TimH said:
Whatever Ron. Any thinking person can clearly see that you failed to answer my questions, and why.
FACT- vCJD cases are decreasing(year to year).
FACT- resarchers don't use muscle tissue homogenate in transmission studies because IF prions are present in muscle tissue, they are not present in high enough numbers to transmit the disease, even through IC injection, never mind oraly.
Ron is well aware of these facts but refuses to acknowledge them.He needs to create fear over BSE/vCJD in order to sell his testing technology.
It's that simple. If he needs to compare selling untested beef to " a drunk behind the wheel", so be it for our Ronnie.
Wake up people and see this P.O.S. for what he truly is.
I travel through Edmonton,from time to time Ron, we should do lunch.
:lol: :lol: :lol: :lol:
lets analyze tims comments ;
timmyboy writes ;
> Whatever Ron. Any thinking person can clearly see that you failed to answer my questions, and why.
> FACT- vCJD cases are decreasing(year to year).
tim, what you fail to understand is that the UK BSE nvCJD theory, is but a small part, of a much larger problem.
we have multiple strains, in multiple species now, some of which even coexist with one another in the same
species, and as they mutate, they seem to become more virulent to humans and other species.
this is why we must stop it in it's tracks now, because if you continue to flounder, you will not be able to stop it,
and that is even debatable now. we must stop the continued amplification and transmission via proven routes,
and the SRM removal is a proven factor. all one has to do is look at the numbers of BSE cases in the UK from
the height of the BSE epidemic, to the low cases now, all do to the feed ban and eradication of documented cases.
THIS is why SRM removal, and the continued vigilance in transmission studies with all tissue in these atypical strains
are paramount.... comprende amigo?
BSE CASES IN RELATION TO FEED BAN SEE MAP OF CURVE
page 11
http://www.defra.gov.uk/animalh/cvo/report/2002/cvo2002.pdf
Creutzfeldt-Jakob Disease, Prion Protein Gene Codon 129VV, and a Novel PrPSc
Type in a Young British Woman
Simon Mead, PhD, MRCP; Susan Joiner, MSc; Melanie Desbruslais, BSc; Jonathan
A. Beck, BSc; Michael O'Donoghue, PhD; Peter Lantos, FRCP; Jonathan D. F.
Wadsworth, PhD; John Collinge, FRS
Arch Neurol. 2007;64(12):1780-1784.
Background Variant Creutzfeldt-Jakob disease (vCJD) is an acquired prion
disease causally related to bovine spongiform encephalopathy that has
occurred predominantly in young adults. All clinical cases studied have been
methionine homozygotes at codon 129 of the prion protein gene (PRNP) with
distinctive neuropathological findings and molecular strain type (PrPSc type
4). Modeling studies in transgenic mice suggest that other PRNP genotypes
will also be susceptible to infection with bovine spongiform encephalopathy
prions but may develop distinctive phenotypes.
Objective To describe the histopathologic and molecular investigation in a
young British woman with atypical sporadic CJD and valine homozygosity at
PRNP codon 129.
Design Case report, autopsy, and molecular analysis.
Setting Specialist neurology referral center, together with the laboratory
services of the MRC [Medical Research Council] Prion Unit.
Subject Single hospitalized patient.
Main Outcome Measures Autopsy findings and molecular investigation results.
Results Autopsy findings were atypical of sporadic CJD, with marked gray
and white matter degeneration and widespread prion protein (PrP) deposition.
Lymphoreticular tissue was not available for analysis. Molecular analysis of
PrPSc (the scrapie isoform of PrP) from cerebellar tissue demonstrated a
novel PrPSc type similar to that seen in vCJD (PrPSc type 4). However, this
could be distinguished from the typical vCJD pattern by an altered protease
cleavage site in the presence of the metal ion chelator EDTA.
Conclusions Further studies will be required to characterize the prion
strain seen in this patient and to investigate its etiologic relationship
with bovine spongiform encephalopathy. This case illustrates the importance
of molecular analysis of prion disease, including the use of EDTA to
investigate the metal dependence of protease cleavage patterns of PrPSc.
Author Affiliations: MRC [Medical Research Council] Prion Unit and
Department of Neurodegenerative Disease, Institute of Neurology, University
College London, National Hospital for Neurology and Neurosurgery, London,
England (Drs Mead, Wadsworth, and Collinge; Mss Joiner and Desbruslais; and
Mr Beck); and Institute of Psychiatry, King's College London (Dr Lantos). Dr
O'Donoghue is now with the Department of Clinical Neurology, Nottingham
University Hospitals NHS [National Health Service] Trust, Nottingham,
England.
http://archneur.ama-assn.org/cgi/content/short/64/12/1780
Possible vCJD case in UK may signal more to come
Lisa Schnirring Staff Writer
Jan 9, 2008 (CIDRAP News) – A British woman who died of a brain disease
suggestive of variant Creutzfeld-Jacob disease (vCJD) had a genetic marker
not seen in any previous vCJD patients, raising the possibility that her
illness represented a new form of the disease that could signal a new wave
of infections, according to a recent research report.
The researchers, who reported their findings in the December 2007 issue of
Archives of Neurology, found that the 39-year-old woman carried the VV
(valine-homozygous) version of the prion protein gene (PRNP), a type
previously thought to confer protection against vCJD.
Past research has linked vCJD to eating meat products contaminated with
brain and spinal cord material from cattle infected with bovine spongiform
encephalopathy (BSE), or mad cow disease. Normal prion proteins in the brain
are corrupted after contact with the BSE agent, eventually causing death in
both cattle and humans. BSE, vCJD, and sporadic CJD—a rare disease of
unknown cause that closely resembles vCJD—are all prion diseases, also known
as transmissible spongiform encephalopathies.
Cases of vCJD began surfacing in the United Kingdom in 1996, in the wake of
a BSE epidemic in cattle. According to the most recent update from the
National CJD Surveillance Unit (NCJDSU) based at the Western General
Hospital in Edinburgh, Scotland, the number of patients in the UK who have
died of confirmed or probable vCJD stands at 163. Until the case described,
all vCJD patients who had been tested had the MM (methionine-homozygous)
version of PRNP.
In early 1999 the patient described started having visual symptoms, followed
by a host of other neurological problems, such as memory and gait
impairments, according to the report. Polymerase chain reaction testing
revealed that the patient had the VV variant of the PRNP gene. The patient
died 14 months later.
Brain autopsy findings included severe gray- and white-matter degeneration
and extensive prion protein deposits in the cortex and white matter, which
the authors wrote is atypical for sporadic CJD. Molecular analysis of the
pathologic prion protein (PrPSc) from the woman's cerebellar tissue showed a
novel type of PrPSc that was similar in some, but not all, respects to type
4, which is seen in vCJD.
The authors wrote that it wasn't clear if the PrPSc typing points to a BSE
cause of the patient's illness or if the finding represents another form of
sporadic CJD.
Though a single case can't be the basis for connecting a novel PrPSc type to
BSE, "it will be important to see whether other similar cases occur in the
United Kingdom and other BSE-exposed countries," the researchers wrote.
Studies in transgenic mice are under way to explore transmission
characteristics related to the woman's case, according to the report.
Simon Mead, the study's lead author, said the findings shouldn't cause
alarm, according to a Jan 5 New Scientist report. "The final conclusion
remains open. It is waving the flag for neurologists to watch for other
cases," said Mead, who is at the Medical Research Council Prion Unit at
University College London.
Mead told New Scientist that patterns of prion disease seem to vary among
people depending on the prion gene variant they have, and incubation period
could be one aspect in which the variants differ. Experts have said CJD is
known to have a long incubation period, perhaps as long as 50 years.
In 2006, another group of British researchers analyzed DNA from three
surgical samples that had previously tested positive in immunohistochemical
studies of vCJD prevalence in the UK (though the patients had no clinical
signs of the disease). Genotype analysis of the patients' PRNP at codon 129
found that two of the samples were of the VV type, providing the first
evidence that patients from this subgroup could be infected. (DNA could not
be extracted from the third sample.) Previously, people who carried at least
one copy of the V variant of PRNP were thought to have no risk of
contracting vCJD.
The authors of the 2006 study suggested their findings might mean that
people who are infected with vCJD and have a VV type may have a prolonged
incubation period, during which the disease could spread either via blood
donations or from contaminated surgical instruments used on the individuals
during the asymptomatic phase of the illness.
Will Hueston, DVM, PhD, director of the University of Minnesota Center for
Animal Health and Food Safety in St. Paul, told CIDRAP News that it's too
soon to say whether the DNA findings from the woman are associated with BSE.
"I think that neurologists are probably attempting to be more cautious," he
said. "This is most likely not BSE, but they [the researchers] want to be
very clear that similar cases should be thoroughly evaluated."
The results of the study could also signify another variant of CJD, which is
already known to occur in various forms, "but they don't know what box to
put it in," said Hueston, adding that classifying prion disease types is
often difficult.
Mead S, Joiner S, Desbruslais S, et al. Creutzfeldt-Jakob disease, prion
protein gene codon 129VV, and a novel PrPSc type in a young British woman.
Arch Neurol 2007 Dec;64(12):1780-74 [Abstract]
http://www.cidrap.umn.edu/cidrap/content/other/bse/news/jan0908vcjd.html
timmyboy also writes here ;
> FACT- resarchers don't use muscle tissue homogenate in transmission studies because IF prions are present in muscle tissue, they are not present in high
> enough numbers to transmit the disease, even through IC injection, never mind oraly.
:shock:
what an idiotic and asinine statement. do you even read what you write tim ?
let's look at all the facts ;
P02.35 Molecular Features of the Protease-resistant Prion Protein (PrPres) in H-type BSE
Biacabe, A-G1; Jacobs, JG2; Gavier-Widén, D3; Vulin, J1; Langeveld, JPM2;Baron, TGM1 1AFSSA, France; 2CIDC-Lelystad, Netherlands; 3SVA, Sweden
Western blot analyses of PrPres accumulating in the brain of BSE-infected cattle have demonstrated 3 different molecular phenotypes regarding to the apparent molecular masses and glycoform ratios of PrPres bands. We initially described isolates (H-type BSE) essentially characterized by higher PrPres molecular mass and decreased levels of the diglycosylated PrPres band, in contrast to the classical type of BSE.This type is also distinct from another BSE phenotype named L-type BSE, or also BASE (for Bovine Amyloid Spongiform Encephalopathy), mainly characterized by a low representation of the diglycosylated PrPres band as well as a lower PrPres molecular mass. Retrospective molecular studies in France of all availableBSE cases older than 8 years old and of part of the other cases identified since the beginning of the exhaustive surveillance of the disease in 2001 allowed to identify 7 H-type BSE cases, among 594 BSE cases that could be classified as classical, L- or H-type BSE. By Western blot analysis of H-type PrPres, we described a remarkable specific feature with antibodies raised against the C-terminal region of PrP that demonstrated the existence of a more C-terminal cleaved form of PrPres (named PrPres #2 ), in addition to the usual PrPres form (PrPres #1). In the unglycosylated form, PrPres #2 migrates at about 14 kDa, compared to 20 kDa for PrPres #1. The proportion of the PrPres #2 in cattle seems to by higher compared to the PrPres #1.Furthermore another PK–resistant fragment at about 7 kDa was detected by some more N-terminal antibodies and presumed to be the result of cleavages of both N- and C-terminal parts of PrP. These singular features were maintained after transmission of the disease toC57Bl/6 mice. The identification of these two additional PrPres fragments (PrPres #2 and7kDa band) reminds features reported respectively in sporadicCreutzfeldt-Jakob disease and in Gerstmann-Sträussler-Scheinker (GSS) syndrome in humans.
FC5.5.1 BASE Transmitted to Primates and MV2 sCJD Subtype Share PrP27-30 and PrPScC-terminal Truncated Fragments
Zanusso, G1; Commoy, E2; Fasoli, E3; Fiorini, M3; Lescoutra, N4; Ruchoux,MM4; Casalone, C5; Caramelli, M5; Ferrari, S3; Lasmezas, C6; Deslys, J-P4; Monaco, S3 1University of Verona, of Neurological and Visual Sciences,Italy; 2CEA, IMETI/SEPIA, France; 3University of Verona, Neurological andVisual Sciences, Italy; 4IMETI/SEPIA, France; 5IZSPLVA, Italy; 6The Scripps Research Insitute, USA
The etiology of sporadic Creutzfeldt-Jakob disease (sCJD), the most frequent human prion disease, remains still unknown. The marked disease phenotypeheterogeneity observed in sCJD is thought to be influenced by the type of proteinaseK-resistantprion protein, or PrPSc (type 1 or type 2 according to the electrophoreticmobility of the unglycosylated backbone), and by the host polymorphic Methionine/Valine(M/V) codon 129 of the PRNP. By using a two-dimensional gel electrophoresis(2D-PAGE) and imunoblotting we previously showed that in sCJD, in addition to thePrPSc type, distinct PrPSc C-terminal truncated fragments (CTFs) correlated with different sCJD subtypes. Based on the combination of CTFs and PrPSc type, we distinguished three PrPSc patterns:
(i) the first was observed in sCJD with PrPSc type 1 of all genotypes,;
(ii) the second was found in M/M-2 (cortical form); (iii) the third inamyloidogenic M/V-2 and V/V-2 subtypes (Zanusso et al., JBC 2004) . Recently, we showed that sCJD subtype M/V-2 shared molecular and pathological features with an atypical form of BSE, named BASE, thus suggesting a potential link between the two conditions. This connection was further confirmed after 2D-PAGE analysis, which showed an identical PrPSc signature, including the biochemical pattern of CTFs. To pursue this issue, we obtained brain homogenates from Cynomolgus macaques intracerebrally inoculated with brain homogenates from BASE. Samples were separated by usinga two dimensional electrophoresis (2D-PAGE) followed by immunoblotting. We here show that the PrPSc pattern obtained in infected primates is identical to BASE and sCJD MV-2 subtype. These data strongly support the link, or at least acommon ancestry, between a sCJD subtype and BASE. This work was supported by Neuroprion(FOOD-CT-2004-506579)
*******************************************************
USA MAD COW CASES IN ALABAMA AND TEXAS
***PLEASE NOTE***
USA BASE CASE, (ATYPICAL BSE), AND OR TSE (whatever they are calling ittoday), please note that both the ALABAMA COW, AND THE TEXAS COW, both were''H-TYPE'', personal communication Detwiler et al Wednesday, August 22, 200711:52 PM. ...TSS
http://lists.ifas.ufl.edu/cgi-bin/wa.exe?A2=ind0708&L=sanet-mg&T=0&P=19779
(May 16, 2007)
TAFS1 Position Paper on Atypical scrapie and Atypical BSE
snip...
Does it represent a risk to human health?
It is too early to tell whether or not it represents a risk to humans. For the moment it is
assumed to be a danger, and is treated like BSE. Results of experimental transmission
to primates remain unpublished. Some scientists suggest that similarities between the
molecular features of H-type BSE and some prion diseases of humans may indicate
that they are related. Care must be exercised in interpreting such preliminary data(8)
specifically with regard to suggestions of a cause and effect.
Transmissibility to cattle has been confirmed, but remains currently unpublished as the
study is incomplete. It may therefore be possible to investigate further, by oral
challenge, whether or not the infectious agent is distributed around the body in a
different way from BSE, possibly infecting tissues that are not considered-infectious
in BSE. This may have implications for risk management and public health.
http://www.tafsforum.org/position_papers/TAFS_POSITION_PAPER_ON_ATYPICAL_SCRAPIE_AND_%20ATYPICAL_BSE_070516.pdf
In FY 2007, 331 scrapie cases have been confirmed and reported by the
National Veterinary Services Laboratories (NVSL), including 59* Regulatory
Scrapie Slaughter Surveillance (RSSS) cases (Figure 5 and Slide 16). In FY
2007, two field cases, one validation case, and two RSSS cases were
consistent with Nor-98 scrapie. The Nor98-like cases originated from flocks
in California, Minnesota, Colorado, Wyoming and Indiana respectively.
Nineteen cases of scrapie in goats have been reported since 1990 (Figure 6).
The last goat case was reported in September 2007.
snip...
see full report here ;
http://www.aphis.usda.gov/animal_health/animal_diseases/scrapie/downloads/yearly_report.pps
P03.141
Aspects of the Cerebellar Neuropathology in Nor98
Gavier-Widén, D1; Benestad, SL2; Ottander, L1; Westergren, E11National Veterinary Insitute, Sweden; 2National Veterinary Institute,Norway
Nor98 is a prion disease of old sheep and goats. This atypical form ofscrapie was first described in Norway in 1998. Several features of Nor98 were shown to be different from classical scrapie including the distribution of disease associated prion protein (PrPd) accumulation in the brain. The cerebellum is generally the most affected brain area in Nor98. The study here presented aimed at adding information on the neuropathology in the cerebellum of Nor98 naturally affected sheep of various genotypes in Sweden and Norway. A panel of histochemical and immunohistochemical (IHC) stainings such as IHCfor PrPd, synaptophysin, glial fibrillary acidic protein, amyloid, and cell markers for phagocytic cells were conducted. The type of histological lesions and tissuereactions were evaluated. The types of PrPd deposition were characterized.The cerebellar cortex was regularly affected, even though there was a variationin the severity of the lesions from case to case.Neuropil vacuolation was more marked in the molecular layer, but affected also the granular cell layer. There was a loss of granule cells. Punctate deposition of PrPd was characteristic. It was morphologically and in distribution identical with that of synaptophysin, suggesting that PrPd accumulates in the synaptic structures. PrPd was also observed in the granule cell layerand in the white matter.
*** The pathology features of Nor98 in the cerebellum of the affected sheep showed similarities with those of sporadic Creutzfeldt-Jakob disease inhumans.
please see these abstract of these studies here ;
http://www.prion2007.com/pdf/Prion%20Book%20of%20Abstracts.pdf
Prion Infection of Muscle Cells In Vitro
Wendy M. Dlakic, Eric Grigg, and Richard A. Bessen*
Department of Veterinary Molecular Biology, Montana State University, Bozeman, Montana 59717
Received 28 November 2006/ Accepted 9 February 2007
The prion agent has been detected in skeletal muscle of humans and animals with prion diseases. Here we report scrapie infection of murine C2C12 myoblasts and myotubes in vitro following coculture with a scrapie-infected murine neuroblastoma (N2A) cell line but not following incubation with a scrapie-infected nonneuronal cell line or a scrapie brain homogenate. Terminal differentiation of scrapie-infected C2C12 myoblasts into myotubes resulted in an increase in the expression of the disease-specific prion protein, PrPSc. The amount of scrapie infectivity or PrPSc in C2C12 myotubes was comparable to the levels found in scrapie-infected N2A cells, indicating that a high level of infection was established in muscle cells. Subclones of scrapie-infected C2C12 cells produced high levels of PrPSc in myotubes, and the C-terminal C2 polypeptide fragment of PrPSc was found based on deglycosylation and PrPSc-specific immunoprecipitation of cell lysates. This is the first report of a stable prion infection in muscle cells in vitro and of a long-term prion infection in a nondividing, differentiated peripheral cell type in culture. These in vitro studies also suggest that in vivo prion infection of skeletal muscle requires contact with prion-infected neurons or, possibly, nerve terminals.
snip...
In an attempt to develop a cell culture model for scrapie infection in muscle, we investigated the murine C2C12 myoblast cell line, since it can undergo myogenic differentiation into nondividing myotubes, which are similar to myofibrils in vivo (59). In this study we demonstrate that scrapie infection and PrPSc production can be established in murine C2C12 myoblasts and myotubes in vitro. The properties of C2C12 cells infected with 22L scrapie were similar to those found in scrapie-infected N2A cells, including a high level of PrPSc resistance to PK degradation, the production of C1 and C2 PrP polypeptide fragments, and the ability to induce scrapie in mice upon intracerebral inoculation. Based on the similarities of scrapie infection in these cell lines, we propose that 22L scrapie-infected C2C12 myoblasts and myotubes will provide a useful tissue culture model for studying prion infection of skeletal muscle in vivo. The recent report of prion infection in muscle of deer with chronic wasting disease indicates a food safety risk to humans who consume venison (2).
In order to establish scrapie infection in C2C12 cells, it was necessary to coculture myoblasts with 22L scrapie-infected N2A cells and then selectively grow the myoblasts in Zeocin-containing growth medium, which does not support N2A cell survival. Attempts to establish scrapie infection in C2C12 myoblasts and myotubes by direct incubation with a scrapie brain homogenate were unsuccessful in four independent experiments even though homogenates contained high scrapie titers that were able to establish scrapie infection of N2A cells. These findings suggested that the 22L scrapie agent, or induction of PrPSc formation, was directly transferred between the infected N2A cells and uninfected myoblasts or myotubes. It is likely that scrapie infection requires direct contact between these cell types, since separation of these cell types by a porous membrane did not result in scrapie infection of the C2C12 myoblasts. The type of cell that can transfer scrapie infection to C2C12 myoblasts and myotubes also appears to be an important factor, since coculture with 22L scrapie-infected SMB cells did not result in infection of C2C12/Zeor cells. However, previous coculture studies demonstrated that SMB cells infected with the 79A strain of the scrapie agent can transfer infection to uninfected SMB cells (31); cell-to-cell contact is not needed to infect SMB cells, since infection can be established using the standard method of incubating the dividing cells with a scrapie brain homogenate (31). These findings suggest that the ability of a scrapie-infected neuronal cell line, but not a nonneuronal cell line, to infect C2C12 myoblasts or myotubes may be due to the formation of a neuromuscular junction in vitro. Both N2a cells and C2C12 myotubes have been demonstrated to form neuromuscular junctions in nerve-muscle cell line cocultures (17, 61). Transynaptic spread of the prion agent has been demonstrated along motor pathways (9). Localization of PrPSc to the neuromuscular junction in prion-infected muscle cells in vivo (39) and of PrPC to both the presynaptic and postsynaptic membranes in the neuromuscular junction provides support for a role of the neuromuscular junction in prion agent spread. The transfer of scrapie infection via exosomes (47) and release of scrapie infectivity into the supernatant (7, 49) represents another possible mechanism for prion infection of C2C12 cells, but this is less likely to have been the case in the current study since we observed an absence of measurable PrPSc in C2C12 cells when they were cocultured with 22L scrapie-infected N2A cells that were separated by a porous membrane. However, we cannot exclude the possibility that these mechanisms of cell-to-cell spread are involved in scrapie infection of C2C12 muscle cells.
In C2C12 muscle cells there was evidence of N-terminal truncation of the full-length prion protein into either a 16-kDa PrPC polypeptide in both mock- and 22L scrapie-infected C2C12 cells or an 18-kDa PrPSc polypeptide in 22L scrapie-infected C2C12 myotubes. The size and cleavage properties of these truncated PrP polypeptides are consistent, but not precisely matched, with the 17-kDa C1 and 21-kDa C2 PrP polypeptide fragments, respectively, that have been described in mock- and prion-infected brain and cell lines (16, 30, 55, 58). The apparent discrepancy in the size of the C2 PrPSc polypeptide between 22L C2C12 myotubes and other scrapie-infected cells or brain tissue could be due to the use of different polyacrylamide gel separation systems or, perhaps, to differences in cleavage specificity among tissue types. Prior studies demonstrated that N-terminal truncation of PrPSc into the C2 polypeptide is mediated by calpain-dependent endoproteolytic cleavage to generate the C2 PrPSc polypeptide (58). Calpains play an essential role in muscle cell differentiation and, in myoblasts, they have been associated with cell adhesion and spreading (23). In C2C12 cells, inhibition of calpains prevented myocyte migration and fusion into multinucleated myotubes (22). High calpain activity during myogenic differentiation is consistent with cleavage into the C2 PrPSc polypeptide in 22L scrapie-infected C2C12 myotubes.
Our findings that scrapie infection can be established in both dividing myoblasts and nondividing myotubes in vitro directly illustrates that the scrapie agent can replicate in muscle cells. This study supports previous reports of direct detection of PrPSc in myocytes of sheep with scrapie, in skeletal muscle cells of the tongue in rodents experimentally infected with the prion agent (24, 39, 53), and in cardiomuscle fibers of humans with Creutzfeldt-Jakob disease (4) or deer with chronic wasting disease (29). The C2C12 cell model for scrapie infection could be used to investigate prion infection of muscle, including the mechanism of scrapie infection in muscle cells, the pathway of PrPSc formation and trafficking in a terminally differentiated cell type, and the relationship of prion infection in muscle cells to prion-induced myopathy that is found in scrapie of sheep and chronic wasting disease of deer and elk.
http://jvi.asm.org/cgi/content/full/81/9/4615
Brevia
Prions in Skeletal Muscles of Deer with Chronic Wasting Disease
Rachel C. Angers,1* Shawn R. Browning,1* Tanya S. Seward,2 Christina J. Sigurdson,4 Michael W. Miller,5 Edward A. Hoover,4 Glenn C. Telling1,2,3
The emergence of chronic wasting disease (CWD) in deer and elk in an increasingly wide geographic area, as well as the interspecies transmission of bovine spongiform encephalopathy to humans in the form of variant Creutzfeldt Jakob disease, have raised concerns about the zoonotic potential of CWD. Because meat consumption is the most likely means of exposure, it is important to determine whether skeletal muscle of diseased cervids contains prion infectivity. Here bioassays in transgenic mice expressing cervid prion protein revealed the presence of infectious prions in skeletal muscles of CWD-infected deer, demonstrating that humans consuming or handling meat from CWD-infected deer are at risk to prion exposure.
1 Department of Microbiology, Immunology and Molecular Genetics, University of Kentucky, Lexington, KY 40536, USA.
2 Sanders Brown Center on Aging, University of Kentucky, Lexington, KY 40536, USA.
3 Department of Neurology, University of Kentucky, Lexington, KY 40536, USA.
4 Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, USA.
5 Colorado Division of Wildlife, Wildlife Research Center, Fort Collins, CO 80526, USA.
* These authors contributed equally to this work.
Present address: Department of Infectology, Scripps Research Institute, 5353 Parkside Drive, RF-2, Jupiter, FL 33458, USA.
Present address: Institute of Neuropathology, University of Zurich, Schmelzbergstrasse 12, 8091 Zurich, Switzerland.
To whom correspondence should be addressed. E-mail:
[email protected]
snip...
further into this study;
Our results show that skeletal muscle as well as CNS tissue of deer with CWD contains infectious prions. Similar analyses of skeletal muscle from BSE-affected cattle did not reveal high levels of prion infectivity (4). It will be important to assess the cellular location of PrPSc in muscle. Although PrPSc has been detected in muscles of scrapie-affected sheep (5), previous studies failed to detect PrPSc by immunohistochemical analysis of skeletal muscle from deer with natural or experimental CWD (6, 7). Because the time of disease onset is inversely proportional to prion dose (8), the longer incubation times of prions from skeletal muscle extracts compared with those from matched brain samples indicated that prion titers were lower in muscle than in the CNS, where infectivity titers are known to reach high levels. Although possible effects of CWD strains or strain mixtures on these incubation times cannot be excluded, the variable 360- to 490-day incubation times suggested a range of prion titers in skeletal muscles of CWD-affected deer. Muscle prion titers at the high end of the range produced the fastest incubation times, which were 30% longer than the incubation times of prions from the CNS of the same animal. Because all mice in each inoculation group developed disease, prion titers in muscle samples producing the longest incubation times were higher than the end point of the bioassay, defined as the infectious dose at which half the inoculated mice develop disease. Although the risk of exposure to CWD infectivity after consumption of prions in muscle is mitigated by relatively inefficient prion transmission via the oral route (9), our results show that semitendinosus/semimembranosus muscle, which is likely to be consumed by humans, is a major source of prion infectivity. Humans consuming or handling meat from CWD-infected deer are therefore at risk to prion exposure.
http://www.sciencemag.org/cgi/content/full/311/5764/1117?ijkey=28178d5c7be66ae4856e454b705f80213966c419
Journal of Virology, July 2004, p. 6792-6798, Vol. 78, No. 13
0022-538X/04/$08.00+0 DOI: 10.1128/JVI.78.13.6792-6798.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.
Prion Infection of Skeletal Muscle Cells and Papillae in the Tongue
Ellyn R. Mulcahy,1, Jason C. Bartz,1 Anthony E. Kincaid,2 and Richard A. Bessen3*
Department of Medical Microbiology and Immunology,1 Department of Physical Therapy, Creighton University, Omaha, Nebraska 68178,2 Department of Veterinary Molecular Biology, Montana State University Bozeman, Montana 597173
Received 29 September 2003/ Accepted 3 March 2004
The presence of the prion agent in skeletal muscle is thought to be due to the infection of nerve fibers located within the muscle. We report here that the pathological isoform of the prion protein, PrPSc, accumulates within skeletal muscle cells, in addition to axons, in the tongue of hamsters following intralingual and intracerebral inoculation of the HY strain of the transmissible mink encephalopathy agent. Localization of PrPSc to the neuromuscular junction suggests that this synapse is a site for prion agent spread between motor axon terminals and muscle cells. Following intracerebral inoculation, the majority of PrPSc in the tongue was found in the lamina propria, where it was associated with sensory nerve fibers in the core of the lingual papillae. PrPSc staining was also identified in the stratified squamous epithelium of the lingual mucosa. These findings indicate that prion infection of skeletal muscle cells and the epithelial layer in the tongue can be established following the spread of the prion agent from nerve terminals and/or axons that innervate the tongue. Our data suggest that ingestion of meat products containing prion-infected tongue could result in human exposure to the prion agent, while sloughing of prion-infected epithelial cells at the mucosal surface of the tongue could be a mechanism for prion agent shedding and subsequent prion transmission in animals.
http://jvi.asm.org/cgi/content/abstract/78/13/6792?ijkey=f816427f81bd333a811bc24e6a245c2728d1d875&keytype2=tf_ipsecsha
(American Journal of Pathology. 2006;168:927-935.)
© 2006 American Society for Investigative Pathology
Detection and Localization of PrPSc in the Skeletal Muscle of Patients with Variant, Iatrogenic, and Sporadic Forms of Creutzfeldt-Jakob Disease
Alexander H. Peden, Diane L. Ritchie, Mark W. Head and James W. Ironside
From the National Creutzfeldt-Jakob Disease Surveillance Unit and Division of Pathology, School of Molecular and Clinical Medicine, University of Edinburgh, Western General Hospital, Edinburgh, United Kingdom
Variant Creutzfeldt-Jakob disease (vCJD) differs from other human prion diseases in that the pathogenic prion protein PrPSc can be detected to a greater extent at extraneuronal sites throughout the body, principally within lymphoid tissues. However, a recent study using a high-sensitivity Western blotting technique revealed low levels of PrPSc in skeletal muscle from a quarter of Swiss patients with sporadic CJD (sCJD). This posed the question of whether PrPSc in muscle could also be detected in vCJD, sCJD, and iatrogenic (iCJD) patients from other populations. Therefore, we have used the same high-sensitivity Western blotting technique, in combination with paraffin-embedded tissue blotting, to screen for PrPSc in muscle tissue specimens taken at autopsy from 49 CJD patients in the United Kingdom. These techniques identified muscle PrPSc in 8 of 17 vCJD, 7 of 26 sCJD, and 2 of 5 iCJD patients. Paraffin-embedded tissue blotting analysis showed PrPSc in skeletal muscle in localized anatomical structures that had the morphological and immunohistochemical characteristics of nerve fibers. The detection of PrPSc in muscle tissue from all forms of CJD indicates the possible presence of infectivity in these tissues, suggesting important implications for assessing the potential risk of iatrogenic spread via contaminated surgical instruments.
http://ajp.amjpathol.org/cgi/content/abstract/168/3/927?ijkey=8380f2361801c6e85f54910bf6156f1ce0fe6490&keytype2=tf_ipsecsha
The 11th BSE case in Japan
PrPSc was also detected in the peripheral nerves (sciatic nerve, tibial nerve, vagus nerve). ...
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 BSE cattle may need to be reexamined.
9/13/2005
179
Page 10 of 17
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 ten specimens positive according to ELISA were subjected to WB/IHC. Seven showed positive by both WB and IHC, all exhibiting the typical electrophoretic profile of a high content of the di-glycosylated molecular form of PrPSc (1-3) and the distinctive granular deposition of PrPSc in neuronal cells and neuropil of the dorsal nucleus of vagus. An ELISA-positive specimen from a 23 month-old Holstein steer slaughtered on September 29, 2003, in Ibaraki Prefecture (Ibaraki case) was sent to the NIID for confirmation. The animal was reportedly healthy before slaughter. The OD titer in ELISA was slightly higher than the 'cut-off' level given by the manufacturer. The histology showed no spongiform changes and IHC revealed no signal of PrPSc accumulation typical for BSE. However, WB analysis of the homogenate that was prepared from the obex region and used for ELISA revealed a small amount of PrPSc with an electrophoretic profile different from that of typical BSE-associated PrPSc (1-3). The characteristics were (i) low content of the di-glycosylated molecular form of PrPSc, (ii) a faster migration of the non-glycosylated form of PrPSc on SDS-PAGE, and (iii) less resistance against PK digestion as compared with an authentic PrPSc specimen derived from an 83-month-old Holstein (Wakayama case) (Fig. 1). Table 1 summarizes the relative amounts of three distinctive glycoforms (di-, mono, non-glycosylated) of PrPSc calculated by densitometric analysis of the blot shown in Fig. 1. As 2.5 mg wet weight obex-equivalent homogenate of the Ibaraki case (Fig. 1, lane 4) gave slightly stronger band intensities of PrPSc than an 8 mg wet weight obex-equivqlent homogenate of a typical BSE-affected Wakayama case (Fig. 1, lane 2), the amount of PrPSc accumulated in the Ibaraki case was calculated to be 1/500 - 1/1000 of the Wakayama case. In the Ibaraki case, the PrPSc bands were not detectable in the homogenates of the proximal surrounding region of the obex. These findings were consistent with the low OD value in ELISA, i.e., 0.2 -0.3 for the Ibaraki case versus over 3.0 for the Wakayama case. The DNA sequence of the PrP coding region of the Ibaraki case was the same as that appearing in the database (GenBank accession number: AJ298878). More recently, we encountered another case that resembled the Ibaraki case. It was a 21-monthold
Holstein steer from Hiroshima Prefecture. WB showed typical BSE-specific PrPSc deposition though IHC did not detect positive signals of PrPSc (data not shown). Though the clinical onset of BSE is usually at around 5 years of age or later, a 20-month-old case showing the clinical signs has been reported (4). Variant forms of BSE similar to our cases, i.e., with atypical histopathological and/or biochemical phenotype, have been recently reported in Italy (5) and in France (6). Such variant BSE was not associated with mutations in the prion protein (PrP) coding region as in our case (5,6). The Ministry of Agriculture, Forestry and Fisheries of Japan (MAFF) announced a ban of feeding ruminants with meat bone meal (MBM) on September 18, 2001, and a complete ban was made on October 15 of the same year. According to the recent MAFF report, the previous seven cases of BSE in Japan were cattle born in 1995 - 1996 and possibly fed with cross-contaminated feed. However, the two cattle in this report were born after the complete ban. Whether contaminated MBM was implicated in the present cases remains to be investigated.
REFERENCES Collinge, J., Sidle, K. C. L., Meads, J., Ironside, J. and Hill, A. F. (1996): Molecular analysis of prion strain variation and the aetiology of 'new variant' CJD. Nature, 383, 685690.
Bruce, M. E., Will, R. G., Ironside, J. W., McConnell, I., Drummond, D., Suttie, A., McCardle, L., Chree, A., Hope, J., Birkett, C., Cousens, S., Fraser, H. and Bostock, C. J. (1997): Transmissions to mice indicate that 'new variant' CJD is caused by the BSE agent. Nature, 389, 498-501.
Hill, A. F., Desbruslais, M., Joiner, S., Sidle, K. C. L., Gowland, I. and Collinge, J. (1997): The same prion strain causes vCJD and BSE. Nature, 389, 448-450.
Matravers, W., Bridgeman, J. and Smith, M.-F. (ed.)(2000): The BSE Inquiry. p. 37. vol. 16. The Stationery Office Ltd., Norwich, UK.
Casalone, C., Zanusso, G., Acutis, P. L., Crescio, M. I., Corona, C., Ferrari, S., Capobianco, R., Tagliavini, F., Monaco, S. and Caramelli, M. (2003): Identification of a novel molecular and neuropathological BSE phenotype in Italy. International Conference on Prion Disease: from basic research to intervention concepts. Gasreig, Munhen, October 8-10.
Bicaba, A. G., Laplanche, J. L., Ryder, S. and Baron, T. (2003): A molecular variant of bovine spongiform encephalopatie. International Conference on Prion Disease: from basic research to intervention concepts. Gasreig, Munhen, October 8-10.
Asante, E. A., Linehan, J. M., Desbruslais, M., Joiner, S., Gowland, I., Wood, A. L., Welch, J., Hill, A. F., Lloyd, S. E., Wadsworth, J. D. F. and Collinge, J. (2002). BSE prions propagate as either variant CJD-like or sporadic CJD-like prion strains in transgenic mice expressing human prion protein. EMBO J., 21, 6358-6366.
9/13/2005
Page 12 of 17 SEE SLIDES IN PDF FILE;
http://www.nih.go.jp/JJID/56/221.pdf
http://www.fsis.usda.gov/OPPDE/Comments/03-025IFA/03-025IFA-2.pdf
Subject: Experimental BSE Infection of Non-human Primates: Efficacy of the Oral Route
Date: September 29, 2007 at 12:50 pm PST
P04.27
Experimental BSE Infection of Non-human Primates: Efficacy of the Oral Route
Holznagel, E1; Yutzy, B1; Deslys, J-P2; Lasmézas, C2; Pocchiari, M3; Ingrosso, L3;
Bierke, P4; Schulz-Schaeffer, W5; Motzkus, D6; Hunsmann, G6; Löwer, J1
1Paul-Ehrlich-Institut, Germany;
2Commissariat à l´Energie Atomique, France;
3Instituto Superiore di Sanità, Italy;
4Swedish Institute for Infectious Disease control, Sweden;
5Georg August University, Germany;
6German Primate Center, Germany
Background:
In 2001, a study was initiated in primates to assess the risk for humans
to contract BSE through contaminated food. For this purpose, BSE brain was
titrated in cynomolgus monkeys.
Aims:
The primary objective is the determination of the minimal infectious dose (MID50)
for oral exposure to BSE in a simian model, and, by in doing this, to assess the risk for
humans. Secondly, we aimed at examining the course of the disease to identify
possible biomarkers.
Methods:
Groups with six monkeys each were orally dosed with lowering amounts of
BSE brain: 16g, 5g, 0.5g, 0.05g, and 0.005g. In a second titration study,
animals were intracerebrally (i.c.) dosed (50, 5, 0.5, 0.05, and 0.005 mg).
Results:
In an ongoing study, a considerable number of high-dosed macaques already
developed simian vCJD upon oral or intracerebral exposure or are at the onset of the
clinical phase. However, there are differences in the clinical course between orally and
intracerebrally infected animals that may influence the detection of biomarkers.
Conclusions:
Simian vCJD can be easily triggered in cynomolgus monkeys on the oral
route using less than 5 g BSE brain homogenate. The difference in the incubation
period between 5 g oral and 5 mg i.c. is only 1 year (5 years versus 4 years). However,
there are rapid progressors among orally dosed monkeys that develop simian vCJD as
fast as intracerebrally inoculated animals.
The work referenced was performed in partial fulfilment of the study "BSE in primates"
supported by the EU (QLK1-2002-01096).
http://www.prion2007.com/pdf/Prion%20Book%20of%20Abstracts.pdf
look at the table and you'll see that as little as 1 mg (or 0.001 gm) caused
7% (1 of 14) of the cows to come down with BSE;
Risk of oral infection with bovine spongiform encephalopathy agent in
primates
Corinne Ida Lasmézas, Emmanuel Comoy, Stephen Hawkins, Christian Herzog,
Franck Mouthon, Timm Konold, Frédéric Auvré, Evelyne Correia, Nathalie
Lescoutra-Etchegaray, Nicole Salès, Gerald Wells, Paul Brown, Jean-Philippe
Deslys
Summary The uncertain extent of human exposure to bovine spongiform
encephalopathy (BSE)--which can lead to variant Creutzfeldt-Jakob disease
(vCJD)--is compounded by incomplete knowledge about the efficiency of oral
infection and the magnitude of any bovine-to-human biological barrier to
transmission. We therefore investigated oral transmission of BSE to
non-human primates. We gave two macaques a 5 g oral dose of brain homogenate
from a BSE-infected cow. One macaque developed vCJD-like neurological
disease 60 months after exposure, whereas the other remained free of disease
at 76 months. On the basis of these findings and data from other studies, we
made a preliminary estimate of the food exposure risk for man, which
provides additional assurance that existing public health measures can
prevent transmission of BSE to man.
snip...
BSE bovine brain inoculum
100 g 10 g 5 g 1 g 100 mg 10 mg 1 mg 0·1 mg 0·01 mg
Primate (oral route)* 1/2 (50%)
Cattle (oral route)* 10/10 (100%) 7/9 (78%) 7/10 (70%) 3/15 (20%) 1/15 (7%)
1/15 (7%)
RIII mice (ic ip route)* 17/18 (94%) 15/17 (88%) 1/14 (7%)
PrPres biochemical detection
The comparison is made on the basis of calibration of the bovine inoculum
used in our study with primates against a bovine brain inoculum with a
similar PrPres concentration that was
inoculated into mice and cattle.8 *Data are number of animals
positive/number of animals surviving at the time of clinical onset of
disease in the first positive animal (%). The accuracy of
bioassays is generally judged to be about plus or minus 1 log. ic
ip=intracerebral and intraperitoneal.
Table 1: Comparison of transmission rates in primates and cattle infected
orally with similar BSE brain inocula
Published online January 27, 2005
http://www.thelancet.com/journal/journal.isa
Research Project: Study of Atypical Bse
Location: Virus and Prion Diseases of Livestock
Project Number: 3625-32000-073-07
Project Type: Specific C/A
Start Date: Sep 15, 2004
End Date: Sep 14, 2007
Objective:
The objective of this cooperative research project with Dr. Maria Caramelli from the Italian BSE Reference Laboratory in Turin, Italy, is to conduct comparative studies with the U.S. bovine spongiform encephalopathy (BSE) isolate and the atypical BSE isolates identified in Italy. The studies will cover the following areas: 1. Evaluation of present diagnostics tools used in the U.S. for the detection of atypical BSE cases. 2. Molecular comparison of the U.S. BSE isolate and other typical BSE isolates with atypical BSE cases. 3. Studies on transmissibility and tissue distribution of atypical BSE isolates in cattle and other species.
Approach:
This project will be done as a Specific Cooperative Agreement with the Italian BSE Reference Laboratory, Istituto Zooprofilattico Sperimentale del Piemonte, in Turin, Italy. It is essential for the U.S. BSE surveillance program to analyze the effectiveness of the U.S diagnostic tools for detection of atypical cases of BSE. Molecular comparisons of the U.S. BSE isolate with atypical BSE isolates will provide further characterization of the U.S. BSE isolate. Transmission studies are already underway using brain homogenates from atypical BSE cases into mice, cattle and sheep. It will be critical to see whether the atypical BSE isolates behave similarly to typical BSE isolates in terms of transmissibility and disease pathogenesis. If transmission occurs, tissue distribution comparisons will be made between cattle infected with the atypical BSE isolate and the U.S. BSE isolate. Differences in tissue distribution could require new regulations regarding specific risk material (SRM) removal.
http://www.ars.usda.gov/research/projects/projects.htm?ACCN_NO=408490
we also know that ;
Results PrPSc was present in the brain tissue of all patients. In
addition, we found PrPSc in 10 of 28 spleen specimens and in 8 of 32
skeletal-muscle samples. Three patients had PrPSc in both spleen and
muscle specimens. Patients with extraneural PrPSc had a significantly
longer duration of disease and were more likely to have uncommon
molecular variants of sporadic Creutzfeldt–Jakob disease than were
patients without extraneural PrPSc.
and we now know that ;
Prions in Skeletal Muscles of Deer with Chronic Wasting Disease
www.sciencexpress.org / 26 January 2006 / Page 1 / 10.1126/science.1122864
we have known that prions were in skeletal muscle ;
http://europa.eu.int/comm/food/fs/sc/ssc/out254_en.pdf
http://www.bseinquiry.gov.uk/files/yb/1990/01/19009001.pdf
Medical Sciences
Prions in skeletal muscle
Patrick J. Bosque*,,, Chongsuk Ryou*, Glenn Telling*,§, David Peretz*,, Giuseppe Legname*,, Stephen J. DeArmond*,,¶, and Stanley B. Prusiner*,,,**
* Institute for Neurodegenerative Diseases and Departments of Neurology, ¶ Pathology, and Biochemistry and Biophysics, University of California, San Francisco, CA 94143
Contributed by Stanley B. Prusiner, December 28, 2001
snip...
Discussion
Our studies demonstrate that mouse skeletal muscle is intrinsically capable of propagating prions, that titers at least as high as 107 ID50 units/g can accumulate in muscle, and most surprisingly, that the efficiency of this accumulation varies markedly among groups of muscles taken from different regions of the body. Our finding of prion accumulation in skeletal muscle seems unambiguous, in that we obtained similar results with two different prion strains in wt mice and in Tg mice expressing PrPC almost exclusively in skeletal muscle. Why prions accumulate more efficiently in certain muscles than in others is not clear. However, different skeletal muscle groups demonstrate differential susceptibility to a number of disease processes, a property that presumably reflects biochemical differences in skeletal muscles of different body regions (45).
Studies in Tg mice (46) and cultured cells (43, 44) have implicated a cellular factor other than PrP, provisionally termed "protein X," that is needed for the efficient propagation of prions. Perhaps only some skeletal muscles have sufficient amounts of protein X to enable the accumulation of high titers of prions. Similarly, the inefficient accumulation of prions in hepatic tissue demonstrated by our studies is further evidence of a role for an auxiliary factor such as protein X in prion propagation.
That high prion titers may be found in skeletal muscle even if central nervous system and lymphatic tissues are carefully excluded from the muscle raises the concern that humans consuming meat from prion-infected animals are at risk for acquiring infection. However, several caveats must be considered when assessing the risk of humans developing disease from prion-tainted meat. First, the efficiency of prion accumulation in muscle may vary with either the host species or the prion strain involved. Indeed, mouse RML prions seem to have accumulated more efficiently in muscle than did hamster Sc237 prions. Second, oral transmission is inefficient compared with the i.c. inoculations used for the bioassays reported in this study. In hamsters, oral exposure is 105- to 109-fold less efficient that the i.c. route (47, 48). Finally, the species barrier must be considered. In many cases, efficient transmission of prions from one species to another requires a high degree of homology in the amino acid sequence of PrP between the two species. However, the degree to which amino acid sequence influences the efficiency of transmission depends on the strain of prion. In the case of new variant Creutzfeldt-Jakob disease (nvCJD) prions, Tg mice expressing bovine PrPC are much more susceptible to nvCJD prions, derived from human brain, than are Tg mice expressing either human or chimeric human-mouse PrPC (refs. 10 and 49; C. Korth and S.B.P., unpublished data).
Previous studies have generally reported low prion titers in muscle tissue. Some of these studies used inefficient cross-species transmissions, which might be responsible for their failure to detect prions in muscle (19). Our investigations reveal another potential explanation for this failure. Because muscle prion accumulation varies between muscle groups or perhaps between specific muscles, previous studies may have failed to sample the muscles bearing the highest prion titers. If prions accumulate in certain muscles of humans with prion disease to levels near those that we found in mice with prion disease, it should be possible to definitively diagnose all forms of CJD and related disorders by using muscle tissue for biopsy. This approach would offer significant advantages over the relatively difficult and morbid brain biopsy procedure, which is currently the only way to definitively diagnose prion disease in humans.
Whether prions accumulate in skeletal muscle of cattle with BSE, of sheep with scrapie, or of deer and elk with chronic wasting disease remains to be established. However, our findings indicate that a comprehensive and systematic effort to determine the distribution of prions in the skeletal muscle of animals with prion disease is urgently needed. The distribution of prions in muscle may vary with the animal species, perhaps even with breeds, varieties, and lines within a species as well as with the strain of prions. Such assays need to be carried out by using sensitive and quantitative techniques, such as bioassays in Tg mice and quantitative immunoassays adapted to PrPSc detection in muscle tissue.
http://www.pnas.org/
CONTINUED
