-lactams including the currently marketed inhibitors and the carbapenems. KPC enzymes have been reported in a variety of Enterobacteriaceae but remain difficult for clinical microbiology laboratories to detect. Ceftazidime (CAZ) + NXL104 disks are being developed to support the clinical development of CAZ/NXL104. We have invented a novel use for NXL104 as a diagnostic reagent for the detection of KPC β-lactamases expressed by strains of Enterobacteriaceae species. Methods: Bacterial isolates tested, six clinical isolates known to harbour KPC-2 or KPC-3 (3 K. pneumoniae, 1 Escherichia coli and 2 Enterobacter cloacae) and five clinical Enterobacteriaceae isolates without KPCs but with ESBLs, including the K. pneumoniae ATCC 700603 (SHV-18). MICs were determined as per CLSI microbroth dilution technique with ceftazidime (CAZ) and imipenem (IPM) alone or combined with NXL104 at a fixed concentration of 4µg/mL. Double disk synergy test for detection of KPC: Mueller-Hinton agar plates were inoculated with a lawn of ~108 CFU/mL adjusted test strains. A 10µg IPM disk was placed at a distance of ~20-22 mm apart from a disk of 30µg CAZ+60µg NXL104. Plates were inverted and incubated at 37°C overnight. Results: Synergy, indicating the presence of carbapenemase, was seen as an expansion of the IPM zone adjacent to the CAZ/NXL104 disk only for strains known to carry a KPC enzyme. These results correlated with the decrease in IPM MICs in the presence of NXL104 (≤0.125 - 2µg/mL) compared to its absence (8 - 64µg/mL). Conclusion: These results demonstrate CAZ/NXL104 double disk synergy test as a potentially useful method for detecting KPCs in Enterobacteriaceae species in clinical microbiology laboratories.
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