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Glass Glass Baby: Birth of Healthy Kittens following Sperm Vitrification for Artificial Insemination

A world leader in small cat reproductive research, our Center for Conservation and Research of Endangered Wildlife (CREW) has successfully produced the first non-human offspring – two kittens – using vitrified sperm (semen preserved as glass instead of ice) for artificial insemination (AI).  Dr. Bill Swanson explains the significance of this breakthrough in this video.

Vito and Elsa, kittens produced from vitrified sperm

Vito and Elsa, kittens produced from vitrified sperm

Cryopreservation of cat semen for assisted reproduction can be challenging, requiring technical expertise and specialized equipment for semen collection, processing and freezing. As a simplified alternative to standard semen cryopreservation methods, CREW scientists have been investigating the use of vitrification – the ultra-rapid cooling of liquids to form a solid without ice crystal formation. This approach essentially preserves semen as glass instead of ice.

For vitrification, cat semen is diluted in a chemically-defined medium containing soy lecithin and sucrose as cryoprotectants and, after a five minute equilibration period, pipetted in small volumes (~30 microliters) directly into liquid nitrogen to form tiny glass marbles of frozen sperm.

In initial studies, we found that cat sperm survived vitrification as successfully as that frozen using standard straw freezing methods and that vitrified sperm were capable of fertilizing cat oocytes in vitro. In our first assessment of in vivo viability, eight of these embryos were transferred into three synchronized females. Although one female appeared to have two early implantations, no offspring were produced.

In a follow-up study, artificial insemination (AI) with vitrified sperm was assessed in three additional females. With our laparoscopic oviductal AI technique (LO-AI), only a couple million sperm are required per insemination, allowing the use of the relatively low sperm numbers that are preserved in vitrified semen pellets. Following LO-AI with vitrified sperm, all three females conceived, with two of the pregnancies progressing to term and culminating in the birth of two healthy kittens in early April. These kittens, a male named Vito (short for vitrification) and a female named Elsa (after the character in the movie Frozen), are the first non-human offspring – of any species – produced with vitrified sperm (although three human babies have been born from earlier research).

Vito and his mother, Ebony

Vito and his mother, Ebony

Elsa and her mother, Ivy

Elsa and her mother, Ivy

Our preliminary results with semen from fishing cats and ocelots indicate that vitrification is effective for preserving post-thaw sperm viability and function across cat species. These findings suggest that this fast and simple cryopreservation method may have broad applicability for semen banking of endangered felids housed in zoos and possibly living in the wild.

Jaci Johnson with Elsa and Vito

Jaci Johnson with Elsa and Vito

The study’s lead author, veterinary student Jaci Johnson, has been selected to present these findings at the upcoming American Association of Zoo Veterinarian’s Annual Meeting in Portland, Oregon. (Funded, in part, by the Procter & Gamble Wildlife Conservation Scholarship program in collaboration with Ohio State University’s College of Veterinary Medicine.)

- Reprinted from the Spring 2015 CREW Progress Report

May 14, 2015   1 Comment

Small Cat Conservation Gets a Big Boost with a Federal Grant

You probably already know that the Cincinnati Zoo is committed to the conservation of lions, tigers and cheetahs, but did you know that we are also leading the way in small cat conservation? And our Small Cat Signature Project just got bigger! Our Center for Conservation and Research of Endangered Wildlife (CREW) recently received a Museums for America Grant from the Institute of Museum and Library Services to improve our ability to maintain healthy captive populations of five small cat species across the country—the Brazilian ocelot, the Pallas’ cat, the black-footed cat, the Arabian sand cat and the fishing cat.

Ocelot (Photo: Cassandre Crawford)

Ocelot (Photo: Cassandre Crawford)

Pallas' cat (Photo: Mark Dumont)

Pallas’ cat (Photo: Mark Dumont)

Black-footed cat (Photo: Kathy Newton)

Black-footed cat (Photo: Kathy Newton)

Sand cat (Photo: Mark Dumont)

Sand cat (Photo: Mark Dumont)

Fishing cat (Photo: Kathy Newton)

Fishing cat (Photo: Kathy Newton)

Unfortunately, none of these small cat populations are considered sustainable through natural breeding alone. That’s where Dr. Bill Swanson, CREW’s Director of Animal Research and the world’s leading expert on small cat reproduction, comes in. Working in partnership with Dr. Jason Herrick of the National Foundation for Fertility Research and the Species Survival Plan coordinators for each species, Dr. Swanson will direct the project with a focus on three goals: 1) Collect and freeze semen from the most valuable cats for each species, 2) Produce viable offspring using artificial insemination in recommended breeding pairs that fail to reproduce naturally, 3) Produce offspring with frozen-thawed semen from genetically valuable or under-represented males.

CREW Scientists perform an artificial insemination procedure on a Pallas’ cat. (Photo: Shasta Bray)

CREW Scientists perform an artificial insemination procedure on a Pallas’ cat. (Photo: Shasta Bray)

Arabian sand cat kittens produced through artificial insemination and embryo transfer.

Arabian sand cat kittens produced through artificial insemination and embryo transfer.

Building on CREW’s decades of ground-breaking research on small cat reproduction, successful completion of this project will greatly enhance the sustainability and stewardship of small cat collections in AZA zoos. Now that’s big news!

October 9, 2014   1 Comment

Another First for CREW: Indian Rhino Posthumously Fathers a Calf

CREW has done it again! We are excited to announce the birth of a female Indian rhino calf produced by artificial insemination (AI) conducted by CREW Reproductive Physiologist, Dr. Monica Stoops, and born on June 5 at the Buffalo Zoo. From a historical standpoint, this is the first offspring for a male rhino who never contributed to the genetics of the Indian rhino population during his lifetime – a major victory for endangered species around the world and a lifetime of work in the making.

Rhino calf Monica

Rhino calf Monica

The father, “Jimmy,” died at the Cincinnati Zoo in 2004.  Over the course of those ten years, Jimmy’s sperm was stored at -320°F in CREW’s CryoBioBank™ in Cincinnati, before it was taken to Buffalo, thawed and used in the AI.

Jimmy the Indian Rhino

Jimmy the Indian Rhino

“We are excited to share the news of Tashi’s calf with the world as it demonstrates how collaboration and teamwork among the Association of Zoos & Aquariums (AZA) organizations are making fundamental contributions to rhino conservation,” said Dr. Monica Stoops. “It is deeply heartening to know that the Cincinnati Zoo’s beloved male Indian rhino, Jimmy, will live on through this calf and we are proud that CREW’s CryoBioBank™ continues to contribute to this endangered species’ survival.”

“Tashi,” the Buffalo Zoo’s 17-year-old female has previously conceived and successfully given birth through natural breeding in both 2004 and 2008.  Unfortunately, her mate passed away and the Buffalo Zoo’s new male Indian rhino has not yet reached sexual maturity. Because long intervals between pregnancies in female rhinos can result in long-term infertility, keepers at the Buffalo Zoo knew it was critical to get Tashi pregnant again and reached out to Dr. Stoops for her expertise.

In February of 2013, Dr. Stoops worked closely with Buffalo Zoo’s rhino keeper Joe Hauser and veterinarian Dr. Kurt Volle to perform a standing sedation AI procedure on Tashi. Scientifically speaking, by producing offspring from non or under-represented individuals, CREW is helping to ensure a genetically healthy captive population of Indian rhinos exists in the future.  This is a science that could be necessary for thousands of species across the globe as habitat loss, poaching, and population fragmentation (among other reasons) threaten many with extinction.

The Buffalo Zoo staff monitored Tashi’s pregnancy over the 15-16 month gestation period and at 3:30 p.m., on June 5, she gave birth to a healthy female calf, weighing 144 pounds.

Rhino calf Monica,  Lead  Rhino Keeper Joe Hauser, CREW Reproductive Physiologist Dr. Monica Stoops

Rhino calf Monica, Lead Rhino Keeper Joe Hauser, and CREW Reproductive Physiologist Dr. Monica Stoops with CryoBioBank

“Without Dr. Stoops’ dedication to the species, and to the development of AI science, there is no doubt this calf would not be here today,” said Hauser. “She has spent countless hours spear-heading research and technology for Indian rhino conservation and the Buffalo Zoo is excited to acknowledge that dedication and announce that the name of the calf is “Monica.”

Tashi’s calf demonstrates that AI science is a repeatable and valuable tool to help manage the captive Indian rhino population. With only 59 Indian rhinos in captivity in North America and approximately 2,500 remaining in the wild, being able to successfully introduce genetics that are non or under-represented in the population is critical to maintaining the genetic diversity necessary to keep a population healthy and self-sustaining.

“We are always thrilled to welcome a new baby to the Buffalo Zoo, but this birth is particularly exciting because the science involved is critical to saving endangered animals,” said Dr. Donna Fernandes, President of the Buffalo Zoo. “This type of professional collaboration among AZA Zoos is vital to the important work we do as conservation organizations and we are honored to play a critical role.”

July 3, 2014   No Comments