Objectives To investigate the genotypic resistance of integron-carrying Salmonella Typhimurium isolates from horses and their genetic relationship. Methods Sixty-one Salmonella isolates were screened for the presence of class 1 integrons by PCR. The gene cassettes of integron-positive isolates were detected by PCR, restriction fragment length polymorphism typing, and sequencing. The potential for the transfer of resistance determinants was investigated by conjugation experiments. The presence of Salmonella genomic island 1 (SGI1) or its variants was studied by PCR and nucleotide sequencing. PFGE was used to genotype the isolates. Results Eight distinct Xba I-PFGE profiles and seven integron types were observed among 26 integron-carrying Salmonella Typhimurium isolates. The gene cassettes detected were dfrA1 , dfrA7 , dfrA14 , aadA1 , aadA2 , aadB and bla PSE . A rare type of integron found in nine isolates carried the dfrA14 and aadA1 gene cassettes. Twelve Salmonella Typhimurium DT104 isolates contained SGI1 or one of its variants (SGI1, SGI1-B and SGI1-C). A novel variant of SGI1, designated SGI1-M, was identified in one isolate in which the aadA2 gene of SGI1 was replaced by the aadB gene. Transfer of integrons and antimicrobial resistance determinants to Escherichia coli K12 via conjugation was possible with nine isolates. Resistance to fluoroquinolones in nine isolates was caused by mutations in the gyrA gene leading to the amino acid changes Ser-83 → Ala and Asp-87 → Asn. Conclusions The integron-positive clinical Salmonella Typhimurium isolates from horses belong to distinct strains. The data demonstrate the capability of Salmonella Typhimurium to acquire additional antibiotic resistance determinants and underline the need for the prudent use of antimicrobials.