cwd tse prion testing PMCA , IHC, tonsil, rectal, biopsy ???
cwd tse prion PMCA , IHC, tonsil, rectal, biopsy
???
spreading cwd around...tss
Project Number: 5P01AI077774-05 Contact PI / Project Leader: SOTO, CLAUDIO
Title: PATHOGENESIS, TRANSMISSION AND DETECTION OF ZOONOTIC PRION DISEASES (P01)
Awardee Organization: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
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: www.cwd-info.org )
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 .
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: njhaley@lamar.colostate.edu
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.
snip...
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:
sgilch@ucalgary.ca 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:
Abstract
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
snip...
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.
snip..
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.
snip..
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
•Detergents
•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”
2014
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:
shonhj@korea.kr) 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
***SINGELTARY SUBMISSION
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
snip...
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.
snip...
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.
snip...
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).
snip...
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.
snip...
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.
snip...
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.
snip...
TSS
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
posted by Terry S. Singeltary Sr. at
10:12 PM
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