Carbapenem resistance in Escherichia coli associated with plasmid-determined CMY-4 beta-lactamase production and loss of an outer membrane protein

Three cefoxitin-resistant Escherichia coli isolates from stool specimens of a patient with leukemia were either resistant, intermediate, or sensitive to imipenem, Conjugation experiments showed that cefoxitin resistance, but not imipenem resistance, was transferable. All isolates were shown by isoel ectric focusing to produce two beta-lactamases with isoelectric points of 5 .4 (TEM-1, confirmed by sequencing of a PCR product) and >8.5 (consistent w ith a class C beta-lactamase), The gene coding for the unknown beta-lactama se was cloned and sequenced and revealed an enzyme which had 99.9% sequence identity with the plasmid-determined class C beta-lactamase CMY-2, The clo ned beta-lactamase gene differed from bla(CMY-2) at one nucleotide position that resulted in an amino acid change, tryptophan to arginine at position 221, We propose that this enzyme be designated CMY-4, Both the imipenem-res istant and -intermediate isolates lacked a 38-kDa outer membrane protein (O MP) that was present in the imipenem-sensitive isolate. The lack of an OR;I P alone did not explain the difference in carbapenem susceptibilities obser ved. However, measurement of beta-lactamase activities (including measureme nts under conditions where TEM-1 beta-lactamase was inhibited) indicated th at the imipenem-intermediate isolate expressed six- to eightfold less beta- lactamase than did the other isolates. This study illustrates that carbapen em resistance in E. call can arise from high-level expression of plasmid-me diated class C beta-lactamase combined with an OMP deficiency. Furthermore, in the presence of an OMP deficiency, the level of expression of a plasmid -mediated class C beta-lactamase is an important factor in determining whet her E, coli isolates are fully resistant to carbapenems.