Related Experiment Video
Updated: Dec 8, 2025
Anti-virulent Disruption of Pathogenic Biofilms using Engineered Quorum-quenching Lactonases
Published on: January 1, 2016
Comparative Kinetic Analysis of OXA-438 with Related OXA-48-Type Carbapenem-Hydrolyzing Class D β-Lactamases
Denise De Belder1,2, Barbara Ghiglione2,3, Fernando Pasteran1
1Servicio Antimicrobianos - National Reference Laboratory in Antimicrobial Resistance (NRLAR), Instituto Nacional de Enfermedades Infecciosas-ANLIS "Dr. Carlos G. Malbrán", Buenos Aires 1282, Argentina.
Abstract:
Novel variants of OXA-48-type enzymes with the ability to hydrolyze oxyimino-cephalosporins and carbapenems are increasingly reported. Since its first report in 2011, OXA-163 is now extensively spread throughout Argentina, and several variants like OXA-247 have emerged. Here, we characterized a new blaOXA-48-like variant, OXA-438, and we performed a comparative kinetic analysis with the local variants OXA-247 and OXA-163 and the internationally disseminated OXA-48. blaOXA-163, blaOXA-247, and blaOXA-438 were located in a 70 kb IncN2 conjugative plasmid. OXA-438 presented mutations in the vicinity of conserved KTG (214-216), with a 2-aa deletion (R220-I221) and a D224E shift (as in OXA-163) compared to OXA-48. Despite Kpn163 (OXA-163), Kpn247 (OXA-247) and Eco438 (OXA-438) were resistant to meropenem and ertapenem, and the transconjugants (TC) remained susceptible (however, the carbapenems minimum inhibitory concentrations were ≥3 times 2-fold dilutions higher than the acceptor strain). TC163 and Eco48 were resistant to oxyimino-cephalosporins, unlike TC247 and TC438. kcat/Km values for cefotaxime in OXA-163 were slightly higher than the rest of the variants that were accompanied by a lower Km for carbapenems. For OXA-163, OXA-247, and OXA-438, the addition of NaHCO3 improved kcat values for both cefotaxime and ceftazidime; carbapenems kcat/Km values were higher than for oxyimino-cephalosporins. Mutations occurring near the conserved KTG in OXA-247 and OXA-438 are probably responsible for the improved carbapenems hydrolysis and decreased inactivation of oxyimino-cephalosporins compared to OXA-163. Dichroism results suggest that deletions at the β5-β6 loop seem to impact the structural stability of OXA-48 variants. Finally, additional mechanisms are probably involved in the resistance pattern observed in the clinical isolates.
Insights
Novel OXA-48-like carbapenemase variants, OXA-163, OXA-247, and OXA-438, are spreading in Argentina. These enzymes show varying resistance to carbapenems and cephalosporins due to mutations, impacting treatment options.
Area of Science:
- Molecular biology
- Microbiology
- Biochemistry
Background:
- OXA-48-type enzymes are a growing concern due to their ability to hydrolyze carbapenems and oxyimino-cephalosporins.
- OXA-163, first reported in 2011, is widespread in Argentina, with variants like OXA-247 also emerging.
- Understanding the enzymatic properties and genetic basis of these variants is crucial for combating antimicrobial resistance.
Purpose of the Study:
- To characterize a new OXA-48-like variant, OXA-438.
- To perform a comparative kinetic analysis of OXA-438 against local variants (OXA-163, OXA-247) and the globally prevalent OXA-48.
- To investigate the impact of specific mutations on enzyme activity and resistance profiles.
Main Methods:
- Genetic characterization of blaOXA-48-like variants.
- Kinetic analysis (kcat/Km) of enzyme activity against various beta-lactam antibiotics.
- Determination of minimum inhibitory concentrations (MICs) in transconjugant strains.
- Circular dichroism to assess structural stability.
Main Results:
- OXA-163, OXA-247, and OXA-438 were located on a 70 kb IncN2 conjugative plasmid.
- OXA-438 possesses mutations near the conserved KTG motif, including a 2-amino acid deletion and a D224E substitution.
- Transconjugants expressing OXA-163 and OXA-48 showed resistance to oxyimino-cephalosporins, while OXA-247 and OXA-438 transconjugants did not.
- All three variants conferred resistance to meropenem and ertapenem, with OXA-163, OXA-247, and OXA-438 exhibiting higher carbapenem MICs than the acceptor strain.
- Mutations in OXA-247 and OXA-438 likely enhance carbapenem hydrolysis and reduce oxyimino-cephalosporin inactivation compared to OXA-163.
- Deletions in the β5-β6 loop appear to affect the structural stability of OXA-48 variants.
Conclusions:
- OXA-438 represents a new OXA-48-like variant with distinct kinetic properties and resistance profiles.
- Mutations near the KTG motif significantly influence the hydrolytic activity against different beta-lactams.
- Additional resistance mechanisms may be involved in the observed resistance patterns in clinical isolates.
More Related Videos
08:37The Application of Open Searching-based Approaches for the Identification of Acinetobacter baumannii O-linked Glycopeptides
Published on: November 2, 2021
12:03Antimicrobial Synergy Testing by the Inkjet Printer-assisted Automated Checkerboard Array and the Manual Time-kill Method
Published on: April 18, 2019
