Monday, May 05, 2014

cwd tse prion testing PMCA , IHC, tonsil, rectal, biopsy ???

cwd tse prion PMCA , IHC, tonsil, rectal, biopsy ???
Description Abstract Text:
DESCRIPTION (provided by applicant):
Transmissible spongiform encephalopathies (TSEs) are a group of fatal neurodegenerative disorders affecting humans and animals. Although rare, the recent outbreak of Bovine Spongiform Encephalopathy(BSE) in cattle and Chronic Wasting disease (CWD) in cervids and the transmission of the disease from cattle to humans have caused great concern. This problem is aggravated by the lack of an efficient, sensitive and early diagnosis as well as many uncertainties surrounding the unprecedented nature of the infectious agent, its mechanism of propagation and the species-barrier that controls prion transmission. The major goals of this Program Project are to understand the mechanism of transmission and pathogenesis of BSE and CWD prions, to estimate the risk of these infectious agents to propagate disease to other animals and especially to humans, to assess the mechanisms and routes of prion dissemination among animals and to develop novel strategies for ante-mortem detection of infected animals. A team of accomplished scientists with widely recognized expertise and track record of contributions in this field will work collaboratively to reach these goals. Project 1 (Juergen Richt, PL) proposes to study in detail mutant and knock out cows and assess the role of genetic forms of BSE in disease transmission, susceptibility and species barrier. For these studies, we plan to generate and characterize knock in transgenic cattle expressing a PrP mutation recently identified in a natural BSE case in USA. We will also use transgenic mice models expressing wild type and mutant bovine PrP and in vitro studies of PrP replication using the PMCA. Project 2 (Glenn Telling, PL) proposes to generate and use transgenic mice models to assess the strength of various species barriers and the influence of prion strains. We will study the susceptibility of various models to be infected by CWD as well as the possibility that deer models may be infected by prions from other species. To address this aim, experiments will be done in transgenic mice models and using in vitro conversion studies. Project 3 (Claudio Soto, PL) proposes to study the replication of CWD and BSE prions in vitro, evaluate tissue distributions of infectious protein, enlighten the routes of transmission and develop a diagnostic assay. For this purpose, we will use the PMCA technology, recently developed in Dr Soto's lab to mimic the prion replication process. The Administration Core (core A) (Claudio Soto, Director) will facilitate and integrate Projects and Cores and take care of all administrative aspects needed for the smooth operation of the Program. The Tissue Core (core B) (Pierluigi Gambetti, Director) will process, analyze and store the tissue samples generated and provide access to the members to a biosafety facility to manipulate BSE infectious material. The findings generated in this Program will have undoubtedly contribute to understand the pathogenesis, route of propagation and early detection of these two worrisome zoonotic prion diseases.
Project Terms: Address; Affect; Animals; Area; Biological Assay; Bovine Spongiform Encephalopathy; Caring; Cattle; Cattle Diseases; cervid; Chronic Wasting Disease; Creutzfeldt-Jakob Syndrome; Deer; Detection; Development; Diagnostic; Disease; Disease Outbreaks; disease transmission; Drug or chemical Tissue Distribution; Early Diagnosis; Experimental Models; Genetic; Goals; Human; In Vitro; Infectious Agent; Knock-in Mouse; Knock-out; Knowledge; member; Methodology; Modeling; mouse model; mutant; Mutation; Nature; Neurodegenerative Disorders; novel strategies; operation; Pathogenesis; Positioning Attribute; Predisposition; prevent; Prion Diseases; Prions; Process; programs; Proteins; PrP; public health medicine (field); Replication-Associated Process; Research; research study; Risk Estimate; Role; Route; Sampling; Scientist; Scrapie; Sheep; Technology; Tissue Sample; Tissues; Transgenic Mice; Transgenic Organisms; transmission process; Uncertainty; Work
cwd tse prion tonsil biopsy ???
The Wisconsin Wildlife Technician told us that there is a live tonsil biopsy test but it is not approved for use by the DNR in Wisconsin. He said it is used mainly for research purposes. The CWD Program Manager told us that while tonsils may be collected from live animals, the sampling data collected by the national reference testing laboratory over approximately 10 years has shown that collection of tonsil samples is not very reliable. “Most often the proper sample (the tonsil) is not correctly collected because it is difficult to reach and visualize the tonsillar tissue located far back in the animal’s throat while the animal has been anesthetized for this procedure,” she said. “Therefore, many times the sample submitted as ‘tonsil’ is found in the laboratory testing process to be an adjacent section of soft tissue oral mucosa - and not suitable for CWD testing
According to the Chronic Wasting Disease Alliance, however, researchers from the USDA and Colorado State University have evaluated and validated another live testing method using rectal-tissue biopsies in captive and wild elk in Colorado. It appears to be nearly as accurate as post-mortem testing.
“The key advantage to the rectal biopsy test is that it can be performed on live animals. Until now, there was no practical live test for CWD in elk,” said Research Wildlife Biologist Dr. Kurt Ver Cauteren with APHIS’ Wildlife Services.
“The use of this new live test in the initial screening, surveillance and monitoring of CWD will greatly aid in the management and control of the disease in the wild, as well as in captive settings,” said Ver Cauteren.
Chronic Wasting Disease Alliance reports that many thousands of captive elk have been killed in the western United States and Canada in order to control CWD, as well as thousands of free-ranging mule deer, white-tailed deer, and elk.
(Learn more about CWDA’s work at: )
Development of an antemortem test for detecting the misfolded prion protein associated with CWD (PrPCWD) in nonclinical animals would be useful for wildlife and captive population management strategies. To date, preclinical testing for PrPCWD utilizes immunohistochemistry (IHC) of the palatine tonsils or rectal lymphoid tissues in cervids.18,21,24 However, IHC does not routinely detect very early cases of CWD in these tissues.17,20
The "gold standard" diagnostic test for CWD is the Immunohistochemistry (IHC) test performed on the obex tissue of the brain (right) or specific lymphoid tissues.
IHC testing is a method utilizing antibody based staining which is evaluated using light microscopy. This test is both sensitive and specific. In addition, the microscopic methods used to detect positive staining also allow confirmation that the correct tissue and location within the tissue are present to detect the earliest accumulations of the prion agent. Other diagnostic tests utilizing enzyme-linked immunosorbent assay (ELISA) and other technologies that allow more rapid testing of larger numbers of samples have been, and continue to be, developed. Several test kits using ELISA and other technologies have been licensed by USDA's Center for Veterinary Biologics for CWD diagnosis in wild elk and deer. CWD testing can only be performed by approved laboratories that are part of the USDA's National Animal Health Laboratory Network system. A rectal associated mucosal lymphoid tissue (RAMALT) biopsy (live-animal) test has been developed by researchers and appears to hold promise for future use in certain CWD monitoring or management situations in farmed cervids. This technique utilizes the current IHC testing technologies described above.
Information on brain (obex) and lymphoid tissue sample collection is found in the APHIS CWD Sample Collection Guidance .
View a map showing the locations of the USDA contract laboratories for CWD, or view the National Veterinary Services Laboratories listing of laboratories and contact points .
 From: Will Laegreid
Sent: Monday, May 05, 2014 4:45 PM
Subject: RE: cwd testing with immunohistochemistry (IHC) of the palatine tonsils
Dear Mr. Singletary,
Thank you for your inquiry about CWD testing.  If I understand your question correctly, you are asking about the accuracy of IHC for early preclinical diagnosis of CWD.  The quote you included in your message is spot on, IHC will not reliably detected very early cases of CWD.  The differences between samples, tonsil versus rectal lymphoid biopsies, is largely affected by two factors, the relative number of lymphoid follicles in each tissue (tonsil > rectal) and the skill of the person taking the biopsy.  It is my opinion that both samples can work, but that tonsil biopsies are more sensitive (probably due to the greater number of follicles for examination).  It also appears that disease progression, affected by prion genotype of the individual animal, influences sensitivity.  I hope this addresses your questions.
Best regards,
William W. Laegreid, DVM, PhD
Director, Wyoming State Veterinary Laboratory
Head, Department of Veterinary Sciences
University of Wyoming
1174 Snowy Range Road
Laramie, WY 82070
(307) 766-9929
(307) 721-2051 Fax
Evaluation and Interpretation of Rectal Mucosa Biopsy Testing for Chronic Wasting Disease within Four White-Tailed Deer Herds in North America
Dr. Bruce V. Thomsen USDA-APHIS-VS, National Veterinary Services Laboratories (NVSL) An effective live animal test is needed to assist in the control of chronic wasting disease (CWD), which has spread through captive and wild herds of white-tailed deer in both Canada and the United States. Rectal biopsy sample testing for CWD has shown promising results in previous studies and rectal biopsy sample testing has also been utilized successfully as a live animal test to diagnose the closely related disease, scrapie in sheep. This study compared the test results of postmortem rectal mucosa biopsy samples to those from conventional postmortem samples of the brainstem at the obex; the medial retropharyngeal lymph node; and the palatine tonsil in four CWD-infected, captive white-tailed deer herds. Three of the herds were located in Canada and one of the herds was from the United States. The effects of age, sex, genotype at prion protein (PRNP) codon 96, and stage of disease progression were evaluated as possible factors that might influence test performance. Test sensitivity for CWD on rectal biopsy samples in white-tailed deer ranged from 63% to 100% in the four herds within this study. Test performance was influenced by genotype at PRNP codon 96 and by 2 stage of disease progression. Test sensitivity was the highest for 96GG deer and lower for 96GS deer. Rectal biopsy test sensitivity was 100% for deer in the later stages of disease progression, as evidenced by abundant immunohistochemical staining for PrPCWD in sections of brainstem. Rectal biopsy test sensitivity was reduced for deer in the earlier stages of disease. Selective use of this test, in conjunction with conventional testing postmortem testing, could provide valuable information during disease investigations of CWD suspect deer herds.
Thursday, May 02, 2013
*** Chronic Wasting Disease (CWD) Texas Important Update on OBEX ONLY TEXTING ***
Prion-Seeding Activity in Cerebrospinal Fluid of Deer with Chronic Wasting Disease
Nicholas J. Haley mail, * E-mail:
Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Alexandra Van de Motter, Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Scott Carver, Affiliation: School of Zoology, University of Tasmania, Hobart, Tasmania, Australia
X Davin Henderson, Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Kristen Davenport, Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Davis M. Seelig, Affiliation: Department of Veterinary Clinical Sciences, University of Minnesota, St. Paul, Minnesota, United States of America
X Candace Mathiason, Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Edward Hoover Affiliation: Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, United States of America
X Published: November 25, 2013 DOI: 10.1371/journal.pone.0081488
Abstract Transmissible spongiform encephalopathies (TSEs), or prion diseases, are a uniformly fatal family of neurodegenerative diseases in mammals that includes chronic wasting disease (CWD) of cervids. The early and ante-mortem identification of TSE-infected individuals using conventional western blotting or immunohistochemistry (IHC) has proven difficult, as the levels of infectious prions in readily obtainable samples, including blood and bodily fluids, are typically beyond the limits of detection. The development of amplification-based seeding assays has been instrumental in the detection of low levels of infectious prions in clinical samples. In the present study, we evaluated the cerebrospinal fluid (CSF) of CWD-exposed (n=44) and naïve (n=4) deer (n=48 total) for CWD prions (PrPd) using two amplification assays: serial protein misfolding cyclic amplification with polytetrafluoroethylene beads (sPMCAb) and real-time quaking induced conversion (RT-QuIC) employing a truncated Syrian hamster recombinant protein substrate. Samples were evaluated blindly in parallel with appropriate positive and negative controls. Results from amplification assays were compared to one another and to obex immunohistochemistry, and were correlated to available clinical histories including CWD inoculum source (e.g. saliva, blood), genotype, survival period, and duration of clinical signs. We found that both sPMCAb and RT-QuIC were capable of amplifying CWD prions from cervid CSF, and results correlated well with one another. Prion seeding activity in either assay was observed in approximately 50% of deer with PrPd detected by IHC in the obex region of the brain. Important predictors of amplification included duration of clinical signs and time of first tonsil biopsy positive results, and ultimately the levels of PrPd identified in the obex by IHC. Based on our findings, we expect that both sPMCAb and RT-QuIC may prove to be useful detection assays for the detection of prions in CSF.
In summary, the present study evaluated two amplification assays – sPMCAb and RT-QuIC – for their ability to amplify PrPd in the CSF of CWD-exposed and naïve white-tailed deer. Results between the two assays correlated well with each other and to IHC results from obex collected at necropsy, albeit with reduced sensitivity. A priori variables, notably date of tonsil biopsy positivity and duration of clinical signs, influenced the likelihood of a sample being positive by either assay, while our post-amplification analyses (slope of curve or peak fluorescence) did not correlate with clinical histories. Based on our findings, we believe that amplification assays hold continued promise in the detection of prion-infection using post- or ante-mortem samples and our future work will continue to evaluate the utility of these assays in detecting seeding activity in these tissues and biological fluids.
AD.11: Early detection of chronic wasting disease prions in urine of pre-symptomatic deer by real-time quaking-induced conversion assay
Theodore R. John,2 Hermann M. Schätzl1 and Sabine Gilch1 1University of Calgary; Calgary, AB Canada; 2University of Wyoming; Laramie, WY USA
Chronic wasting disease (CWD) is a prion disease of captive and free-ranging deer (Odocoileus spp), elk (Cervus elaphus nelsonii) and moose (Alces alces shirasi). For the latter, the first case in Canada was recently diagnosed in a road-killed moose in Alberta. Unlike in most other prion diseases, in CWD infectious prions are found in a wide variety of peripheral tissues and bodily fluids, such as skeletal and heart muscle, antler velvet, blood, saliva, urine and feces. This distribution and the shedding of prions most likely contribute to the horizontal transmission of the disease within and between cervid species upon foraging on contaminated pastures. Since to date, ante-mortem diagnosis is only possible by immunohistochemical detection of protease resistant prion protein (PrPSc) in tonsil or rectoanal mucosa-associated lymphoid tissue (RAMALT) biopsies which requires anesthesia of animals, a non-invasive intra vitam assay is highly desirable. We have used the real time quaking-induced conversion (RT-QuIC) assay for detection of seeding activity in brain homogenates, urine or fecal extracts of orally infected mule or white-tailed deer. Seeding activity was found in fecal extracts, although sensitivity of detection has to be improved. Furthermore, we demonstrate that with this assay CWD prions can be detected in urine of animals in the pre-symptomatic stage of the disease. In summary, we provide first evidence that RT-QuIC can be useful for noninvasive pre-sympomatic diagnosis and surveillance of CWD.
 AD.74: Shedding pattern in PrPCWD in experimentally infected elk and potential mechanism of CWD transmission Jianmin Yang, Tammy Pickles, Sandor Dudas, Catherine Graham and Stefanie Czub Canadian and OIE Reference Laboratories for BSE; Canadian Food Inspection Agency; Lethbridge, AB Canada Background. The dynamics of CWD spread in elk appear to be different than in deer species, potentially related to differences in population structure and behavior, but also to mechanisms and timing of shedding of the agent. In deer, shedding occurs via saliva and urine/feces; and tissues associated with the production of these excrets have been confirmed positive. This is the report of the detection of PrPcwd within tissues involved in the production of saliva, urine and feces in elk experimentally challenged with CWD. Materials and Methods. Tissues were obtained from animals orally challenged with CWD and sequentially sacrificed during incubation period and clinical stages of disease. Animals of both 132LM and 132MM genotype were included; and the following tissues were examined: Rectum; Kidney; Urinary Bladder (UB); Tongue; Parotid Salivary Gland (PSG); Oral Mucosa (OM) and Nasal Mucosa (NM). In addition, Tonsil, Retropharyngeal Lymph Node (RPLN), Brain and feces obtained at necropsy were also tested. Assays employed included a modified western blot (moWB) method using mAb ICSM18 and immunohistochemistry using mAb F99. Results. The earliest detection of clinical signs, weight loss and change in behavior was at approximately 570 d post inoculation (DPI). This was preceded by the detection of CWD in brain by IHC at 300 DPI, while all other tissues examined were negative by both IHC and moWB. At 400 and 525 DPI, Lymphoid Tissues and Brain were positive by IHC and moWB. In contrast, the UB and OM tested positive by moWB but negative by IHC at 400 DPI. At 525 DPI, moWB detected CWD prions within the Tongue, PSG, and NM. Animals with terminal disease displayed intense, widespread distribution within Brain, Lymphoid Tissue, and Peripheral Nervous Tissue of many organs. Animals with 132LM genotype showed a reduced distribution in peripheral tissues as compared with 132MM genotype animals. Conclusions. Based on tissue distribution, shedding of PrPcwd in elk occurs well before the onset of clinical disease. The mechanisms of shedding are likely similar to those in deer species, but may occur much later in the incubation period. This difference has potential impact on the regulation and management of CWD positive elk farms.
Prion. May 1, 2013; 7(3): 253–258. Published online Apr 10, 2013. doi: 10.4161/pri.24430 PMCID: PMC3783112
Early detection of chronic wasting disease prions in urine of pre-symptomatic deer by real-time quaking-induced conversion assay
Theodore R. John, 1 Hermann M. Schätzl, 1 , 2 , 3 and Sabine Gilch 1 , 4 ,* 1Department of Veterinary Sciences; University of Wyoming; Laramie, WY USA 2Department of Molecular Biology; University of Wyoming; Laramie, WY USA 3Faculty of Veterinary Medicine; Department of Comparative Biology and Experimental Medicine; University of Calgary; Calgary, AB Canada 4Faculty of Veterinary Medicine; Department of Ecosystem and Public Health; University of Calgary; Calgary, AB Canada *Correspondence to: Sabine Gilch, Email: Author information ► Article notes ► Copyright and License information ► Received February 7, 2013; Revised March 17, 2013; Accepted March 24, 2013. Copyright © 2013 Landes Bioscience This is an open-access article licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License. The article may be redistributed, reproduced, and reused for non-commercial purposes, provided the original source is properly cited. Go to:
Chronic wasting disease (CWD) is a prion disease of captive and free-ranging deer (Odocoileus spp), elk (Cervus elaphus nelsonii) and moose (Alces alces shirasi). Unlike in most other prion diseases, in CWD prions are shed in urine and feces, which most likely contributes to the horizontal transmission within and between cervid species. To date, CWD ante-mortem diagnosis is only possible by immunohistochemical detection of protease resistant prion protein (PrPSc) in tonsil or recto-anal mucosa-associated lymphoid tissue (RAMALT) biopsies, which requires anesthesia of animals. We report on detection of CWD prions in urine collected from pre-symptomatic deer and in fecal extracts by using real time quaking-induced conversion (RT-QuIC). This assay can be useful for non-invasive pre-symptomatic diagnosis and surveillance of CWD.
Keywords: prion, chronic wasting disease, diagnosis, surveillance, RT-QuIC, urine, feces
Introduction Chronic wasting disease (CWD) is to date the most contagious prion disease and affects captive and free-ranging elk, deer and moose in North America.1,2 The disease is caused by the accumulation of an abnormally folded isoform of the cellular prion protein PrPc, denominated PrPSc.3,4 CWD is the cervid equivalent of bovine spongiform encephalopathy (BSE), scrapie in sheep and goat5 or Creutzfeldt-Jakob disease (CJD) in humans.6 Although transmission studies of CWD prions to humanized transgenic mice or non-human primates suggest a strong species barrier,7-9 recent in vitro studies have demonstrated that human PrP can be converted by CWD prions into PrPSc upon adaptation.10 Therefore, a potential for zoonotic transmission, as exemplified by BSE,11 cannot be completely excluded.
A huge body of evidence suggests that CWD can be efficiently transmitted horizontally within and between cervid species,12 which may be the reason for geographical spread and increase in case numbers. Horizontal transmission is explained by the rather unusual peripheral distribution of prions in CWD affected animals and the high susceptibility to the disease by oral infection.13,14 Unlike in most other prion diseases, CWD prions can be found in a wide variety of tissues, such as skeletal and cardiac muscle15,16 or kidney,17 in addition to the lymphoreticular system and blood.18 Furthermore, they are shed in significant amounts in saliva,18,19 urine19 or feces,20 which enables oral infection of animals by foraging on contaminated pastures. In addition, it has been demonstrated that prions can persist in soil21 and that water in endemic areas can contain CWD-associated PrPSc 22.
Currently, disease surveillance is mainly based on testing hunter harvested animals. Since this testing is not obligatory, it depends on the compliance of hunters. CWD test systems are based on the detection of proteinase K resistant PrPSc, either by immunoblot, ELISA or immunohistochemistry.22 The main materials used for this are brain stem homogenates and tonsil or rectoanal mucosa-associated lymphoid tissue (RAMALT) biopsies.23,24 Therefore, intra vitam diagnosis is only possible by invasive methods that require anesthesia of animals. Ante-mortem and, ideally, pre-symptomatic detection of CWD prions in specimens that can be easily obtained without the necessity of anesthetizing animals is highly desirable in order to simplify diagnosis and surveillance. In vitro methods such as protein misfolding cyclic amplification (PMCA)25 or quaking-induced conversion (QuIC)26-28 assay have been proven very useful for sensitive detection of prions in various samples of different species. Using real time (RT-) QuIC, detection of prion amounts as low as 1 fg in cerebrospinal fluid is possible.26 The assay is based on monitoring the incorporation of the amyloid dye Thioflavin T into fibrils of newly converted recombinant PrP seeded by prions or PrPSc contained in the sample.
Since RT-QuIC offers advantages over PMCA, e.g., it does not require sonication, we have chosen this method for testing its usefulness in the detection of CWD prions in deer urine and feces. We demonstrate that CWD prions are detectable in urine of orally infected deer prior to the onset of clinical symptoms. Furthermore, we show that fecal extracts can be used as a seed in RT-QuIC assays. Thereby, we were able to detect CWD prions in fecal extracts collected at later stages of the disease. This study provides the first evidence that RT-QuIC can be successfully used for the preclinical diagnosis of CWD in specimens that are available by non-invasive methods.
Go to: Results Detection of CWD in tissues or body fluids that are easily available and do not require invasive methods is highly desirable and would enable improved surveillance of the disease in free-ranging cervids. Therefore, our aim was to adapt the RT-QuIC assay for detection of CWD prions in deer urine and feces.
At first, we ...
In summary, we demonstrate that CWD prions can be detected by RT-QuIC in urine of orally infected white-tailed deer and mule deer at a pre-symptomatic stage of the disease.
Overall, we demonstrate that RT-QuIC can be used for the pre-symptomatic diagnosis of CWD in urine. After improving sensitivity, e.g., by adaptation of the eQuIC protocol31,32 or by paramagnetic nanoparticle capture,33 the detection of seeding activity in fecal samples can be a versatile tool for simplifying CWD surveillance and diagnosis.
Diagnostic accuracy of rectal mucosa biopsy testing for chronic wasting disease within white-tailed deer (Odocoileus virginianus) herds in North America: Effects of age, sex, polymorphism at PRNP codon 96, and disease progression
Bruce V. Thomsen,1 David A. Schneider, Katherine I. O’Rourke, Thomas Gidlewski, James McLane, Robert W. Allen, Alex A. McIsaac, Gordon B. Mitchell, Delwyn P. Keane, Terry R. Spraker, Aru Balachandran
Abstract. An effective live animal diagnostic test is needed to assist in the control of chronic wasting disease (CWD), which has spread through captive and wild herds of white-tailed deer (Odocoileus virginianus) in Canada and the United States. In the present study, the diagnostic accuracy of rectal mucosa biopsy sample testing was determined in white-tailed deer from 4 CWD-infected captive herds. Specifically, the current study compared the immunohistochemical detection of disease-associated prion protein in postmortem rectal mucosa biopsy samples to the CWD status of each deer as determined by immunodiagnostic evaluations of the brainstem at the obex, the medial retropharyngeal lymph node, and the palatine tonsil. The effects of age, sex, genotype, and disease progression were also evaluated. Diagnostic sensitivity on rectal biopsy samples for CWD in white-tailed deer ranged from 63% to 100%; the pooled estimate of sensitivity was 68% with 95% confidence limits (95% CLs) of 49% and 82%. However, diagnostic sensitivity was dependent on genotype at prion protein gene (PRNP) codon 96 and on disease progression as assessed by obex grade. Diagnostic sensitivity was 76% (95% CLs: 49%, 91%) for 96GG deer but only 42% (95% CLs: 13%, 79%) for 96GS deer. Furthermore, diagnostic sensitivity was only 36% for deer in the earliest stage of disease (obex grade 0) but was 100% for deer in the last 2 stages of preclinical disease (obex grades 3 and 4). The overall diagnostic specificity was 99.8%. Selective use of antemortem rectal biopsy sample testing would provide valuable information during disease investigations of CWD-suspect deer herds.
Key words: Chronic wasting disease; rectal mucosa biopsy; white-tailed deer.
Friday, August 24, 2012
Diagnostic accuracy of rectal mucosa biopsy testing for chronic wasting disease within white-tailed deer (Odocoileus virginianus) herds in North America
Current CWD Status 2013
•Wild cervids: CWD has been detected in 17 states: CO, IL, KS, MD, MN, MO, ND, NE, NY, NM, SD, TX, UT, VA, WI, WV, WY
•Farmed cervid herds: CWD has been detected in 60 farmed cervid herds (40 elk herds, 19 WTD herds,1 red deer herd) in 13 states: CO, KS, IA, MI, MN, MO, MT, NE, NY, OK, PA, SD, WI
•In 2012: farmed red deer (MN); farmed elk (CO); farmed WTD (IA, PA) wild WTD (KS, MO, WI); wild mule deer (TX)
•14 positive farmed herds remain 7 Elk herds (CO); 3 Elk herds (NE) 1 Red deer herd (MN) 1 WTD herds (IA), hunt facility was depopulated 1 WTD herd (PA) was depopulated
Research Updates
•Detection methods – PMCA, RT-QUIC
•Vaccination – Expt’l studies (U. Saskatchewan)
•CWD experimental transmission studies (IC, oral) Cattle – no evidence of disease Sheep (QQ suffolk) – similar presentation to scrapie Fallow deer – Brain lesions (IC route); None (oral route) Red deer - MM genotype – similar to CWD in elk Reindeer – CWD (oral) – clinical disease by 2 years + pathology
•Scrapie transmission studies to deer (IC, oral) •Important to consider interpretation of experimental findings to relevance to natural disease events 24
Prion Persistence in Soils
•Binding to fomite surfaces, minerals, and soil types (sandy, quartz, clay) – (Pederson,2006+)
•Unknown time duration for environmental persistence, bioavailablity, or to remain infectious CWD reported at least 2.2 years (Miller, 2004) Scrapie reported 16 years (Georgsson, 2006) •Detection is difficult (experimental methods) Bioassays (Intra-cranial, oral inoculations) PMCA (protein misfolding cyclic amplification) RT-QUIC (real time - quaking-induced conversion)
•Degradation – research studies Lichens (serine protease) (Johnson, 2011) Metal oxides (manganese) (Russo, 2009)
Prion Decontamination Methods Chemical Methods
•Acids and bases (1-2 M NaOH)
•Alkylating agents Formaldehyde Glutaraldehyde
•Phenols (Environ LpHTM)
•Halogens (NaOCl -20,000 ppm)
•Organic solvents
•Oxidizing agents
•Minerals /Salts (MnO2)
•Proteolytic enzymes NOTE: No EPA registration / FIFRA amendment pending
Disposal Options
•Freeze carcasses pending CWD test results
•After CWD test results – options:  Burial (on site)  Landfill  Rendering (see FDA guidance)  Incineration  Alkaline Hydrolysis/ “Digestion”
Accuracy of administrative diagnostic data for pathologically confirmed cases of Creutzfeldt-Jakob disease
Article in Press

spreading cwd around...tss


Between 1996 and 2002, chronic wasting disease was diagnosed in 39 herds of farmed elk in Saskatchewan in a single epidemic. All of these herds were depopulated as part of the Canadian Food Inspection Agency’s (CFIA) disease eradication program. Animals, primarily over 12 mo of age, were tested for the presence CWD prions following euthanasia. Twenty-one of the herds were linked through movements of live animals with latent CWD from a single infected source herd in Saskatchewan, 17 through movements of animals from 7 of the secondarily infected herds.


***The source herd is believed to have become infected via importation of animals from a game farm in South Dakota where CWD was subsequently diagnosed (7,4). A wide range in herd prevalence of CWD at the time of herd depopulation of these herds was observed. Within-herd transmission was observed on some farms, while the disease remained confined to the introduced animals on other farms.



spreading cwd around...tss


Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea


Hyun-Joo Sohn, Yoon-Hee Lee, Min-jeong Kim, Eun-Im Yun, Hyo-Jin Kim, Won-Yong Lee, Dong-Seob Tark, In- Soo Cho, Foreign Animal Disease Research Division, National Veterinary Research and Quarantine Service, Republic of Korea


Chronic wasting disease (CWD) has been recognized as an important prion disease in native North America deer and Rocky mountain elks. The disease is a unique member of the transmissible spongiform encephalopathies (TSEs), which naturally affects only a few species. CWD had been limited to USA and Canada until 2000.


On 28 December 2000, information from the Canadian government showed that a total of 95 elk had been exported from farms with CWD to Korea.


These consisted of 23 elk in 1994 originating from the so-called “source farm” in Canada, and 72 elk in 1997, which had been held in pre export quarantine at the “source farm”.


Based on export information of CWD suspected elk from Canada to Korea, CWD surveillance program was initiated by the Ministry of Agriculture and Forestry (MAF) in 2001.


All elks imported in 1997 were traced back, however elks imported in 1994 were impossible to identify.


CWD control measures included stamping out of all animals in the affected farm, and thorough cleaning and disinfection of the premises.


In addition, nationwide clinical surveillance of Korean native cervids, and improved measures to ensure reporting of CWD suspect cases were implemented.


*Total of 9 elks were found to be affected. CWD was designated as a notifiable disease under the Act for Prevention of Livestock Epidemics in 2002.


*Additional CWD cases - 12 elks and 2 elks - were diagnosed in 2004 and 2005.


*Since February of 2005, when slaughtered elks were found to be positive, all slaughtered cervid for human consumption at abattoirs were designated as target of the CWD surveillance program.


Currently, CWD laboratory testing is only conducted by National Reference Laboratory on CWD, which is the Foreign Animal Disease Division (FADD) of National Veterinary Research and Quarantine Service (NVRQS).


*In July 2010, one out of 3 elks from Farm 1 which were slaughtered for the human consumption was confirmed as positive.


*Consequently, all cervid – 54 elks, 41 Sika deer and 5 Albino deer – were culled and one elk was found to be positive.


Epidemiological investigations were conducted by Veterinary Epidemiology Division (VED) of NVRQS in collaboration with provincial veterinary services.


*Epidemiologically related farms were found as 3 farms and all cervid at these farms were culled and subjected to CWD diagnosis.


*Three elks and 5 crossbreeds (Red deer and Sika deer) were confirmed as positive at farm 2.


All cervids at Farm 3 and Farm 4 – 15 elks and 47 elks – were culled and confirmed as negative.


Further epidemiological investigations showed that these CWD outbreaks were linked to the importation of elks from Canada in 1994 based on circumstantial evidences.


*In December 2010, one elk was confirmed as positive at Farm 5.


*Consequently, all cervid – 3 elks, 11 Manchurian Sika deer and 20 Sika deer – were culled and one Manchurian Sika deer and seven Sika deer were found to be positive.


This is the first report of CWD in these sub-species of deer.


*Epidemiological investigations found that the owner of the Farm 2 in CWD outbreaks in July 2010 had co-owned the Farm 5.


*In addition, it was newly revealed that one positive elk was introduced from Farm 6 of Jinju-si Gyeongsang Namdo.


All cervid – 19 elks, 15 crossbreed (species unknown) and 64 Sika deer – of Farm 6 were culled, but all confirmed as negative.


: Corresponding author: Dr. Hyun-Joo Sohn (+82-31-467-1867, E-mail: 2011 Pre-congress Workshop: TSEs in animals and their environment 5






Friday, May 13, 2011


Chronic Wasting Disease (CWD) outbreaks and surveillance program in the Republic of Korea



Friday, November 22, 2013


Wasting disease is threat to the entire UK deer population CWD TSE PRION disease in cervids




The Scottish Parliament’s Rural Affairs, Climate Change and Environment Committee has been looking into deer management, as you can see from the following press release,


***and your email has been forwarded to the committee for information:




Friday, November 22, 2013


Wasting disease is threat to the entire UK deer population



Sunday, July 21, 2013


Welsh Government and Food Standards Agency Wales Joint Public Consultation on the Proposed Transmissible Spongiform Encephalopathies (Wales) Regulations 2013


*** Singeltary Submission WG18417



Sunday, June 23, 2013


National Animal Health Laboratory Network Reorganization Concept Paper (Document ID APHIS-2012-0105-0001)


***Terry S. Singeltary Sr. submission



Singeltary submission ;


Program Standards: Chronic Wasting Disease Herd Certification Program and Interstate Movement of Farmed or Captive Deer, Elk, and Moose


DOCUMENT ID: APHIS-2006-0118-0411


***Singeltary submission




Friday, December 14, 2012


DEFRA U.K. What is the risk of Chronic Wasting Disease CWD being introduced into Great Britain? A Qualitative Risk Assessment October 2012




In the USA, under the Food and Drug Administration’s BSE Feed Regulation (21 CFR 589.2000) most material (exceptions include milk, tallow, and gelatin) from deer and elk is prohibited for use in feed for ruminant animals. With regards to feed for non-ruminant animals, under FDA law, CWD positive deer may not be used for any animal feed or feed ingredients. For elk and deer considered at high risk for CWD, the FDA recommends that these animals do not enter the animal feed system. However, this recommendation is guidance and not a requirement by law.


Animals considered at high risk for CWD include:


1) animals from areas declared to be endemic for CWD and/or to be CWD eradication zones and


2) deer and elk that at some time during the 60-month period prior to slaughter were in a captive herd that contained a CWD-positive animal.


Therefore, in the USA, materials from cervids other than CWD positive animals may be used in animal feed and feed ingredients for non-ruminants.


The amount of animal PAP that is of deer and/or elk origin imported from the USA to GB can not be determined, however, as it is not specified in TRACES. It may constitute a small percentage of the 8412 kilos of non-fish origin processed animal proteins that were imported from US into GB in 2011.


Overall, therefore, it is considered there is a __greater than negligible risk___ that (nonruminant) animal feed and pet food containing deer and/or elk protein is imported into GB.


There is uncertainty associated with this estimate given the lack of data on the amount of deer and/or elk protein possibly being imported in these products.




36% in 2007 (Almberg et al., 2011). In such areas, population declines of deer of up to 30 to 50% have been observed (Almberg et al., 2011). In areas of Colorado, the prevalence can be as high as 30% (EFSA, 2011).


The clinical signs of CWD in affected adults are weight loss and behavioural changes that can span weeks or months (Williams, 2005). In addition, signs might include excessive salivation, behavioural alterations including a fixed stare and changes in interaction with other animals in the herd, and an altered stance (Williams, 2005). These signs are indistinguishable from cervids experimentally infected with bovine spongiform encephalopathy (BSE).


Given this, if CWD was to be introduced into countries with BSE such as GB, for example, infected deer populations would need to be tested to differentiate if they were infected with CWD or BSE to minimise the risk of BSE entering the human food-chain via affected venison.




The rate of transmission of CWD has been reported to be as high as 30% and can approach 100% among captive animals in endemic areas (Safar et al., 2008).




In summary, in endemic areas, there is a medium probability that the soil and surrounding environment is contaminated with CWD prions and in a bioavailable form. In rural areas where CWD has not been reported and deer are present, there is a greater than negligible risk the soil is contaminated with CWD prion.




In summary, given the volume of tourists, hunters and servicemen moving between GB and North America, the probability of at least one person travelling to/from a CWD affected area and, in doing so, contaminating their clothing, footwear and/or equipment prior to arriving in GB is greater than negligible. For deer hunters, specifically, the risk is likely to be greater given the increased contact with deer and their environment. However, there is significant uncertainty associated with these estimates.




Therefore, it is considered that farmed and park deer may have a higher probability of exposure to CWD transferred to the environment than wild deer given the restricted habitat range and higher frequency of contact with tourists and returning GB residents.







Singeltary submission ;


Program Standards: Chronic Wasting Disease Herd Certification Program and Interstate Movement of Farmed or Captive Deer, Elk, and Moose


*** DOCUMENT ID: APHIS-2006-0118-0411




Thursday, May 01, 2014


Missouri DNR CWD prevention and captive cervid farming Update



Tuesday, April 29, 2014


Missouri SB964 Immediate Action Required: Captive Cervids Transfer is Still Alive in Senate



Sunday, April 13, 2014


Mineral licks: motivational factors for visitation and accompanying disease risk at communal use sites of elk and deer


Environmental Geochemistry and Health



Thursday, October 03, 2013


*** TAHC ADOPTS CWD RULE THAT the amendments **REMOVE** the requirement for a specific fence height for captives


Texas Animal Health Commission (TAHC) ANNOUNCEMENT October 3, 2013



Wednesday, September 04, 2013


*** cwd - cervid captive livestock escapes, loose and on the run in the wild



*** These results would seem to suggest that CWD does indeed have zoonotic potential, at least as judged by the compatibility of CWD prions and their human PrPC target. Furthermore, extrapolation from this simple in vitro assay suggests that if zoonotic CWD occurred, it would most likely effect those of the PRNP codon 129-MM genotype and that the PrPres type would be similar to that found in the most common subtype of sCJD (MM1).



Saturday, April 19, 2014

Exploring the zoonotic potential of animal prion diseases: In vivo and in vitro approaches



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