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The Utah Division of Wildlife Resources (UDWR) will host the 3rd International Chronic Wasting Disease (CWD) Symposium in the summer of 2009, reports the Wildlife Management Institute. It will be held July 22-24, 2009, at the Marriott Hotel and Conference Center in Park City, Utah. Utah is the westernmost state to discover CWD in its free-ranging mule deer population.

According to event coordinators Leslie McFarlane, Wildlife Disease Program Coordinator for UDWR, and Mary Conner, Research Assistant Professor at Utah State University, the 2009 Symposium will address the latest CWD research and management findings and will provide a forum for wildlife agencies to present updates on the status, surveillance and management of CWD in North America.

Although the symposium program schedule has not been finalized, an invitation for special session topics and panel or paper presentations will be sent out to CWD experts, managers and researchers within the next two weeks. For more information or to submit session topics, contact Dr. Conner

CWD Update 88 August 31, 2007

State and Provincial Updates

Illinois: Paul Shelton, Illinois Department of Natural Resources provides the following: During July, IDNR identified a CWD-positive deer in LaSalle County after testing an animal showing classic signs of the illness. This was the first instance of the disease in this county. The deer was a 3 year old doe collected by a Conservation Police Officer after someone reported a sick, emaciated deer. The location was south of I-80, about 2 miles west of Grundy County, near the town of Seneca. This represents about a 25 mile distance from the previous southernmost positive in DeKalb County. Staff from the Division of Wildlife Resources are assessing the implications of the finding.

The total number of CWD-infected deer found in Illinois now numbers 189. Prior to this, the disease had been confined to Winnebago, Boone, McHenry, Ogle, and DeKalb counties. More than 28,000 deer have been tested in Illinois during the past 5 years. Illinois DNR CWD information is available at: https://www.dnr.illinois.gov/programs/CWD/Pages/default.aspx.

Editor’s note: This finding in LaSalle County is a significant departure from the previously known distribution in Illinois. The new location is the first deer detected in the Illinois River basin, which winds southwest through Illinois towards St. Louis.

New Mexico: Press Release from New Mexico Game and Fish (August 28, 2007):

LAS CRUCES: New Mexico recorded its 19th case of chronic wasting disease in deer in a sick animal found in the Bishop’s Cap area of the Organ Mountains. Officer Richard McDonald investigated a report of an emaciated deer July 12. The animal was unaware of human presence, chronically thirsty, urinating often, and staying in and near a water source. Officer McDonald followed the state’s protocol for disease surveillance by killing the animal and sending it to the Veterinary Diagnostic Laboratory in Albuquerque for testing. Based on the symptoms and the area from which the deer came, the laboratory was instructed that chronic wasting disease (CWD) was highly probable. Laboratory diagnostic testing confirmed presence of CWD in this deer. This is the 19th deer with confirmed CWD found since it was first detected in New Mexico in 2002. Two elk have also been found with CWD.

This deer was in Game Management Unit 19, where special CWD restrictions already exist for hunters. Anyone who finds a deer or elk that appears unaware of human presence and displays symptoms including droopy ears, emaciation, chronic thirst, frequent urination, and reluctance to leave water, should report their observations to the Department of Game and Fish, Wildlife Management Division, (505) 476-8127. New Mexico Game & Fish CWD information is at: www.wildlife.state.nm.us/conservation/disease/cwd/index.htm.

Press Release is at: www.wildlife.state.nm.us/publications/press_releases/documents/2007/082807releases.htm#CWD.

Recent Publications

Efficient In Vitro Amplification of Chronic Wasting Disease PrPRES

Timothy D. Kurt, Matthew R. Perrott, Carol J. Wilusz, Jeffrey Wilusz, Surachai Supattapone, Glenn C. Telling, Mark D. Zabel, and Edward A. Hoover Journal of Virology, September 2007, p. 9605-9608, Vol. 81, No. 17

Abstract: Chronic wasting disease (CWD) of cervids is associated with conversion of the normal cervid prion protein, PrPC, to a protease-resistant conformer, PrPCWD. Here we report the use of both nondenaturing amplification and protein-misfolding cyclic amplification (PMCA) to amplify PrPCWD in vitro. Normal brains from deer, transgenic mice expressing cervid PrPC [Tg(cerPrP)1536 mice], and ferrets supported amplification. PMCA using normal Tg(cerPrP)1536 brains as the PrPC substrate produced >6.5 x 109-fold amplification after six rounds. Highly efficient in vitro amplification of PrPCWD is a significant step toward detection of PrPCWD in the body fluids or excreta of CWD-susceptible species. jvi.asm.org/cgi/content/abstract/81/17/9605?etoc.

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Ultrasensitive detection of scrapie prion protein using seeded conversion of recombinant prion protein

Ryuichiro Atarashi, Roger A Moore, Valerie L Sim, Andrew G Hughson, David W Dorward, Henry A Onwubiko, Suzette A Priola & Byron Caughey Nature Methods – 4, 645 – 650 (2007)

Abstract: The scrapie prion protein isoform, PrPSc, is a prion-associated marker that seeds the conformational conversion and polymerization of normal protease-sensitive prion protein (PrP-sen). This seeding activity allows ultrasensitive detection of PrPSc using cyclical sonicated amplification (PMCA) reactions and brain homogenate as a source of PrP-sen. Here we describe a much faster seeded polymerization method (rPrP-PMCA) which detects greater than or equal to50 ag of hamster PrPSc (approximately0.003 lethal dose) within 2–3 d. This technique uses recombinant hamster PrP-sen, which, unlike brain-derived PrP-sen, can be easily concentrated, mutated and synthetically tagged. We generated protease-resistant recombinant PrP fibrils that differed from spontaneously initiated fibrils in their proteolytic susceptibility and by their infrared spectra. This assay could discriminate between scrapie-infected and uninfected hamsters using 2-mul aliquots of cerebral spinal fluid. This method should facilitate the development of rapid, ultrasensitive prion assays and diagnostic tests, in addition to aiding fundamental studies of structure and mechanism of PrPSc formation. www.nature.com/nmeth/journal/v4/n8/abs/nmeth1066.html.

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The following two articles are from the June 2007 edition (Volume 1772, Issue 6) of the journal Biochimica et Biophysica Acta (BBA) – Molecular Basis of Disease. This special edition of the journal, edited by Glenn Telling, is devoted to prion-related disorders and is available at: www.sciencedirect.com/science/journal/09254439.

Chronic Wasting Disease (review)

Christina J. Sigurdson and Adriano Aguzzi Biochimica et Biophysica Acta 1772 (2007) 610–618

Abstract: Until recently, chronic wasting disease of cervids, the only prion disease affecting wildlife, was believed to be geographically concentrated to Colorado and Wyoming within the United States. However, increased surveillance has unveiled several additional pockets of CWD-infected deer and elk in 12 additional states and 2 Canadian provinces. Deer and elk with CWD have extensive aggregates of PrPSc not only in the central nervous system, but also in peripheral lymphoid tissues, skeletal muscle, and other organs, perhaps influencing prion shedding. Indeed, CWD is transmitted efficiently among animals by horizontal routes, although the mechanism of spread is unknown. Genetic polymorphisms in the Prnp gene may affect CWD susceptibility, particularly at codon 225 (S/F) in deer and codon 132 (M/L) in elk. Since CWD infects free-ranging animals and is efficiently spread, disease management will be a challenge.

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Motor behavioral and neuropathological deficits in mice deficient for normal prion protein expression

Karah E. Nazora, Tanya Sewarda and Glenn C. Telling Biochimica et Biophysica Acta 1772 (2007) 645–653

Abstract: It has been difficult to reconcile the absence of pathology and apparently normal behavior of mice lacking prion protein (PrP), referred to as Prnp0/0 mice, with a mechanism of prion pathogenesis involving progressive loss of PrPC-mediated neuroprotection. However, here we report that Prnp0/0 mice exhibit significant age-related defects in motor coordination and balance compared with mice expressing wild type Prnp on a syngeneic background, and that the brains of behaviorally-impaired Prnp0/0 mice display the cardinal neuropathological hallmarks of spongiform pathology and reactive astrocytic gliosis that normally accompany prion disease. Consistent with the appearance of cerebellar ataxia as an early symptom in patients with Gerstmann–Sträussler–Scheinker syndrome (GSS), an inherited form of human prion disease, motor coordination and balance defects manifested in a transgenic (Tg) mouse model of GSS considerably earlier than the onset of end-stage neurodegenerative disease. Our results are consistent with a mechanism in which loss of normal PrPC function is an important pathological component of prion diseases.

The rogue proteins that cause chronic wasting disease (CWD) exhibit a dramatic increase in their infectious nature when bound to common soil particles, according to a new study.

Writing in the journal Public Library of Science (PLoS) Pathogens, a group led by University of Wisconsin-Madison prion expert Judd Aiken reports that prions, the protein agents of a family of fatal brain disorders, bind tightly to a common soil mineral and significantly increase the oral transmissibility of the agent.

The finding is important because it may help explain how chronic wasting disease and scrapie persist in the environment and spread efficiently in animal populations.

“We found a huge difference between infectious agent alone and infectious agent bound to these soil particles,” says Aiken, the senior author of the new study and a professor of comparative biosciences in the UW-Madison School of Veterinary Medicine. “We observed an almost 700-fold difference” in the rate of infection.

Prions are an abnormal form of a protein produced normally by the body. Tough as nails, they can persist in the environment for long periods of time and retain their infectious capabilities. It is believed that prions may persist in the soil around the carcasses of dead animals and other locations where infected animals shed the protein in body fluids.

“These disease agents can stay out there for years and stay infectious,” Aiken explains.

And herbivores such as deer and sheep, which are susceptible to prion infection, tend to consume a fair amount of dirt daily as they graze and forage. They are also known to consume soil as a source of minerals. Mineral licks are frequented by many animals, raising the prospect that the agents may become concentrated in the soil.

Relatively little is known about the routes of prion transmission in animals, but the new Wisconsin study may help to resolve one puzzle: Oral transmission of prions, says Aiken, tends not to be very efficient.

“This is a dichotomy in our field, and maybe (the new research) is part of the answer.”

In their studies, the Wisconsin researchers looked at the ability of prions to bind to different types of common soil minerals. One, known as montmorillonite, is a type of clay and prions seem to have a special affinity for latching onto the microscopic particles.

“We expected the binding of the montmorillonite to be the highest among the minerals we examined. However, we were surprised by the strength of the binding,” notes Joel Pedersen, a UW-Madison professor of soil science who helped direct the new study.

The Wisconsin team also looked at the ability of the prion to bind to two other common soil minerals: quartz and kaolinite, another common clay mineral.

“We found binding of the abnormal protein to all three,” says Aiken, “but the binding to montmorillonite was very avid, very tight. We found it very difficult to remove the prions from the montmorillonite.”

Feeding the prion-mineral mix to hamsters, a common animal model for prion disease, Aiken’s team expected to see a lower rate of infection than animals dosed with pure agent. Surprisingly, prions bound to montmorillonite were significantly more infectious than prions alone.

“We thought the binding might decrease infectivity,” Aiken explains. “In each case, you add montmorillonite and we get more animals sicker and quicker than in the absence of montmorillonite clay.”

What is occurring in soils in the woods and on the farm is unknown, says Pedersen, but the new findings may help begin to answer some key questions about how prions survive in the soil and retain their infectious nature, sometimes for years.

In the case of scrapie, the prion disease of sheep, observations of sheep pastures in the United Kingdom and Iceland have shown that animals introduced into pastures that once held infected animals could become infected. Infectivity of prions was also enhanced when they were bound to whole soil.

“Since the 1940s it’s been known that ‘infected pastures’ have the ability to infect new animals,” according to Aiken.

Pedersen notes that soils are a complex mixture of organic and inorganic components that vary across the landscape and that scientists are just beginning to tease out factors in soils that may contribute to transmissibility. The new study implies, he says, “that some soils may promote the transmission of the prion agent more readily than others.”

Why that’s the case is unknown, Pedersen explains, but the Wisconsin team is exploring several hypotheses: that the soil particles might somehow protect the prion from degradation in the digestive system, that prions bound to clay might change the route or degree of uptake of the agent, or that the mineral somehow alters the size of prion aggregates, which have been shown to be more infectious than prions alone.

Aiken emphasizes there’s still much to learn about routes of prion transmission, and the role of soil is just beginning to be explored.

“Soil is a very complex medium and we don’t know what the agent is binding to” in natural or agricultural settings, Aiken says. “We do know that soil is not the only way it transmits. Animal-to-animal transmission is important, too.”

In addition to Aiken and Pedersen, authors of the PLoS Pathogens paper include Christopher J. Johnson, Rick J. Chappell and Debbie McKenzie. The work was supported by a grant from the U.S. Department of Defense.

CWD Update 87 July 6, 2007

State and Provincial Updates

Illinois: Paul Shelton, Illinois Department of Natural Resources provides the following: Illinois Department of Natural Resources staff collected 6,733 usable CWD surveillance samples during the period beginning July 1, 2006. A few additional samples from suspect deer remain untested at this time. Testing yielded 41 CWD-positive deer and 6,692 ‘not detected’. Forty-one positives were detected in four counties in northern Illinois: Winnebago (18), Boone (13), DeKalb (6), and McHenry (4). No new counties were identified as having CWD, and no disease was detected in Ogle County, in which CWD was first identified during the 2005-2006 sampling season. However, CWD was identified in southeastern DeKalb County, more than 20 miles southeast of previously-identified locations. Samples were collected from a variety of sources, including deer check stations in high-risk counties (3,097), sample drop-off locations for archery hunters (175), cooperating meat processors (1,778), suspect deer (25), roadkills in CWD counties (16), and culling efforts (1,642). Cooperating meat lockers were added as a sampling source this year to provide a statewide sampling base outside the identified CWD area. Hunter-harvested deer accounted for 17 (41%) of the positives identified, with the remainder coming from suspect deer (4; 10%); roadkills (1; 2%); and sharpshooting (19; 46%). Illinois DNR CWD information is available at: https://www.dnr.illinois.gov/programs/CWD/Pages/default.aspx.

Montana: Neil Anderson, Montana Fish, Wildlife and Parks provides the following: Cases of CWD in wild elk or deer have been found within 100 miles of the Montana border in Wyoming and the Black Hills of South Dakota and within 150 miles of the border with the Canadian providences of Saskatchewan and Alberta. Montana Fish, Wildlife and Parks (FWP) began surveillance activities in 1998 following a directive from the Governor’s office to work with the Department of Livestock to survey for and manage CWD in Montana. Initially, FWP activities focused on obtaining a broad geographical survey from across the state using existing game check stations. In 1999 surveillance efforts were expanded to include targeted surveillance of symptomatic animals, special collections of wild elk and deer near alternative livestock operations, and broad geographical surveillance of hunter killed deer and elk. The geographical survey area changed in 2000 concentrating on areas bordering Saskatchewan (region 6) and Wyoming and South Dakota (regions 5 and 7) where CWD had been detected in wild populations. This selected geographical survey has been followed through the 2006 big game hunting season.

Since 1998 over 10,600 moose, deer and elk have been tested for CWD. Hunter-harvested animals remain the bulk of samples collected but efforts to enhance collections of targeted and road-killed animals resulted in an increase in the numbers of samples collected in these groups. To date, all tests have produced no evidence of CWD in wild populations. Surveillance efforts have shifted in priority since 1998 when the first statewide geographical survey was conducted. The majority of samples have been collected near the borders of regions 5, 6 and 7 and in areas surrounding the only alternative livestock operation to test positive for CWD near Philipsburg, MT. The number of samples collected during 2003 was nearly double that of previous efforts and has remained consistent through 2005 declining slightly in 2006. Although surveillance efforts have changed, mule deer have consistently comprised the majority of wild Cervids tested for CWD in Montana. To date, a total of 1342 samples have been tested in the 2006 surveillance period which will continues until June 30, 2007. The hunting district could not be determined for 18 samples but were included in the regional summaries. Montana FWP CWD information is available at: fwp.mt.gov/hunting/chronicWD.html.

West Virginia: Paul Johansen, West Virginia Department of Natural Resources provides the following: Three additional free-ranging white-tailed deer in Hampshire County, West Virginia, tested positive for Chronic Wasting Disease (CWD). This brings the total number of CWD-positive deer found in Hampshire County to 13. These most recent samples were collected from a total of 101 adult deer taken in March and April 2007 by DNR personnel as part of an ongoing and intensive CWD surveillance effort. The three CWD-positive deer were collected within the CWD Containment Area located north of U.S. Route 50 in Hampshire County.

When CWD was first confirmed in Hampshire County in September 2005, the DNR immediately implemented its CWD – Incident Response Plan. As part of that plan, the DNR has been engaged in intensive CWD surveillance efforts designed to determine the distribution and prevalence of the disease. These surveillance efforts have included carefully planned and coordinated deer collections within Hampshire County by CWD deer collection teams comprised of Wildlife Biologists, Wildlife Managers and Conservation Officers within the DNR.

The following disease management strategies have been implemented by the DNR within the affected area of Hampshire County.

• Continue CWD surveillance efforts designed to determine the prevalence and distribution of the disease;
• Lower deer population levels to reduce the risk of spreading the disease from deer to deer by implementing appropriate antlerless deer hunting regulations designed to increase hunter opportunity to harvest female deer;
• Establish reasonable, responsible and appropriate deer carcass transport restrictions designed to lower the risk of moving the disease to other locations;
• Establish reasonable, responsible and appropriate regulations relating to the feeding and baiting of deer within the affected area to reduce the risk of spreading the disease from deer to deer.

West Virginia DNR CWD information is available at: wvdnr.gov/Hunting/ChronicWaste.

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ANNOUNCEMENT:

Third International Chronic Wasting Disease Symposium The Utah Division of Wildlife Resources has announced their intent to host the 3rd International CWD Symposium in Park City, Utah during July of 2009. The initial CWD Symposium was held in Denver, Colorado in 2002; the second was held in Madison, Wisconsin in 2005. Details on the 2009 conference will be forthcoming. Many thanks to our colleagues in Utah for their willingness to arrange and host this event.

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Recent Publications

Oral Transmissibility of Prion Disease Is Enhanced by Binding to Soil Particles Christopher J. Johnson, Joel A. Pedersen, Rick J. Chappell, Debbie McKenzie and Judd M. Aiken PLoS Pathogens Vol. 3, No. 7, e93 doi:10.1371/journal.ppat.0030093.

Abstract: Soil may serve as an environmental reservoir for prion infectivity and contribute to the horizontal transmission of prion diseases (transmissible spongiform encephalopathies [TSEs]) of sheep, deer and elk. TSE infectivity can persist in soil for years, and we previously demonstrated that the disease-associated form of the prion protein (PrPTSE) binds to soil particles and that prions adsorbed to the common soil mineral montmorillonite (Mte) retain infectivity following intracerebral inoculation. Here, we assess the oral infectivity of Mte- and soil-bound prions. We establish that prions bound to Mte are orally bioavailable and that, unexpectedly, binding to Mte significantly enhances disease penetrance and reduces incubation period relative to unbound agent. Cox proportional hazards modelling revealed that across the doses of TSE agent tested, Mte increased the effective infectious titer by a factor of 680 relative to unbound agent. Oral exposure to Mte-associated prions led to TSE development in experimental animals even at doses too low to produce clinical symptoms in the absence of the mineral. We tested the oral infectivity of prions bound to three whole soils differing in texture, mineralogy and organic carbon content, and found soil-bound prions to be orally infectious. Two of the three soils increased oral transmission of disease, and the infectivity of agent bound to the third soil was equivalent to that of unbound agent. Enhanced transmissibility of soil-bound prions may explain the environmental spread of some TSEs despite the presumably low levels shed into the environment. Association of prions with inorganic microparticles represents a novel means by which their oral transmission is enhanced relative to unbound agent. pathogens.plosjournals.org/perlserv/?request=get-document&doi=10.1371/journal.ppat.0030093.

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A Natural Case of Chronic Wasting Disease in a Free-ranging Moose (Alces alces shirasi) Laurie A. Baeten, Barbara E. Powers, Jean E. Jewell, Terry R. Spraker and Michael W. Miller Journal of Wildlife Diseases, 43(2), 2007, pp. 309-314.

Abstract: Chronic wasting disease (CWD) was diagnosed in a free-ranging moose (Alces alces shirasi) killed by a hunter in Jackson County, Colorado, USA, in September 2005. The diagnosis was based upon immunohistochemistry (IHC) demonstrating the presence of accumulations of CWD-associated prion protein (PrPCWD) in tissue sections of medulla oblongata at the level of the obex (dorsal motor nucleus of the vagus) and in retropharyngeal lymph node (RPLN); additional testing by IHC revealed deposits of PrPCWD in multiple sections of medulla oblongata and cervical spinal cord as well as palatine tonsil and submandibular lymph node tissues. Western blot confirmed the presence of PrPCWD in RPLN and tonsil tissue. The PrPCWD also was detected via enzyme-linked immunosorbent assay of RPLN tissue. Spongiform encephalopathy was observed in sections of the brainstem and cervical spinal cord, although no clinical signs were noted by the hunter who killed the animal. The affected moose was homozygous for methionine at codon 209 of the prion protein coding region. In October 2006, two additional free-ranging moose were diagnosed with CWD. Epidemiology and implications of CWD in moose remain to be determined. jwildlifedis.org/cgi/content/abstract/43/2/309.

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Susceptibility of Cattle to First-passage Intracerebral Inoculation with Chronic Wasting Disease Agent from White-tailed Deer A. N. Hamir, J. M. Miller, R. A. Kunkle, S. M. Hall and J. A. Richt Veterinary Pathology 44:487-493 (2007).

Abstract: Fourteen, 3-month-old calves were intracerebrally inoculated with the agent of chronic wasting disease (CWD) from white-tailed deer (CWDwtd) to compare the clinical signs and neuropathologic findings with those of certain other transmissible spongiform encephalopathies (TSE, prion diseases) that have been shown to be experimentally transmissible to cattle (sheep scrapie, CWD of mule deer [CWDmd], bovine spongiform encephalopathy [BSE], and transmissible mink encephalopathy). Two uninoculated calves served as controls. Within 26 months postinoculation (MPI), 12 inoculated calves had lost considerable weight and eventually became recumbent. Of the 12 inoculated calves, 11 (92%) developed clinical signs. Although spongiform encephalopathy (SE) was not observed, abnormal prion protein (PrPd) was detected by immunohistochemistry (IHC) and Western blot (WB) in central nervous system tissues. The absence of SE with presence of PrPd has also been observed when other TSE agents (scrapie and CWDmd) were similarly inoculated into cattle. The IHC and WB findings suggest that the diagnostic techniques currently used to confirm BSE would detect CWDwtd in cattle, should it occur naturally. Also, the absence of SE and a distinctive IHC pattern of CWDwtd and CWDmd in cattle suggests that it should be possible to distinguish these conditions from other TSEs that have been experimentally transmitted to cattle. vetpathology.org/cgi/content/abstract/44/4/487.