Global Spread of Carbapenemase-producing Enterobacteriaceae

These resistance traits have been identified among nosocomial and community-acquired infections

Genetic and biochemical characterization of OXA-405, an OXA-48-Type extended-spectrum β-lactamase without significant carbapenemase activity

The epidemiology of carbapenemases worldwide is showing that OXA-48 variants are becoming the predominant carbapenemase type in Enterobacteriaceae in many countries. However, not all OXA-48 variants possess significant activity toward carbapenems (e.g., OXA-163). Two Serratia marcescens isolates with resistance either to carbapenems or to extended-spectrum cephalosporins were successively recovered from the same patient. A genomic comparison using pulsed-field gel electrophoresis and automated Rep-PCR typing identified a 97.8% similarity between the two isolates. Both strains were resistant to penicillins and first-generation cephalosporins. The first isolate was susceptible to expanded-spectrum cephalosporins, was resistant to carbapenems, and had a significant carbapenemase activity (positive Carba NP test) related to the expression of OXA-48. The second isolate was resistant to expanded-spectrum cephalosporins, was susceptible to carbapenems, and did not express a significant imipenemase activity, (negative for the Carba NP test) despite possessing a blaOXA-48-type gene. Sequencing identified a novel OXA-48-type β-lactamase, OXA-405, with a four-amino-acid deletion compared to OXA-48. The blaOXA-405 gene was located on a ca. 46-kb plasmid identical to the prototype IncL/M blaOXA-48-carrying plasmid except for a ca. 16.4-kb deletion in the tra operon, leading to the suppression of self-conjugation properties. Biochemical analysis showed that OXA-405 has clavulanic acid-inhibited activity toward expanded-spectrum activity without significant imipenemase activity. This is the first identification of a successive switch of catalytic activity in OXA-48-like β-lactamases, suggesting their plasticity. Therefore, this report suggests that the first-line screening of carbapenemase producers in Enterobacteriaceae may be based on the biochemical detection of carbapenemase activity in clinical settings

Evaluation of the RAPIDEC® CARBA NP, the Rapid CARB Screen® and the Carba NP test for biochemical detection of carbapenemase-producing Enterobacteriaceae

Objectives The objective of this study was the evaluation of the performance of two commercially available biochemical tests for the rapid detection of carbapenemase-producing Enterobacteriaceae compared with a home-made technique. Methods A collection of 150 enterobacterial isolates, including 132 isolates with decreased susceptibility to at least one carbapenem molecule, were tested for carbapenemase activity using the RAPIDEC® CARBA NP (bioMérieux), the Rapid CARB Screen® (Rosco Diagnostica) and the home-made Carba NP test. This strain collection included 55 non-carbapenemase producers, 21 KPC producers, 21 NDM producers, 17 VIM producers, 11 IMP producers, 16 OXA-48 producers and 9 OXA-48-like producers (OXA-162, OXA-181, OXA-204, OXA-232 and OXA-244). Results The RAPIDEC® CARBA NP detected all carbapenemase producers except a single OXA-244 producer. Using the Rapid CARB Screen®, one KPC-2, two NDM-1, one OXA-48 and five OXA-48 variant producers gave equivocal results and one OXA-244 producer was not detected. Using the Carba NP test, the same OXA-244 producer was not detected and one OXA-181 producer and one OXA-244 producer gave equivocal results. Sensitivity and specificity were 99% (95% CI 94.3%-99.8%) and 100% (95% CI 93.5%-100%), respectively, for the RAPIDEC® CARBA NP test, 89.5% (95% CI 81.7%-94.2%) and 70.9% (95% CI 57.9%-81.2%) for the Rapid CARB Screen® and 96.8% (95% CI 91.1%-98.9%) and 100% (95% CI 93.5%-100%) for the Carba NP test. The impact of the use of an adequate bacterial inoculum for obtaining the optimal performance with the RAPIDEC® CARBA NP was noted. Conclusions The RAPIDEC® CARBA NP possesses the best performance for rapid and efficient detection of carbapenemase-producing Enterobacteriacea

Molecular epidemiology of carbapenem-resistant Acinetobacter baumannii in New Caledonia

ABSTRACTCarbapenem-resistant Acinetobacter baumannii (CR-Ab) ranked third, with a frequency of 24.8%, among 202 strains of multidrug-resistant bacteria isolated from clinical samples in the main hospital of New Caledonia in 2004. All CR-Ab isolates were analysed by isoelectric focusing, conjugation, pulsed-field gel electrophoresis and PCR for the presence of carbapenemase genes. Fifty CR-Ab isolates produced carbapenemase OXA-23. The isolates belonged to a single clone presenting several subtypes, suggesting an endemic situation. This study further illustrates the widespread prevalence of carbapenemase OXA-23-producing CR-Ab isolates in the South Pacific

Characterization of BRP_MBL the bleomycin resistance protein associated with the carbapenemase NDM

The metallo-β-lactamase NDM-1 is among the most worrisome resistance determinants and is spreading worldwide among Gram-negative bacilli. A bleomycin resistance gene, bleMBL, downstream of the blaNDM-1 gene has been associated with resistance almost systematically. Here, we characterized the corresponding protein, BRPMBL, conferring resistance to bleomycin, an antitumoral glycopeptide molecule. We have determined whether the expression of the blaNDM-1-bleMBL operon is inducible in the presence of carbapenems and/or bleomycin-like molecules using quantitative reverse transcription-PCR (qRT-PCR), determination of imipenem and zeocin MICs, and carbapenemase-specific activity assays. We showed that the blaNDM- 1-bleMBL operon is constitutively expressed. Using electrophoretic mobility shift and DNA protection assays performed with purified glutathione S- transferase (GST)-BRPMBL, we demonstrated that BRPMBL is able to bind and sequester bleomycin-like molecules, thus preventing bleomycin-dependent DNA degradation. In silico modeling confirmed that the mechanism of action required the dimerization of the BRPMBL protein in order to sequester bleomycin and prevent DNA damage. BRPMBL acts specifically on bleomycin-like molecules since cloning and expression of bleMBL in Staphyloccoccus aureus did not confer cross-resistance to any other antimicrobial glycopeptides such as vancomycin and teicoplanin

NmcA Carbapenem-hydrolyzing Enzyme in Enterobacter cloacae in North America1

An imipenem-resistant Enterobacter cloacae isolate was recovered from the blood of a patient with a hematologic malignancy. Analytical isoelectric focusing, inhibitor studies, hydrolysis, induction assays, and molecular sequencing methods confirmed the presence of a NmcA carbapenem-hydrolyzing enzyme. This first report of NmcA detected in North America warrants further investigation into its distribution and clinical impact

Worldwide Diversity of Klebsiella pneumoniae That Produce β-Lactamase blaKPC-2 Gene1

TOC summary: Clones harboring different plasmids with identical genetic structure could be the origin of worldwide spread

Evaluation of the RAPIDEC® CARBA NP, the Rapid CARB Screen® and the Carba NP test for biochemical detection of carbapenemase-producing Enterobacteriaceae

The objective of this study was the evaluation of the performance of two commercially available biochemical tests for the rapid detection of carbapenemase-producing Enterobacteriaceae compared with a home-made technique.Methods: A collection of 150 enterobacterial isolates, including 132 isolates with decreased susceptibility to at least one carbapenem molecule, were tested for carbapenemase activity using the RAPIDEC® CARBA NP (bioMérieux), the Rapid CARB Screen® (Rosco Diagnostica) and the home-made Carba NP test. This strain collection included 55 non-carbapenemase producers, 21 KPC producers, 21 NDM producers, 17 VIM producers, 11 IMP producers, 16 OXA-48 producers and 9 OXA-48-like producers (OXA-162, OXA-181, OXA-204, OXA-232 and OXA-244).Results: The RAPIDEC® CARBA NP detected all carbapenemase producers except a single OXA-244 producer. Using the Rapid CARB Screen®, one KPC-2, two NDM-1, one OXA-48 and five OXA-48 variant producers gave equivocal results and one OXA-244 producer was not detected. Using the Carba NP test, the same OXA-244 producer was not detected and one OXA-181 producer and one OXA-244 producer gave equivocal results. Sensitivity and specificity were 99% (95% CI 94.3%–99.8%) and 100% (95% CI 93.5%–100%), respectively, for the RAPIDEC® CARBA NP test, 89.5% (95% CI 81.7%–94.2%) and 70.9% (95% CI 57.9%–81.2%) for the Rapid CARB Screen® and 96.8% (95% CI 91.1%–98.9%) and 100% (95% CI 93.5%–100%) for the Carba NP test. The impact of the use of an adequate bacterial inoculum for obtaining the optimal performance with the RAPIDEC® CARBA NP was noted.Conclusions: The RAPIDEC® CARBA NP possesses the best performance for rapid and efficient detection of carbapenemase-producing Enterobacteriaceae

Neonatal infections with multidrug-resistant ESBL-producing E. cloacae and K. pneumoniae in Neonatal Units of two different Hospitals in Antananarivo, Madagascar

Background: We investigated the molecular mechanism of ß-lactam resistance in extended-spectrum ß-lactamase (ESBL)-producing Enterobacterial strains isolated in neonatal units of different hospitals in Anatnanarivo, Madagascar.Methods: Bacteria were identified by standard biochemical methods, disc diffusion antibiograms and Etest. Resistance genes were sought by PCR. Strains were characterized by Rep- PCR (Diversilab), plasmid analysis and rep-typing.Results: From April 2012 to March 2013, 29 ESBL-producing E. cloacae and 15 K. pneumoniae were isolated from blood culture (n = 32) or gastric samples (n = 12) performed at day 0 or 2 from 39/303 newborns suspected of early neonatal infection. These infants were treated with expanded spectrum cephalosporins, due to lack of carbapenems, leading to a high mortality rate (45 %). Isolates recovered were all, but 4, multidrug resistant, particularly to fluoroquinolones (FQ) except for 21 E. cloacae isolates. Isolates produced TEM-1 and CTX-M-15 ß-lactamases and their genes were located on several self- transferable plasmids of variable sizes sizes that could not be linked to a major plasmid incompatibility group. E. cloacae isolates belonged to 6 Rep-types among which two counted for 11 isolates each. The FQ resistant E. cloacae isolates belonged to one clone, whereas the FQ susceptible E. cloacae isolates belonged to four clones. The K. pneumoniae isolates belonged to 9 Rep-types among which one included five isolates.Conclusion: This study is the first molecular characterization of ESBL- producing isolates from neonatology units in Madagascar, a country with limited epidemiological data. It revealed an important multi-clonal dissemination of CTX-M-15- producing isolates reflecting both the high community carriage and the very early nosocomial contamination of the neonates