Carbapenems as inhibitors of OXA-13, a novel, integron-encoded beta-lactamase in Pseudomonas aeruginosa.

A clinical Pseudomonas aeruginosa strain, PAe391, was found to be resistant to a number of antibiotics including ticarcillin, piperacillin, cefsulodin and amikacin, and a disk diffusion assay showed evidence of pronounced synergy between imipenem and various beta-lactam antibiotics. Cloning and nucleotide sequence analysis revealed the dicistronic arrangement of an aac(6')-Ib variant and a novel blaOXA-type gene between the intI and qacE delta 1 genes typical of integrons, in PAe391, this integron was apparently chromosome-borne. The beta-lactamase, named OXA-13, displayed nine amino acid changes with respect to OXA-10:I in position 10 of OXA-10 to T (I10T), G20S, D55N, N73S, T107S, Y174F, E229G, S245N and E259A, OXA-13 (pIapp = 8.0) showed poor catalytic activity against penicillins as well as cephalosporins, but was efficient in hydrolysing some penicillinase-resistant beta-lactams, such as cefotaxime and aztreonam. It was efficiently inhibited by imipenem (KIapp = 11 nM), and formed a stable complex. While the KIapp value of meropenem was similar (16 nM), the corresponding complex was less stable.

[Identification of a cephalosporinase susceptible to clavulanic acid in a clinical strain of Serratia fonticola]. / Identification dans une souche clinique de Serratia fonticola d'une céphalosporinase sensible à l'acide clavulanique.

We analyzed the beta-lactamase production of a Serratia fonticola isolated for its resistance to cefuroxime (Minimum Inhibitory Concentration > 256 mg/l) in December 1993 from a patient hospitalized in Meaux. The wild strain was resistant to amoxycillin but sensitive to augmentin, that suggested the production of a beta-lactamase susceptible to clavulanic acid. For the wild strain, beta-lactamase production was inducible and only one enzyme with an isoelectric point of 8.12 was detected. beta-lactamase production was 16 mU/mg for non-induced extracts and ranged from 100 to 230 mU/mg in the presence of inducing beta-lactams (enzyme activity was measured with penicillin G as substrate). On a Szybalski gradient a constitutive strain was obtained. Its enzyme production was 13,000 mU/mg. The kinetics and isoelectric points of the enzymes produced by the two strains were identical. This beta-lactamase hydrolyzes penicillins (amoxycillin: Vm = 60 relative to penicillin G = 100, ticarcillin: 15), first generation cephalosporins (cephalothin Vm = 930). However, this enzyme hydrolyzes efficiently oxyimino-cephalosporins: cefuroxime (Vm = 70) and cefotaxime (Vm = 120), but cephamycins are not substrates. Clavulanic acid has a very good affinity for this beta-lactamase (Ki = 0.09 microM) which is inactivated progressively (I50 = 0.045 microgram/ml). These properties shows some similarities with those of the class A beta-lactamases of P. vulgaris RO104 (pI = 8.3), P. penneri 14HBC (pI = 6.65) and the plasmid-mediated extended-spectrum MEN-1 (pI = 8.4).

[Properties of a cephalosporinase produced by Proteus penneri inhibited by clavulanic acid]. / Propriétés d'une céphalosporinase de Proteus penneri inhibée par l'acide clavulanique.

P. penneri produces an inducible cephalosporinase, as many Enterobacteriaceae. Nevertheless this betalactamase is susceptible to clavulanic acid which is an exception also encountered for P. vulgaris. The authors studied the enzyme produced by P. penneri 14HBC resistant to cefotaxime (MIC 16 mg/l) isolated in Spain in 1992. This betalactamase of isoelectric point 6.65 hydrolyzes first generation cephalosporins, amoxycillin and poorly ticarcillin as it occurs for all cephalosporinases. However, this enzyme hydrolyzes strongly oxyimino-cephalosporins: cefuroxime, cefotaxime, cefepime, cefpirome as it occurs with extended-spectrum betalactamases. Cephamycins and imipenem are not substrates. Clavulanic acid has a very good affinity for this betalactamase which is inactivated progressively. These properties are similar to those of the enzyme of P. vulgaris Ro104 of isoelectric point 8.3 which, contrarily to other cephalosporinases, belongs to the structural Ambler's class A.