Our findings do not support the suggestion of a preferential use of azithromycin over clarithromycin in order to limit inducible macrolide resistance. Both compounds provoked a comparable resistance phenotype in M. abscessus. Caution is needed when using either azithromycin or clarithromycin for treatment of M. abscessus infections.

bAntibiotic therapy of infections caused by the emerging pathogen Mycobacterium abscessus is challenging due to the organism's natural resistance toward most clinically available antimicrobials. We investigated the bactericidal activity of antibiotics commonly administered in M. abscessus infections in order to better understand the poor therapeutic outcome. Time-kill curves were generated for clinical M. abscessus isolates, Mycobacterium smegmatis, and Escherichia coli by using antibiotics commonly categorized as bactericidal (amikacin and moxifloxacin) or bacteriostatic (tigecycline and linezolid). In addition, the impact of aminoglycoside-modifying enzymes on the mode of action of substrate and nonsubstrate aminoglycosides was studied by using M. smegmatis as a model organism. While amikacin and moxifloxacin were bactericidal against E. coli, none of the tested compounds showed bactericidal activity against M. abscessus. Further mechanistic investigations of the mode of action of aminoglycosides in M. smegmatis revealed that the bactericidal activity of tobramycin and gentamicin was restored by disruption of the chromosomal aac(2=) gene in the mycobacterial genome. The lack of bactericidal antibiotics in currently recommended treatment regimens provides a reasonable explanation for the poor therapeutic outcome in M. abscessus infection. Our findings suggest that chromosomally encoded drug-modifying enzymes play an important role in the lack of aminoglycoside bactericidal activity against rapidly growing mycobacteria.

Reliable identification of carbapenemase-producing members of the family Enterobacteriaceae is necessary to limit their spread. This study aimed to develop a diagnostic flow chart using phenotypic screening and confirmation tests that is suitable for implementation in different types of clinical laboratories. A total of 334 clinical Enterobacteriaceae isolates genetically characterized with respect to carbapenemase, extended-spectrum ␤-lactamase (ESBL), and AmpC genes were analyzed. A total of 142/334 isolates (42.2%) were suspected of carbapenemase production, i.e., intermediate or resistant to ertapenem (ETP) and/or meropenem (MEM) and/or imipenem (IPM) according to EUCAST clinical breakpoints (CBPs). A group of 193/334 isolates (57.8%) showing susceptibility to ETP, MEM, and IPM was considered the negative-control group in this study. CLSI and EUCAST carbapenem CBPs and the new EUCAST MEM screening cutoff were evaluated as screening parameters. ETP, MEM, and IPM with or without aminophenylboronic acid (APBA) or EDTA combined-disk tests (CDTs) and the Carba NP-II test were evaluated as confirmation assays. EUCAST temocillin cutoffs were evaluated for OXA-48 detection. The EUCAST MEM screening cutoff (<25 mm) showed a sensitivity of 100%. The ETP APBA CDT on Mueller-Hinton agar containing cloxacillin (MH-CLX) displayed 100% sensitivity and specificity for class A carbapenemase confirmation. ETP and MEM EDTA CDTs showed 100% sensitivity and specificity for class B carbapenemases. Temocillin zone diameters/MIC testing on MH-CLX was highly specific for OXA-48 producers. The overall sensitivity, specificity, positive predictive value, and negative predictive value of the Carba NP-II test were 78.9, 100, 100, and 98.7%, respectively. Combining the EUCAST MEM carbapenemase screening cutoff (<25 mm), ETP (or MEM), APBA, and EDTA CDTs, and temocillin disk diffusion on MH-CLX promises excellent performance for carbapenemase detection. In recent years, the emergence of diverse carbapenemases in members of the family Enterobacteriaceae has become a major challenge for health care systems (1). Carbapenemase-producing bacterial isolates pose a severe clinical problem, as nonsusceptibility to beta-lactams is frequently accompanied by coresistance to additional drug classes, e.g., aminoglycosides or quinolones (2, 3). As a consequence, treatment options for carbapenemase producers are alarmingly limited and often drugs displaying significant side effects need to be administered as a last resort (4).␤-Lactamases are classified according to their functional properties and molecular structure by Ambler and Bush (5, 6). Some of these enzymes also display hydrolytic activity toward carbapenems, e.g., Klebsiella pneumoniae carbapenemase (KPC, Ambler/ Bush class A)-, the New Delhi metallo-␤-lactamase (NDM-1)-, VIM-, and GIM-type enzymes (all Ambler/Bush class B) or OXA-48 (Ambler/Bush class D). A key characteristic used to discriminate enzymes belonging to different Ambler/Bush classes is responsiveness to specific inhibitors: class...

bThis study evaluated the performance of the Rapidec Carba NP test, which was introduced recently into the market for the detection of carbapenemase production in a broad spectrum of ␤-lactamase-producing Enterobacteriaceae clinical isolates. In total, 252 clinical Enterobacteriaceae isolates that had been genetically characterized with respect to carbapenemase, extended-spectrum ␤-lactamase (ESBL), and AmpC genes were analyzed; 51/252 isolates (20.2%) were genetically confirmed to be carbapenemase producers, whereas 201/252 isolates (79.8%) were genetically negative for the presence of carbapenemase genes. The Rapidec Carba NP test was applied according to the manufacturer's instructions, and results were read after 30 and 120 min of incubation. The overall sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the Rapidec Carba NP test were 90.2%, 100%, 100%, and 97.6%, respectively, when the manufacturer's instructions were followed. Four of 5 false-negative results occurred with OXA-48-like enzymes. After an incubation time of 30 min, the sensitivity was 49%. The sensitivity increased to 100% when the recommended bacterial inoculum was doubled and the test was read strictly after 120 min of incubation. The Rapidec Carba NP test is a useful tool for the reliable confirmation of carbapenemase-producing Enterobacteriaceae isolates. The test should be read strictly after 120 min of incubation and the inoculum should be larger than recommended by the manufacturer.

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