There are only two males left. Are they the loneliest animals?

There are only two confirmed Yangtze giant softshell turtles left on Earth, both male.

In 2012, when the last Pinta Island tortoise on Earth, "Lonely George", passed away, the title of "the loneliest animal" fell to the Yangtze giant softshell turtle.

The Yangtze giant softshell turtle is a slightly unfamiliar name. It is the largest living turtle in the world, a first-class protected animal in my country, and a critically endangered species of the World Conservation Union. It has appeared many times in "Dream of Red Mansions" and "Journey to the West": the "bald turtle" mentioned by Jia Baoyu in his bet with Lin Daiyu, and the old turtle that once carried Tang Monk and his disciples across the Tongtian River, are all referenced in reality by the Yangtze giant softshell turtle.

Although the number of Yangtze giant softshell turtles is in the single digits, humans have not given up on saving them.

The last known female Yangtze giant softshell turtle has passed away

In April this year, relevant Vietnamese authorities confirmed that a female Yangtze giant softshell turtle had died in Dong Mo Lake.

The Yangtze giant softshell turtle was 1.56 meters long and weighed 93 kilograms. The cause of death is still unclear. It was sampled and marked in 2020 and released into Tongmo Lake. It is currently the only confirmed female Yangtze giant softshell turtle in the world.

This news surprised Rao Dingqi, an expert on amphibians and reptiles at the Kunming Institute of Zoology of the Chinese Academy of Sciences. He has been paying attention to and studying the Yangtze giant softshell turtle since its identity was confirmed in 2002.

"The death of this female Yangtze giant softshell turtle has reduced the known number of Yangtze giant softshell turtles on Earth from three to two. Unfortunately, the remaining two are both males, one lives in Suzhou Zoo in my country, and the other lives in Xuanhan Lake in Vietnam."

For a long time, the Yangtze giant softshell turtle was called "癞头缋" and was considered by scholars to be a type of softshell turtle. However, a closer look reveals that the Yangtze giant softshell turtle and the softshell turtle are very different.

The giant softshell turtle can hardly weigh more than 100 kilograms, but the giant softshell turtle can. The giant softshell turtle is round in shape, while the giant softshell turtle is oval. In addition, the giant softshell turtle has fine yellow spots all over its body and bright patterns on its head. Even though the difference between the two is obvious, people in the past still mistakenly confused the two.

It was not until 1873 that a British scholar realized that the "癞头缋" might be a new species and named it "Sternberg's Softshell Turtle", but it did not attract much attention for more than 100 years. It was not until 1993 that Zhao Kentang, a Chinese biologist, confirmed that the "癞头缋" was an independent species after extensive research and gave it the Chinese name "斑龟". Since then, the identity of this species that has lived on Earth for hundreds of millions of years and once lived next to dinosaurs has finally been clarified.

The Yangtze giant softshell turtle is the largest freshwater softshell turtle in the world. It once lived widely in the Yellow River, Yangtze River, Taihu Lake, Qiantang River, Red River, and the Ma River Basin in Vietnam. However, it was always considered to be another species and did not attract enough attention from humans. As soon as its identity was confirmed, it became endangered. In 2005, a Yangtze giant softshell turtle in the Beijing Zoo died, just one year after people identified it as a Yangtze giant softshell turtle. In 2006, a Yangtze giant softshell turtle died in the Shanghai Zoo, less than three months after it was "identified".

What factors have caused the Yangtze giant softshell turtle to become endangered? Lv Shunqing, a professor at the School of Life and Environmental Sciences of Huangshan University, analyzed the reasons.

First of all, this is related to the survival habits of the giant softshell turtle. The giant softshell turtle is a solitary animal with strong territorial behavior. Territorial behavior is conducive to the animal's feeding, reproduction and raising offspring. "Simply put, a giant softshell turtle has to occupy a large area. Once other giant softshell turtles invade, a population fight will begin."

Secondly, in order to maintain its huge size, the Yangtze giant softshell turtle, a "foodie", is either eating or on the way to find food. Therefore, it needs large freshwater bodies such as Taihu Lake and Red River as its habitat.

With the frequent human activities, the survival situation of the Yangtze giant softshell turtle in the wild is getting worse and worse. As cold-blooded animals, they are particularly sensitive to changes in the surrounding environment. "Since 1972, no Yangtze giant softshell turtle has been found in Taihu Lake. According to the survival needs of the Yangtze giant softshell turtle, the only place in China where wild Yangtze giant softshell turtles may exist is the Red River."

Finally, the idea that "eating turtles can prolong life" has led to the decline of giant softshell turtles. As early as the pre-Qin period, Chinese softshell turtles were used as snacks. According to existing data, there were still dozens of giant softshell turtles in my country in the 1950s. However, under the idea that eating turtles can nourish and prevent diseases, people in the past over-hunted wild turtles.

"Even if turtles can live for hundreds of years, they can't survive being eaten." Lü Shunqing said that in terms of body shape, the young giant softshell turtle looks very similar to the Chinese softshell turtle and the mountain softshell turtle. In addition, it has been misidentified for a long time and cannot be protected. Therefore, the newly born giant softshell turtle may be eaten as the Chinese softshell turtle.

Scientists have tried to "continue the legacy" of the Yangtze giant softshell turtle

Lü Shunqing's right thumb is forever scarred. It was the first time he was bitten by a giant softshell turtle, and probably the last time. Not long after biting Lü Shunqing, the giant softshell turtle, named "Xiang Xiang," died.

This was the only female Yangtze giant softshell turtle whose sex was confirmed in my country at that time. In 2019, it died without any warning during artificial insemination.

When talking about the story of this female Yangtze giant softshell turtle, Lv Shunqing's throat tightened. "It all started when we identified it in 2007."

In January 2007, Lü Shunqing was attending a meeting in Suzhou. He saw several photos of giant turtles that needed to be identified at the Changsha Zoo. He and Australian expert Gerard Cooklin keenly discovered that they might be Yangtze giant softshell turtles. They hurriedly flew to Changsha and confirmed on the spot that this was a female Yangtze giant softshell turtle about 90 years old and still able to lay eggs.

At that time, there were only a few Yangtze giant softshell turtles in the world. The discovery in Changsha was a big surprise, as it meant that the Yangtze giant softshell turtle could reproduce naturally.

At that time, Suzhou Zoo had a male Yangtze giant softshell turtle, and both zoos were reluctant to transport their baby to the other zoo. In order to allow them to mate naturally, after a lot of work, Changsha Zoo finally agreed to "marry off their daughter." In May 2008, the female Yangtze giant softshell turtle, known as "Xiangxiang," arrived in Suzhou.

In 2009, the female Yangtze giant softshell turtle "Xiangxiang" basked in the sun at Suzhou Zoo. Photo by Lü Shunqing

Since the two turtles had not seen each other for a long time, the staff used a fence to separate them to prevent them from hurting each other. On the first day, the two turtles could only stare at each other. On the second day, the fence was opened and the male turtle approached, but Xiangxiang hid in the water and did not show up. On the third and fourth days, Xiangxiang was sunbathing on the shore, and the male turtle swam around her. On the fifth day, they "consummated their marriage".

Everything went smoothly, and Xiangxiang quickly laid dozens of eggs, but disappointment soon followed. None of the eggs were fertilized. People began to analyze the reasons. Was it because the two turtles had been together for too short a time and failed to mate? Or was it because the food was too monotonous? Or was it a water quality problem? Some people also believed that they had mated successfully, but no fertilized eggs were formed for the time being. Since the sperm of male turtles can survive for months or even years after entering the female's body, Xiangxiang may lay fertilized eggs and hatch babies one day in the future.

Since there is no experience in breeding the giant softshell turtle, scientists dare not act rashly and can only wait.

Six years later, the situation remained the same: they mated every year and laid eggs, but none of them were fertilized. Scientists began to doubt the quality of the male Yangtze giant softshell turtle's sperm, after all, it was already over 100 years old.

In the following years, artificial insemination failed. Seeing that the two giant softshell turtles were getting older, the team planned to try again in 2019, but no one expected that the one who died was the relatively healthy and young Xiangxiang.

On April 12, 2019, a team of Chinese and foreign experts collected semen and performed artificial insemination on the Yangtze giant softshell turtle couple. Before the operation, the physical and chemical indicators and ultrasonic health examinations of the two Yangtze giant softshell turtles showed that they were in good health.

Similar to previous artificial insemination experiments, the process went smoothly without any complications. The only minor incident was that when preparing to give Xiangxiang anesthesia, Lv Shunqing was responsible for controlling Xiangxiang's left forelimb, and he was bitten by Xiangxiang, who was blindfolded.

After the operation, everyone waited for Xiangxiang to wake up, but after two hours, she still hadn't woken up. The team began to get nervous, and the veterinarian followed the standard procedures to rescue her. Until noon the next day, Xiangxiang occasionally woke up, but moved a few times and then stopped moving again. On the afternoon of April 13, Xiangxiang died, and all the ladies present shed tears.

Lu Shunqing's hand still ached slightly, but his heart hurt even more.

Chinese scientists successfully decipher the genome of Yangtze giant softshell turtle

After a brief moment of depression, the expert group quickly got back to work. The team collected Xiangxiang's ovarian tissue and stored it in liquid nitrogen for future use; the zoo quickly formed a third-party autopsy team composed of Chinese and foreign experts.

Rao Dingqi also participated in this. In order to avoid the permanent loss of the genetic information of the female Yangtze giant softshell turtle genome and to continue the investigation and protection of the Yangtze giant softshell turtle, he collected tissue samples from Xiangxiang during her pathological anatomy examination.

More than a year later, the research results of Rao Dingqi's team were published. Through gene sequencing technology, they successfully assembled a high-quality reference genome sequence at the chromosome level of Xiangxiang. Then, the team used strategies such as de novo prediction, homology alignment and transcript-assisted annotation to successfully decode 26,646 high-quality protein-coding genes of Xiangxiang, and through chromosome analysis, clarified the sex determination system of the Yangtze giant softshell turtle.

Based on the genetic data set and the phylogenetic tree construction algorithm, the academic community has a better understanding of the giant softshell turtle.

The genome size of female Yangtze giant softshell turtle is about 2.24 billion bases; the Yangtze giant softshell turtle is closely related to the Chinese softshell turtle, and they parted ways 54.4 million years ago; the Yangtze giant softshell turtle population has shown a continuous downward trend in history, which is related to its low genetic heterozygosity and genetic diversity.

Comparative genomics analysis also shows that the longevity and large size of the giant softshell turtle are mainly related to the rapid evolution of core genes related to growth, such as cell autophagy and DNA damage response. In addition, the giant softshell turtle genome lacks genes related to tooth formation, which can also explain why the giant softshell turtle, including turtle species, has no teeth...

Rao Dingqi said that the reason for preserving some of Xiangxiang's tissues was mainly to determine the DNA sequence of the giant softshell turtle, draw the genome map of this species, and decipher its genetic code, which will be of great significance for future field surveys, searches and protection of the giant softshell turtle. "For example, scientists can design more sensitive, specific, and effective genetic molecular probes, which can help us more effectively use environmental DNA methods to detect, search, discover and identify more potential wild giant softshell turtles in field surveys."

If mammals can be cloned, why can't the giant softshell turtle?

Zheng Ping, a researcher at the Kunming Institute of Zoology of the Chinese Academy of Sciences, explained: "Cloning technology is not as simple as everyone imagines. At present, cloning technology is relatively mature for mammals, but Yangtze giant softshell turtles are reptiles that reproduce by laying eggs, so scientists have not yet explored cloning technology for them."

Zheng Ping said that when the population of a species falls below the lower limit, the gene pool of the species is incomplete and cannot be repaired. Even if it is "resurrected" by cloning, it will only revive an individual, not the species.

In fact, the number of wild turtles and tortoises in China has been declining. Although turtles and tortoises are among the most tenacious and long-lived species in the world, the reality is that China's wild turtles and tortoises are becoming endangered. Currently in the wild, the Yunnan box turtle, the golden-headed box turtle, and the black-necked turtle are almost extinct; the four-clawed tortoise, the Pan's box turtle, the Baise box turtle, the mountain softshell turtle, and the concave-shelled tortoise are all endangered or critically endangered.

The last hope may be in Madushan Reservoir

The Yangtze giant softshell turtle has been dubbed a "zombie species" by zoologists because it is unable to produce offspring due to the fact that only one or a few long-lived individuals remain.

But, perhaps there is still a glimmer of hope.

Just one month after the death of the female Yangtze giant softshell turtle in Vietnam, an anxious Rao Dingqi went to the Madushan Reservoir in Yunnan, which is believed to be a place in China where wild Yangtze giant softshell turtles may still exist.

There is evidence to prove the location of the Madushan Reservoir. Vietnam's Tong Mo Lake is connected to the Red River, and the Madushan Reservoir is located in Honghe Prefecture, Yunnan Province. In 2007, it was filled with water and flooded a 50-kilometer-long river bank. This section of the river may be an important habitat for the Yangtze giant softshell turtle in the wild in my country. Shortly after the reservoir was built, local fishermen still saw giant softshell turtles basking in the sun on sunny days.

Rao Dingqi and Lü Shunqing also heard some news. According to local fishermen, there may be one or more giant softshell turtles living in the Madushan Reservoir. Many years ago, when someone was fishing illegally, fishermen saw a giant softshell turtle floating on the water after being electrocuted. People who saw the giant softshell turtle described it as an ancient turtle with a huge body.

The two scientists have always believed that there may still be wild individuals or even populations of Yangtze giant softshell turtles in the Red River Basin. However, they are also worried that if it is a solitary Yangtze giant softshell turtle, it will have no practical significance for the reproduction of the population. If it is a pair, it would be great.

It is indeed difficult to find a giant softshell turtle in a 50-kilometer-long body of water. The two have lost count of how many times they have been there. Since giant softshell turtles spend most of their time underwater, it is unlikely to be seen, and they rarely come ashore to bask in the sun, let alone lay eggs.

For this investigation, Rao Dingqi and his team spent more than 10 days. Whenever he walked by the reservoir, he would stare at the water surface, for fear of missing any sudden abnormal situation.

The team's method of finding the giant softshell turtle is very direct - walking along the coast, setting up observation points and installing cameras where the giant softshell turtle is most likely to show its head, and using telescopes to patrol the water surface or places where it may bask in the sun; the team also samples the water body and uses environmental DNA methods to detect whether there is excrement and shed skin flakes with giant softshell turtle DNA information. Everyone believes that if they are lucky enough, they can take pictures and videos of the giant softshell turtle and leave evidence.

In May 2023, members of Rao Dingqi's team searched for Yangtze giant softshell turtles in the Red River Basin. Photo by Rao Dingqi

"Although it is impossible to cover the entire reservoir, we consulted fishermen, sorted out several sighting points, and focused our main efforts on these most promising places." Rao Dingqi is not worried that the Yangtze River giant softshell turtles in the water will starve, "There are still fishermen fishing in the Red River basin. If there are still Yangtze River giant softshell turtles, they will not lack food."

Rao Dingqi showed reporters a video clip showing a gray figure in a wide river bay, bobbing up and down on the river surface, but it never surfaced, so it was impossible to tell if it was a giant softshell turtle. "It could also be another large softshell turtle, such as a giant softshell turtle or an Asian softshell turtle."

Sometimes Lü Shunqing and Rao Dingqi would go out into the wild together. They likened this work to "searching for the Loch Ness Monster," an ancient marine reptile that is said to live in a deep lake in Scotland, but has not been confirmed so far.

"What we are doing is similar to searching for the Loch Ness Monster, so we call the search for the Yangtze Yangtze Giant Softshell Turtle 'Searching for the Madushan Monster.'" Lu Shunqing said that the most urgent thing now is to find evidence that the Yangtze Yangtze Giant Softshell Turtle is still alive.

Scientists have even made plans for "once the Yangtze giant softshell turtle is found" - once found, first protect and monitor it on the spot, and then look for other companions. If they are of different genders, one can be caught, equipped with a locator, and released into the waters where the other one is, to try to mate and reproduce naturally. If only females are found, cooperation can also be discussed with the Vietnamese side.

But the price is bound to be high: the male turtles in Suzhou have little hope of reproducing successfully naturally, and can only rely on artificial insemination; the current health status of the male turtles in Vietnam is unknown. The future of the Yangtze giant softshell turtle is still full of fog, especially when there are no female turtles at present.

Even though the Yangtze giant softshell turtle is sliding towards extinction, scientists have not given up hope.

"Never think about recovering something when you have lost it, but put more effort into preventing extinction," said Rao Dingqi.

"Extinction means never to be reborn. It sounds sad, but it makes people sober when they think about it," said Lv Shunqing. (Reporter Yue Ranran)