http://neurology.thelancet.com Published online October 31, 2005
Coexistence of multiple PrPSc types in individuals with
Creutzfeldt-Jakob disease
Magdalini Polymenidou, Katharina Stoeck, Markus Glatzel, Martin Vey, Anne Bellon, and Adriano Aguzzi
Summary
Background The molecular typing of sporadic Creutzfeldt-Jakob disease (CJD) is based on the size and glycoform
ratio of protease-resistant prion protein (PrPSc), and on PRNP haplotype. On digestion with proteinase K, type 1 and
type 2 PrPSc display unglycosylated core fragments of 21 kDa and 19 kDa, resulting from cleavage around amino
acids 82 and 97, respectively.
Methods We generated anti-PrP monoclonal antibodies to epitopes immediately preceding the differential proteinase
K cleavage sites. These antibodies, which were designated POM2 and POM12, recognise type 1, but not type 2, PrPSc.
Findings We studied 114 brain samples from 70 patients with sporadic CJD and three patients with variant CJD.
Every patient classified as CJD type 2, and all variant CJD patients, showed POM2/POM12 reactivity in the
cerebellum and other PrPSc-rich brain areas, with a typical PrPSc type 1 migration pattern.
Interpretation The regular coexistence of multiple PrPSc types in patients with CJD casts doubts on the validity of
electrophoretic PrPSc mobilities as surrogates for prion strains, and questions the rational basis of current CJD
classifications.
snip...
The above results set the existing CJD classifications
into debate and introduce interesting questions about
human CJD types. For example, do human prion types
exist in a dynamic equilibrium in the brains of affected
individuals? Do they coexist in most or even all CJD
cases? Is the biochemically identified PrPSc type simply
the dominant type, and not the only PrPSc species?
http://neurology.thelancet.com Published online October 31, 2005
what i been saying for years, that the diagnostic criteria differentiating between the nvCJD (i.e. 'the chosen ones') and the sCJD (i.e. 'the forgotten ones') has been terribly flawed from the beginning. .... full text html at bottom of this email, if you want the full text pdf with all the bells and whistles, please email me and i will send pdf....tss
HUMAN TSE USA 2005
Animal Prion Diseases Relevant to Humans (unknown types?)
Thu Oct 27, 2005 12:05
71.248.128.109
About Human Prion Diseases /
Animal Prion Diseases Relevant to Humans
Bovine Spongiform Encephalopathy (BSE) is a prion disease of cattle. Since 1986, when BSE was recognized, over 180,000 cattle in the UK have developed the disease, and approximately one to three million are likely to have been infected with the BSE agent, most of which were slaughtered for human consumption before developing signs of the disease. The origin of the first case of BSE is unknown, but the epidemic was caused by the recycling of processed waste parts of cattle, some of which were infected with the BSE agent and given to other cattle in feed. Control measures have resulted in the consistent decline of the epidemic in the UK since 1992. Infected cattle and feed exported from the UK have resulted in smaller epidemics in other European countries, where control measures were applied later.
Compelling evidence indicates that BSE can be transmitted to humans through the consumption of prion contaminated meat. BSE-infected individuals eventually develop vCJD with an incubation time believed to be on average 10 years. As of November 2004, three cases of BSE have been reported in North America. One had been imported to Canada from the UK, one was grown in Canada, and one discovered in the USA but of Canadian origin. There has been only one case of vCJD reported in the USA, but the patient most likely acquired the disease in the United Kingdom. If current control measures intended to protect public and animal health are well enforced, the cattle epidemic should be largely under control and any remaining risk to humans through beef consumption should be very small. (For more details see Smith et al. British Medical Bulletin, 66: 185. 2003.)
Chronic Wasting Disease (CWD) is a prion disease of elk and deer, both free range and in captivity. CWD is endemic in areas of Colorado, Wyoming, and Nebraska, but new foci of this disease have been detected in Nebraska, South Dakota, New Mexico, Wisconsin, Mississippi Kansas, Oklahoma, Minnesota, Montana, and Canada. Since there are an estimated 22 million elk and deer in the USA and a large number of hunters who consume elk and deer meat, there is the possibility that CWD can be transmitted from elk and deer to humans. As of November 2004, the NPDPSC has examined 26 hunters with a suspected prion disease. However, all of them appeared to have either typical sporadic or familial forms of the disease. The NPDPSC coordinates with the Centers for Disease Control and state health departments to monitor cases from CWD-endemic areas. Furthermore, it is doing experimental research on CWD transmissibility using animal models. (For details see Sigurdson et al. British Medical Bulletin. 66: 199. 2003 and Belay et al. Emerging Infectious Diseases. 10(6): 977. 2004.)
http://www.cjdsurveillance.com/abouthpd-animal.html
SEE STEADY INCREASE IN SPORADIC CJD IN THE USA FROM 1997 TO 2004. SPORADIC CJD CASES TRIPLED, and that is with a human TSE surveillance system that is terrible flawed. in 1997 cases of the _reported_ cases of cjd were at 54, to 163 _reported_ cases in 2004. see stats here;
p.s. please note the 47 PENDING CASES to Sept. 2005
p.s. please note the 2005 Prion D. total 120(8) 8=includes 51 type pending, 1 TYPE UNKNOWN ???
p.s. please note sporadic CJD 2002(1) 1=3 TYPE UNKNOWN???
p.s. please note 2004 prion disease (6) 6=7 TYPE UNKNOWN???
http://www.cjdsurveillance.com/resources-casereport.html
CWD TO HUMANS = sCJD ???
AS implied in the Inset 25 we must not _ASSUME_ that
transmission of BSE to other species will invariably
present pathology typical of a scrapie-like disease.
snip...
http://www.bseinquiry.gov.uk/files/yb/1991/01/04004001.pdf
ATYPICAL TSEs in USA CATTLE AND SHEEP ?
http://www.bseinquiry.gov.uk/files/sc/seac17/tab03.pdf
Infected and Source Flocks
As of August 31, 2005, there were 115 scrapie infected and source flocks (figure 3). There were 3 new infected and source flocks reported in August (Figure 4) with a total of 148 flocks reported for FY 2005 (Figure 5). The total infected and source flocks that have been released in FY 2005 are 102 (Figure 6), with 5 flocks released in August. The ratio of infected and source flocks released to newly infected and source flocks for FY 2005 = 0.69 :
1. In addition, as of August 31, 2005, 574 scrapie cases have been confirmed and reported by the National Veterinary Services Laboratories (NVSL), of which 122 were RSSS cases (Figure 7). This includes 55 newly confirmed cases in August 2005 (Figure 8). Fifteen cases of scrapie in goats have been reported since 1990 (Figure 9). The last goat case was reported in May 2005.
snip...
full text ;
http://www.aphis.usda.gov/vs/nahps/scrapie/monthly_report/monthly-report.html
SCRAPIE USA JULY 2005 UPDATE
AS of July 31, 2005, there were 120 scrapie infected soure flocks (figure 3). There were 16 new infected and source flocks reorted in July (Figure 4) with a total of 143 flocks reported for FY 2005 (Figure 5). The total infected and source flocks that have been released in FY 2005 are 89 (Figure 6), with 8 flocks released in July. The ratio of infected and source flocks released to newly infected and source flocks for FY = 0.62 : 1. IN addition, as of July 31, 2005, 524 scrapie cases have been confirmed and reported by the National Veterinary Services Laboratories (NVSL), of which 116 were RSSS cases (Figure 7). This includes 76 newly confirmed cases in July 2005 (Figure 8). Fifteen cases of scrapie in goats have been reported since 1990 (Figure 9). The last goat case was reported in May 2005. ...........
snip...
http://www.aphis.usda.gov/vs/nahps/scrapie/monthly_report/monthly-report.html
SCRAPIE USA JUNE 2005 UPDATE
AS of June 30, 2005, there were 114 scrapie infected and source flocks
(Figure 3). There were 14 new infected and source flocks reported in June
(Figure 4) with a total of 123 flocks reported for FY 2005 (Figure 5).
snip...
In addition, as of June 30, 2005, 448 scrapie cases have been confirmed and
reported by the National Veterinary Services Laboratories (NVSL), of which
106 were RSSS cases (Figure 7). This includes 81 newly confirmed cases in
June 2005 (Figure 8). Fifteen cases of scrapie in goats have been reported
since 1990 (Figure 9). The last goat case was reported in May 2005.
snip...end
http://www.aphis.usda.gov/vs/nahps/scrapie/monthly_report/monthly-report.html
From: TSS ()
Subject: SCRAPIE USA UPDATE MARCH - JUNE 2005
Date: August 24, 2005 at 7:03 pm PST
SCRAPIE USA MONTHLY REPORT 2005
AS of March 31, 2005, there were 70 scrapie infected source flocks (Figure
3). There were 11 new infected and source flocks reported in March (Figure
4) with a total of 51 flocks reported for FY 2005 (Figure 5). The total
infected and source flocks that have been released in FY 2005 are 39 (Figure
6), with 1 flock released in March. The ratio of infected and source flocks
released to newly infected and source flocks for FY 2005 = 0.76 : 1. IN
addition, as of March 31, 2005, 225 scrapie cases have been confirmed and
reported by the National Veterinary Services Laboratories (NVSL), of which
53 were RSSS cases (Figure 7). This includes 57 newly confirmed cases in
March 2005 (Figure 8). Fourteen cases of scrapie in goats have been reported
since 1990 (Figure 9). The last goat cases was reported in January 2005. New
infected flocks, source flocks, and flocks released or put on clean-up plans
for FY 2005 are depicted in Figure 10. ...
FULL TEXT ;
http://www.aphis.usda.gov/vs/nahps/scrapie/monthly_report/monthly-report.html
Published online before print October 20, 2005
Proc. Natl. Acad. Sci. USA, 10.1073/pnas.0502296102
Medical Sciences
A newly identified type of scrapie agent can naturally infect sheep with resistant PrP genotypes
( sheep prion | transgenic mice )
Annick Le Dur *, Vincent Béringue *, Olivier Andréoletti , Fabienne Reine *, Thanh Lan Laï *, Thierry Baron , Bjørn Bratberg ¶, Jean-Luc Vilotte ||, Pierre Sarradin **, Sylvie L. Benestad ¶, and Hubert Laude *
*Virologie Immunologie Moléculaires and ||Génétique Biochimique et Cytogénétique, Institut National de la Recherche Agronomique, 78350 Jouy-en-Josas, France; Unité Mixte de Recherche, Institut National de la Recherche Agronomique-Ecole Nationale Vétérinaire de Toulouse, Interactions Hôte Agent Pathogène, 31066 Toulouse, France; Agence Française de Sécurité Sanitaire des Aliments, Unité Agents Transmissibles Non Conventionnels, 69364 Lyon, France; **Pathologie Infectieuse et Immunologie, Institut National de la Recherche Agronomique, 37380 Nouzilly, France; and ¶Department of Pathology, National Veterinary Institute, 0033 Oslo, Norway
Edited by Stanley B. Prusiner, University of California, San Francisco, CA, and approved September 12, 2005 (received for review March 21, 2005)
Scrapie in small ruminants belongs to transmissible spongiform encephalopathies (TSEs), or prion diseases, a family of fatal neurodegenerative disorders that affect humans and animals and can transmit within and between species by ingestion or inoculation. Conversion of the host-encoded prion protein (PrP), normal cellular PrP (PrPc), into a misfolded form, abnormal PrP (PrPSc), plays a key role in TSE transmission and pathogenesis. The intensified surveillance of scrapie in the European Union, together with the improvement of PrPSc detection techniques, has led to the discovery of a growing number of so-called atypical scrapie cases. These include clinical Nor98 cases first identified in Norwegian sheep on the basis of unusual pathological and PrPSc molecular features and "cases" that produced discordant responses in the rapid tests currently applied to the large-scale random screening of slaughtered or fallen animals. Worryingly, a substantial proportion of such cases involved sheep with PrP genotypes known until now to confer natural resistance to conventional scrapie. Here we report that both Nor98 and discordant cases, including three sheep homozygous for the resistant PrPARR allele (A136R154R171), efficiently transmitted the disease to transgenic mice expressing ovine PrP, and that they shared unique biological and biochemical features upon propagation in mice. These observations support the view that a truly infectious TSE agent, unrecognized until recently, infects sheep and goat flocks and may have important implications in terms of scrapie control and public health.
--------------------------------------------------------------------------------
Author contributions: H.L. designed research; A.L.D., V.B., O.A., F.R., T.L.L., J.-L.V., and H.L. performed research; T.B., B.B., P.S., and S.L.B. contributed new reagents/analytic tools; V.B., O.A., and H.L. analyzed data; and H.L. wrote the paper.
A.L.D. and V.B. contributed equally to this work.
To whom correspondence should be addressed.
Hubert Laude, E-mail:
[email protected]
www.pnas.org/cgi/doi/10.1073/pnas.0502296102
http://www.pnas.org/cgi/content/abstract/0502296102v1
From: TSS ()
Subject: Interspecies Transmission of Chronic Wasting Disease Prions to Squirrel Monkeys (Saimiri sciureus)
Date: October 19, 2005 at 8:33 am PST
0022-538X/05/$08.00+0 doi:10.1128/JVI.79.21.13794-13796.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.
Interspecies Transmission of Chronic Wasting Disease Prions to Squirrel Monkeys (Saimiri sciureus)
Richard F. Marsh,1, Anthony E. Kincaid,2 Richard A. Bessen,3 and Jason C. Bartz4*
Department of Animal Health and Biomedical Sciences, University of Wisconsin, Madison 53706,1 Department of Physical Therapy,2 Department of Medical Microbiology and Immunology, Creighton University, Omaha, Nebraska 68178,4 Department of Veterinary Molecular Biology, Montana State University, Bozeman, Montana 597183
Received 3 May 2005/ Accepted 10 August 2005
Chronic wasting disease (CWD) is an emerging prion disease of deer and elk. The risk of CWD transmission to humans following exposure to CWD-infected tissues is unknown. To assess the susceptibility of nonhuman primates to CWD, two squirrel monkeys were inoculated with brain tissue from a CWD-infected mule deer. The CWD-inoculated squirrel monkeys developed a progressive neurodegenerative disease and were euthanized at 31 and 34 months postinfection. Brain tissue from the CWD-infected squirrel monkeys contained the abnormal isoform of the prion protein, PrP-res, and displayed spongiform degeneration. This is the first reported transmission of CWD to primates.
--------------------------------------------------------------------------------
* Corresponding author. Mailing address: Department of Medical Microbiology and Immunology, Creighton University, 2500 California Plaza, Omaha, NE 68178. Phone: (402) 280-1811. Fax: (402) 280-1875. E-mail:
[email protected] .
Deceased.
--------------------------------------------------------------------------------
Journal of Virology, November 2005, p. 13794-13796, Vol. 79, No. 21
0022-538X/05/$08.00+0 doi:10.1128/JVI.79.21.13794-13796.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.
http://jvi.asm.org/cgi/content/abstract/79/21/13794?maxtoshow=&HITS=10&hits=10&RESULTFORMAT=&fulltext=cwd&searchid=1129736446553_4280&stored_search=&FIRSTINDEX=0&volume=79&issue=21&journalcode=jvi
Research Project: Transmission, Differentiation, and Pathobiology of Transmissible Spongiform Encephalopathies
Location: Virus and Prion Diseases of Livestock
Title: Experimental Second Passage of Chronic Wasting Disease (Cwd-Mule Deer) Agent to Cattle
Authors
Hamir, Amirali
Kunkle, Robert - bob
Miller, Janice - ARS RETIRED
Greenlee, Justin
Richt, Juergen
Submitted to: Journal Of Comparative Pathology
Publication Acceptance Date: July 25, 2005
Publication Date: N/A
Interpretive Summary: To compare the findings of experimental first and second passage of chronic wasting disease (CWD) in cattle, 6 calves were inoculated into the brain with CWD-mule deer agent previously (first) passaged in cattle. Two other uninoculated calves served as controls. Beginning 10-12 months post inoculation (PI), all inoculates lost appetite and weight. Five animals subsequently developed clinical signs of central nervous system (CNS) abnormality. By 16.5 months PI, all cattle had been euthanized because of poor prognosis. None of the animals showed microscopic lesions of spongiform encephalopathy (SE) but the CWD agent was detected in their CNS tissues by 2 laboratory techniques (IHC and WB). These findings demonstrate that inoculated cattle amplify CWD agent but also develop clinical CNS signs without manifestation of microscopic lesions of SE. This situation has also been shown to occur following inoculation of cattle with another TSE agent, namely, sheep scrapie. The current study confirms previous work that indicates that the diagnostic tests currently used for confirmation of bovine spongiform encephalopathy (BSE) in the U.S. would detect CWD in cattle, should it occur naturally. Furthermore, it raises the possibility of distinguishing CWD from BSE in cattle due to the absence of microscopic lesions and a unique multifocal distribution of PrPres, as demonstrated by IHC, which in this study, appears to be more sensitive than the WB.
Technical Abstract: To compare clinicopathological findings of first and second passage of chronic wasting disease (CWD) in cattle, a group of calves (n=6) were intracerebrally inoculated with CWD-mule deer agent previously (first) passaged in cattle. Two other uninoculated calves served as controls. Beginning 10-12 months post inoculation (PI), all inoculates lost appetite and lost weight. Five animals subsequently developed clinical signs of central nervous system (CNS) abnormality. By 16.5 months PI, all cattle had been euthanized because of poor prognosis. None of the animals showed microscopic lesions of spongiform encephalopathy (SE) but PrPres was detected in their CNS tissues by immunohistochemistry (IHC) and Western blot (WB) techniques. These findings demonstrate that intracerebrally inoculated cattle not only amplify CWD PrPres but also develop clinical CNS signs without manifestation of morphologic lesions of SE. This situation has also been shown to occur following inoculation of cattle with another TSE agent, scrapie. The current study confirms previous work that indicates the diagnostic techniques currently used for confirmation of bovine spongiform encephalopathy (BSE) in the U.S. would detect CWD in cattle, should it occur naturally. Furthermore, it raises the possibility of distinguishing CWD from BSE in cattle due to the absence of neuropathologic lesions and a unique multifocal distribution of PrPres, as demonstrated by IHC, which in this study, appears to be more sensitive than the WB.
http://www.ars.usda.gov/research/publications/publications.htm?seq_no_115=178318
[Docket No. 03-025IFA] FSIS Prohibition of the Use of Specified Risk Materials for Human Food and Requirement for the Disposition of Non-Ambulatory Disabled Cattle
03-025IFA
03-025IFA-2
Terry S. Singeltary
Page 1 of 17
From: Terry S. Singeltary Sr. [
[email protected]]
Sent: Thursday, September 08, 2005 6:17 PM
To:
[email protected].
Subject: [Docket No. 03-025IFA] FSIS Prohibition of the Use of Specified Risk Materials for Human Food and Requirements
for the Disposition of Non-Ambulatory Disabled Cattle
Greetings FSIS,
I would kindly like to submit the following to [Docket No. 03-025IFA] FSIS Prohibition of the Use of Specified Risk Materials for Human Food and
Requirements for the Disposition of Non-Ambulatory Disabled Cattle
THE BSE/TSE SUB CLINICAL Non-Ambulatory Disabled Cattle
Broken bones and such may be the first signs of a sub clinical BSE/TSE Non-Ambulatory Disabled Cattle ;
SUB CLINICAL PRION INFECTION
MRC-43-00
Issued: Monday, 28 August 2000
NEW EVIDENCE OF SUB-CLINICAL PRION INFECTION: IMPORTANT RESEARCH
FINDINGS RELEVANT TO CJD AND BSE
A team of researchers led by Professor John Collinge at the Medical
Research Council Prion Unit1 report today in the Proceedings of the
National Academy of Sciences, on new evidence for the existence of a
?sub-clinical? form of BSE in mice which was unknown until now.
The scientists took a closer look at what is known as the ?species
barrier? - the main protective factor which limits the ability of
prions2 to jump from one species to infect another. They found the mice
had a ?sub-clinical? form of disease where they carried high levels of
infectivity but did not develop the clinical disease during their normal
lifespan. The idea that individuals can carry a disease and show no
clinical symptoms is not new. It is commonly seen in conventional
infectious diseases.
Researchers tried to infect laboratory mice with hamster prions3 called
Sc237 and found that the mice showed no apparent signs of disease.
However, on closer inspection they found that the mice had high levels
of mouse prions in their brains. This was surprising because it has
always been assumed that hamster prions could not cause the disease in
mice, even when injected directly into the brain.
In addition the researchers showed that this new sub-clinical infection
could be easily passed on when injected into healthy mice and hamsters.
The height of the species barrier varies widely between different
combinations of animals and also varies with the type or strain of
prions. While some barriers are quite small (for instance BSE easily
infects mice), other combinations of strain and species show a seemingly
impenetrable barrier. Traditionally, the particular barrier studied here
was assumed to be robust.
Professor John Collinge said: "These results have a number of important
implications. They suggest that we should re-think how we measure
species barriers in the laboratory, and that we should not assume that
just because one species appears resistant to a strain of prions they
have been exposed to, that they do not silently carry the infection.
9/13/2005
2
Page 2 of 17
This research raises the possibility, which has been mentioned before,
that apparently healthy cattle could harbour, but never show signs of, BSE.
"This is a timely and unexpected result, increasing what we know about
prion disease. These new findings have important implications for those
researching prion disease, those responsible for preventing infected
material getting into the food chain and for those considering how best
to safeguard health and reduce the risk that theoretically, prion
disease could be contracted through medical and surgical procedures."
ISSUED FRIDAY 25 AUGUST UNDER EMBARGO. PLEASE NOTE THAT THE EMBARGO IS
SET BY THE JOURNAL.
FOR FURTHER INFORMATION CONTACT THE MRC PRESS OFFICE ON 020 7637 6011
(OFFICE HOURS) OR 07818 428297 OR 0385 774357 (OUT-OF-OFFICE-HOURS) OR
PROFESSOR JOHN COLLINGE ON 020 7594 3760. PLEASE NOTE THAT OWING TO
TRAVEL COMMITMENTS PROFESSOR COLLINGE WILL ONLY BE AVAILABLE UNTIL 16.30
ON FRIDAY 25 AUGUST AND CONTACTABLE AGAIN ON MONDAY 28 AUGUST VIA THE
MRC PRESS OFFICE. DR ANDREW HILL (A CO-AUTHOR ON THE PAPER) FROM THE
DEPARTMENT OF PATHOLOGY AT THE UNIVERSITY OF MELBOURNE WILL BE AVAILABLE
ON 00 61 3 8344 3995 (DURING OFFICE HOURS) OR 00 61 3 9443 0009
(OUT-OF-OFFICE HOURS). PLEASE NOTE THAT AUSTRALIA IS TEN HOURS AHEAD OF
UK TIME.
NOTES FOR EDITORS
Professor Collinge is a consultant neurologist and Director of the newly
formed MRC Prion Unit based at The Imperial College School of Medicine
at St Mary?s Hospital. He is also a member of the UK Government?s
Spongiform Encephalopathy Advisory Committee (SEAC). The MRC prion unit
is was set up in 1999, and its work includes molecular genetic studies
of human prion disease and transgenic modelling of human prion diseases.
Prions are unique infectious agents that cause fatal brain diseases such
as Creutzfeldt-Jakob disease (CJD) in humans and scrapie and BSE (mad
cow disease) in animals. In some circumstances prions from one species
of animals can infect another and it is clear that BSE has done this to
cause the disease variant CJD in the UK and France. It remains unclear
how large an epidemic of variant CJD will occur over the years ahead.
The strain of prion used here to infect the mice is the Sc237 strain
(also known as 263K) which infects hamsters, and until now was assumed
not to infect mice.
This research was funded by the Medical Research Council and Wellcome
Trust.
The Medical Research Council (MRC) is a national organisation funded by
the UK tax-payer. Its business is medical research aimed at improving
human health; everyone stands to benefit from the outputs. The research
it supports and the scientists it trains meet the needs of the health
services, the pharmaceutical and other health-related industries and the
academic world. MRC has funded work which has led to some of the most
significant discoveries and achievements in medicine in the UK. About
half of the MRC?s expenditure of £345 million is invested in over 50 of
its Institutes and Units, where it employs its own research staff. The
remaining half goes in the form of grant support and training awards to
individuals and teams in universities and medical schools.
The Wellcome Trust is the world's largest medical research charity with
a spend of some £600 million in the current financial year 1999/2000.
The Wellcome Trust supports more than 5,000 researchers, at 400
locations, in 42 different countries to promote and foster research with
the aim of improving human and animal health. As well as funding major
initiatives in the public understanding of science, the Wellcome Trust
is the country's leading supporter of research into the history of
medicine.
http://www.mrc.ac.uk/index/public_interest/public-press_office/public-press_releases_2000/public-mrc-43-00.htm
SNIP...FULL TEXT;
9/13/2005
Page 3 of 17
https://web01.aphis.usda.gov/regpublic.nsf/0/eff9eff1f7c5cf2b87256ecf000df08d?OpenDocument
PNAS | August 29, 2000 | vol. 97 | no. 18 | 10248-10253
Neurobiology
Species-barrier-independent prion replication in apparently
resistant species
Andrew F. Hill*, Susan Joiner*, Jackie Linehan*, Melanie Desbruslais*, Peter L. Lantos , and John Collinge*,
* Medical Research Council Prion Unit and Department of Neurogenetics, Imperial College School of Medicine at St. Mary's, London W2 1PG, United Kingdom; and
Department of Neuropathology, Institute of Psychiatry, London SE5 8AF, United Kingdom
Communicated by Charles Weissmann, Imperial College of Science, Technology, and Medicine, London, United Kingdom, June 23, 2000 (received for review January
20, 2000)
Abstract
Transmission of prions between mammalian species is thought to be limited by a "species barrier," which depends on differences in the primary structure of prion proteins in
the infecting inoculum and the host. Here we demonstrate that a strain of hamster prions thought to be nonpathogenic for conventional mice leads to prion replication to high
levels in such mice but without causing clinical disease. Prions pathogenic in both mice and hamsters are produced. These results demonstrate the existence of subclinical
forms of prion infection with important public health implications, both with respect to iatrogenic transmission from apparently healthy humans and dietary exposure to cattle
and other species exposed to bovine spongiform encephalopathy prions. Current definitions of the species barrier, which have been based on clinical end-points, need to be
fundamentally reassessed.
snip...
Discussion
Implication of Demonstration of Subclinical Prion Infection. In prion diseases, infectious titers in the brain rise progressively
throughout prolonged, clinically silent periods that precede the onset of disease. Thus asymptomatic animals may harbor
significant infectious titers in brain and other tissues. However, there may be subclinical, as distinct from such preclinical, forms of
prion infection, where animals become asymptomatic carriers of infectivity and do not develop clinical disease in their lifetimes (7,
28). Such carrier states are well recognized in other infectious diseases. However, in prion diseases, where incubation periods are
extremely prolonged, distinction between subclinical and preclinical states is more difficult. It certainly can be argued that animals
dying after a typical lifespan without clinical signs of prion disease but harboring high levels of infectivity represent the late
preclinical stage of "transmissions" where the "incubation period" exceeds the normal lifespan (29). The distinction between the
terms subclinical and preclinical is essentially a semantic one in this context. Here we use the term subclinical infection
operationally to refer to animals in which prion replication is occurring but which have not developed clinical signs of prion disease
during a normal lifespan.
We have demonstrated that conventional mice inoculated with Sc237 prions harbor high levels of PrPSc and high prion titers in
their brains without developing clinical signs of prion disease within their normal lifespan. These results imply the existence of
subclinical prion infections that can be induced by challenge with prions from another species. However, whether or not this
infectivity is classified as preclinical or subclinical, it has important public health implications. Iatrogenic transmission could occur
from apparently healthy humans who may harbor high prion titers and many animal species (including sheep, pigs, and poultry)
were exposed to BSE prions via contaminated feed and could have developed subclinical prion infection. It is known that BSE
9/13/2005
Page 4 of 17
prions retain their distinctive strain characteristics after passage in a number of other species including humans (4, 13), arguing
that such BSE passaged in species other than cattle also may be pathogenic to humans. The possibility that subclinical BSE might
be present in other species and thereby present a threat to human health has been raised (30) but not yet rigorously investigated.
Furthermore, these data argue in favor of screening apparently healthy cattle after slaughter to investigate whether significant
levels of subclinical or preclinical BSE are present.
Secondly, because animals can harbor high levels of infectivity without developing clinical signs of prion disease, these results
argue that PrPSc and indeed prions (whether or not they are identical) may not themselves be highly neurotoxic. Such results are
in accordance with earlier findings of a lack of correlation between clinical disease and neuropathological features of prion disease
(31), prion diseases in which PrPSc is barely or not detectable (32-35), and studies in mice with reduced levels of PrPC expression
that have extremely high levels of PrPSc and prions in the brain and yet remain well for several months after their wild-type
counterparts succumb (36). Conversely, Tg20 mice, with high levels of PrPC, have short incubation periods and yet produce low
levels of PrPSc after inoculation with mouse prions (27). In addition, brain grafts producing high levels of PrPSc do not damage
adjacent tissue in PrP knockout (Prnpo/o) mice (37). The cause of neurodegeneration in prion diseases remains unclear. It remains
possible that prion neurodegeneration is related, at least in part, to loss of function of PrPC. That Prnpo/o mice (other than those
associated with overexpression of the Prnp-like gene Prnd; ref. 38) do not develop neurodegeneration could be caused by
compensatory adaptations during neurodevelopment. Complete or near complete ablation of PrP expression in an adult mouse
using conditional gene expression methods has not yet been achieved. An alternative hypothesis is that a toxic, possibly
infectious, intermediate is produced in the process of conversion of PrPC to PrPSc, with PrPSc, present as highly aggregated
material, being a relatively inert end-product. The steady-state level of such a toxic monomeric or oligomeric PrP intermediate then
could determine rate of neurodegeneration. One possibility is that Sc237-inoculated CD-1 mice propagate prions very slowly and
that such a toxic intermediate is generated at extremely low levels that are tolerated by the mouse. The fact that the PrPScnegative
Sc237-inoculated CD-1 mice were the ones culled earlier than those that were PrPSc positive, allows the assumption that
they may have become PrPSc positive had they lived longer. A more detailed study of the time course of accumulation of infectivity
will be necessary to investigate this further.
Transmission of Infectivity from Subclinical Animals. The transmission properties of prions from the subclinical Sc237inoculated
CD-1 mice were remarkable. With respect to transmissions to additional CD-1 or Tg20 mice, the 100% attack rate and
highly consistent incubation periods suggest transmission in the absence of a barrier. However, the incubation periods, notably in
the Tg20 mice, which succumb to RML mouse prions in around 60 days (27), are very prolonged. The 100% attack rate argues
against this being a consequence of low prion titer in the inoculum. Incubation period at end point dilution in Tg20 mice of RML
mouse prions is around 109 days (37). Remarkably, passage in hamsters of this isolate also showed a 100% attack rate and
consistent incubation periods suggestive of transmission in the absence of a barrier. Again, incubation periods were extremely
prolonged and differed markedly from the transmission properties of Sc237/263K prions in hamsters (8, 10, 39). Indeed, the
incubation period seen would correspond to an Sc237 titer in Syrian hamsters of <103 LD50/g brain, which is completely
inconsistent with the titers measured; Sc237 incubation periods at end point dilution in Syrian hamsters are around 130 days (40).
That a 100% attack rate was seen at a 127-day incubation period argues against persistent Sc237 inoculum, rather than newly
formed prions, being responsible for the pathogenicity to hamsters. Together, these data suggested production of novel infectivity,
pathogenic for both mice and hamsters on passage of Sc237 to CD-1 mice.
A recent report has suggested that hamster scrapie (263K) may persist in the brains of inoculated C57BL/10 mice for prolonged
periods without replication (41). Our data are not consistent with infectivity in the PrPSc-positive Sc237-inoculated CD-1 mice
being the result of persistence of residual Sc237 hamster scrapie inoculum. High levels of mouse PrPSc (and no hamster PrPSc)
are detectable on Western blot, and prions pathogenic for mice are generated. Intracerebral inoculation is known to result in wide
distribution of the inoculum outside the brain via the circulation and, presumably as a result of other clearance mechanisms, brain
titers fall to undetectable levels within a few days (42). Prion titers present in the brains of these mice ( 108 LD50/g mouse brain
assayed in hamsters) considerably exceed those inoculated ( 8.5 × 106). Together, these data argue strongly for prion
replication in these mice. It is possible that the prions detected in the brains of the C57BL/10 mice in the earlier study were not
caused by persistence of inoculated 263K, but by propagation of prions with the properties we describe. The species origin of
PrPSc (hamster or mouse) in the 263K-inoculated C57BL/10 mice was not reported. The observation periods postinoculation were
generally much shorter than those we report here. That those mice with the longest survival postinoculation produced the shortest
incubation periods on passage of infectivity into hamsters is consistent with propagation, rather than simply persistence, of prions
in this earlier study (41).
9/13/2005
Page 5 of 17
Re-Evaluation of Species Barriers. Importantly, these data seriously question our current understanding of species barriers. The
assessment of species barriers has relied on the development of a clinical disease in inoculated animals. On this basis there is a
highly efficient barrier limiting transmission of Sc237 prions to mice. However, although not developing a clinical disease, and
indeed living as long as mock-inoculated mice, Sc237-inoculated mice may accumulate high levels of prions in their brains.
Previous studies on the species barrier between hamsters and mice (using the Sc237 or 263K strain) did not report whether PrPSc
and/or infectivity were present in clinically unaffected animals (8, 12) or have attempted passage from mice only up to 280 days
postinoculation (10). The barrier to primary passage appears in this case to be to the development of rapid neurodegeneration and
the resulting clinical syndrome rather than a barrier to prion propagation itself.
The transmission characteristics of prions generated in the brains of Sc237-inoculated CD-1 mice argue that one or more distinct
prion strains have been generated. The finding that Sc237-inoculated CD-1 mice in which PrPSc could not be detected on Western
blot were the ones that had been culled after shorter periods than mice with detectable PrPSc argues that prion propagation is
occurring in all of these mice, but is detectable only after prolonged incubation periods. That high levels of hamster infectivity were
present in the PrPSc-negative Sc237-inoculated CD-1 mouse (examined at 463 days postinoculation) in the absence of detectable
mouse infectivity, whereas very high and relatively comparable titers of both mouse and hamster infectivity were present in the
PrPSc-positive Sc237-inoculated CD-1 mouse (examined at 730 days postinoculation) suggests that more than one strain may be
propagating in these mice, with preferential replication of a strain with higher pathogenicity for hamsters early in the incubation
period. One possibility is that early replication of a prion strain pathogenic only for hamsters is induced in Sc237-inoculated CD-1
mice, then later followed by the generation of a second strain that is pathogenic for mice. More extensive passage studies,
including cloning of strains at end-point dilution in both mice and hamsters, will be required to investigate this further and to
characterize the strain(s) of prions generated in the brains of Sc237-inoculated CD-1 mice.
http://www.pnas.org/cgi/content/full/97/18/10248
Neurobiology of Disease
Subclinical Bovine Spongiform Encephalopathy Infection in
Transgenic Mice Expressing Porcine Prion Protein
Joaquín Castilla,1 Alfonso Gutiérrez-Adán,2 Alejandro Brun,1 Deirdre Doyle,3 Belén Pintado,2 Miguel A. Ramírez,2
Francisco J. Salguero,1 Beatriz Parra,1 Fayna Díaz San Segundo,1 José M. Sánchez-Vizcaíno,1 Mark Rogers,3 and
Juan M. Torres1
1Centro de Investigación en Sanidad Animal, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria, Valdeolmos,
28130 Madrid, Spain, 2Departamento de Reproducción Animal y Conservación de Recursos Zoogenéticos, 28040 Madrid, Spain,
and 3Department of Zoology and Conway Institute for Biomolecular and Biomedical Research, University College Dublin, Belfield,
Dublin 4, Ireland
Abstract
The bovine-porcine species barrier to bovine spongiform encephalopathy (BSE) infection was explored by generating transgenic But the main
mouse lines expressing the porcine prion protein (PrP) gene. All of the porcine transgenic (poTg) mice showed clinical signs of
BSE after intracerebral inoculation with a high-titer BSE inoculum. The protease-resistant PrP (PrPres) was detected in 14% (3 of
22) of the BSE-infected poTg mice by immunohistochemical or immunoblot analysis. Despite being able to infect 42% (5 of 12) of
control mice, a low-dose BSE inoculum failed to penetrate the species barrier in our poTg mouse model. The findings of these
infectivity studies suggest that there is a strong species barrier between cows and pigs. However, after second-passage infection
of poTg mice using brain homogenates of BSE-inoculated mice scoring negative for the incoming prion protein as inoculum, it was
9/13/2005
Page 6 of 17
possible to detect the presence of the infectious agent. Thus, porcine-adapted BSE inocula were efficient at infecting poTg mice,
giving rise to an incubation period substantially reduced from 300 to 177 d after inoculation and to the presence of PrPres in 100%
(21 of 21) of the mice. We were therefore able to conclude that initial exposure to the bovine prion may lead to subclinical infection
such that brain homogenates from poTg mice classified as uninfected on the basis of the absence of PrPres are infectious when
used to reinoculate poTg mice. Collectively, our findings suggest that these poTg mice could be used as a sensitive bioassay
model for prion detection in pigs.
snip...
Discussion
The transgenic mouse lines developed expressed the porcine PrP transgene at different levels. This is characteristic of random transgene integration in the mouse genome
by the microinjection technique. Given that high expression levels promote reduced incubation times for heterologous and homologous prion propagation in mice (Scott et
al., 1997b ; Castilla et al., 2003 ), we selected two poTg lines, poTg001 and poTg027, expressing fourfold and 16-fold, respectively, the levels of PrP protein found in pig
brain.
We observed that mice expressing higher levels of poPrP spontaneously developed clinical signs. A similar neurological syndrome was described previously by Westaway
et al. (1994 ) in older Tg PrP mice expressing high levels of hamster, ovine, or murine PrP transgenes. This phenomenon may be related to the observed toxicity of
overexpressed PrP in certain cell lines, which suggests that lack of physiological PrPC expression may render pathogenic in mice. However, the lifespan of poTg027 mice
was much longer than the time needed by porcine prions to propagate in these animals, and the confirmation of infection could be tested using proteinase K (PK)-resistant
studies. In none of the cases did the noninoculated animals presenting late clinical signs show PK-resistant protein.
We observed substantial evidence of subclinical BSE infection in our poTg mice. PoTg mice inoculated with BSE1 showed no clinical signs of BSE or detectable PrPres
protein. However, subsequent passage of brain homogenates from these mice indicated the high level of infectivity of one of these animals. The presence of subclinical
infection was particularly evident when we used the poTgBSE1-N2 inoculum (first-passage boTgBSE1 in poTg PrPres-negative mice), which led to a mean incubation time
of 269 d and to PrPres that was detectable by Western blotting in two of six mice. The presence of subclinical infection has been reported in other species (Race and
Chesebro, 1998 ; Hill et al., 2000 ). Although there is no evidence of clinical BSE disease in the domestic pig population, pigs are susceptible to BSE, and our
observations raise the possibility of subclinical infection occurring in pigs. The poTg model could be used as an assay for subclinical infection in suspected cases of prion
disease in pigs.
Three inocula (Fig. 3) were used to infect the poTg mice. These inocula are known to efficiently infect transgenic mice expressing the bovine PrP gene (boTg110 line)
(Castilla et al., 2003 ). We used the same vector to express the porcine and bovine PrP genes under the mouse PrP promoter. In the boTg110 model, increasing the
PrPres titer had no effect on the incubation time. When the low-dose BSE1 inoculum was tested in a normal mouse line, the animals showed neurological signs of disease,
and 5 of 12 (42%) scored positive for PrPres. These data indicate that the BSE1 inoculum can cross the bovine-murine species barrier, although the expression level of the
mouse PrPC is approximately half that shown by our transgenic lines.
However, the low-dose BSE1 inoculum provided evidence for a strong bovine-porcine species barrier, because it produced no signs of infection in the poTg001 or poTg027
mice. Survival times were unchanged compared with those observed in control PBS-inoculated poTg001 or poTg027 mice, and no PrPres was detected in any of the 39
inoculated mice (Table 1). In contrast, the higher titer BSE2 and boTgBSE1 inocula were able to breach the bovine-porcine species barrier, and PrPres was detected in 3 of
22 infected poTg mice (14%). Additional evidence for the bovine-porcine species barrier was obtained in second-passage transmission from BSE-infected poTg mice. The
survival time dropped from 488 to 198 d postinoculation (dpi) for poTg001 and from 300 to 177 dpi for the poTg027 mice. The presence of a strong barrier may explain the
resistance to infection shown by pigs during the BSE epidemic in the United Kingdom.
Contrary to the strong species barrier observed when poTg mice were inoculated with BSE, there was little evidence of a species barrier in the opposite direction (i.e., when
we infected boTg110 mice with poTgBSE1). All of the boTg mice infected with this inoculum scored positive for PrPres, suggesting that the barrier has different difficulty
levels depending on the direction of the infection. Western blotting analysis confirmed that the PrPres observed in the bo110Tg mice displayed the same pattern (band size,
glycoform ratio) as the boTgBSE1 or BSE1 inocula but a pattern that is different from that of the newly generated porcine prion (po027Tg) (Fig. 2C). A characteristic feature
of the BSE prion is that it retains its biological properties when transmitted to other species such as humans (Collinge and Rossor, 1996 ; Collinge et al., 1996 ; Will and
Zeidler, 1996 ; Scott et al., 1999 ), sheep (Foster et al., 1993 , 2001 ), or mice (Fraser et al., 1992 ; Lasmezas et al., 1997 ). Thus, the lack of a strong species
barrier observed for transmission in the direction of pig to cow might be explained if the initial BSE inoculum infecting the pig confers BSE-like properties on the porcine
prion, although the primary amino acid sequence of this prion is the porcine one. Alternatively, these results could be explained as follows: (1) the bovine PrP is a very
permissive protein, more easily transformed by other heterologous prions or (2) the new porcine prion is highly infectious compared with others. This second possibility will
be studied using other transgenic mice expressing ovine and human PrP.
The species barrier is related to amino acid sequence differences in the globular domain of the PrP protein, which undergoes a conformational change from -helix to pleated
sheet structures. The porcine PrP shows the most unique amino acid sequence (5) in this domain when compared with the mouse, cow, sheep, hamster, and
human PrP sequences. Figure 6 compares the globular domains of porcine, bovine, and mouse PrP. It may be observed that four of the five unique amino acids occur in
9/13/2005
Page 7 of 17
helix 3, and that there are two additional differences in this helix between the porcine and bovine sequence, I to V and R to K. The K residue is known to alter the length and
quality of definition of helix 3 (Calzolai et al., 2000 ), and it is possible that this combination of amino acid variants alters the structure of helix 3 sufficiently to inhibit
interactions between porcine PrPC and PrPres. Nuclear magnetic resonance analysis indicates that the global architecture of this region is similar for all species analyzed to
date (Riek et al., 1998 ; Lopez Garcia et al., 2000 ; Zahn et al., 2000 ), but individual amino acid changes have been shown to affect local conformation or surface
charge (Lopez Garcia et al., 2000 ). These subtle differences may be sufficient to strengthen or weaken a species barrier.
http://www.jneurosci.org/cgi/content/full/24/21/5063
British Medical Bulletin 66:161-170 (2003)
© 2003 The British Council
Subclinical prion infection in humans and animals
Andrew F Hill and John Collinge
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, London, UK
Transmission of prion diseases between mammalian species is limited by a so-called 'species' or 'transmission' barrier. Recognition of prion transmission usually relies on
the appearance of clinical symptoms in inoculated animals and the interval between inoculation and appearance of clinical disease is designated incubation period. At some
point during this clinically silent period, neuropathological and biochemical changes as well as accumulation of prions in the brain can be detected and this stage can be
called preclinical prion disease. Recently, several lines of evidence have suggested that subclinical forms of prion disease exist, in which high levels of infectivity and
PrPSc are found in animals that do not develop clinically apparent disease during a normal life-span. Such asymptomatic prion 'carrier' states challenge our current
understanding of pathogenesis as well as of the molecular basis of barriers to transmission. Subclinical as well as preclinical/clinical prion disease may be relevant when
analysing the risk to public health of potential sources of prion exposure.
http://bmb.oxfordjournals.org/cgi/content/abstract/66/1/161
Journal of Virology, July 2003, p. 7991-7998, Vol. 77, No. 14
0022-538X/03/$08.00+0 DOI: 10.1128/JVI.77.14.7991-7998.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
Subclinical Prion Disease Induced by Oral Inoculation
Alana M. Thackray,1 Michael A. Klein,2 and Raymond Bujdoso1*
Department of Clinical Veterinary Medicine, Centre for Veterinary Science, University of Cambridge, Cambridge, United Kingdom CB3 OES,1 Prion Research Group,
Institute of Virology and Immunobiology, University of Würzburg, D-97078 Würzburg, Germany2
Received 9 January 2003/ Accepted 30 April 2003
Natural transmission of prion disease is believed to occur by peripheral infection such as oral inoculation. Following this route of inoculation, both the peripheral nervous
system and the lymphoreticular system may be involved in the subsequent neuroinvasion of the central nervous system by prions, which may not necessarily result in
clinical signs of terminal disease. Subclinical prion disease, characterized by the presence of infectivity and PrPSc in the absence of overt clinical signs, may occur. It is not
known which host factors contribute to whether infection with prions culminates in a terminal or subclinical disease state. We have investigated whether the level of host
PrPc protein expression is a factor in the development of subclinical prion disease. When RML prion inoculum was inoculated by either the i.c. or intraperitoneal route, wildtype
and tga20 mice both succumbed to terminal prion disease. In contrast, orally inoculated tga20 mice succumbed to terminal prion disease, whereas wild-type mice
showed no clinical signs. However, wild-type mice sacrificed 375 or 525 days after oral inoculation harbored significant levels of brain PrPSc and infectivity. These data
show that same-species transmission of prions by the oral route in animals that express normal levels of PrPc can result in subclinical prion disease. This indicates that the
level of host PrPc protein expression is a contributing factor to the regulation of development of terminal prion disease. Events that increase PrPc expression may
predispose a prion-infected animal to the more deleterious effects of prion pathology.
* Corresponding author. Mailing address: Centre for Veterinary Science, Department of Clinical Veterinary Medicine, University of Cambridge, Madingley Rd., Cambridge,
United Kingdom CB3 OES. Phone: 44-1223-337655. Fax: 44-1223-337610. E-mail:
[email protected] .
9/13/2005
Page 8 of 17
Journal of Virology, July 2003, p. 7991-7998, Vol. 77, No. 14
0022-538X/03/$08.00+0 DOI: 10.1128/JVI.77.14.7991-7998.2003
http://jvi.asm.org/cgi/content/abstract/77/14/7991
Journal of Virology, March 2002, p. 2510-2517, Vol. 76, No. 5
0022-538X/02/$04.00+0 DOI: 10.1128/jvi.76.5.2510-2517.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.
Chronic Subclinical Prion Disease Induced by Low-Dose Inoculum
*** Alana M. Thackray,1 Michael A. Klein,2 Adriano Aguzzi,3 and Raymond Bujdoso1*
Centre for Veterinary Science, Department of Clinical Veterinary Medicine, University of Cambridge, Cambridge CB3 0ES, United Kingdom,1 University Hospital Basel,
Institute of Pathology, CH-4003 Basel,2 Institute of Neuropathology, University Hospital of Zürich, CH-8091 Zürich, Switzerland3
Received 24 September 2001/ Accepted 16 November 2001
We have compared the transmission characteristics of the two mouse-adapted scrapie isolates, ME7 and Rocky Mountain Laboratory (RML), in tga20 mice. These mice
express elevated levels of PrP protein compared to wild-type mice and display a relatively short disease incubation period following intracerebral prion inoculation. Terminal
prion disease in tga20 mice induced by ME7 or RML was characterized by a distinct pattern of clinical signs and different incubation times. High-dose RML inoculated
intracerebrally into tga20 mice induced the most rapid onset of clinical signs, with mice succumbing to terminal disease after only 58 ± 3 days. In contrast, high-dose ME7
gave a mean time to terminal disease of 74 ± 0 days. Histological examination of brain sections from prion-inoculated tga20 mice at terminal disease showed that ME7 gave
rise to a more general and extensive pattern of vacuolation than RML. Low-dose inoculum failed to induce terminal disease but did cause preclinical symptoms, including
the appearance of reversible clinical signs. Some mice oscillated between showing no clinical signs and early clinical signs for many months but never progressed to
terminal disease. Brain tissue from these mice with chronic subclinical prion disease, sacrificed at >200 days postinoculation, contained high levels of infectivity and showed
the presence of PrPSc. Parallel analysis of brain tissue from mice with terminal disease showed similar levels of infectivity and detectable PrPSc. These results show that
high levels of infectivity and the presence of the abnormal isomer of PrP can be detected in mice with subclinical disease following low-dose prion inoculation.
* Corresponding author. Mailing address: Centre for Veterinary Science, Department of Clinical Veterinary Medicine, University of Cambridge, Madingley Rd., Cambridge
CB3 0ES, United Kingdom. Phone: 44-1223-337655. Fax: 44-1223-337610. E-mail:
[email protected] .
Journal of Virology, March 2002, p. 2510-2517, Vol. 76, No. 5
0022-538X/02/$04.00+0 DOI: 10.1128/jvi.76.5.2510-2517.2002
http://jvi.asm.org/cgi/content/abstract/76/5/2510
Journal of Virology, November 2001, p. 10106-10112, Vol. 75, No. 21
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.21.10106-10112.2001
Long-Term Subclinical Carrier State Precedes Scrapie Replication and Adaptation in a
Resistant Species: Analogies to Bovine Spongiform Encephalopathy and Variant
Creutzfeldt-Jakob Disease in Humans
Richard Race, Anne Raines, Gregory J. Raymond, Byron Caughey, and Bruce Chesebro*
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, Hamilton, Montana 59840
Received 24 May 2001/Accepted 31 July 2001
9/13/2005
Page 9 of 17
Cattle infected with bovine spongiform encephalopathy (BSE) appear to be a reservoir for transmission of variant Creutzfeldt-Jakob disease (vCJD) to humans. Although just
over 100 people have developed clinical vCJD, millions have probably been exposed to the infectivity by consumption of BSE-infected beef. It is currently not known
whether some of these individuals will develop disease themselves or act as asymptomatic carriers of infectivity which might infect others in the future. We have studied
agent persistence and adaptation after cross-species infection using a model of mice inoculated with hamster scrapie strain 263K. Although mice inoculated with hamster
scrapie do not develop clinical disease after inoculation with 10 million hamster infectious doses, hamster scrapie infectivity persists in brain and spleen for the life span of
the mice. In the present study, we were surprised to find a 1-year period postinfection with hamster scrapie where there was no evidence for replication of infectivity in
mouse brain. In contrast, this period of inactive persistence was followed by a period of active replication of infectivity as well as adaptation of new strains of agent capable
of causing disease in mice. In most mice, neither the early persistent phase nor the later replicative phase could be detected by immunoblot assay for protease-resistant
prion protein (PrP). If similar asymptomatic carriers of infection arise after exposure of humans or animals to BSE, this could markedly increase the danger of additional
spread of BSE or vCJD infection by contaminated blood, surgical instruments, or meat. If such subclinical carriers were negative for protease-resistant PrP, similar to our
mice, then the recently proposed screening of brain, tonsils, or other tissues of animals and humans by present methods such as immunoblotting or immunohistochemistry
might be too insensitive to identify these individuals.
* Corresponding author. Mailing address: Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, 903 South Fourth St., Hamilton, MT 59840-2999. Phone:
(406) 363-9354. Fax: (406) 363-9286. E-mail:
[email protected] .
Journal of Virology, November 2001, p. 10106-10112, Vol. 75, No. 21
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.21.10106-10112.2001
http://jvi.asm.org/cgi/content/abstract/75/21/10106
From: TSS ()
Subject: PrPSc distribution of a natural case of bovine spongiform encephalopathy
Date: August 8, 2005 at 12:28 pm PST
PrPSc distribution of a natural case of bovine
spongiform encephalopathy
Yoshifumi Iwamaru, Yuka Okubo, Tamako Ikeda, Hiroko Hayashi, Mori-
kazu Imamura, Takashi Yokoyama and Morikazu Shinagawa
Priori Disease Research Center, National Institute of Animal Health, 3-1-5
Kannondai, Tsukuba 305-0856 Japan
[email protected]
Abstract
Bovine spongiform encephalopathy (BSE) is a disease of cattle that causes
progressive neurodegeneration of the central nervous system. Infectivity
of BSE agent is accompanied with an abnormal isoform of prion protein
(PrPSc).
The specified risk materials (SRM) are tissues potentially carrying BSE
infectivity. The following tissues are designated as SRM in Japan: the
skull including the brain and eyes but excluding the glossa and the masse-
ter muscle, the vertebral column excluding the vertebrae of the tail, spinal
cord, distal illeum. For a risk management step, the use of SRM in both
animal feed or human food has been prohibited. However, detailed
PrPSc distribution remains obscure in BSE cattle and it has caused controversies
about definitions of SRM. Therefore we have examined PrPSc
distribution in a BSE cattle by Western blotting to reassess definitions of
SRM.
The 11th BSE case in Japan was detected in fallen stock surveillance.
The carcass was stocked in the refrigerator. For the detection of PrPSc,
200 mg of tissue samples were homogenized. Following collagenase
treatment, samples were digested with proteinase K. After digestion,
PrPSc was precipitated by sodium phosphotungstate (PTA). The pellets
were subjected to Western blotting using the standard procedure.
Anti-prion protein monoclonal antibody (mAb) T2 conjugated horseradish
peroxidase was used for the detection of PrPSc.
PrPSc was detected in brain, spinal cord, dorsal root ganglia, trigeminal
ganglia, sublingual ganglion, retina. In addition, PrPSc was also detected
in the peripheral nerves (sciatic nerve, tibial nerve, vagus nerve).
Our results suggest that the currently accepted definitions of SRM in
9/13/2005
179
Page 10 of 17
BSE cattle may need to be reexamined.
T. Kitamoto (Ed.)
PRIONS
Food and Drug Safety
================
ALSO from the International Symposium of Prion Diseases held in Sendai, October 31, to
November 2, 2004;
Bovine spongiform encephalopathy (BSE) in Japan
snip...
"Furthermore, current studies into transmission of cases of BSE that are
atypical or that develop in young cattle are expected to amplify the BSE
prion"
NO. Date conf. Farm Birth place and Date Age at diagnosis
8. 2003.10.6. Fukushima Tochigi 2001.10.13. 23
9. 2003.11.4. Hiroshima Hyogo 2002.1.13. 21
Test results
# 8b, 9c cows Elisa Positive, WB Positive, IHC negative, histopathology
negative
b = atypical BSE case
c = case of BSE in a young animal
b,c, No PrPSc on IHC, and no spongiform change on histology
International Symposium of Prion Diseases held in Sendai, October 31, to
November 2, 2004.
Tetsuyuki Kitamoto
Professor and Chairman
Department of Prion Research
Tohoku University School of Medicine
2-1 SeiryoAoba-ku, Sendai 980-8575, JAPAN
TEL +81-22-717-8147 FAX +81-22-717-8148
e-mail;
[email protected]
Symposium Secretariat
Kyomi Sasaki
TEL +81-22-717-8233 FAX +81-22-717-7656
e-mail:
[email protected]
=================================
9/13/2005
--------------------------------------------------------------------------------
--------------------------------------------------------------------------------
Page 11 of 17
From: TSS ()
Subject: Atypical Proteinase K-Resistant Prion Protein (PrPres) observed in an Apparently Healthy 23-Month-Old Holstein Steer
Date: August 26, 2005 at 10:24 am PST
Atypical Proteinase K-Resistant Prion Protein (PrPres) observed in an Apparently Healthy 23-Month-Old Holstein Steer
Jpn. J. Infect. Dis., 56, 221-222, 2003
Laboratory and Epidemiology Communications
Atypical Proteinase K-Resistant Prion Protein (PrPres) Observed in an Apparently Healthy 23-Month-Old Holstein Steer
Yoshio Yamakawa*, KenÕichi Hagiwara, Kyoko Nohtomi, Yuko Nakamura, Masahiro Nishizima ,Yoshimi Higuchi1, Yuko Sato1, Tetsutaro Sata1 and the Expert Committee
for BSE Diagnosis, Ministry of Health, Labour and Welfare of Japan2
Department of Biochemistry & Cell Biology and 1Department of Pathology, National Institute of Infectious Diseases, Tokyo 162-8640 and 2Miistry of Health, Labour and
Welfare, Tokyo 100-8916
Communicated by Tetsutaro Sata
(Accepted December 2, 2003)
*Corresponding author: Mailing address: Department of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 1628640,
Japan. Tel: +81-3-5285-1111, Fax: +81-3-5285-1157, E-mail:
[email protected]
Since October 18, 2001, 'bovine spongiform encephalopathy (BSE) examination for all cattle slaughtered at abattoirs in the country' has been mandated in Japan by the
Ministry of Health, Labour and Welfare (MHLW). 'Plateria' ELISA-kit (Bio-Rad Laboratories, Hercules, Calif., USA) is routinely used at abattoirs for detecting proteinase K
(PK)-resistant prion protein (PrPSc) in the obex region. Samples positive according to the ELISA screening are further subjected to Western blot (WB) and histologic and
immunohistochemical examination (IHC) at the National Institute of Infectious Diseases (NIID) or Obihiro University. If PrPSc is detected either by WB or by IHC, the cattle
are diagnosed as BSE. The diagnosis is approved by the Expert Committee for BSE Diagnosis, MHLW. From October 18, 2001 to September 30, 2003, approximately 2.5
million cattle were screened at abattoirs. A hundred and te
