Gene editing has made pigs immune to a deadly epidemic

When covid-19 began to race around the world, countries closed businesses and told people to stay home. Many thought that would be enough to stop the coronavirus. If we had paid more attention to pigs, we might have known better. When it comes to controlling airborne viruses, says Bill Christianson, I think we fool ourselves on how effective we can be."

Christianson is an epidemiologist and veterinarian who heads the Pig Improvement Company, in Hendersonville, Tennessee. The company sells elite breeding swine to the pork industry, which for the last 34 years has been fighting a viral disease called porcine reproductive and respiratory syndrome PRRS.

The pathogen causes an illness known as blue ear, for one of its more visible symptoms; when it first emerged, in the 1980s, it was simply called mystery swine disease." Once infected with PRRS (pronounced purrs"), a sow is liable to miscarry or give birth to dead, shriveled piglets.

And I'm going to say yes, it's worse for pigs than covid is for us," says Christianson.

To stop PRRS, as well as other diseases, pig farmers employ measures familiar to anyone who has been avoiding covid-19. Before you enter a secure pig barn, you get your temperature taken, shower, and change clothes. Lunch boxes get bathed in UV light, and supplies are fogged with disinfectant. Then there's the questionnaire about your last pig contact"-seen any swine on your day off? Been to a country fair? (Answering yes means a two-week quarantine away from work.)

Despite the precautions, the virus can slip in. Once inside, it quickly spreads in the close quarters. Swift depopulation"-i.e., culling-of the animals is the most effective way to get rid of it. In bad years, American pig farmers lose $600 million to PRRS.

Now Christianson's company, which is a division of the British animal genetics firm Genus, is trying something different. Instead of trying to seal animals off from the environment, it's changing the pigs themselves. At an experimental facility in the central US (the location kept secret for security reasons), the company has a swine IVF center and a lab where pig eggs are being genetically edited using CRISPR, the revolutionary gene scissors.

During a virtual tour, a worker carried a smartphone through the editing lab into the gestation area, where sows spend nine months until giving birth-farrowing" is the farmer's term. Then he led the way to a concrete room where gene-edited piglets grunted and peered at the camera. According to the company, these young pigs are immune to PRRS because their bodies no longer contain the molecular receptor the virus docks with.

Every virus attacks cells by fusing with them and injecting its genetic cargo. With covid-19, the virus attaches to a receptor called ACE-2, which is common on airway and lung cells-the reason the disease causes problems with breathing. With PRRS, it's CD163, a receptor on white blood cells. These experimental pigs don't have a complete CD163 gene because part of it was snipped away with gene editing. No receptor, no infection.

I never thought it would be a light switch ... but it seems to work on all types of pigs and against all the strains of the virus."

Bill Christianson

According to the company's unpublished research, attempts to infect the gene-edited pigs with PRRS have not succeeded. I never thought it would be a light switch," says Christianson. But it seems to work on all types of pigs and against all the strains of the virus."

Notoriously, a similar method has been tried in humans. In a disastrously reckless 2018 outing, Chinese scientists edited human embryos in hopes of conferring resistance to HIV, the cause of AIDS. Those researchers likewise dreamed of halting a disease by removing a receptor. The problem was the technology wasn't ready to do such an ambitious job safely. Although the CRISPR tool is immensely versatile, it lacks precision, and the DNA surgery created something akin to genetic scars in the twins born from the experiment.

In September a high-level international panel said no one should try modifying babies again until it has been clearly established that it is possible to efficiently and reliably make precise genomic changes without undesired changes in human embryos."

But with pigs, the era of genetic modification is now, and its benefits might be visible soon. Genus hopes to win approval to sell its pigs in the US and China as early as 2025. Already, its experimental stations are home to hundreds of gene-edited pigs and thousands of their descendants-likely the largest number anywhere. (Read the sidebar on the regulatory approval of GM food animals.)

To Raymond Rowland, a researcher at the University of Illinois who was involved in creating the first PRRS-proof animals, gene editing is in its largest sense, a way to create a more perfect life" for pigs and their keepers. The pig never gets the virus. You don't need vaccines; you don't need a diagnostic test. It takes everything off the table," he says.

Elite pigs

Aldous Huxley's novel Brave New World begins with a tour of the Central London Hatchery," where children in a future society are being produced through a test-tube process under a sign that reads COMMUNITY, IDENTITY, STABILITY." The signs at Genus's facilities are mostly about temperature checks and hand-washing, but the concept is not so different. Every pig is numbered, monitored, and DNA-tested for its genetic qualities.

The firm manages animals selected to be the healthiest and fastest growing, and to have the largest litters. These animals-what Genus calls elite germplasm"-are then propagated via breeding on multiplier farms" and purchased by producers everywhere from Iowa to Beijing, who breed them still further.

The company has been using DNA sequencing for several years to identify pigs with preferred traits and to steer its breeding programs. In 2015, it signed an exclusive license to gene-edit pigs and cattle using technology from Caribou Biosciences, a company started by Jennifer Doudna of the University of California, Berkeley, who last October shared a Nobel Prize for the development of CRISPR.

SELMAN DESIGN

Because the pig company had no experience in genetic engineering, it began to hire plant biologists. One of them is its chief scientific officer, Elena Rice, a Russian-born geneticist who spent 18 years at Monsanto, mostly developing genetically modified corn plants to grow bigger and resist drought. The plants were never emotional to me," says Rice. The little pig or little cow-it's very emotional. You want to hug them; you want them to be healthy. It's like having a kid. You don't want them to be sick."

The Genus research station is set up to carry out the editing process quickly, on many pigs. Sows are anesthetized and then rolled into a surgical suite, where veterinarians remove eggs from their ovaries. The eggs are moved to the lab, where they are fertilized and the CRISPR molecules are introduced. Two days after editing, the embryos-by then a few cells big-are implanted into surrogate sows.

CRISPR is renowned for its ability to cut DNA at predetermined locations, but in practice, the technology has a random element. Aim it at one spot in a genome and you'll change it in one of several possible ways. Unplanned changes, or off targets," can appear far away in the genome, too.

In plants, this randomness isn't such a problem. A successful genetic change to a single seed (an event," as plant engineers call it) can be multiplied into a million more fairly quickly. In pigs, it's necessary to create identical edits in many animals in order to establish a population of founder pigs for breeding.

In experiments on pig cells, the Genus researchers have tried many possible edits to the CD163 gene, looking for those that occur most predictably. Even with such efforts, the pigs being born have the right edit only about 20 to 30% of the time. Those piglets whose genomes have errors end up in a compost heap. I want to convey that this technology is not simple. You can be good at this technology or bad at it," says Mark Cigan, a molecular biologist with a senior role in the program. We need to be rigorous, because we want a predictable change in all the pigs. It has to be the same change every time."

Eradicating influenza

While PRRS is the big problem in the US, Genus and other companies think they can make pigs immune to other viruses too. They are exploring whether gene editing could create pigs that don't catch African swine fever, a disease that's rampant in China and since 2018 has led to the loss of half that country's pigs. Researchers like Rowland say edited pigs could also have the indirect benefit of lowering the chance that certain viruses will spill over from pigs to humans.

The origins of covid-19 are still undetermined, but the prevailing theory is that the disease is zoonotic, meaning it jumped from animals to people. Since pigs don't catch the new coronavirus, they probably played no part in covid-19's emergence. But pig farms are notorious for starting flu pandemics. Pigs can catch both bird and human influenza, in addition to swine flu. That makes them a dangerous mixing vessel in which flu viruses can swap stretches of DNA with each other.

Such a reassortment of genetic parts can suddenly produce a new flu virus that spreads among people, who will not have immunity. The 2009 H1N1 swine flu carried viral elements from birds, pigs, and humans. In the US there were about 61 million cases: almost 300,000 people ended up in the hospital, and around 12,500 died. The deadly 1918 flu pandemic was accompanied in the US by a hog flu," though the connection between them remains unproven.

Starting last year, Genus has been paying a Kansas State University scientist, Jurgen Richt, to help design pigs resistant to influenza. Richt isn't sure he can render pigs entirely immune to the fast-evolving flu viruses, but he's hopeful he can slow the pathogens down, maybe even enough to lower the odds of another pandemic. If you get less replication, you get less mutation, less reassortment," he says. The end result is less evolution of the virus.

Because the receptors influenza attaches to are so common in the body, no animal could survive their removal, Richt says. So the project aims instead to remove other genes, for proteins called proteases that the flu-and covid-19-require as helper molecules to effectively enter cells. Because there are many types of flu, it will be necessary to remove more than one protease, leading to the question of whether pigs with too many deleted genes can thrive. If a pig is a Jenga tower, just how many blocks can be removed before the animal falls apart?

I don't know the limit to taking out genes. That is why we do trial and error," says Richt. But what we want is to make them resistant to all influenzas, from all walks of life."

It's not clear yet whether the PRRS-resistant pigs, with only one receptor removed, are healthy and otherwise normal. Cigan says the company thinks they are; researchers can't see other differences in their tests, which measure things like how much the pigs eat and gain weight. But unplanned changes could be subtle.

Richt says a decade ago he was involved in making cattle resistant to mad cow disease. After removing one gene, he sensed they were changed. The way they stood up was funny-it was hard to get them back up," he says. The caretaker told me they are stupid, so maybe intelligence was affected." With only a dozen cows, he never was sure, but he suspects the cattle lost a luxury function"-one that wasn't vital to survival but whose removal led to a degradation of the sensory system.

Black Plague

If gene editing is perfected in pigs-a species anatomically so similar to humans that doctors hope to transplant pig kidneys to humans someday-what will be the implications for people? The debate about human genetic modification has often been reduced to asking whether it would be moral to change a child's eye color or intelligence, for instance. But the pig hatchery shows that CRISPR might be able to give people inborn genetic vaccines" against the worst infectious diseases they might encounter.

The scientists in China who edited human embryos to resist HIV were pursuing just such a revolutionary development. And the problems they ran into were similar to those Genus faces: they couldn't control the exact edits they made and couldn't be sure that disrupting one gene (called CCR5) wouldn't have unanticipated consequences. In that experiment, though, there were no second tries. In addition, many questioned whether the risky attempt was medically necessary, since drugs can keep HIV under control for decades.

If gene editing is perfected in pigs-a species anatomically so similar to humans that doctors hope to transplant pig kidneys to humans someday-what will be the implications for people?

Since the Chinese fiasco, the American and British science academies have said that gene editing, when it's safe enough to use in human reproduction, should avoid enhancement" of any kind and instead take on narrower goals, such as preventing people from passing inherited conditions like sickle-cell disease to their children.

Yet others think it's important to master the technology as a possible guard against future pandemics. Removing a receptor from the next generations of humans could be civilization's fallback if society is hit with a super-disease that can't be controlled by vaccines or drugs, and for which we don't develop immunity.

We as a species need to maintain the flexibility, in the face of future threats, to take control over our own heredity," George Daley, the dean of Harvard Medical School, told an audience in Hong Kong in 2018. He listed resistance to global pandemics" as one reason to develop techniques to modify human beings.

Covid-19 shows how a novel germ can explode out of nowhere and spread globally. The overall death rate from an infection with the new coronavirus, perhaps 0.5%, doesn't threaten humanity's existence. But what if the next pandemic is more like the Black Plague, which killed one-third or more of the population of Europe in the Middle Ages? It's a remote possibility, like an asteroid strike. But being able to engineer humans to resist specific germs might be a back-pocket technology worth having.

From what they know of animals, scientists at Genus think editing humans is futuristic but not impossible. Twenty years ago, Rice would have said it was pure fiction. But now we can actually do it for animals," she says. We have the tools."