The answers to some questions are not really answers, but more questions. Elizabeth Davidson, a pathologist at Arizona State University, does research on a lethal salamander virus. She is one of a large group of scientists from around the world who are working together to find out what has triggered the decline of amphibians in places as far and wide as North and Central America and Australia. Her particular study area has turned up a few exciting answers, and enough questions to keep this global group of biologists busy for years.
Davidson's research entered the amphibian conservation realm somewhat serendipitously. She had heard about massive numbers of tiger salamanders dying in Arizona's San Rafeal Valley from fellow ASU biologist Jim Collins, who was studying the salamanders' ecology. Davidson and research assistant James Jancovich, curious about what was killing the animals, accompanied Collins to the San Rafeal Valley during a dieoff in 1995. They found ponds full of dead salamanders apparently killed by a bacterial infection called "red leg" disease.
Assuming that the bacteria were to blame for the deaths, Davidson and Jancovich took samples back to the laboratory for further study. What they would find later would open a minefield of unanswered questions.
In the laboratory, they had no trouble isolating the bacterium causing red leg and plenty of other bacteria. "But we were isolating the same bacteria from healthy animals, too. That was one of my first clues that we were barking up the wrong tree," says Jancovich. If healthy animals harbored the same bacteria, why weren't they dying? Davidson and Jancovich began to suspect that the bacteria were simply opportunists preying on already-sick animals, and that a different agent was the real culprit.
Davidson sought the help of colleague Frank Morado, a marine pathologist at the National Oceanographic and Atmospheric Administration in Seattle. The damage Morado saw in the infected tissue reminded him of a similar infection he had recently seen in salmon and insects. The salamanders, he told Davidson, showed all the hallmarks of attack by a virus.
Eagerly anticipating a breakthrough in their work, Davidson and Jancovich prepared tissue samples for study with an electron microscope. On a Friday, they spent most of the day scanning the tissues for virus particles. To their disappointment, they found nothing. "You're looking for a needle in a haystack," warned the skeptical microscopist who assisted them.
"That day we went home bummed," says Jancovich. "But we came back on Monday morning and looked at each other and we both said ‘we're going to find this thing.' We looked in an insect virus pathology book and realized we had been looking at the wrong magnification. So we went back Monday evening and cranked up the magnification and there it was!"
"It was one of the most exciting times I remember having in the lab," recollects Davidson. "There were virus crystals all over. Just everywhere. We called up Jim Collins and said ‘Do you want to see your virus?' and he said 'Really?!'"
Theirs is a type of iridovirus, a group that typically infects insects and other invertebrates, as well as fish and frogs. The name derives from the iridescent appearance that virus-laden insects acquire in the late stages of infection---at death 75% of insect's body weight may be virus, enough to refract light.
Since its identification in the San Rafeal Valley, the salamander virus has now been found to be responsible for major dieoffs of several subspecies of tiger salamanders, including an endangered subspecies, at two other sites in Arizona. Other closely related viruses have caused salamander deaths in Utah, North Dakota, Wyoming, and distant Saskatchewan.
How the virus is spread from one site to another is still an unsolved mystery. Ponds in a single region may be too distant for salamanders to make the trek, virus in tow. And a promenade from Arizona to Saskatchewan is out of the question.
Davidson likes an idea suggested by doctoral student Danna Schock. Schock discovered the salamander virus in Saskatchewan when doing research for her M.Sc., and is now working on the salamander project at ASU. "Danna is suspicious that the distribution of these diseases follows the migratory flyway of birds. There's no better way to go from one pond to the next in a few hours' time, and within a week's time, perhaps, to fly the entire distance. We suspect that viruses could easily be transported on the feet or feathers of a bird."
But Davidson points out that "they could also very nicely be transported on my boots. Or a net. Or a fisherman's boots. Or on salamanders that are used as fishing bait. Human intervention is an obvious possibility." Davidson and other scientists working at the infected ponds now thoroughly clean hands and boots and equipment with "great buckets of Clorox" to ensure that they don't spread the disease.
There is much to learn about the survival and transmission of the virus over time, as well. Dieoffs occur in cycles: Nearly all the salamanders in a pond become infected and die, and then some time later the ponds become repopulated with healthy animals. The pond may later become reinfected or it may stay healthy. It is not clear how healthy animals are able to survive in previously infected ponds, or how the virus is reintroduced into uninfected areas.
Says Davidson, "there are so many things we don't know about the salamanders, really major questions like where the salamanders go when ponds dry up. The water comes back in the ponds and after a while, poof!, there they are again." Why salamanders are susceptible to the disease, whereas frogs and fish sharing the same water escape infection, is also unclear.
Davidson and Collins became even more concerned about the salamanders' predicament when they learned that other amphibians around the world were mysteriously dying, many of them in some of the world's most pristine habitat. Golden toads have completely disappeared from the highland rainforests of Panama, after being infected not by a virus, but by a fungus. The microscopic water-borne fungus, called a chytrid, has also wiped out frogs in Australia and the National Zoo, and has made recent appearances in frogs in Arizona.
In 1998, Davidson, Collins, and many other scientists alarmed by the amphibian declines met en masse at the National Science Foundation in Washington, D.C. to share observations. They came away convinced that the frog and salamander disappearances were a very real global problem. They synthesized their ideas in a major NSF grant that charges the group with unraveling the causes of the declines. The list of collaborators includes over 20 scientists from 3 countries and pools the skills and perspectives of experts in genetics, ecology, modeling, virology, mycology, and bacteriology, among other disciplines.
One of Davidson's goals as a participant in the project is to sequence the DNA of the salamander iridoviruses. Jancovich, who recently completed his M.S., has stayed on as a technician in charge of the sequencing. The sequences will tell them how the viruses are related, and by manipulating the DNA they hope to identify the genes determining what animals the viruses can infect, what environments let them thrive, and how they cause infections.
The long-term prognosis for the disappearing amphibians is still open to question. Davidson is cautious in her judgment, but she worries that the world's frogs and salamanders may be on the path of an inexorable downfall that will leave only the most disease-resistant species standing. She warns that "loss of biodiversity is happening everywhere. My guess is that, like it or not, the amphibians of the future are going to be the tough guys."