meropenem and dipicolinic-acid

Klebsiella pneumoniae strains co-producing klebsiella pneumoniae carbapenemase (KPC) and verona integron-encoded metallo-beta-lactamase (VIM) are frequently isolated in Greece and have also occurred in other European countries. Conventional combined disc tests exhibit low sensitivity against these emerging pathogens. We have evaluated modifications of the KPC/Metallo-β-Lactamase Confirmation kit (ROSCO) exhibiting high diagnostic value against KPC, VIM and KPC + VIM producers. The key changes were the inclusion of additional combined tablets containing meropenem plus two inhibitors (dipicolinic acid (1000 μg per tablet) for metallo-β-lactamases and a boronic acid derivative for KPCs) and the replacement of aminophenylboronic acid by phenylboronic acid (400 μg per tablet).

Topics: Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Cloxacillin; Europe; Greece; Imipenem; Klebsiella Infections; Klebsiella pneumoniae; Meropenem; Microbial Sensitivity Tests; Picolinic Acids; Reproducibility of Results; Thienamycins

Enterobacteriaceae producing carbapenemases, such as KPC or metallo-β-lactamases (MBLs), have emerged on several continents. Phenotypic tests are urgently needed for their rapid and accurate detection. A novel carbapenemase detection test, comprising a meropenem disk, and meropenem disks supplemented with 730 μg of EDTA, 1000 μg of dipicolinic acid (DPA), 600 μg of aminophenylboronic acid (APBA), or 750 μg of cloxacillin, was evaluated against Klebsiella pneumoniae isolates with KPC (n = 34), VIM (n = 21), IMP (n = 4) or OXA-48 (n = 9) carbapenemases, and carbapenem-resistant Enterobacteriaceae with porin loss in combination with an extended-spectrum β-lactamase (ESBL) (n = 9) or AmpC hyperproduction (n = 5). Commercially available diagnostics tablets from Rosco containing meropenem and the same inhibitors as described above (except EDTA) were also evaluated. An increased meropenem inhibition zone was sought in the presence of each added β-lactamase inhibitor. APBA had excellent sensitivity for detecting K. pneumoniae with KPC enzymes. Isolates with combined AmpC hyperproduction and porin loss were also positive in the APBA test but, unlike KPC producers, showed cloxacillin synergy. Both DPA and EDTA had excellent sensitivity for detection of MBL-producing K. pneumoniae. However, EDTA showed poor specificity, with positive results noted for 1/9 ESBL-producing isolates, for 4/34 KPC-producing isolates, and for 4/9 OXA-48-producing isolates, whereas all of these were negative when DPA was used. The in-house test distinguished accurately between several different mechanisms mediating reduced susceptibility to carbapenems in Enterobacteriaceae. The commercial combination tablets from Rosco performed similarly to the in-house test, with the exception of one false-positive MBL result and one false-positive KPC result among the OXA-48 producers.

Topics: Anti-Bacterial Agents; beta-Lactamases; Boronic Acids; Cloxacillin; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Meropenem; Microbial Sensitivity Tests; Picolinic Acids; Sensitivity and Specificity; Thienamycins

We evaluated the ability of the combination disk test (CDT) and the Modified Hodge Test (MHT) to discriminate between various carbapenemase-producing Pseudomonas aeruginosa isolates (KPC, n = 36; metallo-β-lactamase (MBL), n = 38) and carbapenemase non-producers (n = 75). For the CDT, the optimal inhibitor concentrations and cut-off values were: 600 μg of 3-aminophenylboronic acid (APB) per disk (an increment of ≥4 mm), 1000 μg of dipicolinic acid (DPA) per disk (an increment of ≥5 mm) and 3000 μg of cloxacillin per disk (an increment of ≥3 mm). APB had excellent sensitivity (97%) and specificity (97%) for the detection of KPC enzymes. DPA detected MBL enzymes with a sensitivity and specificity of 97% and 81%, respectively. The MHT resulted in a low sensitivity (78%) and specificity (57%). The CDT could be very useful in daily practice to provide fast and reliable detection of KPC and MBL carbapenemases among P. aeruginosa isolates.

Topics: Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Boronic Acids; Cloxacillin; Disk Diffusion Antimicrobial Tests; Humans; Meropenem; Picolinic Acids; Pseudomonas aeruginosa; Pseudomonas Infections; Sensitivity and Specificity; Thienamycins

We evaluated dipicolinic acid (DPA) as a chelating agent for detection of IMP- or VIM-type metallo-beta-lactamase (MBL)-producing Pseudomonas aeruginosa clinical isolates. Using the broth microdilution testing in the presence or absence of DPA, MBL producers exhibited 100%, 92%, or 100% of >or=8 times (media, 32 times) reduction of MICs in presence of DPA for ceftazidime, imipenem, or meropenem, respectively. In disk diffusion testing, expansion of growth inhibitory zone of these clinical isolates was clearly observed. Thus, DPA could be useful in the detection for MBL-producing P. aeruginosa clinical isolates.

Topics: Anti-Bacterial Agents; beta-Lactam Resistance; beta-Lactamases; Ceftazidime; Chelating Agents; Humans; Imipenem; Meropenem; Microbial Sensitivity Tests; Picolinic Acids; Pseudomonas aeruginosa; Thienamycins