Salmonella isolates resistant to oxyiminocephalosporins due to the production of extended-spectrum β-lactamases (ESBLs) have emerged worldwide since 1992 (2). This has caused concern since cephalosporins are drugs of choice for the treatment of salmonellosis in children, to whom fluoroquinolones must not be administered. Different blaSHV, blaTEM, blaCTX, and blaCMY genes have encoded ESBL production in Salmonella (5, 8, 9). The same genes have been shown to encode resistance in different countries, which could indicate a global spread of these genes. In Denmark ESBL-producing isolates of Salmonella or other gram-negative bacteria have not previously been isolated from food-producing animals. However, in August 2003 the first ESBL-producing Salmonella isolate was isolated from the intestine of a boar imported from Canada. The boar was imported in May 2003 by a Danish breeding station and suddenly died on 4 August.
The isolate was serotyped as Salmonella enterica serovar Heidelberg and was examined for antimicrobial susceptibility by MIC determinations (6). The β -lactam resistance was not transferable to E. coli recipients, whereas transfer was obtained in high frequencies to nalidixic acid-resistant S. enterica serovar Heidelberg, serovar Typhimurium, and serovar Dublin recipients. MICs found for the donor, recipients, and transconjugants as well as detected resistance genes are given in Table 1. Plasmid profiling and hybridization showed that the blaCMY-2 gene was located on a >60-kb plasmid. The pulsed-field gel electrophoresis (PFGE) profile of the isolate was compared to those of 10 other serovar Heidelberg isolates from food products or animals in Denmark and four isolates from humans. PFGE was performed by using XbaI and BlnI as restriction enzymes according to PulseNet protocol, and its results were compared to those of the PulseNet database. No identical isolates were found.
TABLE 1.
ICs found for and occurrence of resistance genes in Serovar Heidelberg isolated from an imported boar in Denmark and MICs found for recipients used for conjugation and transconjugants
| Antimicrobial agent | MIC (μg/ml)
|
Gene encoding resistance | ||||||
|---|---|---|---|---|---|---|---|---|
| Serovar Heidelberg donor | Serovar Heidelberg recipient | Serovar Typhimurium recipient | Serovar Dublin recipient | Serovar Heidelberg transconjugant | Serovar Typhimurium transconjugant | Serovar Dublin transconjugant | ||
| Amoxicillin + Cl | >32b | ≤2 | ≤2 | ≤2 | >32 | >32 | 32 | blaCMY-2 |
| Ampicillin | >32 | ≤1 | ≤1 | ≤1 | >32 | >32 | >32 | blaCMY-2 |
| Apramycin | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | |
| Cefalothin | >64 | ≤2 | ≤2 | ≤2 | >64 | >64 | >64 | blaCMY-2 |
| Cefotaxime | 16 | ≤0.125 | ≤0.125 | ≤0.125 | 8 | 8 | 8 | blaCMY-2 |
| Cefoxitin | 32 | 0.25 | 0.25 | 0.25 | 32 | 32 | 32 | blaCMY-2 |
| Ceftiofur | >8 | ≤0.5 | ≤0.5 | ≤0.5 | >8 | >8 | >8 | blaCMY-2 |
| Ceftriaxone | 16 | ≤0.125 | ≤0.125 | ≤0.125 | 16 | 16 | 16 | blaCMY-2 |
| Cefurixime | 64 | 8 | 8 | 8 | 64 | 64 | 64 | blaCMY-2 |
| Chloramphenicol | >64 | 8 | 4 | 4 | 64 | >64 | >64 | floR |
| Ciprofloxacin | ≤0.03 | 0.25 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | |
| Colistin | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | |
| Florfenicol | >64 | 4 | 4 | 4 | >64 | >64 | >64 | floR |
| Gentamicin | ≤1 | ≤1 | ≤1 | ≤1 | ≤1 | ≤1 | ≤1 | |
| Nalidixic acid | ≤8 | >128 | >128 | >128 | >128 | >128 | >128 | |
| Neomycin | >32 | ≤2 | ≤2 | ≤2 | ≤2 | ≤2 | ≤2 | aphA-2 |
| Spectinomycin | >128 | 64 | 32 | 32 | >128 | >128 | >128 | aadA |
| Streptomycin | >64 | 8 | 8 | ≤4 | >64 | >64 | >64 | aadA |
| Sulphamethoxazole | >1,024 | ≤64 | ≤64 | ≤64 | >1,024 | >1,024 | >1,024 | NDa |
| Tetracycline | >32 | ≤2 | ≤2 | ≤2 | >32 | >32 | >32 | tet(A) |
| Trimethoprim | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | ≤4 | |
ND, not determined.
Boldface indicates resistance according to NCCLS guidelines (6).
Transferable plasmids (60 to 160 kb) with resistance to ampicillin, cefoxitin, ceftiofur, cephalothin, florfenicol, streptomycin, sulfisoxazole, and tetracycline have previously been detected in Salmonella isolates from the animal and human populations in Canada (1) and the United States (4, 9, 10). The blaCMY-2 gene has so far not been reported in serovar Heidelberg from other countries. However, the location on similar plasmids could indicate that the blaCMY-2 gene in the isolates from Canada, Denmark, and the United States is located on the same transferable genetic resistance element.
Serovar Heidelberg is one of the most common serovars causing infections in humans in the United States (7). This serovar is not so far of any major importance in Denmark but clearly has the potential to become an important cause of infections in humans. In addition, since the resistance is located on a transferable plasmid, this resistance might easily spread once it is introduced into an animal population.
The present observation showed that a Salmonella isolate with multiresistance was imported by Denmark through the market of live animals. International spread of multiresistant clones has previously been observed (3), and there are several potential routes where this might occur. Trading in live animals provides a very efficient way to spread both pathogenic clones and resistance genes. Breeding animals constitute the top of the production pyramid, and from these herds clones and genes might easily spread to all other animals in the production system. Recent studies from the United States indicate that similar plasmids with the blaCMY-2 genes can be isolated from several sources, including cattle and swine, indicating that plasmids and genes have spread. Salmonella isolates with ampC-mediated β-lactam resistance have so far not become common in Europe. However, in 2003 an outbreak of serovar Newport with blaCMY-2, associated with eating horse meat, was detected in France (http://www.eurosurveillance.org/ew/2003/030703.asp#2). The global spread of ampC-mediated resistance in Salmonella and other Enterobacteriaceae is one of the emerging problems that we presently face. Thus, studies determining how this spread occurs are urgently needed.
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