Reciprocal chromosome painting and G-banding were used to compare the karyotypes of three Australian marsupials (Sminthopsis crassicaudata, Macropus eugenii, Trichosurus vulpecula) and one South American marsupial (Monodelphis domestica). The results revealed only a limited number of rearrangements between these species and that the four karyotypes can be described as different combinations of fifteen conserved segments. Five chromosomes are totally conserved between M. domestica (pairs 1, 2, 5, 8 and the X) and the presumed 2n = 14 Australian ancestral karyotype, while M. domestica pairs 3 and 6 and 4 and 7 would have been involved in fusion/fission rearrangements. Chromosome comparisons are presented in a chromosome homology map. Although the species studied diverged 70 million years ago, the karyotype of Monodelphis domestica is highly conserved in relation to those of Australian marsupials.