Epidemiological survey of amoxicillin-clavulanate resistance and corresponding molecular mechanisms in Escherichia coli isolates in France: new genetic features of bla(TEM) genes.

Amoxicillin-clavulanate resistance (MIC >16 microg/ml) and the corresponding molecular mechanisms were prospectively studied in Escherichia coli over a 3-year period (1996 to 1998) in 14 French hospitals. The overall frequency of resistant E. coli isolates remained stable at about 5% over this period. The highest frequency of resistant isolates (10 to 15%) was observed, independently of the year, among E. coli isolated from lower respiratory tract samples, and the isolation rate of resistant strains was significantly higher in surgical wards than in medical wards in 1998 (7.8 versus 2.8%). The two most frequent mechanisms of resistance for the 3 years were the hyperproduction of the chromosomal class C beta-lactamase (48, 38.4, and 39.7%) and the production of inhibitor-resistant TEM (IRT) enzymes (30.4, 37.2, and 41.2%). By using the single-strand conformational polymorphism-PCR technique and sequencing methods, we determined that 59 IRT enzymes corresponded to previously described IRT enzymes whereas 8 were new. Three of these new enzymes derived from TEM-1 by only one amino acid substitution (Ser130Gly, Arg244Gly, and Asn276Asp), whereas three others derived by two amino acid substitutions (Met69Leu and Arg244Ser, Met69Leu and Ile127Val, and Met69Val and Arg275Gln). The two remaining new IRTs showed three amino acid substitutions (Met69Val, Trp165Arg, and Asn276Asp and Met69Ile, Trp165Cys, and Arg275Gln). New genetic features were also found in bla(TEM) genes, namely, bla(TEM-1B) with either the promoters Pa and Pb, P4, or a promoter displaying a C-->G transversion at position 3 of the -35 consensus sequence and new bla(TEM) genes, notably one encoding TEM-1 but possessing the silent mutations originally described in bla(TEM-2) and then in some bla(TEM)-encoding IRT enzymes.