Contribution of porin deficiency to the ceftazidimase profile in Escherichia coli strains producing CTX-M beta-lactamases.

GHIGLIONE BÁRBARA; RODRIGUEZ MARGARITA; PEREZ GARÓFALO MAGALÍ; RADICE MARCELA; DI CONZA JOSÉ; POWER PABLO; GUTKIND GABRIEL

Congreso; FEMS 2013 - 5th Congress of European Microbiologists; 2013

Background: CTX-M beta-lactamases confer resistance to cefotaxime while they poorly hydrolyze ceftazidime. Nevertheless, diversification of the CTX-M family led to the emergence of variants responsible for decreased susceptibility to ceftazidime, being the D240G mutation the most prevalent among the so called ceftazidimases. Objectives: To analyze the influence of the D240G mutation in wild type and OmpC/OmpF porin-deficient E. coli strains expressing CTX-M beta-lactamases. Methods: Representative parental blaCTX-M-genes and their corresponding D240G mutants from CTX-M clusters 1, 2, 8, 9 and 25 were PCR-amplified from native strains or generated by site-directed mutagenesis. Genes were cloned in pK19 under regulation of the vector´s promoter and transformed in E. coli XL1blue. ESBL screening and susceptibility tests were performed according to CLSI recommendations. Recombinant plasmids from cluster 1 were also transformed in E. coli K12 and OmpC/OmpF deficient strains, and susceptibility profiles determined. Conclusions: D240G mutation leads to an increase in ceftazidime resistance in CTX-M-producing XL1blue clones of all clones when expressed under the same genetic background, and cefotaxime resistance was significantly higher than ceftazidime resistance. CTX-M-12 and -96 producing E. coli K12 and E. coli lacking OmpC clones remained susceptible to ceftazidime, while those lacking OmpF were resistant, suggesting the importance of this porin in the entrance of ceftazidime. In absence of both OmpF and OmpC, resistance to ceftazidime is slightly higher. In absence of efficient kinetic hydrolysis of ceftazidime, the term ceftazidimases for these enzymes needs to be avoided as resistance arises by a combination of different mechanisms.