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Twenty-five MRSA ST398 isolates from cases of bovine clinical mastitis and two isolates from farm personnel collected from 17 dairy farms in Germany were investigated for genetic relatedness, antimicrobial resistance and virulence properties.Genomic relationships were determined by ApaI PFGE, spa typing, SCCmec typing and dru typing. Antimicrobial resistance phenotypes were determined by broth microdilution. Resistance and virulence genes were detected via a diagnostic DNA microarray and specific PCRs.Nine major ApaI PFGE patterns were detected. Three spa types (t011, t034 and t2576) and two SCCmec types (IV and V) were identified. Five different dru types were seen with dt11a being predominant. All isolates were negative for Panton–Valentine leucocidin, enterotoxin and exfoliative toxin genes. Ten resistance patterns were observed with 11 (40.7%) isolates being resistant to only β-lactam antibiotics and tetracyclines. Several resistance genes were detected: blaZ (penicillin resistance); tet(M), tet(K) and tet(L) (tetracycline resistance); erm(A), erm(B), erm(C) and erm(T) (macrolide/lincosamide/streptogramin B resistance); aacA–aphD, aphA3, aadD and spc (aminoglycoside or aminocyclitol resistance); fexA (phenicol resistance); dfrK (trimethoprim resistance); and vga(A) and vga(C) (pleuromutilin/lincosamide/streptogramin A resistance). The two human isolates were indistinguishable in their genotypic and phenotypic characteristics from the mastitis isolates of the same farm.As previously described for ST398 from swine, isolates of this sequence type from cases of bovine mastitis also demonstrated a high degree of variability when ApaI PFGE profiles and other genotypic and phenotypic characteristics were compared. A uniform virulence gene pattern appeared to be conserved between ST398 isolates from both animal species.