Devils Hole in the context of National Wildlife Refuge

Pupfish are a group of small killifish belonging to ten genera of the family Cyprinodontidae of ray-finned fish. Pupfish are especially noted for being found in extreme and isolated situations. They are primarily found in North America, South America, and the Caribbean region. As of August 2006, 120 nominal species and 9 subspecies were known. Several pupfish species are extinct and most extant species are listed. In the U.S., the most well-known pupfish species may be the Devils Hole pupfish, native to Devils Hole on the Nevada side of Death Valley National Park. Since 1995 the Devils Hole pupfish has been in a nearly steady decline, where it was close to extinction at 35–68 fish in 2013.

The common name is said to derive from the mating habits of the males, whose activities vaguely resemble puppies at play; Carl L. Hubbs, a prominent ichthyologist and one of the first people to take an interest in them, coined the name after he observed their "playful" circling and tussling, which is actually the aggressive behavior of territorial males.

The Devils Hole pupfish (Cyprinodon diabolis) is a critically endangered species of the family Cyprinodontidae (pupfishes) found only in Devils Hole, a water-filled cavern in the US state of Nevada. It was first described as a species in 1930 and is most closely related to C. nevadensis and the Death Valley pupfish (C. salinus). The age of the species is unknown, with differing analyses offering ranges between one thousand and sixty thousand years. It is a small fish, with maximum lengths of up to 30 mm (1.2 in). Individuals vary in coloration based on age and sex: males are bright metallic blue while females and juveniles are more yellow. A defining trait of this species is its lack of pelvic fins. The pupfish consumes nearly every available food resource at Devils Hole, including beetles, snails, algae, and freshwater crustaceans, with diet varying throughout the year. It is preyed on by the predaceous diving beetle species Neoclypeodytes cinctellus, which was first observed in Devils Hole in 1999 or 2000. Reproduction occurs year-round, with spikes in the spring and fall. Females produce few eggs and the survivorship from egg to adult is low. Individuals live 10–14 months.

Devils Hole is more than 130 m (430 ft) deep, though pupfish are only found in the upper 24 m (80 ft). The water is a constant temperature of 33 °C (91 °F) and dissolved oxygen levels are low. A small, shallowly submerged rock shelf provides critical feeding and spawning habitat for the pupfish. Nearby agricultural irrigation in the 1960s and 1970s caused the water to drop in Devils Hole, resulting in less and less of the shelf remaining submerged. Several court cases ensued, resulting in the Supreme Court case Cappaert v. United States, which determined that the preservation of Devils Hole as a National Monument in 1952 implicitly included preservation of adequate groundwater to maintain the scientific value of the pool and its fauna. Other threats faced by the species include flash floods, earthquakes, and vandalism.