Although no natural hybrids are known, fertile laboratory hybrids were readily obtained from Xenotoca eiseni and X. melanosoma in forced crosses in which a conspecific mate was not available. In choice crosses with males and females of both species, sympatric stocks never hybridized, but allopatric fishes frequently mismated. Ethological data reveal differences in courtship behavior and discriminatory ability in sympatric fishes not observed for allopatric conspecific stocks nor in fish from two populations of the congener X. variata. These differences, which prevent interbreeding, are offered as evidence for the perfection of premating isolating mechanisms in sympatry. Hybrid inferiority, essential for divergence in sympatry, was reflected in reduced survival and inability to compete for mates.