All killifish lay eggs, unlike the live-bearing topminnows of the related family Poeciliidae (see live-bearer). Certain South American and African killifish live in pools subject to an annual drying out and bury their eggs in the bottom, where they remain dormant until water refills the pond. These fishes are called annual fishes because their life cycle from birth to mating and subsequent death usually does not exceed one year.

Many killifish, including some of the annuals, are attractively coloured and are kept in home aquariums. A number of genera have been popularized, among them Aphyosemion (lyretails and others), Epiplatys, and Rivulus. Killifish are also of value as bait for anglers and as a form of mosquito control because they eat the larvae of these insect pests.

North American killifishes, sometimes called pupfishes (Cyprinodon), inhabit the California coastal waters and the perimeters of certain western salt lakes. One well-known species, the tiny (1.5-inch 【0.6-centimetre】) Tecopa pupfish (C. nevadensis), long-considered endangered, was declared extinct in 1981. Other pupfish are on the official list of endangered species.