Sharks are frequently depicted in movies as fearsome predators with a thirst for human blood. In reality, people are exponentially more harmful to sharks than they are to us: Human pressures kill tens of millions of sharks per year, pushing many species to the brink of extinction and spurring scientists to come up with innovative new ways to conserve these vital ocean animals.
Case in point: Researchers led by Jennifer Wyffels and the South-East Zoo Alliance for Reproduction & Conservation have completed the largest effort to artificially inseminate sharks—an experiment that produced the first shark hatchlings ever fertilized by cold-stored semen.
The unprecedented project yielded 97 baby sharks, with parents that were located thousands of miles apart in some cases, demonstrating that genetic material could be moved between institutions “to maintain gene diversity in a population without the need to transport an animal,” according to a study published on Thursday in Scientific Reports.
As you might imagine, transporting sharks across continents so that they can breed with ideal mates is an expensive and difficult task. While previous experiments have shown that shark eggs can be artificially fertilized by freshly collected semen, Wyffels and her colleagues set out to see if it was possible to pull off the same feat with semen that had been cold-stored at temperatures of 1 to 4 °Celsius for up to 48 hours, enabling it to be transported across huge distances.
“It was never a goal to become the largest shark insemination effort but rather we wanted to investigate several semen parameters or conditions for artificial insemination including using cold-stored or fresh semen,” Wyffels, who is now a postdoctoral researcher at the University of Delaware, said in an email. “Those different treatments required including more females than had been included in previous reports of shark artificial insemination.”
While the most iconic sharks tend to be the gigantic great whites or the distinctive hammerheads, Wyffels’ team worked with a much more manageable species, called the whitespotted bamboo shark. With an adult length of about three feet, these sharks are common in zoos, and even home aquariums, because they are relatively easy to feed and breed in captivity.
“Bamboo sharks were chosen as a model for this research for several reasons,” Wyffels said. “They are a smaller shark species and easier to handle for reproductive procedures. Many aquariums have bamboo sharks and that was important so that we could exchange semen between institutions.”
“Another consideration was reproductive mode, live-bearing versus egg-laying,” she continued. “It is much easier to monitor eggs for embryo development than it is to examine a pregnant female shark. It is always nice to start with a model that is ‘easier’ and work up to more challenging species.”
Whitespotted bamboo sharks are also interesting to researchers because females can reproduce without any help from males through a process called parthenogenesis, resulting in full clones of the mother, known as parthenotes, derived from only an egg and no sperm.
For this experiment, Wyffels and her colleagues collected sperm samples from 19 male sharks and artificially inseminated 20 females. The sperm was primarily gathered using a technique called manual expression, in which were pushed from their sexual organ, the cloaca, by applying pressure to their pelvic area.
Since female whitespotted bamboo sharks can store sperm from multiple males in their reproductive tracts over long durations, the team examined the eggs of their prospective mothers for six weeks to ensure that they were not hoarding any sperm from previous sexual encounters. Once the females cleared that test, they were sedated and inseminated during a 10-minute procedure.
Months later, the incubated eggs hatched, producing dozens of artificially bred baby sharks. Freshly collected semen samples fertilized eggs in 27.6 percent of cases, producing 80 hatchlings.
Meanwhile, semen that was cold-stored for 24 or 48 hours had respective success rates of 28.1 percent and 7.1 percent, producing a total of 17 hatchlings. The results showed, for the first time, that cold-stored samples could not only produce viable offspring, but that they had about the same success rate as fresh samples within a 24-hour period.
“Our Australian colleagues transported semen between institutions and used it later on the same day for artificial insemination,” Wyffels said. “We built upon their success and extended the time between collection and insemination up to two days. I believe longer storage is possible but we did not test it in this study because two days is enough time to transport semen most places in the world.”
In one case, cold-stored semen from a male at Tampa’s Florida Aquarium was shipped overnight to Ripley’s Aquarium of the Smokies in Gatlinburg, TN, where the sample was used to fertilize the eggs of three females. The biggest distance traveled by these cold-stored samples was nearly 2,500 miles, as the crow flies, between a male at the Adventure Aquarium in Camden, NJ, to a female at the Aquarium of the Pacific in Long Beach, California, resulting in two fertilized eggs that successfully hatched.
The study also revealed another fascinating twist: Two of the inseminated females produced clutches that included parthenotes, suggesting that the mothers didn’t use all of the sperm that they were given to reproduce.
“One of the most surprising findings was that three hatchlings developed from parthenogenesis, after females were artificially inseminated,” Wyffels said. “We know almost nothing about the basic reproductive mechanisms that control sperm storage and release, sperm competition, gamete compatibility and parthenogenesis in sharks. What determines the fate of an egg and if it will develop via parthenogenesis or be sexually fertilized? These are just a few of the questions this study brings to mind for future work.”
Now that Wyffels and her colleagues have shown that it’s possible to push the limits of artificial insemination in sharks, they hope to build on their findings to help vulnerable populations recover and thrive, both in captivity and in the oceans. For instance, the team is currently working to apply lessons from their new study to aid reproduction in sand tiger sharks, which are critically endangered in some parts of their range and infrequently reproduce in aquariums.
“We hope artificial insemination will be one of many tools used to support reproduction for sharks in aquariums and eventually even wild populations,” Wyffels said, adding that it will be especially important for species with “fragmented and small populations and those with a high risk of extinction.”
“There is a lot of potential for exchange of genes via semen between aquaria and even between wild populations and aquariums,” she concluded.