Last year, a rare endangered horse named Ollie was born. Following his brother, Kurt, delivered in 2020, Ollie became the newest Przewalski’s (pronounced “sheh-vahl-skees”) horse at the San Diego Zoo Safari Park. This was noteworthy on its own. Although herds of this wild equine—known for its stocky body, zebralike mane, and ocher coat—once roamed throughout Europe and central Asia, by 1969 their numbers had dwindled so low that they were declared extinct in the wild. While this is no longer true—thanks to conservation, approximately 2,000 are alive today, half of which live in Mongolia and China—the species is genetically bottlenecked. Nearly all living individuals descend from just 12 animals. As clones, Kurt and Ollie offer much-needed diversity to the population.
Thanks to the Frozen Zoo, the largest biobank for living cell cultures, these two Przewalski’s colts are copies of a male named Kuporovic, whose cells were collected in 1980. At the time, Oliver Ryder, director of conservation genetics at San Diego Zoo Wildlife Alliance, visited Minnesota to examine Kuporovic. Part of his job was to analyze the pedigrees of Przewalski’s horses and recommend how to best preserve genetic diversity, as inbreeding had become a serious concern. Ryder traveled North America to examine hundreds of individuals of this unique species and asked breeders to contribute biological samples, like hoof trimmings, to the Frozen Zoo. In a laboratory next to the 1,800-acre Safari Park, researchers grew cultures of Kuporovic’s DNA, which they cryogenically preserved and stored in pressurized tanks at -320 degrees Fahrenheit. The cultures remained frozen in suspended animation for 38 years.
Then in 2018, a partnership between San Diego Zoo Wildlife Alliance, the conservation group Revive & Restore, and the company ViaGen Pets & Equine led to the effort to clone Kuporovic. Increasingly, cloning is being used as a tool for conservation of endangered animals. This is different from de-extinction, the push to try to bring back long-dead creatures like the dodo or the woolly mammoth from fossils or other remains. Rather, this process uses living material to help a species on the brink of extinction continue to survive. Of the 450 Przewalski’s horse samples in the Frozen Zoo, Kuporovic provided the most genetic variation, making him the best candidate to copy.
To clone Kuporovic, scientists harvested an egg cell from a female quarter horse and stripped it of its DNA. Then they took the nucleus from one of Kuporovic’s somatic cells and inserted it into the empty egg. An embryo formed, which was implanted into a quarter horse’s womb to proceed as in a normal pregnancy. The results, Kurt and Ollie (who is named after Ryder), are genetically identical copies. When they reach sexual maturity, their potential offspring could help revitalize the Przewalski’s species.
For Ryder, who remembers examining Kuporovic 44 years ago, the experience is “dreamlike.”
“I’ve been thinking about that concept [of cloning endangered species], but I tell you it’s a very different thing to see a living, breathing, beautiful young colt,” he says. “It’s like a dream realized. You know, you didn’t dare to dream, but there was this possibility, and now it’s manifest. And it’s very astounding.”
Cloning endangered species is an emotional issue. It’s closely tied to human involvement with animals, which includes our love and affection for them as well as our guilt at the harm we sometimes cause them. For almost 50 years, the Frozen Zoo has attempted to alleviate some of that harm by collecting genetic material from as many birds, mammals, reptiles, amphibians, and fish as possible. So far, the zoo’s all-female staff of four has banked more than 11,000 samples representing 1,300 species and subspecies. Each vial of cells represents hope for the future. However, as climate change accelerates extinction, there’s also urgency in the task.
“Our goal is to bank everything while we can,” says Marlys Houck, curator of the Frozen Zoo. “As animals are dying, as extinction rates are increasing, we’re going to lose these animals. Each individual carries genetic diversity, and if we don’t bank it…it’s lost forever.”
For Houck, there’s a sense that her work at the Frozen Zoo will live on after her. Biotechnology is advancing at a rapid pace, and what used to feel like science fiction is now reality. Already, specimens have been used for in vitro fertilization, artificial insemination, embryo transfer, and, of course, cloning. In addition to the Przewalski’s horses, the Frozen Zoo took part in the first cloning of a U.S. endangered species, the black-footed ferret. Elizabeth Ann, born in 2020, is a copy of Willa, a ferret that died 36 years ago. Houck’s predecessor, Arlene Kumamoto, froze Willa’s cells in 1988.
“Arlene ended up passing away suddenly of cancer in 2000, which was really tragic,” says Houck. “And she never knew what those cells would go on to produce. So here she was, doing her daily work, you know, we’re freezing cells every day.… We have no idea what those will be used for in the future. When Arlene died, cloning was not really a possibility. I mean, it wasn’t very realistic. Yet those cells she froze went on to produce a cloned ferret and will bring back the genetic diversity of Willa, the one that was lost.”
The black-footed ferret has a sinuous, pale body, dark legs, and a mask over its eyes that, combined with curious ears and a frowning mouth, gives it a somewhat concerned expression. It once thrived throughout the Great Plains, feasting on prairie dogs. Its population dropped when European settlers started poisoning that food source, then plummeted when the black-footed ferret came into contact with bubonic plague. The Black Death, as it used to be called, was first detected in the United States in 1900 when a man died from it in San Francisco’s Chinatown. The disease was likely carried into the country by rat-infested ships from Asia.
While the illness was contained among humans, a version of it continues to thrive among weasels and rodents. The effect on black-footed ferrets was so catastrophic that they were considered extinct in 1979. When a small pocket was discovered in Wyoming in 1981, the remaining animals—including Willa—were taken into captivity. Today, the 500 or so living black-footed ferrets are all descended from seven individuals. Like the Przewalski’s horses, the species is genetically bottlenecked.
Willa was unrelated to all other living ferrets. Her identical copy, Elizabeth Ann, was brimming with the prospect of genetic diversity. She was born generally healthy and “feisty,” as she was repeatedly described to me, and sent to the National Black-Footed Ferret Conservation Center in Colorado. When she matured and came into estrus, she rejected the four male ferrets presented to her, refusing to let them into her burrow and even biting one on the nose. “She was very mean to them, and that’s not uncommon,” says Ben Novak, lead scientist with Revive & Restore, headquartered in Sausalito. “She just didn’t like any of them. They have a number of females every year that do that, and that’s because we’re forcing these pairings. It’s not like in nature, [where] the male has to impress them and they have their pick.”
As breeding wasn’t happening, researchers decided to impregnate Elizabeth Ann through artificial insemination. They soon discovered something worrying. Her uterine horn was swollen and filled with fluid, an incurable condition called hydrometra. When signs of infection emerged, Elizabeth Ann had to have a hysterectomy. While she’s healthy, she’ll never produce babies, a situation Novak calls “incredibly, incredibly disheartening.” Two new clones of Willa, Noreen and Antonia (after the protagonist of Willa Cather’s novel My Ántonia), are both likely to mate this year, hopefully with better results.
“It’s a miracle that Elizabeth Ann didn’t like any of the males she was paired with,” says Novak. “Had she bred with one of them and not gotten pregnant, we would have never poked inside to see what was going on, because this condition has zero symptoms. They just would have found her dead one day, which has happened twice before. Two natural-born black-footed ferrets died of the advanced progression of this condition. They don’t display any discomfort or anything like that. So yeah, she saved her own life by being picky.”
Elizabeth Ann has other physical issues as well. She was born with only one kidney, uterine horn, and ovary. This begs the question: Were the deformities caused by cloning, or did they have something to do with Willa’s genetic history? It’s difficult to say, although Elizabeth Ann’s identical sisters appear normal, so far.
It’s commonly believed that clones are unhealthy and tend to die young. Certainly, there have been tragedies. The first cloned endangered species, a bovine called a gaur, died after two days of life. The second, a banteng, another rare type of cattle, survived seven years, not even half its lifespan. A clone of the extinct Pyrenean ibex, a wild goat, copied from the cells of the last living individual, suffocated within minutes of being born because of a deformed lung.
However, these cases aren’t the norm. While data is scant, there have likely been thousands of cloned animals. For every failure, plenty of clones survive to old age, such as the first cloned mouse, cow, and deer. The concern about health started with Dolly the sheep, the first-ever cloned mammal. Born in 1996, she had severe arthritis and passed away at six, which is only middle-aged for a sheep. She also spent her life indoors, an unnatural state that may have affected her well-being. Dolly produced six normal lambs, and her latter-born siblings were healthy.
“There’s these really high-profile cases where the clones have problems, and that’s what people see and go, Ah, cloning must be bad,” says Novak. “But when you pull away and look at the majority of cloning, across 56 different species to date, it’s actually a very low efficiency, but it’s a highly effective technology.… The vast majority of clones will be healthy and fertile. The majority will live to an average age. In fact, we know that there’s almost no discernible difference between clones and normal-born offspring.”
Efficiency is another issue. It takes many tries to get a living clone, and only a small fraction of attempts result in live births. This often means the surrogate mother miscarries, raising the question of whether it’s ethical to cause one animal to suffer in pursuit of creating another. The good news is that the process seems to be getting better over time. Dolly the sheep was one of 277 cell fusions. Snuppy, the first cloned dog, took 1,095 tries in 123 different surrogates. But this was 28 and 19 years ago. Compare that with the recent Przewalski’s horse clones—Kurt was one of four embryos, and Ollie was one of seven.
Clones are also expensive. For financial reasons alone, it’s unlikely an endangered clone would be released into the wild, where it could be picked off by the first predator it met. It would take several generations of breeding, and enough offspring to spare, before its progeny would be allowed into nature—and only after they showed they could survive on their own. That’s a lot of ifs. And none of it would matter if there had been no attempts to address the larger problems that threatened the species in the first place. Opponents of cloning believe that resources are better spent on tackling habitat loss, poaching, disease, toxins, and other issues that endanger animals.
In some cases, a clone could be genetically modified to help it survive. While scientists can vaccinate black-footed ferrets to protect them from plague, that’s not practical for wild populations. However, it may be possible to give them a “genetic vaccine.” The ferret’s genes could be modified to produce an anti-plague antibody to help its immune system fight the illness. This trait would be passed down, inoculating future generations against the disease.
While this technology doesn’t yet exist, it’s under development. Revive & Restore has created transgenic mice that “express antibodies against plague in their germline, and can be passed on from generation to generation,” molecular biologist Bridget Baumgartner told the online publication proto.life. Other projects aim to genetically modify wild animals to protect humans. For example, researchers at the Massachusetts Institute of Technology are trying to make the white-footed mouse resistant to the bacteria that causes Lyme disease, which would slow its spread to people. At UC San Diego, scientists are attempting to suppress malaria-carrying mosquitoes.
But these technologies aren’t without risk. It’s easy to imagine the trouble that could result if genetically modified creatures got into nature before they were fully understood. It could have disastrous consequences.
When writing about these issues, I occasionally find myself thinking about Margaret Atwood’s 2003 novel, Oryx and Crake. The story is set in a postapocalyptic landscape full of GMO hybrid creatures, such as rakunks (raccoon-skunk crosses), spoats (spider-goats), and pigoons (pigs used to grow human organs). People are an endangered species, and new humanoids are taking over the planet. Atwood, who comes from a family of scientists, is well aware of the harm humans have inflicted on the environment. What might happen if new technologies allow us to affect nature on its most incremental, genetic scale, she asks?
Sometimes, when I hear about ranchers cloning cows from prime ribeye steaks, as happened in Texas around 2014, or jellyfish genes being used to make chickens glow under a blue light, I see Atwood’s concern. The strangeness of biotech seems ominous. It’s unnatural, I’ll think, and then wonder what I mean by that. After all, humans have always manipulated the natural world, albeit in slower, clumsier ways. Maybe I just mean that it feels like science fiction.
Since the beginning, writers have shaped our views of science. At this point, it’s impossible to divorce public perception of cloning from science fiction, which often portrays clones as soulless copies or evil twins. Perhaps the technology wouldn’t be so controversial if, back when Dolly the sheep was born, it was described with a less exciting word. Maybe it should have been a nondescript, serious-sounding term like “germ-line transmission”—a dull name for a mind-boggling technique for replicating birds.
It turns out birds can’t be cloned. The egg, a giant cell, poses too many challenges. The yolk is the nucleus of the cell, where a minuscule DNA speck floats in a viscous fluid, making it challenging to manipulate. Germ-line transmission is a work-around technology that, similar to cloning, may allow one species of bird to give birth to another.
In theory, here’s how it works: Primordial germ cells, which are embryonic precursors to the sperm or egg, are taken from a developing donor bird and replicated in a petri dish. Researchers then choose a surrogate to reproduce the donor bird. Unlike in cloning, which uses adult animals, the surrogate is still a chick in an egg. Germ cells are injected through the eggshell into the bloodstream, where they circulate and become part of the baby’s reproductive system. The baby is now a chimera that will carry the donor cells for the rest of its life. When it matures and mates with another chimera, it will ideally lay eggs containing only the donor offspring.
So far, this doesn’t quite work. Chimeras contain DNA from both the donor and the surrogate, and when they lay eggs, some belong to the donor species and some belong to the surrogate’s. Researchers are trying to create a “sterile” chimera that will produce only donor birds. With funding from Revive & Restore, a team at the Rockefeller University has performed research using two types of zebra finches. Other scientists in Los Angeles and Ohio are working on experiments involving sterile quail.
In the end, germ-line transmission may lead to a common bird, like a chicken, reproducing an endangered species like the California condor. It could also lead to the hybrid reproduction of the extinct passenger pigeon.
However, neither germ-line transmission nor cloning for conservation is likely to become a common practice anytime soon. They’re simply too complicated. The endangered animal’s breeding habits must be understood and its genome unlocked. A closely related domestic animal must be available as a surrogate. Then there’s the cost, partnership, and political will to take into account. Even if all that is in place, there’s another piece of the puzzle that can’t be manufactured: the genetic material from the original animal has to exist. Without it, cloning makes no sense.
“Sadly, when we look beyond Przewalski’s horses and black-footed ferrets for other species where these criteria would create beneficial cloning, we didn’t find anything,” says Novak. “And that’s partly because there just are no cell lines or tissue samples from the past for bottlenecked species or endangered species that have any value to their current populations. It’s extremely rare.”
But as the Frozen Zoo continues to bank as many animals as possible, and as technology advances alongside it, new opportunities to help struggling species should emerge. Like nature itself, changes happen in unexpected bursts.
Consider the American chestnut. These large, graceful trees were once plentiful on the East Coast, numbering in the billions. Then, around 1900, an airborne fungus from Japan killed almost every single one. It was a catastrophic loss that must have seemed hopeless at the time. And yet today, several genetically engineered strains of the American chestnut are becoming increasingly resistant to the fungus, meaning they’re “on the verge of making a dramatic comeback into the eastern deciduous forests of North America,” according to Beth Shapiro’s book How to Clone a Mammoth. Over 1,000 American chestnuts are growing in New York. Someday in the future, Americans may be able to roast native nuts by the fire again. Which is to say, sometimes our ecological damage isn’t forever. Occasionally, we can make things better.•
Joy Lanzendorfer’s first novel, Right Back Where We Started From, was published in 2021. Her work has appeared in the New York Times, Washington Post, Raritan, the Atlantic, and Ploughshares as well as on NPR and for the Poetry Foundation, among others.
