Insectivore-like animals are traditionally believed among the first eutherian mammals that have appeared on the earth. The modern insectivores are thus crucial for understanding the systematics and phylogeny of eutherian mammals as a whole. Here cross-species chromosome painting, with probes derived from flow-sorted chromosomes of human, was used to delimit the homologous chromosomal segments in two Soricidae species, the common shrew (Sorex araneus, 2n = 20/21), and Asiatic short-tailed shrew (Blarinella griselda, 2n = 44), and one Erinaceidae species, the shrew-hedgehog (Neotetracus sinensis, 2n = 32), and human. We report herewith the first comparative maps for the Asiatic short-tailed shrew and the shrew-hedgehog, in addition to a refined comparative map for the common shrew. In total, the 22 human autosomal paints detected 40, 51 and 58 evolutionarily conserved segments in the genomes of common shrew, Asiatic short-tailed shrew, and shrew-hedgehog, respectively, demonstrating that the common shrew has retained a conserved genome organization while the Asiatic short-tailed shrew and shrew-hedgehog have relatively rearranged genomes. In addition to confirming the existence of such ancestral human segmental combinations as HSA 3/21, 12/22, 14/15 and 7/16 that are shared by most eutherian mammals, our study reveals a shared human segmental combination, HSA 4/20, that could phylogenetically unite the Eulipotyphlan (i.e., the core insectivores) species. Our results provide cytogenetic evidence for the polyphyly of the order Insectivora and additional data for the eventual reconstruction of the ancestral eutherian karyotype.