This study aimed to investigate the effects on nutrient intake and digestibility, milk yield (MY) and composition, milk fatty acids profile, and microbial protein synthesis caused by feeding lactating dairy cows four different forage cactus genotypes. Eight Girolando cows (5/8 Holstein × 3/8 Gyr), weighing 490 ± 69.0 kg (means ± standard deviation), and producing 15.5 ± 1.0 kg/d of milk during pretrial were distributed to two contemporaneous 4 × 4 Latin squares. The cows were fed a total mixed ration composed of sorghum silage (385 g/kg of dry matter (DM)), concentrated mix (175 g/kg DM), and forage cactus (440 g/kg DM). The experimental treatments consisted of different cactus genotypes, such as Gigante cactus (GC), Miúda cactus (MC), IPA Sertânia cactus (SC), and Orelha de Elefante Mexicana cactus (OEMC). The feeding of MC provided a higher intake of DM, organic matter (OM), and total digestible nutrients, as well as higher MY, energy-corrected milk, and microbial protein synthesis in comparison with those resulting from the other genotypes tested. The GC promoted lower DM and OM, and the apparent digestibility of neutral detergent fiber. The cows fed with OEMC showed lower MY and milk protein yield and content, and higher unsaturated over saturated fatty acid ratio in milk. Miúda forage cactus increased nutrient intake, digestibility of DM and OM, and microbial synthesis without impairing the milk fatty acid profile.