Recent morphological, biochemical, ecological and reproductive studies have revealed that there are four sympatric biological species of sea urchins in Echinometra from Okinawa, which are currently referred to as Echinometra species A, B, C and D (e.g. Uehara et al., 1990,1991; Arakaki & Uehara, 1991; Matsuoka & Hatanaka, 1991; Palumbi & Metz, 1991; Nishihira et al., 1991). Of these four species, E. sp. A (Ea) and E. sp. C (Ec), while being most closely related to each other genetically (Matsuoka & Hatanaka, 1991), can also be distinguished from each other by differences in adult morphology and microhabitat preference. No studies have addressed the mechanisms that maintain the independence of these species despite their close affinity and sympatry.

Experimental hybridisation conducted between two genetically very divergent species of Okinawan Echinometra, E. sp. A (Ea) and E. sp. D (Ed), revealed the presence of prezygotic isolation (Asian & Uehara, 1997). To test whether the closely related Okinawan Echinometra spp. are isolated by pre- or postzygotic mechanisms, we carried out experimental hybridisation by using much closer species, Ea and Ec, through a series of cross-fertilisation experiments and rearing of the resulting hybrids. The percentage of successful heterogametic fertilisation was high when eggs of Ec and sperms of Ea were involved, whereas it was considerably lower with eggs of Ea and sperms of Ec. This reduction, at least among some heterogametic crosses, indicates the presence of a gamete recognition protein binding system, as reported by Metz et al. (1994) and Metz & Palumbi (1996), which might eventually lead to gametic incompatibility and reproductive isolation.