The two most common haemotropic Mycoplasma of cats, Mycoplasma haemofelis and ‘ Candidatus Mycoplasma haemominutum’ have been identified using molecular techniques in all continents, except Antarctica. We report the first molecular characterization in South America of a dual infection with M haemofelis and ‘ Candidatus Mycoplasma haemominutum’ in three domestic cats. The 16S ribosomal RNA gene was amplified in three anaemic cats in which haemoplasma organisms were seen attached to the erythrocytes in the peripheral blood smear. Bands of the expected size for M haemofelis and ‘ Candidatus Mycoplasma haemominutum’ were observed in all three cats. The 393 bp segment of one of the amplicons had a similarity value of 100% to M haemofelis, whereas the other amplicon, a 192 bp segment, was 100% similar to ‘ Candidatus Mycoplasma haemominutum’. After diagnosis, two cats received blood transfusion and they were all treated with doxycycline. All three cats recovered uneventfully.