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Emerging parasites

The exterminator of amphibians

Animal trafficking may spread fungus that is lethal for toads and frogs

Fabio ColombiniDendrobates tinctorius, colorful and sensitiveFabio Colombini

An unexpected deviation from the route proved just how harmful the fungus Batrachochytrium dendrobatidis (or Bd) can be for Brazilian amphibians. In June 2006, Cátia Dejuste de Paula was looking for parasites and microorganisms in amphibians for her PhD research at the School of Veterinary Medicine at the University of São Paulo (USP) when a request arrived for her to examine the possible cause of death of two different species of 50 amphibians  in a private zoo in the São Paulo State .

After weighing up all other possibilities the conclusion that she, José Luiz Catão Dias (associate coordinator of USP’s Laboratory of Compared Wild Animals Pathology – Lapcom), and collaborators from the United States reached is that Bd must have been responsible for the death of a large part of the animals examined. This is a direct indication, perhaps for the first time, of the lethal nature of this fungus in Brazil. For years Bd has been associated with the reduction (at times, elimination) of populations of toads and frogs worldwide; Brazilian species seemed immune to this microorganism.

This episode is evidence, and perhaps the first evidence, that animal traffic may favor the dissemination of fungi in amphibians kept in captivity, since the toads had been apprehended before being sent to a private zoo with the authorization of federal government bodies. Detailed in the journal, Diseases of Aquatic Organisms, the study suggests the possibility of contagion between animals kept in captivity, reinforcing the need for preventive measures, such as quarantine and being bathed in antifungal substances.

It is also a clear indication that some species are sensitive to the fungus, while others seem to be resistant. 
Cátia detected the fungus in various stages of development on the skin of 20 of the 30 Dendrobates tinctorius, or dyed dart frog, a colored frog with a mottled black skin with green or blue spots, albeit poisonous. However, there were no signs of the fungus on any of the 20 animals of the other species, Adelphobates galactonotus, also colored and poisonous, which must have died for some other reason.

LUIS FELIPE TOLEDOAdelphobates galactonotus is indifferent to the devastating Bd fungusLUIS FELIPE TOLEDO

It was already known that some species may be resistant to the fungus, but this capacity has not yet been seen as clearly as with Adelphobates galactonotus. The large-bodied American bullfrogs (Lithobates catesbianus), which can reach 20 centimeters in length and weigh 1.5 kg., are generally green or brown in color, is usually  the most quoted species as an example of peaceful coexistence with the microorganism, and is now being considered as a possible transmitter to other species, the researchers warn.

Imported since the 1930s from the United States to be bred commercially, but abandoned after few people showed any interest in buying the skin and meat, the bullfrog spread throughout the country, since it multiplies easily and adapts to both moist and dry environments. Nevertheless, it is not the only involuntary carrier of fungi. In December 2011, in Diseases of Aquatic Organisms, researchers from McGill University in Canada warned that lizards and snakes were transmitters of this organism, thus helping to spread it.

“Although it can be an animal parasite, Bd does not need them as hosts, because it can live on organic material in aquatic environments, like other fungi that survive in the soil, on tree bark or in decaying leaves,” comments Selene Dall’Acqua Coutinho, a professor from Paulista University (Unip) who has been working with fungi for 30 years and who carried out the molecular biology analyses that complemented the examinations of Cátia and Catão. “Bd is probably widely disseminated in the environment.”

This is what Cátia, currently a researcher with the Wildlife Conservation Society in Brazil, observed: “Wherever you look for the fungus you find it.” In her PhD, concluded in 2011, she looked for microorganisms and parasites in 120 animals of 33 different species of toads and frogs collected in the Boraceia biological station, a humid forest area covering 100 hectares that belongs to USP, in the municipality of Salesópolis. She found parasites – mainly in the intestine – in almost half (55) the animals she examined. The most common microorganism found in 22 toads (19.1% of the total) was the Bd, identified by means of microscopic and DNA examinations from a sample provided by Alan Pessier, from the San Diego Zoo, in California.

Fewer toads in the wild
“We saw the fungus but the animals seemed healthy, without skin lesions or alterations,” says Cátia. Neither she nor the other researchers found sick amphibians in Boraceia or in other places, but they observed that the diversity of species and number of amphibians were smaller than a few years ago. Bd sticks to the fine, moist skin of the amphibians and, as the researchers in the United States and Australia indicated in 2009, changes the balance of muscle electrolytes (ions), making some animal species die suddenly from cardiac arrest.

Catia Dejuste de Paula and Luiz Catão-Dias / USP The arrows indicate the fungus on a toad’s skin: infected animals show no signs of being sickCatia Dejuste de Paula and Luiz Catão-Dias / USP

A biologist and professor at USP, Miguel Urbano Trefaut Rodrigues says that years ago toads of the genus Hylodes were common in the Caparaó National Park, on the border between the states of Espírito Santo and Minas Gerais. Last year he walked around there and noted that they had disappeared: “I was shocked.” Toads of the genus Allobates have also grown scarcer in the forests of Espírito Santo and Rio de Janeiro. “A Dendrobatidae, Anomaloglossus, was abundant in the Tepequém Hills in Roraima; we used to see 10, 20 beside waterfalls. Now there are none.”

In 2005, after examining 96 toads of 25 species collected at 10 different points in the Atlantic rainforest, Ana Carnaval from the University of California in Berkeley, Rodrigues, his students and biologists from the University of Costa Rica and Rio de Janeiro significantly expanded the geographic area of the spread of the fungus, which they concluded can live at altitudes that vary from 100 to 2,400 meters. In January of this year, a North American team from Cornell University, in collaboration with researchers from Paulista State University (Unesp) and the State University of Campinas (Unicamp) indicated that the fungus may live in a variety of environments and also at low altitudes in the Atlantic rainforest. All they can do for the time being is to follow the traces of the fungus, since it is difficult to cultivate this organism in a laboratory to study its behavior in detail.

“The amphibian species that are disappearing live in preserved areas in Brazil, Central America and Oceania, where there are no dirty or polluted streams that could be fatal,” says Catão. Brazil is one of the countries with the biggest diversity of amphibians, with almost 900 species already identified, of which 16 are considered to be threatened with extinction and one is already extinct. The reduction in the populations of amphibians could mean an increase in populations of the insects that transmit diseases such as dengue fever, malaria and yellow fever, “considering merely an anthropocentric view,” says Catão. In broader terms, it could make the survival of other animals, such as birds and reptiles that feed on toads and frogs, difficult.

According to Catão, the fungus is one of the causes of the mass extinction of amphibians, which is equivalent to others that have occurred in the history of the Earth, although it is not the only responsible factor: “We have to keep an open mind to other possible causes.” The main ones are the loss or reduction of the habitat and the proliferation of other parasites, such as the ranavirus, a viral group that is lethal to tadpoles, which are normally resistant to the fungus.

“Fungi are opportunistic and advance more easily when the hosts are under stress or some type of environmental pressure,” comments Selene, offering a similar human example: candidiasis, which manifests itself when the organism’s defenses are weakened. She believes that Bd is not yet as well adapted to the different environments as the dermatophyte fungi that cause mycoses in animals and people and spread through domestic environments.

Last year a veterinary student at Unip, Sândara Pimentel Sguario, isolated colonies of fungi of another species, Mycroscoporum canis, in a cat that her boyfriend had left with her and that she kept in her bedroom. Selene suggested to her student to collect samples of material not only from the cat but also from the rug, the bed, her computer chair, and the entire bedroom. It turned out that these contained the same fungus that had infested the cat. “I’ve not yet read anything that indicates that Bd can infect people or other mammals,” says Selene. “From an evolutionary perspective it will perhaps take time before it affects people, or maybe it never will.”

The Project
Compared pathology of selected infections of anuran amphibians that live in the wild in the Atlantic rainforest biome: prospective and retrospective study (nº 2009/52638-3); Modality Regular Research Project Funding; Coordinator José Luiz Catão Dias – FMVZ/USP; Investment R$ 47,371.09

Scientific articles
DE PAULA, C.D. et al. Batrachochytrium dendrobatidis in illegal wildlife trade confiscated amphibians used in ex situ breeding program 
in Brazil. Diseases of Aquatic Organisms. v. 98, n.2, p. 171-75. 2012.
GRÜNDLER, M.C. et al. Interaction between breeding habitat and elevation affects prevalence but not infection intensity of Batrachochytrium dendrobatidis in Brazilian anuran assemblages. Diseases of Aquatic Organisms. v. 97, n. 3, 
p. 173-84. 2012.
CARNAVAL, A.C.O.Q. et al. Amphibian chytrid fungus broadly distributed in the Brazilian Atlantic rain forest. EcoHealth. v. 3, n. 1, p. 41-48. 2006.

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