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Six multiply resistant isolates of Salmonella typhimurium var. copenhagen with high-level resistance to fluoroquinolones (e.g., MIC of ciprofloxacin: 32 micrograms/ml) were isolated from human patients (n = 3) and from cattle (n = 3). The isolates were examined by complementation tests using a set of broad-host-range plasmids, which carry either the gyrA+ or the gyrB+ genes or a combination of both from Escherichia coli K-12. The results indicated a combination of gyrA and gyrB mutations in all isolates. Subsequent direct sequencing of PCR-generated internal DNA fragments of gyrA revealed an identical double mutation in all six isolates (Ser-83-->Ala and Asp-87-->Asn). In addition, the results of phenotypic (i.e., phagetype, biotype, serotype) and genotypic characterization [i.e., ribotyping and polymerase chain reaction fingerprinting (PCR-fingerprinting)] were identical for all six isolates and were distinguishable from a quinolone-susceptible strain of the same serovar and an unrelated isolate of S. typhimurium. These data indicate the clonal identity of the fluoroquinolone-resistant strains of S. typhimurium isolated from men and cattle in Germany.