Brood defense behaviour of parental Midas cichlids, Amphilophus xiloaensis, was observed in Lake Xiloá, Nicaragua. The research was conducted during two study periods separated by 22 years, 1972–73 and 1995. Role-differentiation was observed between the male and female during brood defense. The male defended the territory from adult conspecifics, while the female defended the brood from predators. These observations were consistent in two different study periods 22 years apart and at two different locations within the lake. Attack rates increased as the brood matured, contradicting laboratory findings on this species. These findings reinforced the use of brood defense behavior as a possible diagnostic tool to differentiate sibling species. The pattern of brood care and role differentiation remained stable over many generations, and should be a useful character for distinguishing sibling species.