Programme Hospitalier Recherche Clinique, Institut Pasteur, Inserm, French Public Health Agency.

Escherichia coli usually produces only very small amounts of a constitutive AmpC beta-lactamase, but clinical strains overproducing this enzyme have been isolated. Three different ampC promoters of E. coli clinical strains were cloned upstream of the chloramphenicol acetyltransferase (CAT) gene in the pKK232-8 reporter plasmid and their relative strengths were compared by two different methods. The strength of the promoters from AmpC hyperproducers was 70- to 120-fold higher than those from a low-level AmpC producer. One of the strong promoters, which differs from strain K12 at bases -88, -82, -42, -18, -1 and +58, was mutated to abolish the -42 mutation. This change resulted in a 43-fold decrease in CAT concentration. In another promoter, with eight different mutations at positions -88, -82, -32, -18, -1, +5, +24 and +58, the -32T-->A transversion, which created perfect homology with the -35 consensus sequence, was reverted; this led to a 13-fold decrease in CAT concentration. The -42 and -32 mutations play an important role in E. coli resistance to beta-lactams by increasing ampC transcription.

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During a 1-year survey of Shiga toxin-producing Escherichia coli (STEC) prevalence in central France, 2,143 samples were investigated by PCR for Shiga toxin-encoding genes. A total of 330 (70%) of 471 fecal samples collected from healthy cattle at the Clermont-Ferrand slaughterhouse, 47 (11%) of 411 beef samples, 60 (10%) of 603 cheese samples, and 19 (3%) of 658 stool specimens from hospitalized children with and without diarrhea were positive for thestx gene(s). A STEC strain was isolated from 34% (162 of 471) of bovine feces, 4% (16 of 411) of beef samples, 1% (5 of 603) of cheese samples, and 1.5% (10 of 658) of stool specimens. Of the 220 STEC strains isolated, 34 (15%) harbored thestx 1 gene, 116 (53%) harbored thestx 2 gene, and 70 (32%) carried both thestx 1 and stx 2 genes. However, 32 (14.5%) were not cytotoxic for Vero cells. Theeae gene, found in 12 (5%) of the 220 strains, was significantly associated with the stx 1 gene and with isolates from children. Sequences homologous to ehxAwere found in 102 (46%) of the 220 strains. Thirteen serotypes, OX3:H2, O113:H21, O113:H4, OX3:H21, O6:H10, OX178:H19, O171:H2, O46:H38, O172:H21, O22:H16, O91:H10, O91:H21, and O22:H8, accounted for 102 (55%) of 186 typeable isolates, and only one strain (0.5% of the 186 STEC isolates from cattle), belonged to the O157:H7 serotype. We showed that the majority of the STEC isolates from cattle, beef, and cheese were not likely to be pathogenic for humans and that the STEC strains isolated from children in this study were probably not responsible for diarrheal disease. Finally, the strains associated with hemolytic-uremic syndrome in the same geographical area were shown to belong to particular subsets of the STEC population found in the bovine reservoir.

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