First Release Trial to Help Pave the Way for Reintroduction Programs for Critically Endangered Frogs

Ninety Limosa harlequin frogs (Atelopus limosus) bred in human care are braving the elements of the wild after Smithsonian scientists sent them out into the Panamanian rainforest as part of their first-ever release trial. The study, led by the Panama Amphibian Rescue and Conservation Project, aims to determine the factors that influence not only whether frogs survive the transition from human care to the wild, but whether they persist and go on to breed.

“Only by understanding the trials and tribulations of a frog’s transition from human care to the wild will we have the information we need to someday develop and implement successful reintroduction programs,” said Brian Gratwicke, international program coordinator for the rescue project and Smithsonian Conservation Biology Institute (SCBI) amphibian conservation biologist. “Although we are not sure whether any of these individual frogs will make it out there, this release trial will give us the knowledge we need to tip the balance in favor of the frogs.”

The Limosa harlequin frogs released at the Mamoní Valley Preserve, have small numbered tags inserted under their skin so that researchers can tell individuals apart. The scientific team also gave each frog an elastomer toe marking that glows under UV light to easily tell this cohort of frogs apart from any future releases. Smithsonian-Mason School of Conservation Ph.D. student Blake Klocke is currently monitoring the frogs daily at the site, collecting information about survivorship, dispersal, behavior and whether the warm micro-climate in the area provides any protection against disease.

The study is also looking at whether a “soft release” boosts the frogs’ ability to survive. Thirty of the newly released frogs spent a month at the site in cages, acclimating to their surroundings and foraging on leaf-litter invertebrates. Eight of these frogs, and eight that were released without the trial period, are wearing miniature radio transmitters that will give Klocke and team a chance to look at differences in survival and persistence between the two groups. The researchers also collected skin bacteria samples from the soft release frogs to measure changes during their transition from captivity to the wild.

“The soft release study allowed us to safely expose captive-bred frogs to a more balanced and varied diet, changing environmental conditions and diverse skin bacteria that can potentially increase their survival in nature,” said Angie Estrada, Ph.D. student at Virginia Tech and a member of the team leading the soft release, which was funded through a Smithsonian Tropical Research Institute (STRI) grant and support from the National Science Foundation. “It allowed us to monitor health and overall body condition of the animals without the risk of losing the frogs right away to a hungry snake.”


Limosa harlequin frogs are especially sensitive to the amphibian chytrid fungus, which has pushed frog species to the brink of extinction primarily in Central America, Australia and the western United States. The Panama Amphibian Rescue and Conservation Project brought a number of individuals into the breeding center between 2008 and 2010 as chytrid swept through their habitat. The Limosa harlequin frogs in this release trial are the first captive-bred generation of the species, and only part of the rescue project’s total insurance population for the species.

“After all the work involved in collecting founder individuals, learning to breed them, raising their tadpoles, producing all their food, and keeping these frogs healthy, the release trial marks a new, exciting stage in this project,” said Roberto Ibáñez, in-country director of the rescue project and STRI scientist. “These captive-bred frogs will now be exposed to their world, where predators and pathogens are ever-present in their environment. Their journey will help provide the key to saving not only their own species, but Panama’s other critically endangered amphibian species.”

The Panama Amphibian Rescue and Conservation Project is a project partnership between the Cheyenne Mountain Zoo, the Houston Zoo, Zoo New England, the SCBI and STRI. This project received additional support from the Friends of the National Zoo, Holohil, The Woodtiger Fund, Mamoni Valley Preserve and Earth Train.

SCBI plays a leading role in the Smithsonian’s global efforts to save species from extinction and train future generations of conservationists. SCBI spearheads research programs at its headquarters in Front Royal, Va., the Smithsonian’s National Zoo in Washington, D.C., and at field research stations and training sites worldwide. SCBI scientists tackle some of today’s most complex conservation challenges by applying and sharing what they learn about animal behavior and reproduction, ecology, genetics, migration and conservation sustainability.

First-Time Breeding of Frog Suggests Hope for Critically Endangered Species

First captive-bred Craugastor evanesco photo by R. Ibanez.

When researchers discovered Craugastor evanesco in the rainforests of Panama, they called it the vanishing robber frog to signify just how quickly the deadly infectious amphibian disease chytridiomycosis had devastated its population. By the time the researchers had published about the new species in 2010, the vanishing robber frog had already disappeared from the park where they had discovered it.

Now, however, the vanishing robber frog may have a fighting chance at a future thanks to the Panama Amphibian Rescue and Conservation Project, which in December became the first program to breed the species in human care. After multiple attempts at breeding the species since 2015, a single pair has now produced one offspring—a success that has encouraged a cautious optimism that the rescue project can replicate the effort.

“A single individual doesn’t make a successful captive breeding program, but demonstrates that it can be done,” says Brian Gratwicke, an amphibian conservation biologist for the Smithsonian Conservation Biology Institute and rescue project international coordinator. “Every journey begins with the first step and this is a critical first step, not just for this species, but potentially for other endangered amphibians with similar reproductive needs.”

The rescue project, a world-class amphibian center run by SCBI and the Smithsonian Tropical Research Institute, currently has a founding population of 20 males and 20 females of the vanishing robber frog. Conservationists collected the frogs from a lowland site in central Panama where the rescue project is working with the support of Minera Panama S.A. to conserve amphibians in the area. But bringing a new and critically endangered species into human care requires learning its own unique husbandry and reproductive needs before it blinks out of existence—sometimes resulting in insurmountable challenges.

“Piecing together a species’ natural history with artificial systems, we can recreate to the best of our abilities an environment where the animals feel comfortable enough to breed,” said Heidi Ross, STRI’s director of El Valle Amphibian Conservation Center, whose expertise and persistence led to the successful first-time breeding of the species. “If we can get them to this point, to become sexually active in our artificial habitat, then we can simply tweak the system based on what worked, what did not work, and what materials are at our disposal. What we arduously do day in and day out is make sure we are providing the basic needs to the animals so that they help us help them from going extinct in the wild.”

The Craugastor group of frogs has a unique reproductive system called direct development—they bury eggs in wet sand and fully formed miniature adults hatch from the eggs. Understanding the frogs’ reproductive cues, special dietary needs and how to emulate the natural environment is essential to successful breeding, Ross says.

“Given the current difficult situation for amphibians in our region, this project represents scientific and biological hope, not only for this species of frog, but also for the recovery of Craugastor evanesco within its distribution range,” said Blanca Araúz, biologist and biodiversity superintendent of Minera Panamá. “As one of the species of interest for our project Cobre Panamá, its reproduction in captivity is important. Because the deadly infectious disease acts fast, experienced scientists can control the infection in these frogs and breed them under better conditions.”

Although scientists are still occasionally finding individual vanishing robber frogs in the field, they have not found a viable, self-sustaining population. Chytridiomycosis has been linked to dramatic population declines of amphibian species worldwide. This particular group of frogs in the Craugastor rugulosus series are particularly susceptible to chytridiomycosis with three closely related species in Panama having disappeared, putting extra pressure on ensuring the survival of Craugastor evanesco.

“It’s all a learning curve,” Gratwicke says. “I’m hopeful that we’ll be able to replicate this breeding event to develop a sustainable breeding program. If we can do that, we’ll be able to get this species back out in the wild as soon as we figure out how to safely do so. If we can do that, it’ll be time to celebrate.”

The Panama Amphibian Rescue and Conservation Project is a partnership between the Houston Zoo, Cheyenne Mountain Zoo, Zoo New England, SCBI and STRI.

Earth optimism: Frogs

What’s Working in Conservation

The global conservation movement has reached a turning point. We have documented the fast pace of habitat loss, the growing number of endangered and extinct species, and the increasing speed of global climate change. Yet while the seriousness of these threats cannot be denied, there are a growing number of examples of improvements in the health of species and ecosystems, along with benefits to human well-being, thanks to our conservation actions. Earth Optimism is a global initiative that celebrates a change in focus from problem to solution, from a sense of loss to one of hope, in the dialogue about conservation and sustainability.

The Smithsonian will host an Earth Optimism Summit in Washington DC April 21-23, 2017
Please register if you are coming in person or stream the presentations online here: https://earthoptimism.si.edu/calendar/summit Sunday is a free public day.

Dr Brian Gratwicke will present the Panama Amphibian Rescue and Conservation Project on Saturday April 22 5:15pm on the panel Science on the Edge

Love potion for frogs

Scientists at the Smithsonian Institution and partners have published a paper that optimizes sperm collection protocols from the critically endangered Panamanian Golden Frog Atelopus zeteki. It also improves our understanding of reproduction in endangered harlequin frogs. The research, to be published published 15 March 2017, in Theriogenology, was conducted by Dr. Gina DellaTogna, a Panamanian biologist who studied this charismatic animal at the National Zoological Park in Washington DC. The study characterizes the dose-response patterns for several artificial hormone treatments and describes the sperm morphology for the first time in this species.

Atelopus zeteki spermatazoan

“This study is important, because it contributes towards the basic understanding of reproduction of a highly endangered group of frogs in Latin America,” said DellaTogna, who performed the experiments for her PhD at the University of Maryland. “This study has already helped us to solve critical reproduction problems in captive Atelopus collections in Panama and allowed us to repeatedly collect high-quality sperm samples for genome resource banking at any time of the year, without harming the frogs.”

“Basic reproductive research is something that has yielded huge conservation dividends for the successful care and management of other endangered species like Pandas and Black Footed Ferrets,” said Pierre Comizzoli, a co-author of the paper and reproduction specialist at the National Zoo. “Gina’s research opens the door to develop methods like sperm freezing and storage to preserve the long term genetic integrity and diversity in small populations.”
The research is particularly relevant to current amphibian conservation efforts in Panama where the Panama Amphibian Rescue and Conservation Project has captive-breeding colonies of five species of Atelopus that are threatened with extinction from the deadly fungal disease chytridiomycosis.

Roberto Ibáñez, and Gina DellaTogna working on hormonal stimulation of frogs at the Panama Amphibian Rescue and Conservation Project

“Successful reproduction is key to any captive assurance program,” said Roberto Ibáñez, the director of the Panama Amphibian Rescue and Conservation project at the Smithsonian Tropical Research Institute in Panama. “Gina has already begun applying what she has learned to successfully help us to produce offspring from four other endangered harlequin frog species. I hope that she will eventually extend it to species with different modes of reproduction that are also difficult to breed”.

The research was made possible with assistance from the Maryland Zoo in Baltimore who manage the Golden Frog Species Survival Plan. Funding was provided from the Panamanian Government’s Secretaría Nacional de Ciencia y Tecnología (SENACYT), The WoodTiger Fund, the Smithsonian Endowment for Science and the University of Ottawa Research Chairs Program.

Della Togna G, Trudeau VL, Gratwicke B, Evans M, Augustine L, Chia H, Bronikowski EJ, Murphy JB, Comizzoli P. 2017 Effects of hormonal stimulation on the concentration and quality of excreted spermatozoa in the critically endangered Panamanian golden frog (Atelopus zeteki). Theriogenology. http://dx.doi.org/10.1016/j.theriogenology.2016.12.033 

More Good Reasons Not to Lick a Toad

A New Review of Chemicals Produced by the Toad Family, Bufonidae

Cane toad (Rhinella marinus)

As human diseases become alarmingly antibiotic resistant, identification of new pharmaceuticals is critical. The cane toad and other members of the Bufonidae family produce substances widely used in traditional folk medicine, but endangered family members, like Panama’s golden frog, Atelopus zeteki, may disappear before revealing their secrets. Smithsonian scientists and colleagues catalog the known chemicals produced by this amphibian family in the Journal of Ethnopharmacology highlighting this largely-unexplored potential for discovery.

“We’re slowly learning to breed members of this amphibian family decimated by the chytrid fungal disease,” said Roberto Ibañez, Panamanian staff scientist at the Smithsonian Tropical Research Institute (STRI) and in-country director of the Panama Amphibian Conservation and Rescue (PARC) project. “That’s buying us time to understand what kind of chemicals they produce, but it’s likely that animals in their natural habitats produce an even wider range of compounds.”

15 of 47 frog and toad species used in traditional medicine belong to the family Bufonidae. For millennia, secretions from their skin and from glands near their ears called parotid glands, as well as from their bones and muscle tissues have been used as remedies for infections, bites, cancer, heart disorders, hemorrhages, allergies, inflammation, pain and even AIDS.

Toxins of two common Asian toad species, Bufo gargarizans and Duttaphrynus melanostictus, produce the anticancer remedies known as Chan Su and Senso in China and Japan, respectively. Another preparation used to treat cancer and hepatitis, Huachansu or Cinobufacini, is regulated by the Chinese State Food and Drug Administration. In Brazil, the inner organs of the toad, Rhinella schneideri, are applied to horses to treat the parasite Habronema muscae. In Spain, extract from the toad Bufo bufo is used to treat hoof rot in livestock. In China, North and South Korea, ranchers use the meat of Bufo gargarizans to treat rinderpest.

Only a small proportion of the more than 580 species in the Bufonidae family have been screened by scientists. “In Panama, not only do we have access to an amazing diversity of amphibian species,” said Marcelino Gutiérrez, investigator at the Center for Biodiversity and Drug Discovery at Panama’s state research institute, Instituto de Investigaciones Cientificas y Servicios de Alta Tecnologia (INDICASAT),  “we’re developing new mass spectrometry and nuclear magnetic resonance spectroscopy techniques to make it easier and cheaper to elucidate the chemical structures of the alkaloids, steroids, peptides and proteins produced by these animals. We work closely with herpetologists so as not to further threaten populations of these species in the wild.” Their efforts to catalog chemicals produced by the Bufonidae included researchers from the University of Panama, Vanderbilt University, in Tennessee, U.S.A. and Acharya Nagarjuna University in Guntur, India.

Most of the chemicals produced by frogs and toads protect them against predators. Atelopus varius contains tetrodototoxin. Chiriquitoxin is found in Atelopus limosus, one of the first species that researches succeeded in breeding in captivity as well as in Atelopus glyphus and Atelopus chiriquiensis. An atelopidtoxin (zetekitoxin) from the Panamanian golden frog, Atelopus zeteki, appears to consist of two toxins. Toxins from a single frog skin can kill 130-1000 mice.

The golden frog, A. zeteki, Panama’s national frog, is the only species of the genus Atelopus that secretes zetekitoxins. Threatened by the chytrid fungal disease that infects the skin and causes heart attacks, with collection for the exotic pet trade and by habitat destruction, if golden frogs were to disappear, they would take this potentially valuable chemical with them.

More than 30 percent of amphibians in the world are in decline. Racing to stay ahead of the wave of disease spreading across Central America, Panama is leading the way in conservation efforts. The Smithsonian’s Panama Amphibian Rescue and Conservation project (PARC) identified several Atelopus species in danger of extinction, and are learning how to create the conditions needed to breed them in captivity. Not only do animal caretakers at their facilities in Gamboa and El Valle, Panama experiment to discover what the frogs eat, they also recreate the proper environment the entire frog life-cycle: egg laying, egg hatching and tadpole survival, to successfully breed Atelopus. Each species has unique requirements, making it an expensive challenge to create this Noah’s ark for amphibians.

The chemical building blocks amphibians use to create toxic compounds come from sources including their diet, skin glands or symbiotic microorganisms. Toads in the genus Melanophryniscus sequester lipophilic alkaloids from their complex diet consisting of mites and ants. Researchers found that toxins found in a wild-caught species of Atelopus could not be isolated from frogs raised in captivity: another reason to conserve frog habitat and to begin to explore the possibility of releasing frogs bred in captivity back into the wild.

Learn more about amphibians by visiting the PARC blog and the Panama’s Fabulous Frogs exhibit at the Smithsonian’s Culebra Point Nature Center in Panama.

The Smithsonian Tropical Research Institute, headquartered in Panama City, Panama, is a part of the Smithsonian Institution. The Institute furthers the understanding of tropical nature and its importance to human welfare, trains students to conduct research in the tropics and promotes conservation by increasing public awareness of the beauty and importance of tropical ecosystems. Website. Promo video.

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Rodriguez, Candelario, Rollins-Smith, Louise, Ibanez, Roberto, Durant-Archibold, Armando, Gutiérrez, Marcelino. 2016. Toxins and pharmacologically active compounds from species of the family Bufonidae (Amphibia, Anura). Journal of Ethnopharmacology, doi:10.1016/j.jep.2016.12.021

A Golden Day to Fight for Frogs

Golden Frog Day parade (Photo by Brian Gratwicke)

Golden Frog Day parade (Photo by Brian Gratwicke)

Each year around August 14, the streets of El Valle de Anton in Panama fill up with golden frogs. Though they’re not the real amphibians—the Panamanian golden frog is extinct in the wild—school-age children dress up as the animals in a spirited celebration of what has become a popular national holiday: Panamanian Golden Frog Day.

“Panamanian Golden Frog Day is about being thankful for the gift of life that we are able to experience each and every day,” says Katie Uckele, a volunteer at Punta Culebra Nature Center, one of the participants in the celebrations. “The Panamanian golden frog reminds us to cherish the gift of life and celebrate biodiversity in the world.”

Kids pledge to protect frogs on Golden Frog Day. (Photo by Brian Gratwicke)

Kids pledge to protect frogs on Golden Frog Day. (Photo by Brian Gratwicke)

In 2010—just one year after the last confirmed observation of a Panamanian golden frog in the wild—Panama’s National Assembly declared August 13 National Golden Frog Day, passing a law that made the Panamanian golden frog one of Panama’s official cultural and ecological symbols. Since then the holiday has grown from the mere acknowledgement of the National Assembly’s declaration to an entire week full of frog-focused events for children and adults across the country.

This year’s Golden Frog Day started August 13, ran through August 20 and included two family days, a race for frogs, an open house at the Gamboa Amphibian Research and Conservation Center, a book fair with a live frog exhibition and a parade in El Valle.

Golden Frog Day came near the end of the Panama Amphibian Rescue and Conservation Project’s #FightforFrogs campaign, during which time Golden Frog—a global online services provider—matched donations to the rescue project, helping raise money critical to the Zoo’s fight for frogs. The successful digital campaign brought in $21,800 in donations. With Golden Frog’s initial donation of $10,000 and their generous commitment to match up to an additional $20,000, we’ve raised a total of $51,800 for frogs.

Frog jumping competition on Golden Frog Day (Photo by Brian Gratwicke)

Frog jumping competition on Golden Frog Day (Photo by Brian Gratwicke)

“I’m very hopeful for the future of golden frogs and several other highly endangered frogs in Panama,” says Brian Gratwicke, international coordinator for the Panama Amphibian Rescue and Conservation Project. Gratwicke adds that he will be baking golden frog cupcakes for his co-workers in celebration of Panamanian Golden Frog Day. “We have a fantastic team of dedicated conservationists working at maintaining and breeding frogs, and conducting the research needed to put them back in the wild.”

Though no longer found in the wild, the golden frog is a beloved icon in Panama, where local markets sell thousands of enamel-painted terracotta and hand-carved tagua nut golden frog statues, and hand-stitched fabric works of art called molas with the likeness of the amphibian. Last year Panamanian Golden Frog Day celebrations even kicked off with a golden frog-themed national lottery ticket.

Golden Frog Day float (Photo by Brian Gratwicke)

“Panamanian golden frogs mean hope,” says Angie Estrada, a Ph.D. student at Virginia Tech and a native Panamanian. “Hope that Panamanians can reclaim and protect their rivers, streams and forest. Hope that we can collaborate with people from different countries and backgrounds when the goal is larger than our own interests. Hope that we will be able to find more frogs out in the wild, and that if we don’t we will keep looking. Hope that if they disappeared, we will be able to say that we did everything we could to help them out.”

The Panama Amphibian Rescue and Conservation Project is a project partnership between the Cheyenne Mountain Zoo, the Houston Zoo, Zoo New England and Smithsonian Institution. You can follow the Fight for Frogs campaign on Twitter using the #FightForFrogs hashtag or on the rescue project’s Facebook page.