This freshwater fish is facing an identity crisis

Last fall, researchers released a special brood of bright blue- and red-striped fish into the upper Kanawha Falls watershed in West Virginia. Bred at White Sulphur Springs National Fish Hatchery, these purebreds were tasked with an important job: boosting the threatened population of their wild counterparts by mating with their own species in the wild.

These finger-length fish are called candy darters, and they’re only found in a few waterways in West Virginia and Virginia. Their already-small populations are dwindling thanks to interbreeding with a similar but more robust species: the variegate darter. The resulting “candigate” hybrids have gradually pushed candy darter genes out of the ecosystem and now endanger the future of the species, which is why researchers are now attempting a two-fold genetic rescue. They’re trying to boost pure populations by preventing enough candy darters from producing hybrids with variegate darters while still retaining the genetic diversity within the candy darter genome.  

“The main goal right now is to try and preserve that unique genotype that we have in its entirety,” says Nate Owens, a non-game fish biologist with the West Virginia Division of Natural Resources. He says the focus is making sure they collect enough individuals to maintain the full candy darter genome, because if variegate darters infiltrate the DNA of enough fish, “we would lose the amount of adaptation that can occur through time, and they may go extinct.”  

According to Owens, these conservation efforts include genotyping candy darter individuals caught in the wild to ensure their genetic purity. If they meet the standard, they can take part in the hatchery breeding program or be translocated to rebuild populations further upstream where variegate darters can’t reach them. However, if variegate darters continue invading their habitats, researchers might have to reevaluate the 100% genetic purity standard, or could start deliberately encouraging some hybridization to ensure that at least most of the candy darter’s genome lives on as climate change and human activities transform Appalachian streams.

“It just gets really interesting philosophically,” says Amy Welsh, a conservation geneticist at West Virginia University. “Are we creating a new species at that point? Or are we just helping the candy darter?”

Until the 1980s, candy darters and variegate darters were never seen in the same waterways. But then, researchers think, fishermen started dumping buckets of variegate darters into candy darter territory after using them for live bait fishing. Variegates are larger and faster, so when they were introduced into candy habitats, they beat out male candies to mate with female candies, creating the hybrids. Since then, the area occupied by the hybrids has grown. A study by the United States Geological Survey mapped out the hybridization zone in 2004, but a follow-up survey in 2014 showed that the zone grew as variegate darters continued swimming upstream into candy darter territory. 

Hybridization, or breeding between two species, is not an uncommon phenomenon in the wild, particularly in freshwater fish. Fish breed externally, so it’s easier for the eggs of one species to be fertilized by the sperm of a different species unintentionally due to competition for spawning territory and close proximity. However, hybridization can become an issue when human activities bring non-native species into the habitat of a native species. In the case of the candy darter, this leads to genetic swamping, meaning that the genes of the variegate darter have overtaken those of candy darters in the ecosystem. Over time, variegate darters have diluted the candy darter population, with each hybrid generation becoming more like variegates and less like candies, leaving only four genetically pure populations left at the time of the 2014 survey. 

This hybridization, plus habitat loss and environmental degradation, kickstarted a push to get the candy darter listed for protection under the Endangered Species Act. The U.S. Fish and Wildlife Service initially listed the fish under the lesser classification of “threatened” in 2018 because the agency said candy darters were still in about half of their historical range. But Welsh says that’s true only if you count hybrids as well as pure candy darters. After she and other experts pointed this out, the agency changed the fish’s status to “endangered”. The more urgent classification spurred conservation efforts like the breeding, translocation and monitoring programs, to which Welsh’s lab contributes. She uses samples of genetic material from individuals taken from those pure populations to figure out if they’re suitable to mate with other candy darters.

“We will screen them to make sure that they are pure candy darter individuals before they’re released into the new location so that we are ensuring that we’re creating a pure population,” Welsh says. 

In collaboration with Welsh’s lab and other partners, the West Virginia Department of Natural Resources is establishing pure populations of candy darters in their historic range in the upper area of Kanawha Falls where they’ve previously been extirpated from, according to Owens. By monitoring these populations using genetic analysis, the researchers can analyze how they are faring.   

While hybridization is the biggest threat to candy darters in West Virginia, populations across the border in Virginia are suffering from a different gene flow issue. They are isolated from one another due to dams and the development of farmland, so individuals from different populations aren’t mating and exchanging genetic material, leaving them more vulnerable to environmental changes like climate change and development.

“They’re losing genetic variation that might have adaptive value,” says Eric Hallerman, a professor of fish conservation at Virginia Tech. The more gene flow there is between populations, the better equipped they are to survive and adapt to changes in the environment. Different genetic combinations in the offspring of unrelated individuals can lead to traits that could be helpful for a species. Without this variation, Virginia candy darters are at risk of losing genetic viability and their ability to potentially adapt to new conditions. 

The Department of Wildlife Resources in Virginia is investigating how to encourage those isolated populations to breed with each other again, including possibly removing small dams and translocating candy darters from one population to another to increase genetic diversity. However, these plans are still in early stages.

“You have to do a lot of homework before you move into putting these animals just anywhere,” says Mike Pinder, a non-game and endangered fish biologist in the Virginia Department of Wildlife Resources. “You have to find out and make sure that where you’re putting them is not already occupied by the species, and that where you’re going to put them has suitable habitat for the species. Otherwise, you’re just putting good money on bad.”

Welsh also monitors genetic diversity in West Virginia’s candy darters. In addition to determining whether individuals are 100% pure candy darter, she also analyzes how related one fish is to another. This way, she can monitor populations’ genetic makeup to make sure there isn’t too much inbreeding between related individuals. 

“We can use genetics to do a parentage analysis to see who reproduced with who out in the wild and who was successful,” she says. “So we’ll monitor all those things because they can have an impact on genetic diversity and the long term adaptive potential of the population.”

As for the “candigate” hybrids, there’s no clear answer about their classification. Visually, a hybrid that is the offspring of a pure variegate and pure candy will look like a mix of both, but as that hybrid mates with another hybrid or another variegate, they begin to look less like candy darters while still retaining some candy darter genetic material. The same goes for individuals that look like candy darters that have even a little variegate DNA – they’re not pure enough to be classified as candy darters and take part in the breeding program. 

But even if the proliferation of hybrids is threatening the survival of the candy darter as a unique species, it might mean that at least some version of the species might survive in a rapidly warming ecosystem in the long run. Candy darters need colder water, while variegates thrive in warmer water. This leads Welsh to speculate that if candy darters with specific variegate genes were selected to breed, they might be able to survive.

As for the remaining pure candy darter populations, they’re being monitored and supplemented with more pure individuals as conservation efforts identify them. The locations for the establishment of these populations were carefully selected to ensure no variegates or hybrids would sneak in, but these are only a small part of their historic range. Getting candy darters back to their full range in the watershed would require completely removing variegates and hybrids from the area. 

“I think it’s really going to come down to the progression of science,” West Virginia’s Owens says. “If we’re going to get these established [populations] back into their historic range where [there are] no barriers to prevent the variegate darters from moving into their populations, we’re going to have to… try something different to try and eliminate those individuals from the communities.”

Once the areas with candy and variegate darters have become completely hybridized, the Division of Natural Resources may use the fish’s own genes against them to try and get the population to die out by creating what’s called a “Trojan male.” These Trojan male darters would be genetically modified to only produce male offspring, eventually leading to no fertile females for reproduction in the population. But for now, efforts remain focused on conservation techniques based on genetics, as well as educating the public about the importance of not dumping live bait or removing native species. 

“If we don’t stop moving fishes around, we’re going to end up with large homogenous communities of the same fish and we’re going to lose biodiversity,” Owens says.

This would be a huge blow for the Appalachian region, as it is historically one of the most biodiverse areas on the planet with 80,000 rare species, according to The Nature Conservancy.

“In southeastern North America, we have one of the two biodiversity hotspots globally for [temperate] freshwater fishes. It’s us and eastern China,” Virginia Tech’s Hallerman says. “It’s something precious that most people don’t know about.”