The effect of a grain-based concentrate supplement on fatty acid (FA) intake and concentration of milk FA in early lactation was investigated in grazing dairy cows that differed in their country of origin and in their estimated breeding value for milk yield. It was hypothesized that Holstein-Friesian cows of North American (NA) origin would produce milk lower in milk fat than those of New Zealand (NZ) origin, and that the difference would be associated with lower de novo synthesis of FA. In comparison, increasing the intake of concentrates should have the same effect on the FA composition of the milk from both strains. Fifty-four cows were randomly assigned in a factorial arrangement to treatments including 3 amounts of concentrate daily [0, 3, and 6 kg of dry matter (DM)/cow] and the 2 strains. The barley/steam-flaked corn concentrate contained 3.5% DM FA, with C18:2, C16:0, and C18:1 contributing 48, 18, and 16% of the total FA. The pasture consumed by the cows contained 4.6% DM FA with C18:3, C16:0, and C18:1 contributing 51, 10, and 10% of the FA, respectively. Pasture DM intake decreased linearly with supplementation, but total DM intake was not different between concentrate or strain treatments, averaging 16.2 kg of DM/cow, with cows consuming 720 g of total FA/d. Cows of the NA strain had lesser concentrations of milk fat compared with NZ cows (3.58 vs. 3.95%). Milk fat from the NA cows had lesser concentrations of C6:0, C8:0, C10:0, C12:0, C14:0, and C16:0, and greater concentrations of cis-9 C18:1, C18:2, and cis-9, trans-11 C18:2, than NZ cows. These changes indicated that in milk from NA cows had a lesser concentration of de novo synthesized FA and a greater concentration of FA of dietary origin. Milk fat concentration was not affected by concentrate supplementation. Increasing concentrate intake resulted in linear increases in the concentrations of C10:0, C12:0, C14:0, and C18:2 FA in milk fat, and a linear decrease in the concentration of C4:0 FA. The combination of NA cows fed pasture alone resulted in a FA composition of milk that was potentially most beneficial from a human health perspective; however, this would need to be balanced against other aspects of the productivity of these animals.