In a basement laboratory adjacent to an 1,800-acre wildlife park in San Diego, California, Marlys Houck looks up and sees a man in uniform holding a blue insulated lunch bag, filled with small pieces of eyes, trachea, feet and feathers.
“Aha,” she says softly. “Here are today’s examples.”
The bag in question contains small pieces of soft tissue collected from animals that died of natural causes in the zoo. Today, the monsters include a leaf frog and a starling.
The man holding the bag is James Boggeln, a volunteer at the zoo, who hands it over to Houck, the curator of this laboratory known as the “Frozen Zoo.” She and her team will begin the process of turning these pieces of tissue into a research and conservation bank for the future. They will put the tissue in flasks where enzymes digest it, and then the lab members will slowly incubate it over the course of a month, creating an abundance of cells that can be frozen and eventually resuscitated for future use.
The nearly 50-year-old Frozen Zoo is home to the world’s oldest, largest and most diverse repository of living cell cultures – more than 11,000 samples representing 1,300 different species and subspecies, including three extinct species and more very close to extinction.
Today, the Frozen Zoo is run by an all-female team of four, who watch over a huge collection of hand-marked bottles with labels like ‘giraffe’, ‘rhino’ and ‘armadillo’, all stored in huge circular tanks filled with liquid nitrogen. In a world suffering from a climate and biodiversity crisis, putting species on ice offers a way to be hopeful about the future.
The work being done here has always felt meaningful, but an accelerating extinction crisis has increased the pressure on Houck and her team. It’s a race against time to stop monsters in the Frozen Zoo before they escape the world outside the laboratory. The women who hold these jobs see it as their duty to secure the future.
The work can be painstaking; samples from birds, mammals, amphibians and fish, for example, all require different processes. But because the stakes are so high, Houck describes it with a kind of sacred reverence.
She feels the pressure of the role: when her predecessor was in charge, a mechanical failure led to the loss of 300 samples, a year’s worth of work. So her mind is focused on keeping the samples the zoo has frozen safe, Houck says — “but then combined with the excitement and this joy, because it’s an honor to be able to do this.”
‘Collect things for reasons you don’t yet understand’
The zoo was founded in 1972 by a German-American pathologist named Kurt Benirschke, who began collecting animal skin samples in his laboratory at the University of California San Diego and moved them to the San Diego Zoo a few years later. At the time, there was no technology to go beyond basic chromosome research, but Benirschke often quoted the American historian Daniel Boorstin: “You have to collect things for reasons you don’t yet understand.”
That quote still hangs on a poster at the Frozen Zoo, where Houck pulls out bottles of liquid nitrogen tanks that resemble giant silver thermoses the size of a human. The tanks are pressurized to -320F, a temperature that prevents cells from moving or changing – leaving them alive, but in suspended animation. From this temperature, the cells can be revived and continue to live as if decades – or centuries – had not passed.
No two species are exactly the same, and some groups are more difficult to preserve than others. The Frozen Zoo started with mammals and then expanded to cryobanking birds, reptiles and amphibians. The success rate in mammals is close to 99%, Houck says. “In amphibians it was around 1% for a number of years, and now I think we’re maybe at 20 to 25%. Birds are quite high.”
Racks in the nitrogen tanks hold 100 vials each, and each vial contains 1 to 3 million living cells. Those cells – a giraffe, a lemur or something even more endangered like a vaquita – contain possible solutions to a range of existing and future problems.
Ultimately, the cells could be used to bring back completely extinct species – but that’s not the main goal. Instead, the material is typically used to save existing species that are struggling. In 2020, the Frozen Zoo used cryopreserved DNA to clone a black-footed ferret, the first endangered species in the United States to be cloned. Last year, frozen cells cryopreserved 42 years ago were used to clone two critically endangered Przewalski’s wild horses, returning valuable genetic diversity to the living population that could make it more resistant to new diseases or environmental threats. One of the foals was named Kurt, after the zoo’s founder.
San Diego’s Frozen Zoo’s work is part of a global movement to cryobank everything from animals to seeds. Today, there are about a dozen wildlife-based cryobanks around the world, mainly located in North America and Europe.
We are losing species too quickly for science to keep up. The least we can do is try to get that material
Sue Walker, Nature is safe
The work done in San Diego has been particularly groundbreaking, says Sue Walker, chief science officer at Chester Zoo, and co-founder and vice-chairman of the British nonprofit Nature’s Safe, a cryobank that collects living cells, sperm and eggs. She says that in a few decades it could be possible to convert these cells into pluripotent stem cells, which can be reprogrammed to produce sperm and eggs.
In an ideal world, species could be preserved in the wild, but in reality this is not the case. “We are going extinct too quickly for science to keep up,” she says. “So the least we can do is try to save that material.”
It is difficult to obtain permits to bring in animal tissues from other countries, so the hope is to increase capacity in other places to do local cryobanking, especially near conservation centers in Africa, South America and Southeast Asia. But that means building the capacity to process and maintain the cell population in a uniform way. It’s expensive and complex work, but also necessary, Walker says.
“I think we should do everything we can to save some of these species that are on the brink of extinction,” she says. “It’s about the hope in the banking system.”
Time machines to the past and future
Working on the cell cultures can be like operating a time machine. Houck was once studying rhino chromosomes and opened a bottle with her predecessor Arlene Kumamoto’s handwriting on it. Kumamoto had frozen the cells the same month Houck graduated from high school. “I just thought: oh my god… she was freezing cells that I am now using for my studies. If she hadn’t done that, I wouldn’t be doing what I do,” Houck says. “What are we doing today that will be used in the future?”
Julie Fronczek, who has worked at the zoo for 24 years, looks up from her microscope and comes up with a theory about why a group of women are leading this work at the Frozen Zoo. “We take care of the cells. They are living beings and they need to be fed and cared for, and then you need to know what they want, when they want it,” she says. “It looks a bit like babies.”
The team adds approximately 250 to 350 specimens to the zoo each year. The leaf frog that arrived today is a high priority, Houck says. The starling that also arrived? Less critical. Such decisions weigh heavily on her. Because each new animal that arrives has to be bred and preserved, taking up space in the giant containers, she has to consider how many of that species are already represented, and how many opportunities there are to get more of them. “I would prefer not to have to turn anything down. It would be better if we could accept every sample that came in, because they are all important.”
The collection includes three extinct species: the po’ouli or Hawaiian honeycreeper, Rabbs’ tree frog and the Saudi gazelle. They collectively hold their breath as they watch for more species in their collection to become extinct. “The next ones will probably be the white rhino and the vaquita,” says Houck.
The space in which the tanks are located is full, but the tanks are not yet at full capacity. That’s why the team continues to grow and preserve the cells that could mean life or death for endangered animals in the future.
Going forward, the lab will need to update its methods – those handwritten vials will eventually be scannable barcodes. New scientists are also needed to keep a close eye on the growing zoo on the ice.
“I think we’re all very protective of the frozen zoo and the legacy and the legacy of Dr. Benirschke,” Houck says. “And I hope we can teach a generation that will pass it on from us.”