Comparative Kinetic Analysis of OXA-438 with related OXA-48-Type Carbapenem-Hydrolyzing Class D β-Lactamases.

Novel variants of the 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 in 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): 2-aa deletion (R220-I221) and 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, the transconjugants (TC) remained susceptible (although the carbapenems MIC were ≥3 times 2-fold dilutions higher than acceptor strain). TC163 and Eco48 were resistant to oxyimino-cephalosporins, unlike TC247 and TC438. kcat/Km value for cefotaxime in OXA-163 was slightly higher than the rest of variants, accompanied by a lower Km for carbapenems. For OXA-163, OXA-247 and OXA-438, addition of NaHCO3 improved kcat values for both cefotaxime and ceftazidime; carbapenems kcat/Km were higher than for oxyimino-cephalosporins. Mutations occurring near conserved KTG in OXA-247 and OXA-438 are probably responsible for 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 in the structure stability of OXA-48 variants. Finally, additional mechanisms are probably involved in the resistance pattern observed in the clinical isolates.