Characterization of AmpC-mediated resistance in clinical Salmonella isolates recovered from humans during the period 1992 to 2003 in England and Wales

Abstract

The increase in AmpC-mediated resistance in salmonellae constitutes a serious public health concern, since these enzymes confer resistance to a wide range of beta-lactams. One hundred six isolates were selected from 278,308 Salmonella isolates based on resistance to ampicillin and cephalosporins and were subjected to further characterization. Nine isolates had a cefoxitin inhibition diameter < or = 17 mm and were proven to be AmpC positive by multiplex PCR. Sequence analysis revealed the presence of bla(DHA-1), bla(CMY-2), and bla(CMY-4) genes. All nine isolates presented different pulsed-field gel electrophoresis restriction profiles. The AmpC genetic determinants were present in transferable plasmids of around 11, 42, 70, 98, and 99 MDa. A combination of size and restriction fragment length polymorphism (RFLP) analysis showed that all the bla(CMY) plasmids investigated in our study were different, which suggests that bla(CMY) may be located in different plasmid environments. Some United Kingdom isolates linked to foreign travel showed RFLP plasmid patterns consistent with plasmids previously seen in the United States, which suggests that bla(CMY-2) has also been disseminated through plasmid transfer. The fact that two of the domestically acquired United Kingdom isolates presented previously unseen RFLP plasmid patterns could indicate that these strains have followed routes different from those prevalent in North America or other parts of the world. This study represents the first report of bla(CMY) genes in Salmonella isolates in the United Kingdom and the first report of CMY-4 in Salmonella enterica serotype Senftenberg worldwide.

FIG. 1.

Image generated by Bionumerics software showing the XbaI PFGE restriction profile for nine Salmonella AmpC-positive isolates. The sizes (in kb) of the fragments generated were calculated by comparison to a Salmonella serotype Braenderup PulseNet universal control strain (H9812). The clustering analysis was conducted with the following parameters: band tolerance, 1%; band optimization, 0%.

FIG. 2.

bla_CMY-2 Southern hybridization after HpaI digestion of bla_CMY-2-harboring plasmids from Salmonella isolates. The images were analyzed using Bionumerics software, and the sizes of the hybridizing fragments were calculated by comparison to DNA molecular weight marker II (Roche).