Genetic and Biochemical Characterization of OXA-535, a Distantly Related OXA-48-Like β-Lactamase

Dabos, Laura; Jousset, Agnès B; Bonnin, Rémy A.; Fortineau, Nicolas; Zavala, Agustín; Retailleau, Pascal; Iorga, Bogdan; Naas, Thierry

doi:10.1128/aac.01198-18

Cited by 12 publications

(14 citation statements)

References 24 publications

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“…The in silico analysis showed that this assay should be able to detect all the IMP, NDM, VIM and KPC variants described as of December 2018. Among the OXA-48-like group, OXA-54 and OXA-535, described in the chromosome of Shewanella species [15,16] and OXA-436 identified on a plasmid in Enterobacter species, Citrobacter species and K. pneumoniae in Denmark [17] are unlikely to be detected.…”

Section: Discussionmentioning

confidence: 99%

Rapid detection of IMP, NDM, VIM, KPC and OXA-48-like carbapenemases from Enterobacteriales and Gram-negative non-fermenter bacteria by real-time PCR and melt-curve analysis

Mentasti

Prime

Sands

et al. 2019

Eur J Clin Microbiol Infect Dis

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Carbapenemase producing microorganisms are increasingly isolated and often associated with treatment failures and outbreaks. The need for reliable and timely detection and/or confirmation of carbapenemase production is paramount, therefore a real-time PCR assay targeting IMP, NDM, VIM, KPC and OXA-48-like carbapenemases was designed and validated. All available allele variants of the above carbepenmases were downloaded from the Beta-Lactamase DataBase (http://bldb.eu/), aligned with Clustal Omega and primers designed using Primer-BLAST. Real-time PCR monoplexes were optimized for the QuantStudio 6-Flex (Applied Biosystems) using the PowerUp SYBR Green Master Mix (Life Technologies) and validated using a panel of 204 characterised strains carrying a wide range of beta-lactamases, sometimes in combination. Melt-curve analysis was used to confirm positive results. The in silico approach allowed primers to be designed in conserved regions of the KPC and NDM alignments, while three primer sets for IMP and two for VIM were necessary to ensure amplification of the different variants. One primer set was designed for OXA-48-like, however it is unlikely to detect all variants. Expected results were obtained for all 204 tested strains, with 100% sensitivity and specificity. Melt-curve analysis showed consistent Tm results for KPC, NDM and OXA-48-like; differences were instead noted for IMP and VIM as likely consequence of higher variability in the PCR target regions. Inhibition was not observed. The assay is rapid, easy to perform and implement. It enables unequivocal detection of IMP, NDM, VIM, KPC and OXA-48-like carbapenemases even when more than one type is present simultaneously.

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Section: Discussionmentioning

confidence: 99%

Rapid detection of IMP, NDM, VIM, KPC and OXA-48-like carbapenemases from Enterobacteriales and Gram-negative non-fermenter bacteria by real-time PCR and melt-curve analysis

Mentasti

Prime

Sands

et al. 2019

Eur J Clin Microbiol Infect Dis

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“…Additionally, the Ω-loop (residues 143-165) and β5-β6-loop (residues 213-218) bordering the active site seem to be important determinants of OXA-48 activity, as discussed below. According to the β-lactamase database, at least 15 plasmid-encoded OXA-48 β-lactamases have been identified and validated (with further variants chromosomally encoded mainly in different Shewanella species 30,31 ). These variants differ from wild-type OXA-48 by certain amino acid substitutions or deletions.…”

Section: Introductionmentioning

confidence: 99%

Antimicrobial Resistance Conferred by OXA-48 β-Lactamases: Towards a Detailed Mechanistic Understanding

Hirvonen

Spencer

Kamp

2021

Antimicrob Agents Chemother

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OXA-48-type β-lactamases are now routinely encountered in bacterial infections caused by carbapenem-resistant Enterobacterales. These enzymes are of high and growing clinical significance due to the importance of carbapenems in treatment of healthcare-associated infections by Gram-negative bacteria, the wide and increasing dissemination of OXA-48 enzymes on plasmids, and the challenges posed by their detection. OXA-48 confers resistance to penicillin (which is efficiently hydrolyzed) and carbapenem antibiotics (more slowly broken down). In addition to the parent enzyme, a growing array of variants of OXA-48 is now emerging. The spectrum of activity of these variants varies, with some hydrolyzing expanded-spectrum oxyimino-cephalosporins. The growth in importance and diversity of the OXA-48 group has motivated increasing numbers of studies that aim to elucidate the relationship between structure and specificity and establish the mechanistic basis for β-lactam turnover in this enzyme family. In this review we collate recently published structural, kinetic, and mechanistic information on the interactions between clinically relevant β-lactam antibiotics and inhibitors with OXA-48 β-lactamases. Collectively, these studies are starting to form a detailed picture of the underlying bases for the differences in β-lactam specificity between OXA-48 variants, and the consequent differences in resistance phenotype. We focus specifically on aspects of carbapenemase and cephalosporinase activities of OXA-48 β-lactamases and discuss β-lactamase inhibitor development in this context. Throughout the review, we also outline key open research questions for future investigation.

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“…All OXA-48 carbapenemase variants were correctly identified, including difficult-to-detect variants (OXA-244 and OXA-519), using biochemical tests such as Carba NP (12,13). It also detects very recently identified variants that have never been tested by ICA (OXA-484, -505, -517, and -793; Table 1) and the distantly related OXA-535 variant (14), which is the progenitor of OXA-436, a distantly related OXA-48 variant responsible for an outbreak associated with several enterobacterial species in Denmark (14-16). Moreover, OXA-163 and OXA-405, two OXA-48 variants that lack significant imipenemase activity (17) but with strong expanded-spectrum hydrolytic activity, were consistently detected.…”

mentioning

confidence: 96%

Improvement of the Immunochromatographic NG-Test Carba 5 Assay for the Detection of IMP Variants Previously Undetected

Volland

Girlich

Laguide

et al. 2019

Antimicrob Agents Chemother

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Here, we evaluated the immunochromatographic assay NG-Test Carba 5v2 (NG-Biotech), with improved IMP variant detection on 31 IMP producers, representing the different branches of the IMP phylogeny, including 32 OXA-48, 19 KPC, 12 VIM, 14 NDM, and 13 multiple carbapenemase producers (CPs), 13 CPs that were not targeted, and 13 carbapenemase-negative isolates. All tested IMP variants were accurately detected without impairing detection of the other carbapenemases. Thus, NG-Test Carba 5v2 is now well adapted to countries with high IMP prevalence and to the epidemiology of CP-Pseudomonas aeruginosa, where IMPs are most frequently detected.

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Genetic and Biochemical Characterization of OXA-535, a Distantly Related OXA-48-Like β-Lactamase

Cited by 12 publications

References 24 publications

Rapid detection of IMP, NDM, VIM, KPC and OXA-48-like carbapenemases from Enterobacteriales and Gram-negative non-fermenter bacteria by real-time PCR and melt-curve analysis

Rapid detection of IMP, NDM, VIM, KPC and OXA-48-like carbapenemases from Enterobacteriales and Gram-negative non-fermenter bacteria by real-time PCR and melt-curve analysis

Antimicrobial Resistance Conferred by OXA-48 β-Lactamases: Towards a Detailed Mechanistic Understanding

Improvement of the Immunochromatographic NG-Test Carba 5 Assay for the Detection of IMP Variants Previously Undetected

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