by In a basement lab tucked into an 1,800-acre wildlife park in San Diego, California, Marlys Houck looks up to see a man in uniform holding a blue insulated lunch bag full of tiny pieces of eyes, trachea, legs and feathers.
“Ah,” she says, softly. “These 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, examples include the leaf frog and starling.
James Boggeln, a zoo volunteer, is the man with the bag, who gives it to Houck, the keeper of this laboratory, known as the “Frozen Zoo”. She and her team will begin the process of turning these pieces of tissue into research and preservation banks for the future. They’ll put the tissue into flasks where enzymes break it down, then the lab members will slowly incubate it over a month – growing an abundance of cells that can eventually be frozen and revived for future use.
At almost 50 years old, the Frozen Zoo has the world’s oldest, largest and most diverse repository of live cell cultures – more than 11,000 samples representing 1,300 different species and subspecies, including three extinct and more that are very close to extinction.
Today the Frozen Zoo is operated by an all-female team of four, who tend to a large collection of vials hand-marked with labels such as “giraffe”, “rhino” and “armadillo”, all stored in huge circular tanks filled with liquid nitrogen. In a world suffering from climate and biodiversity crisis, putting species on ice is one way to be optimistic about the future.
The work here has always been meaningful, but a looming crisis has put a lot of pressure on Houck and his team. It’s a race against time to get samples into the Frozen Zoo before they slip away from the world outside the lab. The women who hold these jobs see it as their duty to keep the future alive.
The work can be demanding – samples from birds, mammals, amphibians and fish, for example, require different processes. But with such high stakes, Houck describes it with holy reverence.
She feels the pressure of the role – when her predecessor was in charge, 300 samples, a year’s work, were lost due to mechanical failure. So her mind is focused on keeping the samples frozen at the zoo safe, Houck says – “but then with this excitement and 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 started a collection of animal skin samples in his laboratory at the University of California at San Diego and then moved to the San Diego Zoo a few years later. At the time there was no technology to use it 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 in the Frozen Zoo, where Houck pulls vials from liquid nitrogen tanks that look like giant, human-enlarging money. The tanks are pressurized at -320F, a temperature that stops cells from moving or changing – keeping them alive but in suspended animation. From this temperature, the cells can be revived and continue to live as if years or centuries had not passed.
No species is exactly the same, and some groups are more challenging to preserve than others. The Frozen Zoo started with mammals, and then expanded to cryobanked birds, reptiles and amphibians. The success rate with mammals is close to 99%, says Houck. “With amphibians, it was close to 1% for several years, and now I think we’re maybe 20 to 25%. Birds are quite tall.”
Racks in the nitrogen tanks hold 100 vials each, and each vial contains 1m to 3m live cells. Those cells—a giraffe, a lemur, or something more endangered like a vaquita—hold potential solutions to a range of existing and future problems.
Eventually, the cells could be used to bring back extinct species – but that’s not the main goal. Instead, the material is generally used to rescue 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 wild Przewalski’s horses, returning valuable genetic diversity to the living population that will make it more resilient to new diseases or environmental threats. One of the foals was named Kurt, after the founder of the Zoo.
The work done by San Diego’s Frozen Zoo 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, mostly located in North America and Europe.
We are losing species too fast for science to keep up. The least we can do is try to put that matter in the bank
Sue Walker, Save the Nature
The work done in San Diego is very innovative, says Sue Walker, head of science at Chester Zoo, and co-founder and vice-chairman of UK non-profit Nature’s Safe, a cryo bank that collects live cells and sperm and eggs. She says that in a few decades it may be possible to turn 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’re losing species too fast for science to keep up,” she says. “So the least we can do is try to bankroll that.”
It is difficult to obtain permits to import tissue from animals in other countries, so it is hoped that the capacity will increase in other places to use cryobank locally, especially near conservation hubs in Africa, South America and in southeast Asia. But that means increasing the ability to process and preserve the cell population in a uniform way. It’s expensive and complicated work – but it’s also necessary, says Walker.
“I think we have to throw everything at it to save some of these endangered species,” she says. “It’s about banking hope.”
Time machines in the past and future
Working on cell cultures can be like operating a time machine. Houck was once studying rhinoceros chromosomes when she opened a vial with her predecessor Arlene Kumamoto’s handwriting on it. Kumamoto put the cells into a deep freeze the same month Houck graduated from high school. “I thought, oh my god… she’s freezing the cells I’m using now for my studies. If she hadn’t done that, I wouldn’t be doing what I’m doing,” says Houck. “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 to offer a theory about why a group of women is leading this work at the Frozen Zoo. “We are feeding the cells. They are living creatures and they need to be nurtured and cared for, then you have to find out what they need when they need it,” she says. “It’s kind of like children.”
The staff add around 250 to 350 specimens to the zoo each year. The leaf frog that arrived today is a high priority, says Houck. Did the starling come too? Less critical. Such decisions are of great importance. Since each new animal that comes in has to be nurtured and preserved, and takes up space in the huge containers, she has to consider how much of that species is already represented, and how much potential there is for more of them. to get. “I wish I didn’t have to turn anything over. It would be better if we could accept every example that came in because they are all important.”
There are three extinct species in the collection: the po’ouli or Hawaiian honey tree, the Rabbs tree frog, and the Arabian gazelle. They are holding their collective breath as they watch more species in their collection go extinct. “The white rhino and the vaquita are probably next,” says Houck.
The room where the tanks are full – but the tanks are not yet at full capacity, so the team continues their work to cultivate and preserve the cells that could give life or death to endangered animals in the future.
In the future, the lab must update its methods – those handwritten vials will become scannable barcodes. He also needs new scientists to come and keep a watchful eye on the growing zoo on the ice.
“I think we’re all very protective of the frozen zoo and Dr. Benirschke’s legacy and legacy,” says Houck. “And hopefully we can raise a generation that will carry it on from us.”
