A novel tetracycline resistance determinant named Tet 39 was found in unrelated Acinetobacter strains isolated from freshwater trout farms (n=4) and sewage (n=6) in Denmark, and from a clinical specimen in the Netherlands (n=1). The determinant was located on transferable plasmids and consisted of tetA(39), most likely conferring resistance by active efflux, and a putative repressor gene tetR(39).