Seminal Work

When is a tragedy also a triumph? When you’re saving an endangered species.

Of all the characters who would eventually take part in that tragic fall day at the Cincinnati Zoo, Nikki was the first awake. An 18-year-old, 4,300-pound Indian rhinoceros, Nikki was 487 days pregnant, and in the early hours of October 26, 2010, she was one restless rhino. She paced through the hay in her stall in the rhino reserve as NPR murmured quietly in the background.

“I got the call around 2:20 [a.m.] from our night watch saying she’s been up and active,” says Randy Pairan, the zoo’s head rhino keeper. Pairan told them he was on his way. He’d stopped for a cup of coffee when the night watch called again to report there was fluid on the floor of Nikki’s stall. Maybe it was urine, maybe something else.

Pairan called Dr. Monica Stoops, the zoo’s reproductive physiologist, and asked her to get on her computer. During Nikki’s pregnancy, her stall had been fitted with cameras. Stoops accessed these from home, pulling up the recent footage to look at the liquid. She thought Nikki’s water must have broken, and she told Pairan she was coming in, too. On her way, Stoops called Paul Reinhart, another keeper, and Dr. Terri Roth, director of the zoo’s research programs.

Before dawn, the quartet converged at the zoo in front of a video feed of Nikki, where they could monitor the delivery remotely, without disrupting it. They’d waited a year and four months for this day, when they hoped to see the world’s first live birth of a rhino conceived by artificial insemination. They were still reviewing the night’s footage when Roth checked the live camera and saw Nikki beginning to give birth. It was 6:06 a.m.

Reinhart and Pairan ran over to the rhino building while Roth and Stoops watched on the live feed as Nikki delivered the calf. They knew immediately something was wrong: the baby wasn’t moving. Entering the building, Pairan knew it too. Nikki stood over her calf, nudging it with her snout. “She was pushing on it, mouthing on it, trying to get some movement out,” Pairan says. “I could sense that she was heartbroken.”

He led Nikki into a separate stall, then began to inspect the calf. When Stoops and Roth raced into the building, Pairan was holding what seemed to be a stillborn male rhino. It wasn’t Nikki’s first calf; she’d been successfully inseminated before and gave birth in January 2008—another first. But that calf had died during delivery, and now that horrible scene was being repeated. “I’m an emotional mess,” Pairan recalls. But as he held the calf, he saw the faintest movement in its neck. “Monica bent down and realized it had a heartbeat,” he says. “She said, ‘We’ve gotta get air into it.’ ”

That’s when Pairan did something he’d never done to an animal before—he started mouth-to-mouth resuscitation. With 45 minutes of CPR and the help of dopram, a drug used to stimulate respiration, the calf started breathing. It was trying to stand up, and started making noises—the staff described it as somewhere between a cow’s “moo” and a sheep’s “baa.” Nikki heard from her stall, let out cries of her own, and the mother and child began calling back and forth.

The staff milked Nikki, who pushed her leg back for easier access, just as she would have if her calf had the strength to suckle. “She was a trooper,” Stoops says. “It’s like she knew what we were doing.” They fed the calf with a tube, then Pairan carried it onto a scale: the newborn was 117 pounds. “There was no doubt in my mind that this calf was going to grow up and do fine,” he says. All four stayed with the animal throughout the day, and when evening came, zoo staff gathered to make plans for the night. Pairan told everyone it would be a long road to recovery. But moments later, that road came to an end.

“It seemed that we all just stop-ped talking at once, looked at the calf, and watched the last breath,” Pairan says. This time, resuscitation failed; its heart had stopped. Thirteen hours after its birth, the first live Indian rhino calf conceived with artificial insemination was dead.

Technically, the birth was a success—another milestone for the zoo and its Lindner Center for Conservation and Research of Endangered Wildlife (CREW), which tackles  some of the thorniest reproductive challenges of captive animals with innovative techniques, using everything from in vitro fertilization in ocelots to artificial insemination in rhinos. But this accomplishment was profoundly bittersweet. A graduate of the Cincinnati Zoo Academy, Pairan has seen plenty of death in his 20 years at the zoo. This one was different; he still fights back tears when he talks about it. “Out of everything I’ve dealt with,” he says, “that was the hardest one ever.”

CREW—the zoo’s conservation research arm—was formed in 1981 to produce healthy, genetically diverse populations of endangered animals in captivity, as a safeguard against extinction. It’s part of a long tradition; as far back as the 1880s, the zoo was breeding trumpeter swans and sea lions. Today CREW conducts valuable research into the reproductive physiologies of some of the world’s most threatened and least-studied animals. The group’s three signature projects are rhinos, small cats, and endangered plants.

When Stoops joined CREW as a post-doctoral candidate in 2002, the zoo had recently won itself a reputation in the (admittedly small) world of rhino reproduction. CREW Director Terri Roth had been studying the reproductive patterns and behavior of Sumatran rhinos since she came to the zoo in 1996; in 2001, Roth’s research culminated in the first calf of that species to be born in captivity in 112 years.

The Sumatran rhino is one of the world’s most endangered species, with fewer than 270 living in the rainforests of Southeast Asia. Maintaining a viable captive population of that species was imperative to protect against its extinction, but it is incredibly difficult to breed these animals in a zoo. Bull rhinos can become aggressive during breeding, so timing the introduction of a female had to be precisely calculated to ensure the best chance of conception. And female rhinos do not give up their reproductive secrets readily. Roth developed techniques for using ultrasound to understand the rhinos’ unique patterns of estrus—that is, when they go into heat. She also determined the necessary dosage of hormones to help a pregnant rhino carry her calf to term. The success of that program is evident: three calves have been born as a result, and one of them, a male named Andalas, has been sent back to a reserve on Sumatra.

Indian rhinos are a relative triumph of wildlife conservation. In the early 1900s, there were as few as 200 remaining in Northeast India and Nepal. Strict hunting bans and the formation of national parks and reserves eventually helped protect them. And while poachers remain a threat—an Indian rhino horn can fetch $30,000 per kilo on the black market—their wild population has grown to about 2,800. But Indian rhinos are still listed as a vulnerable species, and Roth believes their population needs to be managed carefully. When the zoo started looking into Indian rhino reproductive programs in 1998, Roth was able to apply much of what she had learned with their Sumatran kin. “We had this nice template of hormone monitoring and ultrasound,” Roth says. “Now we said, ‘Let’s do that with the Indian Rhino.’”

There are just over 50 Indian rhinos in zoos in North America and Australia, the region that Stoops monitors. Of that population, half descend from just four animals—the best reproducers. With such little genetic variation, the captive rhino population is at risk of becoming inbred. Complicating that is the fact that Indian rhinos, even more so than Sumatran rhinos, can be very aggressive during mating. Sheer size makes Indian rhinos deadly, but they are also fitted with a dental inventory that includes large canine teeth. “The males use those aggressively with females,” says Stoops. “They’ll actually slash females on the side.” Several females have died from wounds they sustained when breeding in captivity. So it makes sense to use the less-aggressive males in breeding programs. Unfortunately, that’s not best for bio-diversity. “We don’t want to lose those genes,” says Roth. “It may be precisely those genes that help it survive in the wild. Whatever we do in zoos, we want to make sure we’re maintaining a population that can survive if we’re putting animals back out, which ultimately we’d like to do.”

Due to the sheer logistics of transporting a 4,000-pound animal with violent tendencies, CREW started looking into artificial insemination as a way to impregnate their female rhinos. But no one had ever artificially inseminated an Indian rhino before, and the process involved some trial and error. The zoo had already developed conditioning techniques to get female Sumatran rhinos to enter a narrow chute, which made it much easier to perform ultrasounds and check hormone levels, and offered an alternative to anesthetizing the animal, which Roth had found to interfere with ovulation. Randy Pairan successfully trained Nikki to enter the chute. “Rhinos are really food-motivated, so it helps,” Stoops says.

CREW also needed a way to acquire the sperm. Stoops and Roth drew from the closest example they could find—the cattle industry, where for decades cattle ranchers have relied on artificial insemination to avoid the inconvenience, expense, and danger of breeding with live bulls.

The method in which semen is collected from a bull—and this is where the story gets a little PG-13—involves a process called “electro-ejaculation.” A probe is inserted into the rectum of an anesthetized bull to stimulate the prostate with a low-voltage electrical current, causing the animal to ejaculate. However, although the technology was in place, the instruments of the cattle industry are insufficiently proportioned for minivan-sized Indian rhinos. So Stoops and her team at CREW had to innovate. Using some PVC tubing, epoxy, and copper wire, the CREW team created a hefty, yard-long probe with a slight curve at the end to accommodate a rhino’s anatomy (Roth joked that the probe resembled the head and neck of the Loch Ness Monster). The device stimulates the prostate and other sex glands with up to nine volts of electricity, which, in a good sample from a healthy male, produces 200 milliliters (about a cup) of semen.

Stoops and Roth have a hit-list of the males with the least-represented genes in the North American captive population. Periodically they pack up the probe and travel to a zoo to collect a sample. The sperm they used to inseminate Nikki came from a male named Vinu in the Bronx Zoo, which Roth and Stoops “probed” in 2005.

The nerve center of CREW’s work is a small room in the basement of their building at the zoo where all these semen samples are stored. It’s called the CryoBioBank, and it includes 11 steel tanks of valuable genetic material sitting in liquid nitrogen at a brisk 320 degrees below zero. The tanks contain gametes—mostly sperm—from multiple individuals of about 75 species of animals. Three of the tanks, dubbed the Frozen Garden, contain seeds and shoots from 150 plant species. This genetic repository offers the zoo the chance to manage genetically diverse populations decades, perhaps even centuries, into the future.

Roth says the applications for a successful Indian rhino artificial insemination program will be far-reaching. By creating a repeatable model, CREW is helping zoos around the world to diversify the genetics of their captive populations. But she also says that, in the future, scientists may be able to use a captive rhino’s sperm to artificially inseminate females living on small reserves in the wild, which also face the risk of inbreeding. Further still, Roth hopes that one day conservationists will be able to collect sperm from a wild bull, and use it to impregnate a wild female in another reserve to keep the gene pool from going stagnant.

Most people still make the distinction between a captive population and a wild population of animals. Roth sees the animal world differently. Most “wild” Indian rhinos are managed to some extent—protected by Indian and Nepalese rangers, relocated to parks that have lost their rhino populations, and, when necessary, kept out of human-dominated areas for their own safety. “It’s actually just a continuum of managed populations,” says Roth. “There’s really no wild anymore.”

Pairan, Stoops, Roth, and the rest of CREW are hoping that the third time will be the charm for Nikki. Stoops says the science is sound behind the process of artificial insemination, hormone regulation, and ultrasound monitoring, so the procedure will remain the same. For Nikki’s next delivery, however, she’ll be sure to have a neonatal specialist on hand.

The zoo added another Indian rhino in 2009, a 5-year-old female named Manjula who came from The Wilds, a 10,000-acre wildlife reserve 20 miles outside of Zanesville, Ohio. Pairan is in the process of training Manjula to be comfortable in the chute, and Stoops hopes to inseminate her soon.

After losing her calf, Nikki went through the expected physiologic response. Her hormone levels dropped, and it took a week for her milk to dry up and some time for her to begin ovulating again. But on December 7, she was inseminated once again. This time the sperm is from Assam, a male at The Wilds, who has not yet fathered a rhino in the captive population. At press time Stoops hadn’t yet confirmed a pregnancy, but the staff was encouraged. “She’s done really well,” Stoops says. “She’s done better than we have.”

Nikki certainly seems content. When Pairan approaches her with a bucket of treats on a recent afternoon in her stall in the rhino reserve, she sticks her head out of a slot and gladly engulfs his hand with her prehensile lip, crunching down the biscuits and entreating him for more. It’s not difficult to see the fondness Pairan has for this animal, whose calf he held in his arms in an ultimately futile effort to save its life. When the treats are gone, Nikki sidles up to the bars of her stall and lies down so that Pairan can scratch her belly. Her thick skin folds back like overlapping plates of armor, but her keepers say she can feel a fly landing on her back.

Stoops and the others hope to see Nikki become a rhino mom; she certainly seems to be ready for motherhood. “Will we be ready?” Stoops asks. “I don’t know.”

“I never want to go through another day like that Tuesday,” Terri Roth says. “But on the other hand, I’ve never seen anything so amazing in my life. That calf started breathing and moving and making noise. We had it for 13 hours. That’s so much better than never having it take a breath at all.”

Originally published in the February 2011 issue.
Illustration by Peter Ryan

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