Protecting biodiversity through understanding amphibian diseases

Amphibians are currently the most threatened group of vertebrates on the planet because of habitat destruction, poaching to support an insatiable international pet trade, pollution, and, increasingly, disease.

One disease, the chytrid fungus Batrachochytrium dendrobatidis (Bd), which infects the skin of frogs, is particularly worrisome. Over the past two decades, Bd has been blamed for mass die-offs and species extinctions of amphibians from Australia to South America, and it is now causing problems in Canada.

Pierre Echaubard mesuring a Northern Leopard Frog

Pierre Echaubard, with a Northern Leopard Frog, during fieldwork at Ontario’s Bruce Peninsula. Credit: Magalie Baril

Of major concern is the endangered Northern Leopard Frog in British Columbia and Alberta. The fungus has infected these populations since the late 1990s. The frogs can die when infected, and attempts to reintroduce them have met with limited success. Curiously, the fungus does not have the same virulence with the Northern Leopard Frog in eastern Canada, where populations are so stable that they are not even listed as a species at risk.

Bruce Pauli, an Environment Canada biologist specializing in ecosystem health, is trying to get to the bottom of this mystery with the help of genomics-based tools. One possibility is a subtle genetic difference between the two frog populations. With funding from the Genomics Research and Development Initiative (GRDI), Pauli is collaborating with a U.S. laboratory that is working up the genome of the two populations to see whether this is the case.

Another possibility is that there could be something in the western habitats that is stressing the frogs and weakening their immune systems, making them unable to fight off infection. Or there could be a genetic difference in the western strain of the fungus, making it more virulent, although Pauli admits that we currently have little information on the genetic variability in the fungus.

Pauli’s overarching focus, however, is on charting the distribution, prevalence and virulence of amphibian disease in Canadian ecosystems, and in addition to the chytrid fungus, he has Ranavirus, a group of deadly viruses, in his sights. He has noticed that the strain of Ranavirus that infects amphibians is particularly hard on Wood Frogs in certain locations. What the two diseases have in common, though, is that they are thought to be relatively recent additions to Canadian ecosystems.

In North America, chytrid fungus has been traced to the use of African Clawed Frogs in pregnancy testing starting in the late 1930s. As Pauli describes it, research has shown that Bd was established in African Clawed Frogs in South Africa by 1938. Around this time, it was discovered that these frogs were useful for human pregnancy testing, with the result that enormous quantities of them were shipped around the world, along with the Bd that was infecting them.

It was only a matter of time before the fungus was introduced and distributed across the entire North American continent. In fact, after the earliest record of Bd in South Africa, the next earliest case was discovered in Canada, in another project Pauli was involved with, that found the fungus in two green frogs collected by the Canadian Museum of Nature in 1961 in Quebec’s St. Lawrence Valley. The problem with these introduced invasive pathogens is that the native animals may have no natural immunity to them, and the effects can be deadly.

The genomics techniques developed and field-tested during the Environment Canada’s GRDI project are allowing Pauli to track the two diseases much more efficiently than with traditional methods. For example, the polymerase chain reaction used to amplify DNA requires just a tiny sample from an animal. A key result of Pauli’s research confirmed that sampling for the skin-borne chytrid fungus could be done simply by rubbing a cotton swab on the skin of the amphibian. Even Ranavirus, which is a blood-borne disease, can be picked up with a swab test, although not with the same efficacy.

This non-invasive testing provides big benefits from a conservation perspective. Foremost is that no animals have to be sacrificed, or at a minimum, have a couple of toes clipped off to provide an adequate sample. Although the jury is still out as to whether toe clips weaken an animal, Pauli explains, "The swabs don’t add any possible stressor into your study population, as the toe clips might, and you definitely would not want to use invasive techniques on endangered species."

He is also interested in seeing what happens when a disease moves through an area. In the summer of 2011, GRDI funding supported his participation in a large field study of amphibian health in New Brunswick ponds, and the research team investigated the mortality of Wood Frogs. Samples taken as it unfolded enabled the group to prove that it was a Ranavirus-related mass mortality event. As it was the second year of the study, they had baseline information from the first year’s sampling program.

As a result, these ponds provide a perfect natural laboratory, and Pauli will monitor them to see what happens to the impacted frog populations. He hopes to find out how disease influences the structuring (that is to say the numbers of frogs at different ages and the ratio of males to females) and viability of these populations.

Pauli’s research is providing a first time look at chytrid fungus and Ranavirus from the perspective of their prevalence, distribution and impact on amphibian populations in Canada. One of the most surprising results he says has been how ubiquitous both diseases are. "Everywhere we sample, we find them."

All the more reason, Pauli concludes that, "We definitely need to learn more about the dynamics of these diseases and what impact they may be having on our native amphibian species."

Building the next generation of researchers

Pierre Echaubard completed his PhD thesis at Laurentian University. During his PhD, he investigated viruses that infect amphibians, specifically what factors modulate the virulence of Ranavirus on its host. In 2009, on the recommendation of his Laurentian supervisor, Dr. David Lesbarrères, he joined Bruce Pauli’s GRDI-funded project team. Echaubard says the link with Environment Canada proved to be a valuable one. "It exposed me to more applied research and led to two major research projects that form part of my thesis."

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