September 9, 2009

Prions found in feces of deer asymptomatic for chronic wasting disease

Scientists have discovered that deer asymptomatic for a fatal brain condition known as chronic wasting disease excrete the infectious prions that cause the disease in their feces. The finding, they say, suggests a plausible explanation for transmission of the disease among deer and, possibly, elk and moose in the environment.

The study is reported as an advance online paper on September 9, 2009 in the journal “Nature.”

While the study reveals that prions are shed in feces of symptomatic deer as well, the discovery that the infected deer shed prions (PREE-ons) in their feces many months before they show clinical symptoms has particularly provocative implications, according to the research team, at University of California, San Francisco and the Colorado Division of Wildlife’s Wildlife Research Center.

Deer, elk and moose inadvertently consume feces and soil in the course of their daily grazing. Given this, the team set out to determine whether the animals could develop chronic wasting disease through long-term consumption of contaminated feces. They did so by measuring the amount of prions contained in the feces of orally infected deer up until the time they became symptomatic and then calculated whether prolonged exposure to the concentrations of prions in these feces would be enough to cause the disease.

“Prion levels in feces samples of asymptomatic deer were very low compared to those in the brains of the same deer at the time of death,” says the lead author of the study, Erdem Tamguney, PhD, an assistant professor at the Institute for Neurodegenerative Diseases, based at UCSF. “However, the total number of prions excreted over time was sufficiently high enough to cause disease in other deer.”

The susceptibility of animals to infection, he says, might be increased by the simultaneous ingestion of clay soil, which is thought to enhance the infectivity of prions, possibly by slowing their clearance from the gastrointestinal tract.

“Our findings suggest that prolonged fecal prion excretion by infected deer provides a plausible explanation for the high level of transmission of chronic wasting disease within deer herds, as well as prion transmission among deer and other cervid species. Our work may also explain transmission of scrapie prions among sheep and goats,” says senior author and Nobel laureate Stanley B. Prusiner, MD, UCSF professor of neurology and director of the UCSF Institute for Neurodegenerative Diseases.

The study did not examine whether chronic wasting disease could be transmitted to humans via exposure to deer feces. To date, transgenic mouse studies have indicated that chronic wasting disease does not transmit to humans, but scientists remain open to the possibility that it could.

“We can only say that prions of chronic wasting disease have not transmitted to mice genetically engineered to carry the normal, healthy form of human prion protein in earlier studies,” says Prusiner. “That said, we do not know for sure that deer or elk prions cannot be transmitted to humans.”

The prion is an infectious form of the normal prion protein, which has been found in all mammals examined, including humans. The lethal, infectious form induces the normal protein to twist into a malconformation, initiating a disease process that ravages the brain. Prion diseases, seen in cervids, sheep, cows and humans, are also referred to as spongiform encephalopathies. Prusiner won the Nobel Prize in Physiology or Medicine in 1997 for the discovery of prions as a new biological principle of infection.

The new study sheds light on a question that has puzzled scientists for a number of years: how chronic wasting disease spreads so widely through herds and species in contrast to bovine spongiform encephalopathy in cattle, in which horizontal transmission between animals is rare.

First detected in a research facility in Colorado in 1967, chronic wasting disease has since been detected in fourteen states in the U.S. and in two Canadian provinces, predominantly in the West, both in the wild and on commercial farms. In wild herds, it can sometimes be found in up to 30 percent of animals; in captivity nearly entire herds can be affected.

Studies have shown that the disease can be transmitted orally – deer experimentally fed infected brain tissue become sick – but the animals are not carnivores, nor cannibalistic. And while prions have been reported in the saliva, blood, muscle, urine and antler velvet of symptomatic animals with late-stage disease, there is little evidence that these sources are responsible for high incidence of the disease within herds.

Epidemiological findings argue that chronic wasting disease spreads efficiently across populations. When healthy deer grazed on pastureland previously used by deer with chronic wasting disease, the healthy deer eventually develop the disease. But there has been no clear mechanism of transmission of prions in this way.

The UCSF scientists teamed up with co-investigators led by Michael Miller, DVM, PhD, of the Colorado Division of Wildlife’s Wildlife Research Center, to investigate the issue. Five deer were orally infected with one gram of brain tissue from a deer that had died of chronic wasting disease. Then fecal samples from the animals were collected at five time points – once before they were infected (as a control group), and then post infection at 3-4 months, 9-10 months, 13-14 months and 16-20 months (when animals show symptoms of the disease). Symptoms include inability to hold the head erect, excessive salivation, unsteady gait and poor grooming skills.

The UCSF scientists irradiated the deer feces to kill all bacteria and viruses, and inoculated the fecal material into the brains of mice genetically engineered to over-express the normal, healthy version of the elk prion protein.

An analysis showed that feces collected before infection and at 3-4 months post infection did not transmit the disease to the mice. At all subsequent time points, however, many fecal samples did transmit the disease. Prion infectivity was found in 14 of the 15 samples collected from the five deer as early as 7-11 months before neurological symptoms developed.

To measure the prions in feces, the team conducted a separate experiment in which they correlated the time required for mice to become ill from various prion concentrations in infected elk brain tissue (brain samples were diluted with varying amounts of water). Then they compared this data to the incubation times of the mice inoculated with contaminated feces, determining the amount of infectivity of the samples at different time points. While the number of prions in individual samples was low, the amount that accumulated over a 10-month period was similar to that in brain at the end-stage of the disease.

The findings offer strong evidence, says Prusiner, that prion contamination of forest, shrub-steppe and grassland habitats may be largely responsible for the high incidence of the disease both within and between cervid species and account for geographic spread as deer move between seasonal ranges.

Other co-authors of the study are Lisa L. Wolfe, MS, DVM, and Tracey M. Sirochman, BA, of the Colorado Division of Wildlife, Wildlife Research Center; David V. Glidden, PhD, professor of epidemiology and biostatistics, Christina Palmer, MS, Azucena Lemus, BS, of pathology, and Stephen J. DeArmond, MD, PhD, professor of pathology, all of UCSF.

The study was funded by the Larry L. Hillblom Foundation, the Colorado Division of Wildlife, grants from the U.S. Department of Defense National Prion Research Program and the National Institutes of Health.

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