The first genome sequences of desert pupfish from the American Southwest, including the rare Devils Hole pupfish, have been assembled by researchers. The genomes of the eight Devils Hole pupfish sequenced contained an astonishing number of identical stretches of DNA, amounting to 58% of the genome, making them among the most inbred vertebrates known. Surprisingly, five of the 15 gene deletions involved adaptation to hypoxia, or low oxygen levels, which are common in the pupfish’s environment.
The Devil’s Hole pupfish, as the name suggests, lives in a truly hellish environment.
263 of them are confined to a single deep limestone cave in Nevada’s Mojave Desert, where the water temperature is around 93 degrees Fahrenheit year round, food resources are scarce, and oxygen levels are so low that most other fish would die immediately. The pupfish, Cyprinodon diabolis, has the smallest habitat of any vertebrate known to science.
New research has revealed the extreme impact that these harsh and isolated conditions have had on the genetic diversity of this fish.
In a paper published this week in the journal Proceedings of the Royal Society B, University of California, Berkeley, biologists report the first complete genome sequences of eight pupfish species from the American Southwest — 30 individuals in all, including eight Devils Hole pupfish. Astoundingly, the Devils Hole pupfish is so inbred that 58% of the genomes of these eight individuals are identical, on average.
High levels of inbreeding are associated with a higher risk of extinction, and the inbreeding in the Devils Hole pupfish is equal to or more severe than levels reported so far in other isolated natural populations, such as the Isle Royale wolves in Michigan, mountain gorillas in Africa and Indian tigers.Christopher Martin
“High levels of inbreeding are associated with a higher risk of extinction, and the inbreeding in the Devils Hole pupfish is equal to or more severe than levels reported so far in other isolated natural populations, such as the Isle Royale wolves in Michigan, mountain gorillas in Africa and Indian tigers,” said lead researcher Christopher Martin, UC Berkeley associate professor of integrative biology and curator of ichthyology in the campus’s Museum of Vertebrate Zoology. “Although we were not able to directly measure fitness, the increased inbreeding in these pupfish likely results in a substantial reduction in fitness.”
The researchers discovered that other pupfish species are also inbred, but only 10% to 30% of their genomes are identical.
According to the study’s lead author, graduate student David Tian, the level of inbreeding in Devils Hole pupfish is equivalent to what would happen if four to five generations of siblings mated with one another. This tends to burn in or fix harmful mutations rather than weed them out, potentially dooming a population to extinction through mutational meltdown. The Devils Hole pupfish species is currently doing well in the wild and in captive or “refuge” populations, but the species’ low genetic diversity may pose a problem as climate change and human impacts become more severe.
In the face of these potential threats, the new genome sequences will help scientists and conservationists assess the health of native pupfish populations and potentially intervene in refuge populations to increase the genetic diversity of these species – the Devils Hole pupfish, in particular.
Population decline and rescue
Pupfish species are found all over the world and prefer isolated lakes and springs, often in harsh conditions that would kill most fish. In California and Nevada, about 30 species live in warm, salty desert springs and streams. Martin has studied various pupfish populations, including several on the Bahamas’ San Salvador Island, to better understand the genetics of their adaptation to extreme conditions and unusual ecological niches.
The Devils Hole pupfish, on the other hand, is unique in its small range and perilous existence, according to Martin, making conservationists concerned about its fluctuating population in the wild.
“Part of the question about these declines is whether they may be due to the genetic health of the population,” Martin said. “Maybe the declines are because there are harmful mutations that have become fixed because the population is so small.”
The small population is partly a result of human incursions into their habitat, Martin noted. Local ranchers and developers pumped groundwater in the region in the 1960s and ’70s that drastically reduced the water level in Devils Hole, leading to a drop in population levels. A 1976 Supreme Court ruling that allowed the federal government to limit groundwater pumping saved Devils Hole and the resident population, while captive breeding at a nearby 100,000-gallon pool in the Ash Meadows National Wildlife Refuge rescued the species. Nevertheless, a decline in the 1990s led the wild population to its nadir in 2013: 35 individuals. The wild population has since recovered, while the refuge population has ballooned to about 400, twice the wild population.
However, humans are not entirely to blame for the Devils Hole pupfish’s lack of genetic diversity. The UC Berkeley team also sequenced the genome of a pupfish that was collected in 1980 and is now housed at the University of Michigan. It exhibited inbreeding and a lack of genetic diversity similar to that found in recently collected individuals, the majority of whom died naturally. This implies that the pupfish has likely experienced frequent population bottlenecks over hundreds, if not thousands, of years.
Martin and Tian discovered that 15 genes have vanished entirely from the Devils Hole pupfish genome as a result of this. Five of them appear to play a role in adaptation to low-oxygen or hypoxic environments.
“These deletions are a paradox, because this is a habitat where you’re most exposed to hypoxia,” Martin said. “It could have something to do with the stability of the habitat over time. But it looks to us like the hypoxia pathway is broken. Once you break one gene, it doesn’t really matter if you break additional genes in that regulatory pathway. Our future work is to actually look at what these deletions do. Do they increase tolerance of hypoxia? Do they decrease tolerance of hypoxia? I think those two scenarios are equally plausible at this time.”
Selective breeding within a captive population of Devils Hole pupfish could help increase the diversity and perhaps save the species from eventual extinction, he said. And to restore genes already lost, CRISPR genome editing could add them back.
The fact that the genome of the fish collected in 1980 was about as inbred as today’s fish is “maybe good news,” according to Martin, because the population has historically been highly inbred with very low genetic diversity, implying that the recent decline in the ’90s, with population bottlenecks to only 35 fish in 2013 and 38 fish in 2007, didn’t seem to have had much of an effect.
Tian is currently analyzing approximately 150 complete genome sequences of nine species of American pupfish to gain a better understanding of the deleterious mutations and gene deletions found in the various Southwestern populations. The study, he believes, exemplifies what conservation genomics can do for endangered and possibly inbred populations around the world.