The Devils Hole pupfish inhabits a very horrific environment, as its name suggests.

263 of the pupfish are confined to a single deep limestone cave in the Mojave Desert of Nevada, where the water temperature is almost constantly around 93 degrees Fahrenheit, food is so scarce that they are constantly on the verge of starvation, and oxygen levels are so low that most other fish would die right away. Of all known vertebrates, the pupfish Cyprinodon diabolis inhabits the tiniest habitat.

The significant impact that these severe and isolated settings have had on this fish’s genetic diversity is now supported by new studies.

University of California, Berkeley biologists reveal the first complete genome sequencing of eight species of pupfish from the American Southwest—30 individuals in total, including eight Devils Hole pupfish—in a paper that was published this week in the journal Proceedings of the Royal Society B. Surprisingly, the Devils Hole pupfish has undergone such intense inbreeding that, on average, 58% of the genomes of these eight individuals are identical.

According to lead researcher Christopher Martin, an associate professor of integrative biology at UC Berkeley and curator of ichthyology in the campus Museum of Vertebrate Zoology, “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.” The researchers were unable to test fitness directly, the increased inbreeding in these pupfish probably causes a significant decline in fitness.

The researchers discovered that other pupfish species are likewise inbred, but only 10% to 30% of their genomes are identical.

The Devils Hole pupfish exhibit an amount of inbreeding comparable to what would occur if four to five generations of siblings interbred, according to graduate student David Tian, the study’s lead author. As a result, dangerous mutations are more likely to be fixed rather than eliminated, potentially driving a population to extinction through a mutational meltdown. Although populations of Devils Hole pupfish in the wild and in captivity, or “refuge” populations, are currently thriving, the species’ low genetic diversity could cause problems if climate change and human influences increase.

The new genome sequences will assist researchers and conservationists in evaluating the health of native pupfish populations in the face of these potential threats and, if necessary, intervening in refuge populations to increase the genetic diversity of these species—the Devils Hole pupfish in particular.

With the help of the new genomic data, Tian said, “there is a lot of potential to look at not just genetic diversity and how these species are related to one another phylogenetically, but also look at inbreeding and mutation load to get an idea of what their current status is, how evolutionary history may have influenced their current genetic variation, and think about where the population is going and what we should do, if anything, to preserve these species.”

Pupfish species are dispersed over the world and frequently found in remote lakes and springs with harsh conditions that most fish would not be able to survive. Warm, saline desert springs and streams in California and Nevada are home to about 30 different species. Martin has researched a number of pupfish populations, including a few in the Bahamas’ San Salvador Island, to better understand the genetics underlying their capacity to survive in harsh environments and specialized ecological niches.

Martin said that the Devils Hole pupfish is special due to its restricted range and precarious existence, which makes environmentalists concerned about its varying wild population.

The possibility that these declines may be related to the population’s genetic health is one of the questions surrounding them. The decreases are caused by deleterious mutations that, because the population is so small, have gotten fixed.

Martin pointed out that human intrusions into their habitat had contributed to the reduced number. In the 1960s and 1970s, local ranchers and developers tapped groundwater in the area, substantially lowering the water level in Devils Hole and causing a population decline. Devils Hole and its inhabitants were preserved by a 1976 Supreme Court decision allowing the federal government to prohibit groundwater extraction, and the endangered species was protected by captive breeding at a nearby 100,000-gallon pool at the Ash Meadows National Wildlife Refuge. However, a reduction in the 1990s brought the wild population to a low of 35 individuals in 2013. Since then, the wild population has returned, but the refuge population has grown to be about 400, which is double the wild number.

However, not all of the genetic variety in the Devils Hole pupfish can be attributed to humans. The genome of a pupfish that was captured in 1980 and kept at the University of Michigan was also sequenced by the UC Berkeley researchers. It displayed inbreeding and a lack of genetic variety similar to those observed in newly captured individuals, the majority of whom passed away naturally. This suggests that over hundreds or even thousands of years, the pupfish population has probably experienced frequent bottlenecks.

Martin and Tian discovered that 15 genes have completely vanished from the Devils Hole pupfish genome as a result of this. Five of them appear to play a role in adjusting to life in hypoxic or low-oxygen settings.

“Because this is a habitat where you’re most exposed to hypoxia,” Martin said, “these deletions are a paradox.” The researchers said that it might have something to do with the habitat’s long-term stability. However, it appears that the hypoxic pathway is damaged. It doesn’t really matter if you break more genes in that regulatory pathway after you disrupt one gene. The researchers’ upcoming task is to investigate what these deletions accomplish in detail. Are they able to tolerate hypoxia better? Do they reduce the body’s tolerance to hypoxia? Both of those situations, in their opinion, are currently equally likely.

The researchers suggested that selective breeding could assist the promotion of diversity and possibly prevent the extinction of the Devils Hole pupfish species. Additionally, CRISPR genome editing could re-add missing genes to the body.

The finding that the fish gathered in 1980 had a genome that was roughly as inbred as fish today is perhaps excellent news, in that the population has traditionally been extremely inbred with very low genetic diversity, suggesting that the recent decrease in the ’90s—with population bottlenecks to only 35 fish in 2013 and 38 fish in 2007—doesn’t appear to have much of an effect.

Tian is currently examining 150 full genome sequences from nine species of American pupfish in order to have a better understanding of the harmful mutations and gene deletions that have occurred in the distinct populations of the Southwest. He views the findings as a demonstration of what conservation genomics may accomplish for globally threatened and possibly inbred populations.

In terms of collecting genomic data and applying it to conservation, the researchers are on a very exciting cusp, especially in a period when it’s an issue that’s likely only to get worse with changing climate, more habitat fragmentation, and just manmade changes.”

However, Tian is wary of genetic treatments because it is unclear how genes affect a species’ physical and behavioral traits and how this relates to fitness and adaption to a particular environment. Still, conservation need to be a top focus.

The solution is still more money for these populations, safeguarding ecosystems, pursuing legal means of protecting these species, and figuring out methods for people and these endangered animals to cohabit on this earth.

Sources:

David Tian et al. (2022). Severe inbreeding, increased mutation load and gene loss-of-function in the critically endangered Devils Hole pupfish, Proceedings of the Royal Society B: Biological Sciences. DOI: 10.1098/rspb.2022.1561

https://phys.org/news/2022-11-endangered-devils-hole-pupfish-inbred.html