Detailed karyological surveys of the ant Myrmecia pilosula species group, which is characterized by the lowest chromosome number in higher organisms (2n=2), were attempted. We revealed that this species has developed highly complicated chromosomal polymorphisms. Their chromosome numbers are in the range 2n=2, 3, and 4, and six polymorphic chromosomes are involved, i.e., two for chromosome 1 (denoted as SM1 and ST1), three for chromosome 2 (A2, A′2, and M2), and M(1+2) for the 2n=2 karyotype. We suggested that these chromosomes were induced from a pseudo-acrocentric (A1M) and A2 as follows: (1) A1M→SM1 or ST1 by two independent pericentric inversions; (2) A2→A′2→M2 by chromosomal gap insertion and centromere shift; and (3) ST1+A2→M(1+2) by telomere fusion, where (3) means that the 2n=2 karyotype was derived secondarily from a 2n=4 karyotype. It is a noteworthy finding that active nucleolus organizer (NOR) sites, in terms of silver staining, are tightly linked with the centromere in this species, and that both the centromere and NOR of A2 were inactivated after the telomere fusion.