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From: TSS ()
Subject: Re: BSE June 9: USDA roundtable discussion on beef safety open to public (PRION ROUNDTABLE USA DEC., 11, 2003)
Date: May 29, 2005 at 6:08 am PST

Date: Sun, 29 May 2005 08:03:10 -0500
Reply-To: Bovine Spongiform Encephalopathy
Sender: Bovine Spongiform Encephalopathy
From: "Terry S. Singeltary Sr."
Subject: Re: BSE June 9: USDA roundtable discussion on beef safety open to
public
##################### Bovine Spongiform Encephalopathy #####################

Greetings,

Found this in some of my old files and thought it might be interesting for
some to read how these old BSE Round Table Events took place and what has
been accomplished over the years. What officials thought and said then,
compared to what they are saying now. This is about 62
pages...TSS


Completely Edited Version
PRION ROUNDTABLE


Accomplished this day, Wednesday, December 11, 2003, Denver, Colorado


Dr. Max Thornsberry: The question was asked: What brought this roundtable
about? I was fortunate enough to listen to Dr. Prusiner's lecture by
satellite at Washington State University on prion-related diseases. I was
fascinated by the information he presented and the research he had
administered. A number of livestock producers in the West this fall have
found deer and elk on their property that have tested positive for chronic
wasting disease. During a joint telephone conversation representing South
Dakota Stockgrower’s Association, Montana Stockgrower’s Association,
Missouri Stockgrower’s Association and others, the question was asked: Is
chronic wasting disease infectious to cattle? Is it possible for cattle to
get BSE (Mad Cow Disease) from deer and elk? From that series of questions
came this meeting.
I'm going to go on the program first and go over Dr. Prusiner's lecture—the
information I took down and thought was most pertinent. We want to have a
basic understanding of what the human medical field thinks in terms of
prion-relation diseases. Dr. Prusiner has won the Nobel Prize in Medicine
for making the discovery that BSE is caused by an abnormal protein, which he
calls prion Scrapie (PrPSc). For many, that's kind of a generic term for an
aberrant prion. It may not mean it's the literal Scrapie prion, so he
refers to all aberrant prions as Scrapie prions.
There are normal cellular prions, and Dr. Prusiner believes that sometimes,
in some particular animals, the normal prion is converted into a prion
Scrapie. That Scrapie prion is acquired at a certain level, and somehow
stimulates the cell to start producing more of this abnormal prion. A
conversion process takes place within the cell that converts normal cellular
prions into abnormal Scrapie prions. The cell cannot clear these abnormal
prions fast enough, resulting in the eventual death of the cell.
BSE is also called new variant Creutzfeldt-Jakob Disease (CJD) in human
patients. Aberrant metabolism and the resultant accumulation of this BSE
prion protein is what causes BSE or CJD in people.
The normal prion is soluble in a non-denaturing detergent, but prion
pre-Scrapie is not soluble. This is a very tough piece of protein. Where
the normal prion is readily digested by protease enzymes, the Scrapie prion
is radically resistant to protease digestion.
Dr. Prusiner said BSE in humans is caused by consuming tainted beef.
Apparently he is totally convinced of that. He also made a very poignant
comment about a young woman, about nineteen, who has Creutzfeldt-Jakob
Disease. They can find nothing in her history that would indicate she ever
consumed beef. She was raised as a vegetarian. However, she was a consumer
of gelatin in the form of Jello salads. He believes she picked that prion
up through gelatin, and he's convinced that the normal processes that are
used to denature protein and produce gelatin have no negative effect on the
Scrapie prion and don't destroy the infective nature of this Scrapie prion.
He also says it's possible the normal prion is converted to the Scrapie
prion. Something triggers the conversion process within the cell structure
itself. It becomes, then, what he called, an inherited prion disease.
There are some people who have, within their metabolism, the ability for
this to occur. When it does occur, it somehow gets the cell oriented to
producing more of the abnormal protein, and then it literally kills itself.
I'm sure you're familiar with a lot of these prion diseases, such as Kuru
from cannibalism. Dr. Prusiner lists six ways for prion diseases to be
transmitted from person to person:
1. Kuru—Ritualistic cannibalism of brain and nervous tissues.
2. Improperly sterilized brain electrodes (transfers the disease from person
to person). He knows of no effective way to sterilize a brain probe.
3. Cornea transplants.
4. Use of human pituitary-gland proteins (growth hormone, HCG)
5. Dura mater graphs (surgery for spinal bifida)
6. Protein X—a factor defined by molecular genetic studies that binds the
normal protein and facilitates prion pre-Scrapie formation
Great Britain has had one million cattle die of BSE and the average
incubation time was five years. Dr. Prusiner has documented that brain
extracts from cows with BSE will cause BSE-like disease if injected in the
brain of cattle, sheep, mice, pigs and mink. It can occur in a little time
as 200 to 300 days. The theory that this is a long-term incubation disease
depends on the exposure. If the exposure is into the nervous system, then
the incubation period appears to be much shorter.
There is no reliable specific test for prion diseases in a live animal that
is known or available to the scientific community at this time.
Dr. Prusiner's theory is that BSE accumulated in Great Britain because of a
major change in the manufacture of meat and bone meal. They went from a
heat-type extraction to a chemical solvent extraction. When this change was
made, they were able to solubalize and accumulate much more fat, and with
that fat goes neural tissue. As they fed cattle to cattle and more cattle
were slaughtered, it became an epidemic.
They are still seeing cases of BSE in those countries, but nothing like
they were. Several new cases this past year have occurred in much younger
animals that were not exposed to meat and bone meal and were born from
animals not fed meat and bone meal. Where are these animals getting exposed
to BSE prions? That is an unanswered question at this point.
Twenty-five percent of mice were able to develop CJD in 60 days when brain
tissue from young people with new variant CJD was injected into the brains
of susceptible mice. These prions are considered to be extremely infective.
Dr. Prusiner believes new variant CJD is an infectious disease. If we get
these prions into an environment where they can be consumed or come in
contact with people, they are extremely infectious. His theory is that
humans get new variant CJD from consuming BSE prions from cows.
He also made some comments about sheep and Scrapie. He said sheep with
Scrapie may have a few BSE prions, but when the rendering process takes
place, there is a transformation from the Scrapie prion or those prions die
off or are inactivated. The only ones that survive are the BSE prions. Dr.
Prusiner's theory is that when they were using sheep for meat and bone meal,
the Scrapie prion was inactivated, but the BSE prion was much more resistant
to inactivation, so in the processing of sheep for meat and bone meal, they
actually concentrated the prion that he believes causes BSE.
He also developed a few therapeutic approaches to treating people with BSE.
They have discovered these therapies through cell cultures. They've found
that thorazine, chlorpromazine, and acepromazine have some effect in slowing
down the progression of the disease. But the most interesting discovery was
with a synthetic quinine product, Quinacrine. In cell culture, they have
been able, literally, to slow the progression of prion diseases. They've
had some clinical improvement in CJD patients. They have to use it at such
high levels, a toxicity problem occurs.
In a minority of patients where the disease progression is slowed, some
patients actually have a measured decrease in PRP Scrapie levels.
He classified prion diseases into three classes: Infectious (BSE, chronic
wasting disease), sporadic, and genetic. He believes there are
prion-related diseases such as ALS and Parkinson's disease.
Dr. Prusiner commented a number of times that prions are infectious. This
is not an end-stage disease. He talked about the potential for spreading it
through after-birth and the licking of newborn calves and possibly
decomposing carcasses.
Prions contain protein. They are miss-formed proteins and they have no RNA
or DNA. Somehow they are able to get a cell to reproduce themselves without
the presence of genetic material. Prions also do not trigger the immune
system, so there is no way to vaccinate against this disease, in his
opinion. PRP Scrapie prions are natural at low levels in human patients, but
the cells are able to clear that abnormal protein in a normal situation.
Dr. Prusiner said we don’t know the function of a normal PRP Scrapie prion
in cells, but he believes there is a normal level of abnormal prions in
cellular tissue of human patients. We don't know what that level is or when
it reaches the point where it kills the cell. He believes there is some PRP
Scrapie being formed all the time from normal prions, but the cell has a
mechanism to clear the abnormal prion before it ever accumulates to a
dangerous level.
The question was asked following the lecture: "How did BSE start in cows?"
Dr. Prusiner believes in two theories. One is the sheep Scrapie theory
mentioned previously. He does not believe the BSE prion is the sheep Scrapie
prion. He believes they are two distinct prions, and the sheep Scrapie prion
is much easier to inactivate than the BSE prion.
He also believes in a theory of spontaneous BSE or sporadic BSE. I thought
Pasteur disproved spontaneous generation a long time ago, but evidently it's
come back in vogue because Dr. Prusiner believes, and stands by the
hypothesis that, for some reason, a cow just starts producing these abnormal
BSE prions. He believes that cow died and ended up in the meat and bone
meal and then the process started. That's the theory he outlines as his
most significant hypothesis.

Dr. Spraker: If all people and animals always have a small amount of this
abnormal prion, then why would you need a theory of spontaneous disease?
Why not say something allows the abnormal to accumulate?

Dr. Thornsberry: Dr. Prusiner made that comment. He listed several human
prion-related diseases and what the percentage was in the population. He
feels that some of the prion-related diseases are genetic and some are
spontaneous in a certain percentage of human patients. That was his
hypothesis. I was surprised at the idea of spontaneous generation being
brought up. That was the big theory before Pasteur came along—that maggots
spontaneously came out of a piece of rotting meat. But that was exactly
what he said: Spontaneous or sporadic BSE caused this disease to start to
England

Dr. Bartz: I think of it as a stochastic process in humans. Sporadic CJD,
to me, is real. You get sporadic production of an infectious agent. The
way I look at it, the reason you don't see it in cattle or in sheep is a
time factor. People come down with the clinical forms of sporadic CJD in
their fifties and sixties. Cattle and sheep don't live that long. The
theory is that as PrPC is being produced in the cell, it mis-folds into the
Sc conformation randomly. If you're unlucky enough that this happens to you
at an early stage of your life, it can progress and cause clinical signs.

Dr. Thornsberry: He believes that that occurs in everybody a little bit and
then for some reason some individuals are not able to clear that abnormal
prion?

Dr. Bartz: That I'm not sure about. I think sporadic CJD and familial
CJD—something with mutations in the prion protein of the host—you increase
the probability of this miss-folding.

Dr. Thornsberry: Here's another question—How does a prion get from the
intestinal tract to the brain? Dr. Prusiner says it takes one billion or
more prions orally to cause BSE to multiply in the lymphoid cells. From
there, it goes to the brain. It may travel up the nerves, the spinal cord,
or by the blood, the theory being that its primary means of getting to the
brain is by traveling through spinal tissue.
The question was asked "How do you disinfect medical instruments?" His
answer to that question was "Throw them away." He has found no effective
way to disinfect them from prions.
There is no reliable blood test for prion-related diseases at this time.
Conversion of PRP normal to PRP Scrapie occurs on the cell membrane with the
relationship to cholesterol-rich domains on the cell membrane. This was the
comment he made about natural conversion taking place.
"How do they multiply?" New prion pre-normal proteins are being formed and
degraded into prion pre-Scrapie. As the Scrapie increases, more of it is
somehow stimulated to be degraded into the Scrapie prion. In a situation
where the Scrapie prion is there, as normal prion is being converted into
abnormal prion, then the abnormal prion somehow triggers more abnormal prion
to be converted until it builds up enough that it kills the cell.
Dr. Prusiner made some comments about Parkinson's disease. He said a
protein being improperly handled in the substantia nigra cells is what
causes Parkinson's disease because it causes a decrease in dopamine
production. His theory is that Parkinson’s disease is a prion-related
problem, that Parkinson's disease is the direct result of those cells being
killed because of prion accumulation that he believes is a familial
transfer.
"Can it be transmitted by blood transfusions?" His answer is that We don't
know. We believe that ALS, or Lou Gehrig's disease, is a prion-related
disease.
The last question that was asked of Dr. Prusiner was a simple question.
The person said, "Dr. Prusiner, do you eat meat?" He answered immediately
"Not in Europe." He was asked to elaborate, and he said these prion-related
diseases are all over Europe. He said wherever you go, if you eat meat, you
have the potential of picking them up. He believes the prions are in the
muscle tissue. He does not believe they're just in the spinal column.
My son is an M.D. and they are getting a lot of information on these
prion-related diseases and most of it is coming from Dr. Prusiner. The
medical community has an understanding about prion-related diseases that's
much different from that of the veterinary community, from what I can
determine

At this point, I'd like each of you to introduce yourself and give a short
summary of what you do.


Dr. Linda Detwiler: I formally worked for the USDA in the area of
Transmissible Spongioform Encepthalopathy diseases. I work currently as an
independent consultant to the food service industry, particularly, McDonalds
Corporation, ….rest of comments inaudible…….

Dr. Susan Keller: After graduating from veterinary school, I had practiced
for ~10 years as a predominately large animal veterinarian in North Dakota
before becoming the Deputy State Veterinarian with the North Dakota Board of
Animal Health and the North Dakota Department of Agriculture in 1997.
Current responsibilities include serving as the Designated Scrapie
Epidemiologist for ND and have been certified at Plumb Island as a Foreign
Animal Disease Diagnostician.

Dr. Jason Bartz: I received a PhD from the University of Wisconsin in 1998
and I worked with Drs. Judd Akin and Dick Marsh. I studied mechanisms of
inter-species transmission of prion diseases. I was a post-doctoral fellow
with Richard Besson. I studied pathogenesis, asking questions about how the
agent gets from the periphery to the central nervous system. In September,
I started a faculty position at Creighton University and I'm setting up a
lab to continue study of prion pathogenesis.

Dr. Terry Spraker: I graduated from vet school in 1972. I have a PhD in
pathology and I'm board-certified in veterinary pathology. I saw CWD as a
graduate student and didn't know what it was at that time. I've been
working with it since then, looking at pathogenesis of the disease.

Dr. Thornsberry: I feel honored to have you all here and I hope we'll have
an interesting dialogue. We want to make this information available to the
cattle industry and to veterinarians, so anything you think is pertinent,
please tell us.


TRANSMISSION OF BSE

Dr. Jason Bartz: Dr. Thornsberry asked me to talk about what is the risk of
transmission of chronic wasting disease in cattle? Unfortunately, I cannot
directly answer that. What I'm going to talk about is some of the basis
science behind it and what we know about interspecies transmission. You can
have five factors affecting interspecies transmission in the natural
setting. The first is the question "is infectivity shed into the
environment?" Can PrPSc survive in the environment? Can PrPSc reach a new
host species? Is the agent transferred between deer and cattle? I'm going
to spend most of the time on these last two points: "how does the agent get
from the periphery to the central nervous system where it causes disease?"
Finally, I'll talk about the species barrier effect. "Can PRPSC from one
species convert to the PRPSC from the host into a new PRPSC molecule?"
What are prion diseases? All prion diseases are transmissible. They're
neurodegenerative diseases and they have long incubation periods of months
to decades. They're characterized by the spongiform degeneration.
Clinically, they have motor and/or behavioral/cognitive deficits. There's
no effective therapeutic treatment. They're all inevitably fatal. There's
no adaptive immune response and there are no antibodies produced.
The animal prion diseases include Scrapie, found in sheep and goats. It's
been known in the United Kingdom for over 200 years. It was first identified
in the US in 1947. It has a worldwide occurrence with the exception of New
Zealand and Australia
Transmissible mink encephalopathy is a prion disease of ranch-raised mink.
It was first identified in the 1940s. There have been three major
occurrences in the United States. The latest outbreak was in Wisconsin in
1985. It's a rare disease.
Bovine spongiform encephalopathy was first identified in the UK in 1986.
Clinically it's characterized by an aggressive behavior and it's named mad
cow disease. It was appreciated that it was a food-borne disease, so in
1988, a bovine feed ban was instituted. Amidst concerns about the
transmission of BSE to humans, a specified risk materials ban was put in
place in 1990. This banned tissues such as central nervous system tissues
and spleen, which harbor high level of infectivity in cattle, from going
into the human food chain. In 1996, a variant Creutzfeldt-Jakob disease was
identified. It's distinct from the classical forms of CJD. First of all,
the age of onset of variant CJD is much younger. The first few cases of
variant CJD were people in their teens and twenties. Normally this disease
is in people in their fifties and sixties.
The neuropathology is very distinct in variant CJD compared to other forms
of CJD. Stains reveal distinctive amyloid plaques that are not observed in
other forms of CJD. In the original study that identified variant CJD, an
epidemiological study was performed and the only common risk factor that was
identified was the emergence of BSE in the UK at that time. This suggested
that BSE was able to infect people, resulting in variant CJD.
Two studies support this hypothesis. The first is experimental
transmission of BSE to macaques. The neuropathology of BSE in infected
macaques is indistinguishable from that of variant CJD. The second series
of experiments uses a technique called lesion profile. Lesion profiling is
a technique where you can distinguish distinct prion strains by passage in
mice, by quantifying their neuropathological features. In this series of
experiments, mice were infected with either BSE, variant CJD, or sporadic
CJD. The mice became clinically ill. Their neuropathological features were
quantified and represented as lesion profiles. Importantly, the lesion
profile of mice infected with BSE is the same as that of mice infected with
variant CJD and it's distinct from that of mice infected with sporadic CJD.
This indicates that the same prion strain that causes BSE also causes
variant CJD and is consistent with the hypothesis that BSE was able to
infect humans, resulting in variant CJD.
I'd like to speak a little bit about chronic wasting disease. It was first
identified in 1967 in Ft. Collins, Colorado. It has an expanding
geographical range. It's been found in several states in captive herds and
has also been found in wild deer and elk. The clinical signs include
excessive salivation, reduced fear of humans, and distribution of
infectivity in the host. It seems to differ in deer infected with chronic
wasting disease, PRPSC seems more predominant in the secondary
lymphoreticular tissues compared to elk.
There have been quite a few experimental transmission studies of CWD. It's
been successfully transmitted by the IC route in goats, mink, ferrets,
cattle, and squirrel monkeys.

Physical Properties and Etiological Agents
This agent is highly resistant to UV radiation. UV radiation historically
is used to inactivate agents, and a mechanism it uses is to destroy the
nucleic acid of the infectious agent. However, prion diseases are very
resistant to UV inactivation. One of the practical aspects of this is that
for persistence of an agent in an environment, sunlight is a very good means
of handling it by destroying nucleic acid genomes.
The prion agent is very stable in acid and alkali. Little infectivity is
lost between ranges of pH 2.5 and 10. However, you can lose infectivity in
protein denaturance or chemotropic agents such a phenol or high
concentrations of urea. RNAase and DNAase do not have any elicitable effect
on infectivity.
Treatments that destroy protein and nucleic acids do not destroy
infectivity. This has ramifications in what the etiological agent really is.
Another physical property is that it can survive in the environment. This
is a paper by Brown in 1991 where he buried the prion in flower pots in his
garden for three years. When he dug up the flower pots, there was still
infectivity detected in the soil but not in the sub-soil. The two points we
want from this paper are that (1) the agent can survive in the soil for
three years and (2) based on this study, it does not seem to be leaching
out.
In another paper from Brown in 2000, he took either fresh brain or formalin
fixed brain and treated it at different temperatures ranging from 150
degrees Celsius to 1000 degrees Celsius. When he treated brain material at
600 degrees Celsius, the brain material actually caught on fire and all that
was left was black ash. When he took this material and inoculated it back
into recipient animals by the IC route, these animals came down with
disease, indicating how resistant this agent is to different environmental
and physical treatments.


What Do We Know About the Agent?

When you enrich the amyloid deposits or amyloid plaques that occur in the
CNS of prion-infected animals and examine them by electronmicroscopy, you
see these plaques are comprised of linear non-branching amyloid fibrils.
When you do biochemical analysis of this amyloid fibrils, the main component
of the amyloid fibrils is the prion protein. When you enrich the plaques
from the brain and the fibrils from the plaques and the protein from the
fibrils, you enrich for infectivity. Subsequent studies have shown that the
prion protein is encoded by the host, and this is important because it
eliminates the possibility that the prion protein is a viral by-product.
This suggests the prion agent is comprised solely of protein. The idea for
a self-replicating protein is not new. It was first proposed in 1967 by a
mathematician name Griffith. In 1982, following the identification of the
prion protein, Stan Prusiner developed the prion hypothesis that states that
the etiological agent for these diseases is comprised solely of the prion
protein.
The prion protein exists in two forms. It's encoded by the host. The PRPC
is for cellular and the PRPSC is for Scrapie. PRPC has a high ________
content. It's sensitive to proteinase digestion. It's soluble in
detergents. It's not associated with infectivity. The function of PRPC
right now is not known.
PRPSC is post-translationally-derived from PRPC. It has a high beta sheet
content and this is consistent with PRPSC being an amyloid protein. It has a
core that is highly resistant to protease digestion. PRPSC is a soluble in
detergent and it is thought to be the sole component of the infectious
agent, although I should point out this has not been formally proven.
How does a protein replicate? I think replication is a poor word for what
prions do. "Conversion" is a much better descriptive word. One mechanism
that's been proposed is nucleation dependent polymerization where in the
infectious form of the disease, where the PRPSC of aggregate is being
inoculated into the host, it encounters a PRPC molecule from the host, and
it is incorporated into the growing amyloid fibril. Based on this model,
PRPC should be very limiting in this conversion step. This has been borne
out by transmission studies wherein normal mice, if you IC inoculate, that
incubation period is about 130 days. If you take out PRPC, this is in
transgenic mice that do not express PRPC, these are resistant to infection.
Then if you make transgenic animals that over-express PRPC, these animals
have a much shorter incubation period.
This is a paper showing that RNA is enhancing PRPC to PRPSC. What we know
where the conversion occurs in the cell, there's not a good likelihood of
RNA being located there. I don't know if RNA has specific properties that
aid in conversion that another molecule in the cell may have.

Strain Diversity in Prions
Historically, prion strains have been characterized by differences in
incubation period, and most importantly, differences in neuropathological
features in the central nervous system. Sometimes they even have different
clinical signs. One of the main arguments against the prion hypothesis has
been "how do you have strain diversity in the absence of the nucleic acid
genome, since, according to biological dogma, the differences in the genome
of an infectious agent result in strain differences? It's recently becoming
increasingly clear, that the conformation of PRPSC can encode for strain
diversity. Different strains have different properties such as
susceptibility to protease digestion, migration, polypeptide migration on
western blot analysis. FTIR analysis has been used to demonstrate that the
hyper and drowsy strains of TME have different ratios of alpha and beta
sheet structure.
Investigators have also used a technique called glycoform ratio where they
quantify the three different glycoforms of PRPSC and come up with a distinct
ratio that correlates with strain properties as well. Based on these and
many other studies, it seems that the conformation of PRPSC is actually the
molecule that encodes for strain diversity in prion diseases. So here's a
mechanism to try to explain how an agent in the absence of nucleic acid
genome can have strain diversity.
To summarize the agent, there's no nucleic acid genome. It seems to be
comprised solely of PRPSC. It's very resistant to physical degradation. The
strains of prions are encoded by the conformation of PRPSC.

Pathogenesis
How does the agent get from the periphery to the central nervous system
where is causes disease? I'll talk about oral exposure, since there's
experimental and epidemiological evidence that this likely occurs in kuru
through the practice of ritualistic cannibalism, and also in TME and BSE
through contaminated foodstuffs. People have speculated it happens in
variant CJD through the consumption of BSE-contaminated food, and also in
chronic wasting disease and Scrapie in sheep and goats.
Infectivity, or PRPSC, is first detected in secondary lymphoreticular
tissues, such as spleen and lymph nodes. Within spleen and lymph nodes,
PRPSC accumulates in follicular dendritic cells. One the agent replicates
in secondary LRS tissues, it's next detected in the central nervous system.
There is rapidly replicates in the CNS. It's thought this replication kills
neurons, leading to the onset of clinical signs and eventual death of the
host.
In lymphoreticular system tissues, PRPSC is mainly detected in follicular
dendritic cells, but also to a limited extent in tingible body macrophages.
This and other studies have fairly clearly demonstrated that FDCs can
replicate prions, and right now it's unclear if the tingible body
macrophages are replicating agents or simply just phagocytosing PrPSc
Two pathways of neuro-invasion have been identified. One utilizes
sympathetic internervation. This was an experiment performed by Kimberlin
in 1989. This was following intra-gastric infection of rodents with prions.
At selected time points, tissues were collected and inoculated back into
recipient mice to assay for infectivity. The first place that infectivity
was detected was in secondary LRS, tissues, such as Peyer's patches and
lymph nodes. The first place in the central nervous system that infectivity
was detected was in the thoracic spinal cord, and once it reached the
thoracic spinal cord, it could travel caudally and cranially, eventually
reaching the brain and killing the host. From this and numerous other
studies, it seems that infectivity can spread via the enteric and
sympathetic nervous system to the thoracic spinal cord and then eventually
reach the brain.
A second route of neuro-invasion has been identified that utilizes
para-sympathetic internervation. From experiments in hamsters, it seems
that following oral infection, the agent can be transported directly to the
brain stem via the vagal nerve to the DMNV.
To summarize these two routes of infection, once the agent gets of the GI
tract, it can go up sympathetic internervation to the thoracic spinal cord
and travel to the brain, or it can get into parasympathetic internervation
and be directly transmitted to the brain stem.
This part of the pathway has been fairly well worked out. So what's
happening in the gut to get it into the nerve? This is not so well worked
out. One study looks at transporting it across the gut lumen into the host.
It seems that M cells are able to transport PRPSC across the lumen. Once it
gets across the lumen, it's thought it associated with follicular dendritic
cells, replicates, and then can neuro-invade.
So how does the agent get from the follicular dendritic cell into the
nerve? That's another black box. Recently there's been a paper where they
use transgenic animals, and in these transgenic animals, the distance
between the follicular dendritic cells and the sympathetic internervation
has been reduced. When you reduce the distance between the nerves and the
FDCs, you reduce the incubation period. This suggests there's a direct
transfer between infectivity from FDCs and to the nerves.
Some of the work I've been involved in is figuring out alternative routes
following oral exposure. Once the agent gets into the gut, it neuro-invades
via sympathetic or parasympathetic internervation. I was interested in
alternative routes along the alimentary tract. A tissue that is highly
internervated and also has associated LRS tissue is the tongue. But as you
know, the tongue has an epithelial layer that prevents infectious agents
from crossing it. The epithelial lay can become damaged. Infections of the
oral cavity can damage the epithelial layer. Microbial agents such as
thrush, which are common in infants but also can occur in many compromised
adults can degrade the epithelial layer of the oral cavity.
More common than tongue infections is probably lesions in the oral cavity.
In animals, especially ruminant species, cuts in the oral cavity are quite
common. We wanted to test the hypothesis that if the agent can get in the
tongue, it can cause disease. To do that, we inoculated hamsters in the
lingual muscles of the tongue with a hamster-adaptive strain of TME called
hyper-TME. We found that it can cause disease and the incubation period is
79 days. To put this into context, if you inoculate animals directly into
the brain, the incubation period is 59 days. If you inoculate in the
peripheral nervous system, in the sciatic nerve, it's slightly longer—68
days. When you go extra-neurally, the incubation period is much longer.
Intraperitoneal is 101 days; IV is 118; IM is 142. When you take the same
dose of agent and dry it on a food pellet and feed it to an animal, this is
a long incubation period and very inefficient. Only one out of six animals
comes down with clinical signs.
A short incubation period does not necessarily mean an efficient
establishment of infection. To test for efficiency, we did the
dose-response curve. In this experiment, we took 10-fold serial-dilutions of
agent and inoculated them into the lingual muscles of the tongue. What we
found was that, as you dilute out the agent, you extend the incubation
period. You get about 107 dilution and only one of the five animals comes
down with this very long incubation period. From this we can calculate the
titer. The titer by this route is 108.4 LD-50s per gram of inoculum. When
we do the same sort of dose-response experiment, this time directly
inoculating into the brain, the relative titer is 109.8 LD-50s per gram of
inoculum and you can see that inter-tongue infection is only 10100-fold less
efficient than a direct inoculation to the brain.
When we do this same experiment drying the inoculum on a food pellet and
feeding it, it's a very inefficient route of infection, 103.4 LD-50s per
gram, and this indicates that inter-tongue infection is 100,000-fold more
efficient at causing disease than ingestion of the agent.
Next we wanted to ask the question, "Can lesions on the surface of the
tongue increase the susceptibility of the host to prion infection?" Here we
had four experimental groups. Our positive control was direct inoculation
of the tongue, and as expected they came down with a short incubation
period, 182 days. All the animals got the disease.
Our second control group was per os. Here, three of the 15 animals came
down with the clinical signs at 185 days. For our third group, we placed
inoculum on the surface of an unlesioned tongue. In these animals, five of
the 15 developed clinical signs with an incubation period of 184 days,
similar to the per os. In our experimental group, a small lesion was placed
on the surface of the tongue. The same dose of inoculum was placed on that
lesion, and in this group, all fifteen animals developed clinical signs with
an incubation period of 161 days. Interestingly, these first five animals
had an incubation period similar to direct inoculation of the tongue. This
experiment demonstrates that a lesion on the surface of the tongue can
increase the susceptibility of a host to prion infection.
To summarize the transmission data, inoculation into the tongue is
100,000-fold more effective than ingestion, at least in hamsters. Lesions
on the surface of the tongue increase the susceptibility of the host to TME
infection.

Routes of Efficiency
Why is this route so efficient? If we go back to our general schematic of
prion replication, in peripheral routes, prions replicate in secondary LRS
tissues prior to neuro-invasion. We set up a temporal study. Following
tongue infection, we collected spleen and the sub-mandibular lymph node (the
lymph node that drains the oral cavity, including the tongue). In the
spleen, we were unable to detect PRPSC by western blot out to 10 weeks
post-infection. This was just prior to the onset of clinical signs. As a
positive control, we took a spleen from an animal that was intra-cerebrally
inoculated and we could detect PRPSC in the spleen.
In the sub-mandibular lymph node, we were able to detect PRPSC at one week
post-infection. It increased in abundance to three weeks post-infection and
then plateaued out at five weeks post-infection. Based on this data, we
would think the sympathetic route was involved in transport of the agent to
the central nervous system. Based on this model, following inter-tongue
infection, the agent drains to the sub-mandibular lymph node, replicates in
this lymph node, travels via sympathetic internervation through the superior
cervical ganglia, down the sympathetic chain. It should first be detected
in the central nervous system in the thoracic spinal cord.
To test this, we set up a temporal study following inter-tongue infection
and we collected the spinal cord and did western blot analysis. At seven
weeks post-infection, we could not detect PRPSC in the cord. At eight weeks
post-infection, we could detect PRPSC in the spinal cord. We detected it
not only on the thoracic segments but also in the cervical segments. At
nine and 10 weeks post-infection, PRPSC levels accumulate. This data was
inconclusive in that not only was it in the thoracic, but in the cervical.
Next we asked the question, "Where in the brain or brain stem do you first
see PRPSC following IT infection?" This time we used immunohisto-chemistry.
We found that the first place PRPSC is identified is in the hypoglossal
nucleus. By six weeks post-infection, PRPSC staining was wide-spread
throughout the hypoglossal nucleus. The significance of this staining is
that the hypoglossal nucleus contains only motor neurons, and the only
function of the motor neurons in the hypoglossal nucleus is to internervate
the tongue.

Dr. Detwiler: Do you say that the prion goes from the tongue to the
sub-mandibular lymph node through nerves or through the lymphatic flow?

Dr. Bartz: We think it gets to the lymph node just via the lymphatic flow.
There's no neural connection between the lymph node and the tongue.

Dr. Thornsberry: So when the lymphatic flow gets to the lymph node, then it
dumps into the blood.

Dr. Bartz: Right. But why do we see PRPSC in the sub-mandibular lymph node
at one week post-infection, but by the time clinical disease occurs, we
can't detect it in the spleen? If it's in the blood, it should seed all the
lymphatics in the host, but it doesn't

Dr. Spraker: Deer lymph nodes are 10 times better than spleen.

Dr. Detwiler: In sheep with sheep Scrapie, on this widespread distribution
from multiple tissues at multiple sites, within the incubation period, how
would you explain that from the spread of either the sympathetic or the
para-sympathetic without some kind of circulatory process?

Dr. Bartz: It could get in the blood and it could be neuro-invading by 10
different routes. We don't know. But this is the main route, so the first
place we see it is in the hypoglossal nucleus. It's the only place we see
it in the brain until four weeks post-infection.
This mapping study is consistent with previous reports. When people use
trans-neural viral tracers such as rabies and they inject the hypoglossal
nerve, they find that it goes to the hypoglossal nucleus, and then the
second order neurons that project afferents onto the hypoglossal nucleus in
this reticular formation and all the different structures we identify PRPSC,
so it seems to be transynaptically transported to second-order neurons, the
same as other trans-neural viral tracers.
To summarize inter-tongue, as we think of inter-tongue infection, the agent
is getting into the hypoglossal nerve and is being retrogradily transported
at the hypoglossal nerve to the hypoglossal nucleus. It can replicate in
the hypoglossal nucleus and then be transynaptically transported to
second-order neurons, and eventually we think that PRPSC gets to the
clinical target areas, resulting in clinical signs and death of the host.

Replication and Neuro-Invasion
The hypoglossal nucleus was not the first place we identified PRPSC in the
host. It was in the sub-mandibular lymph node. We wanted to ask the
question, is replication in the LRS system required for neuro-invasion
following inter-tongue infections? To investigate this, we used mMT mice,
which are transgenic mice that has a disruption in the immunoglobulin chain
and they lack B lymphocytes. Since they lack B lymphocytes, which secrete
factors that support FDC maintenance, they do not have follicular dendritic
cells.
Previous studies using these in prion diseases have shown that they're
susceptible to IC infection, and this means that once the agent gets to the
brain, it can replicate and cause disease. But following a IP infection,
they don't come down with disease. If you look in secondary LRS tissues,
there's no detectable infectivity or PRPSC.
Using these mice following inter-tongue infection, if the animals come
down, that would mean that replication in the secondary LRS tissues is not
required for neuro-invasion and to set up this experiment, we had two
different sets of mice. We had the mMT mice, which cannot replicate agent
in the secondary LRS tissues, and we used C-57 black six mice, which can
replicate prions in secondary LRS tissues. We inoculated them directly into
the brain and had an incubation period of 160 days. We did an IP
inoculation. Wild types come down at 216 days. The mMT’s are resistant.
All of this is consistent with previously-published studies. We did an IT
inoculation into wild type mice with 213 days incubation. All the mice came
down. We did an IT inoculation with the mMT mice and got a similar
incubation period—217 days with five of the six mice developing clinical
signs. We did western blot analysis on secondary LRS tissue in all these
mice, and consistent with what other people found, we were unable to detect
PRPSC in the spleen or sub-mandibular lymph node. This data is consistent
with the hypothesis that replication in secondary LRS tissues is not
required for neuro-invasion following inter-tongue infection.
To summarize the pathogenesis, in general, following a peripheral
infection, you get transport and replication in secondary LRS tissues. Once
it replicates in these tissues, neuro-invasion occurs and you get a rapid
accumulation of PRPSC in the brain. So far there have been three routes of
neuro-invasion identified following oral infection, two of which originate
in the gut using either sympathetic or parasympathetic internervation. The
third are the studies with motor neurons—the oral cavity directly
transporting agent to the brain.

Question: You said that's the normal progression of infection—that it
normally goes into those tissues and then makes its way to the nervous
system. But you've proven that's not required?

Dr. Bartz: I think the LRS is an amplifier of the agent in the periphery.
You probably need a certain dose. You either need the agent to be in
proximity to the nerve or replicate to a high enough dose it can get there.
You can probably overcome this with a high enough dose, but if you give a
low dose, you probably need replication in the host. So I think the LRS is
a spot in the host where the agent can replicate in the periphery to
facilitate neuro-invasion. The LRS tissues that prions replicate in—as far
as I know, there's no deficit in these tissues.

Comment: With deer, the hypoglossal nucleus is one of the latter nuclei to
be infected. In deer, 99% of the lymphoid tissue will be infected in early
cases, even before the brain is infected. On the opposite side, there's a
percentage of elk infected that do not have PRP in lymph nodes. So an
alternate route of infection has been postulated.

Dr. Bartz: I'm not making any sort of assertion that this happens in
natural cases. The only reason I think it's interesting to discuss is
because it's so efficient.

Question: Why couldn’t you have multiple routes?

Dr. Bartz: You could. If it occurs in one out of every thousand cases,
would we find it? Maybe, maybe not.

Question: If you look at lymphoid tissue only for diagnosing CWD, you're
going to miss a few. If there are several different route of infection, one
of them could be straight to the nerve in heavy doses.

Dr. Bartz: The last think I want to touch upon is interspecies
transmission. What are the outcomes of inter-species transmission? First,
it cannot convert the host PRPC. There's no infection, no clinical disease.
End of story. The second thing that can happen is the agent can convert the
host PRPC and you get clinical disease. Or the agent can convert host PRPC
and there's no clinical disease, and this is persistence.
I'm going to start with the occurrence of clinical disease and break this
up into two parts. What is the species barrier effect? An example is if
you take mink that have transmissible mink encephalopathy and do an IC
inoculation into the same host species, the incubation period is four
months. But if you go into a new host species, this time the ferret, and do
an IC inoculation, you get a huge extension of the incubation period of 28
to 30 months. The mink and ferret are both mustalids and differ by only two
amino acids.
When you keep passaging the TME in the same host species—the ferret,
eventually you see a shortening of the incubation period and eventual
stabilization of the agent for the new host species. I break down the
species barrier effect into two components. The first is the inter-species
transmission and the second is adaptation of the agent for the new host
species.
One way to look at the species barrier effect is a technique called
cell-free conversion. This technique was developed in Byron Caughey's lab
at NIH. It takes PRPC that's radio-labeled and growing tissue culture. You
purify it and incubate it with brain-derived PRPSC, and when you co-incubate
these two molecules, if the PRPSC can convert the radio-labeled PRPC to a PK
resistant form, when you put this onto a piece of film, the only thing that
should be radioactive is the converted radioactive PRPSC, which is
proteinase-K sensitive to a PK-resistant form.
We know that PRPC is resistant to proteinase-K digestion. When you treat
PRPSC with PK, it all goes away. If you incubate the radio-labeled mouse
PRPC with mouse PRPSC, you'll get mouse PRPSC converting the mouse PRPC to a
PK-resistant form. What's nice about this system is that you can use it to
start to ask questions. What is important between the host PRPC and the
agent PRPSC as far as inter-species transmission? This example uses hamster
PRPSC. We know that if you inoculate hamster PRPSC into mice, there are no
clinical signs. If you take hamster PRPSC, incubate it with the
radio-labeled mouse PRPC, and treat it with proteinase-K, consistent with
the biological properties, there's no conversion.
Now you can use this methodology and map out areas of PRPC that are
important for inter-species transmission. Hamster PRPSC cannot convert.
When you start to use these different chimeras, you can use it to start to
map out regions of the molecule that are important for interspecies
transmission.
You can also use it to try to predict species barriers where you don't know
where transmission data is. This is a paper from Raymond in 2000. Here you
can take PRPSC from sheep Scrapie with a certain genotype and incubate it
with radio-labeled PRPC of sheep with the same genotype, and that's said to
be 100% conversion because we know it's the same PRPSC going by
intra-species transmission. You can start doing modifications where you use
different genotypes of sheep PRPC, and you get a reduction in conversion,
and then you can go to different species. Here they looked at mule deer,
white tail deer or elk. You get a reduction in conversion.
You can also look at human PRPC, and there's a very low conversion. This
low conversion is thought to be consistent, at least in sheep and humans.
Epidemiologically, there's no evidence of sheep Scrapie transmitting to
humans. You can also use this to assess the transmission of the white-tail
deer PRPSC again using PRPC from a mule deer or a white tail deer of the
same genotype. You can start modifying the genotypes of either deer or elk
PRPC, and you get either an increase or decrease in conversion. You can ask
questions about converting bovine PRPC and human PRPC. You get these
much-reduced abilities to convert in this assay.
This reduction in conversion, then, one would predict would correlate with
the transmission studies if they were done.

Inter-Species Transmission
The host genotype is important in whether the host is susceptible or not.
I want to talk about the subsequent transmissions in the same host species.
Why are you getting a reduction in the incubation period and adaptation or
the intra-species transmission? When you have PRPSC from one host species
interacting with PRPC from another, you get conversion to PRPSC. But I
don't think you always get just one prion strain. On this inter-species
transmission, you can get multiple prion strains. Data to support this
hypothesis comes from some work I did using transmissible mink
encephalopathy where we took biologically-cloned transmissible mink
encephalopathy agent and transmitted it into hamsters. The bottom line is
when we took the single strain and put it into hamsters, we were able to get
the hyper and drowsy strains out. On first interspecies transmission,
incubation periods are very long and there's quite a range—219 days to 501
days. They ranged anywhere from 130 days out to over 600 days.
In passage line B, 219-day incubation period, PRPSC was similar to that of
hyper TME. The second passage has a reduction to 67 _____________

[tape change]

On western blots, we can tell hyper and drowsy apart. Otherwise, you have
to do lesion profiling. So this is a nice system for trying to work this
out.
In this passage line A, this first inter-species transmission, PRPSC is
like drowsy TME. Drowsy is clinically characterized by lethargy. Hyper is
clinically characterized by hyper-excitability ataxia. When you passage
this, if you passage it at a high titer, now you have mixture clinical signs
but you also have PRPSC that looks like a mixture of both hyper and drowsy.
By fourth passage, you get the emergence of hyper. If you passage this
animal at a low titer, you can get the emergence of drowsy. What we think
is going on here is that inter-species transmission of the single TME strain
in hamsters produces at least the hyper and drowsy strains, and we think,
depending on the ratio of the strains produced in this first inter-species
transmission, will result in either the rapid emergence or a longer
emergence of the prion disease.

Dr. Detwiler: How would you explain that biochemically?

Dr. Bartz: When PRPC is converted to PRPSC, it's misfolded. There have to
be many different stable energy states for the misfolded protein. I would
hypothesize that mink PRPSC, when it interacts with hamster PRPC, it can
fold into several different stable PRPSC molecules. So initially you get
the mink interacting with hamster, and then you get a strain produced. I
think early on in those first few rounds of replication, whatever strain is
produced is probably going to be the predominant one because it has a jump
start on the rest of them. On this really complicated western blot, we are
mixing hyper and drowsy at known ratios, and basically we can mimic these
effects. So it really is the ratio of hyper/drowsy produced by interspecies
transmission that's causing this sort of effect.
To summarize inter-species transmission, we have PRPSC interacting with the
new host PRPC molecule to change it into PRPSC. We think that, in certain
instances, multiple strains can be produced. Intra-species transmission
results in competition between these strains and eventual emergence of a
predominant strain. We think the initial ratio of strains is important and
affects this whole passage history. Probably the replication properties of
strains is important. We think that drowsy is the predominant strain
produced, but hyper replicates so much faster, it has an advantage.
One really important thing I want to point out here is that strain
properties can change upon inter-species transmission. Chronic wasting
disease doesn't cause disease when you passage it in a hamster, but if you
passage CWD into ferrets, and then take that ferret passage tissue, it can
cause disease in hamsters. So inter-species transmission can expand the
host range. Also, with the hyper and drowsy, the more hamster passages you
do, if you back-passage the inoculum into mink, hyper loses pathogenicity
for mink quite quickly, where drowsy retains pathogenicity for mink.
The important point I want to make is that, when you're assessing
inter-species transmission and you do a transmission study and it's
negative, you have to be careful in saying it's negative for the strains you
looked at. With this example, it's clear you could take hyper TME, inoculate
mink, and they don't come down with the disease, so you might assume hamster
prions don't cause disease in mink. That strain doesn't. You have to be
careful assessing negative transmission results based on what's known about
the strain properties.
The last thing I want to talk about is persistence. This would be the case
where PRPSC interacts with the host PRPC and you get really slow
replication. The replication agent is so slow that the animal dies of old
age before clinical signs can occur.
This study is from Rick Race at NIH, transmitting hamster PRPSC into mice.
He collected animals post-infection out to 782 days. None of these animals
had clinical signs of prion disease, which is consistent with everything we
knew about this species barrier. But when he went back and looked for PRP
residue in these animals, he couldn't detect hamster PRP residue, but in a
few of these animals with very long times post-infection, he could detect
mouse PRP residue.
When he did the second passage, into either hamsters or mice, clinical
signs appeared in the second passage. The point is that first inter-species
transmission may not cause clinical signs, but you still can get replication
to agent that subsequently, when you passage it into the same host species,
results in clinical signs of the disease.
In the cell-free conversion studies, hamster PRPSC could not convert mouse
PRPSC. Every sort of assay has limitations. The cell-free conversion said
it couldn’t replicate. It could, but it was so slow and so long that the
assay could not detect them.
I think persistence is very important. If you have inter-species
transmission occurring and it doesn't cause clinical disease, and if you
take the tissue and keep feeding it to that same host species, you’re going
to get amplification and potentially emergence of the disease.
Is PRPSC shed in the environment? I have no idea. Terry can talk about
that. Does PRPSC survive in the environment? The studies on deer PRPSC
have not been done, but if deer PRPSC behaves like any other PRPSC, yes it
can survive in the environment. Can PRPSC reach a new host species? I
don't know. If they share common pastures, it's a possibility. Can PRPSC
get to the central nervous system? Clearly, cattle are susceptible to oral
infection, so that's yes. Can deer PRPSC convert cattle PRPC to the host
PRPSC? Self-reconversion experiments would say yes, but very inefficiently.
But really, the gold standard is the transmission studies, and there are two
of these ongoing right now. One is at the USDA at Ames, and this is
intra-cerebral inoculation. They are susceptible to IC infections. This
means that once the agent reaches the brain, it can cause disease, but
obviously in the field, that's not the natural route. Beth Williams is
doing some oral infection studies, but I'm not sure of the status of those.

Dr. Thornsberry: So what you’re saying is that, inter-cerebrally, we can
get CWD/PRPSC conversion, but that has not occurred, to anyone's knowledge,
in the natural route.

Dr. Bartz: Right. IC inoculation is used because it has a short incubation
period. It only tells us that replication can occur once the agent reaches
the brain.

Dr. Thornsberry: Let's hypothesize that I had some cattle on the eastern
slope and they were in the same pasture with elk with CWD. If a cow had
been exposed to the PRP Scrapie and it did develop disease four years later,
would that look like BSE? Would there be a way to determine if it came from
CWD?

Dr. Bartz: The IC studies in cattle indicate it does not look like BSE. The
clinical signs of the IC/CWD cattle are more like downer cattle, and not
aggressive. As far as finding the source of a bovine TSE, the gold standard
is the lesion profile study where you take cattle tissue and inoculate it
into mice with appropriate controls, wait until the mice come down, and do
the lesion profiling.

Dr. Thornsberry: There were two cases in Japan, but they indicated that
tissue was not classical BSE as seen in Europe. Have you heard anything
about that?

Dr. Bartz: This is based on differences on migration and the glycoform
ratio of PrPSc.

Dr. Detwiler: Canada based that question because the herd that that animal
came from was in Saskatchewan, in an area with CWD. That was one of the
questions they faced right off the bat: is this BSE or is this some kind of
transmission from CWD-infected elk in the area? Not only the histological
lesions were classic BSE lesions, but clinically it's very difficult because
if you miss the other behavioral changes, which this owner did. It was
someone who had been a catfish farmer. He missed the early signs. The
animal presented to slaughter as a down animal, non-responsive. Clinically
it looked like just a down cow, but they did send that on to the United
Kingdom and they did do some comparison glycoform patterns. Those haven’t
been validated, but at least on preliminary work, it looked like classical
BSE.
The Japanese case was a 23-month-old which was born in October. Their
scientists say the western blot pattern looked different. The most recent
case, which was a 21-month-old, looked more like classic BSE. The Italian
cases were older animals, 15 and 16 years of age. But is it methodology?
Is it really standard? That has to be sorted out before too much can be
said.

Dr. Bartz: Glycoform ratio is dependent on very technical matters, what
antibodies you use, what detection system you use. Those have to be
standardized before you can start comparing from one lab to another.

Dr. Detwiler: The Japanese used a western blot they'd developed in their
lab. It can't be compared across laboratories.

Dr. Bartz: That's problematic.

CHRONIC WASTING DISEASE

Dr. Terry Spraker: Max asked me to give an overview of CWD and then talk
about the research being done at CSU. CWD was first noticed in the Colorado
Division of Wildlife pens in Ft. Collins. CWD in the literature was first
said to be seen in captive deer pens in 1967. I remember being there and
those deer pens weren't even built until 1968, so the early history of CWD
is confusing, but CWD was at least seen in the very early 1970’s as far as I
can remember.
Beth Williams verified that is was truly a spongiform encephalopathy in the
deer. She and Stuart Young described CWD in deer and elk in Colorado and
Wyoming. George Bear A big Game Biologist for the Colorado Division of
Wildlife found a sick elk in Rocky Mountain National Park in 1981 and I
posted it for him and it turned out to be a case of CWD. At that time,
there was lots of interchange from the Colorado Division of Wildlife with
the animals from Rocky Mountain National Park. They took animals from the
deer facilities and released them into the park to study food habits, and
brought the deer and elk back.
We found the first case in mule deer in 1984 about half a mile west of the
Colorado Division of Wildlife deer pens. In 1985, we found the first case
in white-tail deer in Loveland, about 30 miles south of Ft. Collins. There
has in the past been a question whether CWD started in the CDOW deer pens
or came from the wild. The man who built the deer pens in 1968 did put deer
and sheep together. These animals were placed on starvation trials. He
obtained the sheep from CSU. This is where this theory has come about that
one biologist started CWD in the Colorado Division of Wildlife deer pens. He
still has sheep today from the original group, and he's never had a case of
scrapie. There is no real evidence that this was the start of CWD.
The other part of the history of CWD was when it was first found in captive
elk in 1996. The man who had CWD in Canada has gotten elk from a place in
South Dakota that had CWD for probably 10 years, but it was not diagnosed.
The year after it was diagnosed in Canada, it was diagnosed on his farm in
South Dakota. He claims he received deer from Colorado. So there is a
fairly good link from deer and elk originally from Colorado and spreading
northward.
The natural host for CWD is only three cervids: mule deer, elk and
white-tail deer. Recently, CWD has been found everywhere north of
Interstate 70 in Colorado. Now a case has been found south of I-70 and one
in central Utah. We just looked at several deer from Mexico and those were
negative. CWD has been found in wild populations in Wyoming, Nebraska,
Wisconsin, Utah, South Dakota, New Mexico, and Illinois.

Dr. Thornsberry: I have a classmate who deals with the Wisconsin Department
of Natural Resources, and he's done a lot of work with them on CWD. How did
we get this hot pocket of CWD in Wisconsin? Did they trace that to somebody
introducing it, or did it just show up?

Dr. Spraker: No one knows. People bring carcasses back to their homes.
I've had calls from so many states and people want to know what to do with
the carcass. One guy in Maine called me after he'd already thrown the
carcass away. One person has taken six cases of CWD to British Columbia.
The last couple he said he fed to the crabs—threw them in the ocean. There
is a lot of movement of animals and lots of movement of animal parts. There
was a big trade of Wisconsin deer that somehow went to Missouri and then
were released in Texas.

Dr. Spraker: The captive herds are primarily in the same areas where
there's lots of CWD in the wild. I'm sure there's some evidence that it
goes both ways. What are some of the epidemiological observations? There
is evidence of transmission of the disease in captive animals to wild and
vice versa. In the wild, there's natural expansion.
Clinical signs include emaciation. These clinical signs are not manifested
until the terminal aspects of the disease, so an animal might have the
disease for two or three years without showing clinical signs. You'll see
emaciation and occasionally you'll see hair loss. There are similarities to
Scrapie, although you don’t get the weight loss with Scrapie.
The clinical signs are obvious only in the terminal stages. They are
depressed and you'll find them lying down. If you approach them, they do
not get up.

Dr. Thornsberry: Is the chronic weight loss due to lack of neural functions
of the muscles, or is it due to being mentally affected and they just don’t
eat?

Dr. Spraker: I don’t know. The rumens are always full, so they are eating
right up to the end. Because of the olfactory system that's heavily hit in
these animals, I think that, at least in the wild, they lose their sense of
smell. Deer are very dependent on smell for which forages to eat. Prion is
present in the retina, so their eyes are affected also.

Dr. Detwiler: In Ohio, we did see a wasting condition associated with
sheep. I don't think anyone's ever done a study, but there also seems to be
an association, at least in flocks, with the abomasal emptying disorder.

Dr. Spraker: We have looked at a few cases of this abomasal emptying
syndrome in sheep. We check them for prion disease, because it looks like a
classical manifestation of an unusual manifestation of scrapie. We've never
been able to show anything. We see abomasal ulcers in deer, but never in
elk. With captive deer, you see a slowing of the emptying of the rumen and
the animals will drink a lot of water. A clinical sign in captive deer is
the sloshing of the rumen when the animal would run.
One clinical sign deer will manifest is excessive salivation. We saw this
early in animals in Estes Park. We see animals eating out of food bins,
salivating into them, and then other deer would come and eat out of that
same feeder. One possible transmission is through the saliva. But when you
find the prion in the lymphoid tissue of the gut, it could easily go out the
feces also. Excessive salivation is a common clinical sign in terminal
cases.

This is a gross necropsy of a deer, fairly typical. You'll see there's no
fat at all in this animal. This anterior ventral aspect of the apical lobe
is pneumonic. This is typical of aspiration pneumonia and you see this in
both elk and deer. It's most likely associated with innervation of
swallowing. The animals have trouble swallowing and they'll aspirate food
into the lung. Actually, one of the first cases of CWD in captive elk in
the US was seen in 1995, a year before the cases found in Canada, and these
pathologists in South Dakota missed three or four cases of CWD in elk
because they saw the pneumonia and stopped there. This went to court. The
elk rancher felt it cost him money because the pathologist missed it. The
court ruled in favoroft the pathologist because it had not been diagnosed.
The bottom line is that CWD has been missed because people have found
pneumonia and have stopped the necropsy. When we see pneumonia in deer or
elk, the first thing we think of is CWD.
The clinical signs and gross lesions in the elk are identical to the deer
and white tail deer. The histological lesions are similar in all of these
TSEs, at least in the ruminants where you have the vacuolization of neuropil
cytoplasm, vacuolization of gray matter and the neurons, vacuolization
within the neurons, and neural degeneration. You see plaques in a lot of
these diseases. They’re not as florid as you see in humans. They are areas
of accumulation of prion.
In the literature, you see absence of inflammation. I don't believe this
anymore because of the proliferation of the glial cells. The early lesions
you see with CWD are the vacuolization within the neuropil. The neurons
look good and there's no evidence of neuronal degeneration.
The immunohistochemical stain has been helpful with CWD. At first we were
using H&E to diagnose CWD and it was very difficult. With the advent of the
antibodies made for scrapie, we showed that the CWD agent also stained with
the scrapie agent.

Dr. Thornsberry: If it doesn't stimulate an immune response, how do you get
an antibody to it?

Dr. Spraker: The antibody is made in a different animal like a mouse or
rabbit, but most of these antibodies are made in cell cultures. The animals
do make antibodies or the proteins from other hosts.

Dr. Thornsberry: So the statement that there's no immune response to it is
not true.

Dr. Spraker: Deer do not make antibody to their own PRP, but if you put in
one that's not recognized as self, it will make antibody. There are
hundreds of antibodies made to PRP.

Dr. Thornsberry: So you get a hamster immunized against a deer prion.
You're developing a hamster antibody to a deer prion. But you can’t develop
a deer antibody to a deer prion.

Dr. Spraker: That's right. People take small segments of the prion and
make a specific antibody for that amino acid sequence. This antibody is
made for a six-amino-acid sequence. It's made in cell cultures. This
antibody has worked very well.
Another boon was that, early on, we found lymphoid tissue being positive
in deer and elk. This was an important finding. It was confusing at first.
People have ingested deer that had positive lymphoid tissue and the brain
being negative. At that time the positive lymphoid testing was quested by
some agencies and it was considered not a valid test. Hundreds of people
have ingested deer that had positive lymphoid tissues.
Katherine O'Rourke developed a third eyelid test in domestic sheep where
they stained for the lymphoid tissue in the third eyelid. They can find
positive follicles in the third eyelids of sheep and we've shown you can do
this with the tonsils of deer. An article by Margaret Wild has shown you
can diagnose CWD a couple of years before the animals come down with
clinical signs by doing a tonsilar biopsy.

Dr. Detwiler: I think it's important to note the test could be negative or
there could be lack of lymphoid tissue in the third eyelid. You see that
especially in sheep. As they get older, it atrophies. So a negative is not
definitive. But the positive is very significant. We shouldn’t undersell
the fact that the third eyelid had added a new tool for us in the field and
in the production side as well as the regulatory side so we can detect the
disease in a live animal in the pre-clinical stage. We can get into flocks
earlier and start to eliminate animals. It's very good for flock-screening
and it's helped us identify flocks in an earlier timeframe. It's added
diagnostic capability for us.

Dr. Spraker: Yes, that's very important. A negative test doesn't tell you
anything, but a positive test tells you a lot.

Dr. Spraker: So far, we've found a couple of false positives with elk, but
that's when we were playing with different techniques and not using PK.
With the standard tests that have been published and used now, to my
knowledge there have been no false positives. If you do not destroy all the
PrP-cellular, you're going to get positive staining. These antibodies are
made for the PrP-cellular, but part of the testing is that you are supposed
to destroy all the PrP-cellular. There are several different ways of
destroying the PrP-cellular.
Here's a suggested pathogenesis for CWD in the deer. I think we're just at
the tip of the iceberg with pathogenesis. It appears, at least with deer
and elk, there could be an ingestion of the prion. The prion does enter the
Peyer's patches and some of this was shown by Christina Sirgudsen. I
personally think there is an adaptive phase in the lymphoid tissue. It
takes awhile to adapt, and then it goes, I think, up the vagal nerve. It
enters the vagus nucleus and then there's an adaptation in the brain and
then, after it adapts in the brain, it spreads throughout the brain fairly
rapidly. Because of high prevalence of stainable PrP in the lymphoid tissue
of the small intestines and colon, I wouldn’t be surprised if it does not go
out through the feces and through the saliva because of the tonsil being so
heavily infected. There is so much lymphoid involvement so close to the
outside of the body. At least that's one way the prion may go out of the
body.

The Research
I know only a little bit about what's happening with the Department of
Wildlife. It's fairly well known that they're doing susceptibility trials
with cattle. Beth Williams and Mike Miller are the two primary
investigators on this. They have put 10 or 15 cattle in deer pens that have
very high infections of CWD. With this, they're studying environmental
contamination. I think Mike is putting some dead deer in certain areas and
letting them rot and then later they'll put in other deer to see if they can
feed in that area and come down with CWD. The problem with this is that we
have so much CWD all around Fort Collins, it would be hard to determine the
source. They're also doing a project feeding CWD material to mountain
lions. This work is being done in Wyoming and Colorado.
At Ames, they've injected 13 cattle with CWD brain tissue. Five have died.
The first three had some vague clinical signs. Prion was found in those
cattle, but they had very little spongiform change. The author is Ahamir
Hamir.
Dr. Lynn Creekmore is doing a project in Ft. Collins. She's with the
National Center for Animal Health. She has purchased 20 fallow deer and has
put them in one of the pens where there's a degree of CWD. They're being
monitored. At the two-year mark, they show no evidence of CWD. It's a
five-year study.
At CSU, there are three groups working. Ed Hoover is doing quite a bit of
work on some of the pathogenesis. Barb Powers and I are doing some work
with CWD and Mo Solomon in the Department of Environmental Health has done
investigation into some of these ELISA tests.
Ed Hoover's graduate students showed that that oral transmission of CWD
worked, and they found PRP in intestinal and cranial lymph nodes 42 days
after ingestion. These were fawns that received five grams of the brain
tissue. They've also done some work with dendritic follicular cells and
they've shown that the B cells and the dendritic follicular cells play a
part with recognition of prion. They’re also looking at normal location of
cellular PRPC throughout the whole body of a deer.
They have a group of white-tail deer and they're going to look at
transmission of saliva, feces and repeating the brain experiments,
specifically seeing if saliva and feces and be used for transmission of CWD.
They're using mule deer material going into white tail deer. I don't know
how much of a species jump that's going to be. For all practical purposes,
it doesn't look like there should be. Katherine O'Rourke has shown there's
quite a bit of difference in the genetics of the PrP gene within white-tail
deer and within mule deer.
Barb Powers is working with the Bio-Rad ELISA test. They did a tremendous
amount of work last year in validating this test for elk and mule deer in
Colorado. It was a political nightmare. Mo Solomon actually started it,
doing comparison of Bio-Rad and Prionics. They took the tests that were
used for BSE and tried to modify them to work with CWD. The people who
produced Prionics and Bio-Rad sent their primary scientists to try to make
the tests work. Bio-Rad was the better test of the two, so it was chosen
for the extensive surveillance that happened last year that resulted in the
validation of this test for CWD in the US for white tail, mule deer and elk.
But it's only in the lymphoid tissue.

Dr. Detwiler: Since then, there have been two additional tests.

Dr. Spraker: The tests now are Idexx, validated for white tail deer, and
VMRD, validated for white tail and mule deer. It's still only lymphoid
tissue.
We found some of the first early cases and we showed that the captive
animals and the wild animals had the same disease. That was a big question
at first. We found you could not tell the difference between captive and
wild mule deer when you looked at the lesions in the brain and at the
patterns of lymphoid staining throughout the body.
We developed the IHC test with Katherine O'Rourke that's currently used
throughout the US and Canada, and we mapped the prion in mule deer. These
were all hunter-killed animals so the time of exposure for CWD was not
determined. But we saw definite patterns of prion distribution. We had
only the heads. When you do staining only in the tonsil and when you do
serial sections of the brain, there's no evidence of prion in the brain.
Then we had an animal with a positive tonsil and a group that had prion only
in the vagas nucleus. Then it would spread to the solitarius. What was odd
was that the next area where the prion appeared was the hypothalamic region.
We're starting to do this same mapping now in elk and the picture looks the
same so far. The thing to notice here is that the cerebellum is not
affected until the very end. I don't know how it is in sheep, but this is a
characteristic we've seen in the deer.
When you're doing any kind of surveillance, if you used brain tissue, you
have to get this vagas nucleus. If you don’t, you can miss a lot of the
early cases.
We had a chance to work with the state and federal people when they were
trying to eliminate CWD from the US and Canada. Between the US and Canada,
12,000 to 14,000 ranch elk were killed. We looked at a lot of the animals.
We wanted to look for patterns and also compare the stainability of the
lymphoid tissue, because in deer, it seems consistent that the lymphoid
tissue was affected with the brain and you would have early cases where
you'd have lymphoid staining and no brain staining. In mule deer, we found
one case where there was staining in the brain and not staining in the lymph
nodes. I think they've also found that in sheep, and we've found a little
more of that in elk.
For all this testing we've used IHC. Two different machines are used —the
Nexus and the Benchmark. We've used the antibody produced by Katherine
O'Rourke. I know this antibody is being produced by VMRD and it's sold, but
we have not had very good luck with this commercial antibody.
These are some of the different stages in the elk. We're starting the map
the whole brain, so you may be able to look at one section to obex and
predict where the prion is in the rest of the brain. A grade one is a
negative animal and a grade two is an animal that had negative staining in
the brain but positive lymphoid tissue. Grade three is where you may or may
not have lymphoid staining, but you have what we call a one-plus staining in
the obex. The first area you see affected is the lateral aspect of the
middle third of the vagas. In cattle, one of the first areas to be affected
is the solitarius nucleus. Another very important difference between BSE
and CWD is that we would miss probably 30 to 40% of the cases if we did not
use IHC. From what I'm told, with BSE, the immunostaining is not as good in
cattle as it is in deer and elk. You can do very well with good H&E
sections.
With elk, you see very little intra-nuclear staining, whereas with cattle,
you see much more intra-nuclear staining of the prion.
A grade four means that the vagas nucleus is mostly affected. A brain plus
three means it's filling the nucleus and the solitarius and it's beginning
to spill out into some of the other nuclei. At this stage, the rest of the
brain will begin to have prion in it.
We've found that the redex is the vagas is affected later than the nucleus,
so the prion accumulates in the nucleus before you can see it in the redex
of the vagas. The redex of the vagas is the conduit of nerves coming in and
out.
With this grading of elk, we have a variance of CWD staining in the
lymphoid tissue. It's not as neat as it is in deer. Even in the more
terminal stages, we found a couple of elk that did not have staining. Some
of this may be due to not cutting enough lymphoid tissue, but we've really
looked at some of these animals and we have not been able to verify
immunostaining in these lymphoid tissues. It's not as clean as it is in
white tail deer.
We're trying to map the rest of the elk brain and do the same thing with
white tail deer, and them compare them to what a single section of the obex
would be. Also, we have a project looking at some of the
micronutrients —copper, manganese, molybdenum, selenium and zinc—and seeing
if that correlates with CWD. We're also looking at lymphoid tissue in
really early cases of CWD.

Fear Factor
Another problem that has come up because of the rampant fear factor in the
US and in Europe is disposal of carcasses, tissues, and reagents that have
come in contact with TSE. In some places, you can't even think about using
landfills to dispose of carcasses if they have CWD or Scrapie. At CSU, we
have a sodium hydroxide digester that brings the temperature up to 300˚F
with 60 pounds of pressure, and you cook this material for six to right
hours. This digester cost about a half a million dollars and we've sunk
more money into it. Disposal is a big problem. Some communities have
hampered the building of incineration facilities with the claim you can get
CWD by breathing the air. These thoughts are being propagated and causing
us a tremendous amount of trouble.

Dr. Detwiler: Even when you do non, do you still get an odor from the
digester?

Dr. Spraker: Yes, especially when you dump it. If you put it into the
sewer, it's okay. But our city charges for BOD (biologic oxygen demand).
So it costs us $3000 to $4000 a month to dump it in the sewer, and they just
came up with a half a million dollar surcharge for having the privilege to
dump it. We can't afford it, so we've tried to make a dehydrator. The
fragrance of the high ammonia escapes.
The rendering companies in Ft. Collins are going out of business. We were
forced to go somewhere and incineration was very difficult, so we went to
this system.

Dr. Bartz: Is the operating cost high?

Dr. Spraker: No, it's really not. But you have to take into account all
the other problems.
This is the big question. This is the hypocrisy of this disease. We've
had to reconvert all these buildings to make them where all the water goes
into the digester because we couldn’t let it go down the sewer any more.

Dr. Detwiler: Was this EPA's idea?

Dr. Spraker: That's where it first started, yes.

Dr. Bartz: The digester is just for CWD-infected tissue?

Dr. Spraker: It's for TSE, Scrapie and CWD.

Dr. Bartz: What about rodents?

Dr. Spraker: If it's a TSE, it has to go in here, yes. This is the only
way we dispose of animals now, but it's mandatory if it's a TSE—even gloves.
This is data we have to date. People ask Can humans get CWD? You never
say never, but this is what we've seen so far: humans have been heavily
exposed by handling and consuming CWD-positive deer and elk for the minimum
of 35 years. I don't even want to think about how many deer we consumed
from the deer pens when I was a graduate student.
This is very important: Prusiner says it and others have, too—that CWD can
go to human and there's just not been enough exposure yet. This is my
opinion: in Britain, when a cow had BSE it was ground up and diluted and
spread. When a hunter kills a deer or elk with CWD, the immediate family
eats the deer. They're exposed for a year. Most people take a year and a
half to eat an elk. So if CWD is going to go to people, those people would
have a high exposure rather than a diluted exposure. Exposure to brain is
there, as some of these people do brain tanning. To date, there have been
no unusual neuro-degenerative diseases found in people, and they have
looked.
The fear factor is unreal. I had a man come to the office in a panic, and
he was sincere. He said he'd gotten some drops of blood on his pickup from
a deer that came from the CWD area. He thought the back of his pickup was
contaminated. He asked if he should sell his pickup
Another man came in with a deer to dispose of it, and he knew it was
positive, but when he looked at it, he decided to take it home.
I had a case where a hunter had shot an elk or deer and Game and Fish
called and got his wife and told her the deer had CWD. The next day they
had a party and fed the deer to their guests. A nasty divorce was going on,
and after the party, the wife told her husband about the CWD. This was a
group of MDs. As of now, all of them are fine.

Dr. Detwiler: If someone calls you and says he has a known positive animal
and wants to know if he should eat it or not, what would you tell him?

Dr. Spraker: I'd tell him "no." I've thrown away a deer and an elk and I
hated to do it, but I would not knowingly eat it. I'm not afraid to go and
hunt in that area and test, but the test will miss early cases.

Dr. Bartz: Do you have an estimate of the numbers of people you think have
been exposed?

Dr. Spraker: No, but I can tell you this year they found 200 positive deer
and elk. On a smaller scale, just in Colorado, we’d average 100 to 125
positive cases of CWD. But no testing was done in these other states. A
lot of people have eaten these deer for a long time, and they have gone
throughout the United States. There have been claimed cases of hunters
having CWD, but they were all other conditions.

Dr. Keller: The cause of death is not always known when people die of
neuro-degenerative disease.

Dr. Spraker: Yes, but right now they are really paying close attention to
that.

Dr. Detwiler: After being hammered by the public health community that we
don’t test nearly enough cattle, they still ask how many full brains really
get done. The human community does not do a systematic look at brains of
neuro-degenerative diseases.

Dr. Keller: Animal health agencies are being encouraged to determine
prevalence of CWD in cervid populations.

Dr. Spraker: But if we had this much CWD in this small of an area and this
many people consumed it, I think they would have picked up something.
Something would have spiked up. So far there's been no unusual incidence of
any kind of clinical neuro-degenerative condition.

Dr. Thornsberry: Dr. Bartz, hearing that from Dr. Spraker, what is an
explanation? You've probably done the studies to show that these prions are
in the muscle tissue as well.

Dr. Bartz: Prions or PrPSc have been detected in muscle homogenates from
prion-infected mink (Marsh et al., 1969), hamsters (Bartz et al., 2002,
Thormzig et al., 2003) and humans (Glatzel et al., 2003). Additionally,
immunohistochemical data has shown PrPSc to be associated with myocytes in
humans with CJD (Kovacs et al., 2004) and in prion-infected hamsters
(Mulcahy et al. from the lab of Dr. Richard Bessen, Journal of Virology in
Press). It is still unclear if prions replicate in muscle cells, but the
presences of PrPSc in muscle cells suggests that prion infectivity and PrPSc
that is detected in muscle homogenates is not entirely from innervation of
the muscle tissue.

Dr. Thornsberry: What would be an explanation to having that kind of
exposure to prions and not developing into a Scrapie-type prion that we call
disease?

Dr. Spraker: I don’t know, but I know people have consumed it.

Dr. Thornsberry: Fox news reported on a military man stationed in the
Middle East who consumed the traditional breakfast in those Moslem
countries—sheep brains scrambled with eggs. He's not be diagnosed with
Creutzfeldt-Jakob type disease. They're trying to get military pay for him
because of hazard of duty. They think he picked this prion up from
Scrapie-infected sheep or goats, not cattle. Yet we know that the ability
of that to go to humans is supposed to be minimal. Why are there not
outbreaks all through the Middle East of Creutzfeldt-Jakob disease?

Dr. Spraker: We ship a lot of sheep brains over there. It's a complex
situation. I think CWD is a unique spongiform encephalopathy. Whether it
came from sheep or a spontaneous mutation, I don’t know. At the time,
there's no evidence it can be naturally transmitted to any of these animals.
If CWD is found to be naturally transmitted to cattle or sheep, there's
going to be a lot of change.

Dr. Thornsberry: We do know you can give an intra-cranial injection of this
prion and cause lesions? They've documented that. But what about intense
natural exposure like the eating of elk placenta or the licking of an elk
carcass?

Dr. Spraker: When you have captive animals, you see a lot higher incidence
of CWD than in any wild population. A man in Nebraska fenced in some white
tail deer, and they ended up killing them and 52% had CWD. When you
concentrate them, whatever way they transmit it, you're going to have a lot
higher incidence. This is one of the reasons they're killing so many deer,
thinking they're going to reduce the number of animals and thus reduce the
incidence of the disease. They'll never take it away.

Dr. Thornsberry: Is there a high prevalence of prions in placental tissues?

Dr. Spraker: We've had several elk and several deer that died of terminal
CWD that were pregnant. We've not been able to demonstrate PRP in the
placenta or any of the fetal tissues with IHC or with Bio-Rad. We can
demonstrate IHC in the placentones of domestic sheep. A man out of
Katherine O'Rourke's lab has a nice article on demonstrating PRP in the
placenta, and Race has an article about finding PRP in the placenta and
showing how the PRP of the placenta has different glycoforms than the PRP of
the brain. In sheep, they've definitely shown that. For a long time, they
said that Scrapies transmitted from the ewe to the lamb during lambing, but
it does not go trans-placentally. But this does not seem to occur in elk
and deer.

Dr. Detwiler: the important thing with sheep is not only PRP, but they've
actually correlated the PRP detection of infectivity with detection of
PRPSC, which added on to Patterson's work back in the sixties. In sheep
they actually fed placenta to goats and caused the disease by oral ingestion
of the placenta from Scrapie-infected animals.

Dr. Spraker: I'm trying to get some money to do that with deer because I
have some sheep placentas.

Dr. Thornsberry: So to summarize, you do not believe, in the data you've
examined and your experience in pathology, that there is much risk of
transfer of CWD from a cervid to a bovine.

Dr. Spraker: There's been no evidence, but I haven't done any of that work.

Dr. Thornsberry: They're saying,”Is the prion going to be there forever?”

Dr. Spraker: That's the concern that some people are voicing about this.
It's been a hot topic.

Dr. Spraker: There's been a project started by Jim Voss and Dan Guald.
They talked a group of ranchers living within the enzootic area for CWD into
turning in all the heads of their cull cows. The cattle came from enzootic
areas for CWD. They had about 270 culled cows that had been in contact with
deer from six to 12 years. They did histopath and IHC on these cattle and
all were negative for any evidence of a TSE.

Dr. Thornsberry: With that knowledge, where in the world are these BSE
cases coming from? Is this spontaneous generation theory going to hold up?
If that's possible, then you should be able to get BSE anywhere in the
world.

Dr. Spraker: They say that 85% of the CJD cases fall into the sporadic
category. That means they cannot find any evidence of a source of the
disease. Agussi is beginning to think that sporadic CJD is from sheep, but
he doesn't have evidence. If that's the case, why don’t Australia and New
Zealand have sporadic CJD? As far as I understand, with the sporadic cases,
there's a deterioration or a degeneration of the prion that weakens some of
the links and it turns or it changes its configuration.

Dr. Thornsberry: We were shown data that brain tissue heated to 600˚C was
still infectious.

Dr. Spraker: To me it's easier to think maybe it's associated with a metal
and it comes alongside and causes the normal prion to change.

Dr. Thornsberry: We've established pretty well that CWD probably is not
infecting cattle. It's probably not infecting humans. But we have
established that it's very infective with deer and elk, and it’s a problem.
Whatever transmission is taking place, it is infective. It is a dangerously
infective prion within the species that it's established in. And yet we
keep hearing this information, as veterinarians, that BSE is not infective.
It's a terminal disease. The animal gets it and dies. We know that's not
the case with many of these prion diseases.

Dr. Spraker: You also hear that CWD is always fatal. I don’t believe that.
I see too many cases of animals that are big and fat and healthy-looking,
killed by a hunter, and they have so much prion in the brain it's
unbelievable. Even in later cases, there are a lot of them that do not show
clinical signs. The degree of spongiform degeneration is not nearly as high
in some animals as in others. We're slowing pushing in that direction to
see what the difference is.

Dr. Bartz: One of the hallmarks of inter-species transmission, at least in
rodents, is that the spongiform degeneration is way out of proportion in the
first transmission to what you see in subsequent transmission. If you just
look at brain sections, you can always pick out the animal that's the first
passage because the spongiform lesions are so much more intense.

Dr. Spraker: And the amount of accumulated prion is less, right?

Dr. Bartz: It can be less. That hasn't been looked at thoroughly enough.

Dr. Spraker: We see animals that have had this disease for three years and
we see hardly any spongiform change, but yet you see lots of accumulation of
PRP. But when they tell you that every animal that gets CWD dies, how do
you prove that?

Dr. Thornsberry: If that animal doesn’t die and he's a plus three or plus
four, he's shedding prions all over creation. So you have an infectious
disease with an infectious agent in the environment.

Dr. Spraker: The state vets don't like to hear that. No one can prove it.
But just with casual observation over the years, I don't believe that
statement anymore. It might take a lifetime to prove it, but I think that
not even thinking about it is more dangerous than trying it and not being
able to prove it. That might help to explain all these mysterious crop-ups
of CWD. If it does come from sheep, that would help, but it's not a simple
thing. There are two or three cogs that have to go and prions are just one
of them. You can make it work with one in the lab, but natural, it's going
to take two or three.

Dr. Keller: Do you remember what the requirements are in Canada after
finding a positive case of CWD on a cervid premises? I believe they do not
let ruminants back into the same facility for three years or five years?

Dr. Spraker: It varies, but I think it’s around five. South Dakota is
five.

Dr. Detwiler: They're changing again. They just recently discussed it.


THE REGULATORY POINT OF VIEW

Dr. Linda Detwiler: I'm going to take a different approach. I'm going to
talk about scientific findings and how they're pertinent and why they're
important to regulatory policy. We know there have been about 150 deaths
from variant CJD from 1986. These are very tragic indeed, however, there
are a lot more deaths from the flu and other diseases, yet this is what's
made headlines and created a panic.
The objective is to review the research significant to regulatory issues
and what's know and—equally important—what's not known. We'll look at the
BSE case in Canada and what happened there and what may have happened in
their feed cycle. We'll look at implications for the US and possible next
steps.
It's important for us to know the similarities, and equally important to
know the differences. The differences between species for control purposes,
eradication, and prevention are so critical. If you don't know the
differences in regards to transmission, pathogenesis, etc., you won't be
able to control the diseases appropriately. For example you cannot control
Scrapie like you do BSE with a feed ban.
My mantra is that you have to remember these long incubation diseases with
limits to the pre-clinical. You have to be thinking. If you knew the
disease was going to be here tomorrow, what should you have done yesterday?
If you wait until the first time you find the disease, you are years too
late. You might be one incubation cycle too late or multiple incubation
cycles too late. The population you're concerned with, be it cows or sheep
or elk or humans, it's already been infected and you're just going to live
out the clinical manifestation of the disease.
Prevention is best, but if you know it’s been introduced, what can you do
to limit the transmission? Europe has had to play it out.
The UK in 1988 put the ruminant-to-ruminant feed ban on, but they did not
ban the total use. It's banned only for ruminants, so they had too much to
use. It's not prohibited to go other places, so the meat and bone meal is
sold to the continent. It dumps into France and starts to move throughout
Europe. Add another five years incubation period, plus surveillance, and
you see the manifestation of clinical detection in the Benelux countries.
Europe then started to restrict the use of meat and bone meal, and sale of
it went on to other countries.
There are a lot of countries that probably have received product, but are
not doing surveillance. This is something the US needs to watch out for. We
don't classify countries like we do for other diseases, like foot and mouth.
For BSE, we'll stop trade if a country is not okay, which is just the
opposite from the policy for foot and mouth. This is something the US needs
to change. We need to evaluate the countries of the world. Canada has done
this and Canada will trade only if the BSE risk is at an acceptable level.
We can't forget other animal species are susceptible, such as cats.
Domestic cats and large zoo cats are susceptible to the BSE agent, as well
as TSEs of exotic ruminants. If exposed to the BSE agent, bison are
susceptible.
Why is the distribution of infectivity important for us? There are several
reasons from a regulatory standpoint. If you know where the infectivity is,
it helps you eliminate high risk tissue, especially ones that might be a
zoonotic risk. That is, What tissues might be more infectious to humans if
it’s a zoonotic disease or out of the rations of other animlas if they’re
susceptible. Also, it provides direction as to what tissues you might
target for diagnostic purposes.
In natural cases of BSE, infectivity was in retina, brain and the spinal
cord.
In the experimental challenges, calves were fed 100 grams of BSE-infected
brain tissue. At certain intervals, some were sacrificed. Forty-some
tissues from these calves were put into mice to be bio-assayed. The British
were criticized, saying the mouse bio-assay is not as sensitive because of
the species barrier, and that you should go back and repeat this in
cattle—at least for the most important tissues. This study in cattle had
actually generated some additional data { that is additional tissues where
infectivity has been found}. The trigeminal ganglia, the dorsal ganglia,
the distal ileum and the bone marrow were all from the original mouse
pathogenesis study. Recently, tonsil has been found to have infectivity by
a calf bio-assay.
Pre-clinically, the distal ileum was six months post-inoculation, and the
tonsil was about 10 months post-inoculation. To date, cattle muscle samples
have been put into mice and also back into cattle, and no infectivity has
been found.
Scrapie infectivity has been identified in brain, spinal cord, tonsil, the
peripheral lymph nodes, nasal mucosa, placenta, liver, and the length of the
intestine from the esophagus to the rectum. Nora Hunter has published work
where both whole blood and buffy coat were taken from sheep and inoculated
back into sheep from New Zealand, and those sheep did come down with TSE.

Dr. Spraker: How did they demonstrate nasal mucosa?

Dr. Detwiler: Bill Hadwell collected nasal mucosa from Suffolk sheep and
inoculated it into mice.
In baby ruminants, the cells of the distal ileum are a different cell type
up until about nine months of age. The cell type is such that they allow
the passage of large molecules to transported across the intestine. By
about nine months, that function wanes. It's probably for colostral
purposes, for colostral antibodies to transfer over. But some have asked if
that's why there's an ease of transmission across the gut at a younger age.
Horrigan's work at Mission, Texas, showed that after nine months, it
appeared to be more difficult to transmit Scrapie. It may be there's a
correlation with the change of cell types.
This is a chart I made for the OIE, the Office of International Epizootics
to look at secretions and excretions in sheep and goats where, to date, no
infectivity had been found in feces, urine, saliva, colostrum, milk and
semen. Intra-cranial inoculation into mice resulted in no infectivity. But
I want to point out that I don't think we should stop looking. Even with
the blood. If you had asked someone prior to 2000 if infectivity was ever
found in sheep's blood, he would have said "no," because work done to that
time had not shown infectivity in sheep blood. But that work was all blood
intra-cranially into mice. How much blood, feces and saliva can you put
intra-cranially into a mouse?

Dr. Bartz: With BSE into mice, they estimated a 104 difference in
sensitivity between the cow and the mouse.

Dr. Detwiler: How about nasal discharge? Is that a possibility for shed of
the agent? We have to keep asking these questions, even for cattle. If
it's in the lymphoid tissue of the distal ileum, the tonsil, or the lymphoid
tissue of the third eyelid, would it not make sense that, at least at some
low level, there might be some lymphoreticular distribution similar to the
other animals with TSE. I think that's a valid question to ask, and the
calf pathogenesis studies are not finished. I think we have to be careful
definitively in saying it's limited to only two three peripheral tissues.

Dr. Detwiler: Dr. Prusiner is examing muscle from cattle with BSE, and
looking for presence of PRP and probably infectivity.

Dr. Thornsberry: He's very concerned about muscle tissue.

Dr. Bartz: There's one comment I'd like to make on the muscle infectivity
studies. We need to remember that a lot of these are taking a biopsy. If
you take a biopsy of muscle, what's in that? It's obviously muscle cells,
but it's also nervous tissue and LRS tissue. It's in a muscle homogenate,
which is consumed, but the question remains what cell type in that
homogenate is the agent really in?

Dr. Detwiler: I asked that of Dr. Prusiner. How did they ascertain that it
was muscle cells versus nerve tissue, etc. He said they were very careful
in how they took the biopsy.

Dr. Thornsberry: Doesn't every muscle biopsy have some neural tissue in it?

Dr. Detwiler: There's some innervation going to those muscle groups.

Dr. Detwiler: Parenterally, BSE does go into pigs. It was through three
routes: intra-cranial, IV and IP with incubation times of 69 to 150 weeks.
Seven out of 10 came down with BSE. Orally, it did not go after 84 months.
It's important to note that infectivity was not confined only to the CNS,
but also in stomach, jejunum, distal ileum and pancreas.
With chickens, both parenterally and orally, they did not find evidence of
disease. They tried to sub-passage the chickens by taking brain from
animals that had been exposed, to see if it adapted. I think you may have
to do this multiple times. What the research in the UK did do was look at
high-risk tissues from pigs and chickens that had been exposed to BSE, and
put them into susceptible mice. Again, we have to be careful to say there's
no residual infectivity at all. If continually fed to a species over time ,
there may be some kind of passage or adaptation over time. That's why, in
other countries, they take out high-risk tissues and don't allow them to be
fed at all. In Europe, they've gone to the extreme of taking out all animal
tissue being fed to food-producing animals, to break this cycle.

Lateral Transmission
Scrapie and CWD spread animal to animal. In both embryo and semen studies,
infectivity for transmission of BSE was not detected.
Two cases of disease with fairly significant numbers have been caused by
making vaccines from tissues from animals incubating the disease. Just
recently, a Mycoplasma aglacia vaccine that was given to sheep and goats in
Italy caused infection in 600 to 700 premises.
Contaminated feed appears to be the primary if not sole source of
infectivity. Animals that are incubating the disease get slaughtered,
rendered, and fed back to other cattle, and they get sick in an average of
three to six years.
This is significant to me when we talk about the British attack rate
studies. In this experiment, infected brain was given orally. One gram
killed seven of 10, so the British knew that had to go down to lower doses.
They've now looked at a tenth of a gram and a hundredth and a thousandth of
a gram. This is still underway, but at about 60 months, a tenth of gram has
killed three of 15 animals affected with BSE, and a hundredth of a gram has
killed one of 15 thus far at a 50-plus-month incubation. It doesn’t take
too much brain material. The British message is to not underestimate the
possibility of cross-contamination in the feed cycle. In the United States
we allow mills and plant to use the same equipment when processing ruminant
and non-ruminant material. Although a system of cleaning is required by the
FDA, the science questions how effective the flush methodologies could
really be. The same trucks may haul ruminant and non-ruminant feeds. Those
are potentials for cross-contamination.

Dr. Keller: What about risk from feather meal/poultry litter?

Dr. Detwiler: Feather meal can pick up infectivity from other feeds.

Dr. Keller: Whenever you feed a ruminant product to chickens and then the
feathers/litter are collected, they're collecting the ruminant material
directly in the spillage.

Dr. Detwiler: Hold on to that thought and I'll address it later.
A big unknown for the whole world but important to answer involves sheep.
BSE goes into sheep orally and maintains BSE characteristics. In research,
it looks like Scrapie clinically and histologically. To date, the only
valid way to differentiate BSE and Scrapie in sheep is a mouse bio-assay.
Would it spread? If it does go naturally into sheep, would it spread like
Scrapie through some kind of contagious nature?
So far, the distribution of infectivity is looking identical to Scrapie.
The significance of this is that the worst case scenario would be BSE in
sheep naturally. You would have a potentially zoonotic disease that could
spread from sheep to sheep and could not be differentiated from a common
endemic disease, would have a widespread tissue distribution, peripheral
nerve, blood, so no tissue of the sheep would really be safe. What would
countries do if they found this? That's about 40 million sheep.

Dr. Bartz: Do you know if anyone is taking material from the BSE-infected
sheep and inoculating non-human primates, to see if it looks like variant
CJD?

Dr. Detwiler: It seems a good idea, but I'm not aware of it.
Another important thing to get across to the public is that the negatives
do not guarantee absence of infectivity. The animal could be early in the
disease and the incubation period. Even sample collection is so important.
If you're not collecting the right area of the brain in sheep, or if
collecting lymphoreticular tissue, and you don't get a good biopsy, you
could miss the area with the PRP in it and come up with a negative test.
There's a new, unusual form of Scrapie that's been detected in Norway. We
have to be careful that we don’t get so set in the way we do things that we
forget to look for different emerging variations of disease. We've gotten
away from collecting the whole brain in our systems. We're using the brain
stem and we're looking in only one area. In Norway, they were doing a
project and looking at cases of Scrapie, and they found this where they did
not find lesions or PRP in the area of the obex. They found it in the
cerebellum and the cerebrum. It's a good lesson for us. Ames had to go
back and change the procedure for looking at Scrapie samples. In the USDA,
we had routinely looked at all the sections of the brain, and then we got
away from it. They've recently gone back.

Dr. Keller: Tissues are routinely tested, based on which tissue provides an
‘official’ test result as recognized by APHIS
.

Dr. Detwiler: That's on the slaughter. But on the clinical cases, aren’t
they still asking for the brain? But even on the slaughter, they're looking
only at the brainstem. We may be missing certain things if we confine
ourselves to one area.

Why Do We Do Surveillance?
Is the disease present? You absolutely have to do surveillance to find out
if the disease is present. It will also tell you if your prevention has
been successful. If you have the disease, it will also give you indication
that your controls are working. That's how Britain knew the feed ban had a
big effect. They continued to monitor disease and they saw it peak and they
saw it come down about five years after the implementation of the feed ban.
You also need surveillance for buyer confidence and for trade confidence
with our international partners. BSE is found in older animals, over 24
months of age, high risk, those exposed to contaminated feed, neurologically
ill cattle, any fallen stock and emergency slaughter. This is important to
collect samples through active surveillance as owners will sometimes miss
subtle signs.
In regards to inactivation, Paul Brown did some work and had to repeat it
because people didn't believe it. He found that, at 600˚ degrees Celius,
there was survival in the ash. However, there was none in air emission or
the residue. At 1000˚ Celius, he found no survival.
Canada has actually had two cases of BSE. The first was an imported case
in 1993—a UK import. This animal had a broken leg, but because it was a UK
import, the owner called the government veterinary staff and they had it
looked it. The Canadians had to do extensive tracings on all their UK
imports because of this case.
The native case was, of course, in May 2003. This animal was six years old,
an Angus type, presented for slaughter as down with signs of pneumonia. It
was condemned at slaughter, so it did not go for human consumption. The
head was taken for BSE surveillance, and the rest of the carcass went to
rendering.
The 2003 animal was not related to any of the imports. Was it spontaneous,
or was it a case of chronic wasting disease Epidemiologically, the animal
was born in the spring of 1997 and was most likely infected at that time.
Canada did import 182 head of cattle from the United Kingdom between 1981
and 1990, and there were 11 in the high-risk period, possibly incorporated
into the animal feed chain. They also did a risk assessment, which showed
they had a low risk of introduction.
They did trace-backs, and that helped to keep the public calm. There was
no crisis. In fact, beef consumption went up in Canada. But they tried to
find the herd of birth, which is really key in BSE. They narrowed it down
to three herds, and all of these were put under quarantine. The cattle in
those groups were pretty much depopulated.
When the positive animal was identified, CFIA did trace-backs, and that
helped to keep the public calm. There was no crisis. In fact, beef
consumption went up in Canada. But they tried to find the herd of birth,
which is really key in BSE. They narrowed it down to three herds, and all
of these were put under quarantine. The cattle in those groups were pretty
much depopulated.
They do have a national ID system, but it had been in place for only two
years. So they had to use other types of identification to trace back the
animal. Subsequent to the depopulations, one herd was identified as the
herd of birth.
One thing the OIE requires is that, for a country to resume trade in
animals and animal products, they should depopulate progeny if it's a
BSE-infected cow, along with birth cohorts. They had to depopulate over 100
head in these trace-forward herds.
The positive cow itself went to the slaughter plant and was condemned. The
head went to the lab and the remaining carcass went to a renderer. From the
renderer, the material could have gone to three farms. Two of these were
poultry farms. The poultry farms also had cattle, and there was an
admission that the cattle were fed the same material as the poultry. Again
this is prohibited by the feed ban, but it happens on the farm. These three
farms were depopulated in their eintirety.
This renderer did not have dedicated lines, so he made ruminant feed and
non-ruminant, prohibited and non-prohibited. They used a barley flush.
After the positive cow was processed, the barley flush went to one of these
farms and could have been fed to cattle. That was another breakdown in the
feed system. That's not supposed to happen, but it did.
It went to two pet-food plants. Material from the renderer at the time the
animal might have been processed also went to a feed mill that had 1800
clients, but none of these was depopulated. There were violations. The
message is that there are not a lot of resources for checking on-farm
compliance with the feed ban. The rubber hits the road on the farm.
I already talked about the epidemiological investigation, the
trace-forwards and the trace-backs. CFIA depopulated 2700 animals. They
tested approximately 2000 of those. They also brought in an international
review team made up of scientists from Switzerland, the US and New Zealand,
who reviewed the epi investigation and found it to be thorough and complete.
They made recommendations that Canada really had to get an SRM ban to take
the high-risk materials out of the human good chain now. Canada has done
that.
The committee told CFIA they needed to increase surveillance which examined
high risk cattle in large numbers. The review team also said they had to
find mechanisms to prevent this cross contamination in the animal food
chain. Take out high-risk materials from the animal food chain period.
Have dedicated mills, plants, etc.
The media was kind. There was no panic. However, rendering, slaughter and
the producer level impacts are still significant. Initially, there was no
place for the meat and bonemeal to go. The whole system started to back up.
The renderers couldn’t sell it, so they filled their silos. The feed-mills
filled their silos. They stopped slaughtering older cows because the system
couldn’t take any more. The government started to help, and some things
started to go to landfill.
Canada did put the SRM ban in place for public protection, but not yet for
animals. CFIA is working on increasing surveillance, and they need to look
at changes in the feed rule.

More Surveillance
North America needs Canada to increase surveillance. Right now, it's a
black box. We need to know if it's a single, isolated case or not. With
the way BSE works, that's not likely. Or are the ones that were exposed
dead and long gone? If there were to be more cases, how many? And very
important, if there are additional cases, how old are they? If they were
all born prior to the feed ban, that lets us know the feed ban has been very
effective. If not, we have had leaks.
This year (2003) for surveillance in Canada, they'll come to about the same
level they had last year, about 3700 brains tests. They really need to kick
this up. The recommendation by the international committee and even by
their own government is up to the level of about 40,000.
Think about the whole task of on-farm compliance. You have hundreds of
thousands of farms. How do you get 100% compliance? That's very difficult.
You can’t be on all farms at all times. My philosophy is choke infectivity
out high. If you take your infectivity out of the system at the highest
point, what leaks through on the farm won’t matter. Take your high-risk
material, your brains and spinal cords and your deads and downs, and get
them completely out of the animal food chain. If people inadvertently feed
the wrong material on the farm, it won't have the infectivity to keep the
cycle going.
It doesn’t take much to introduce BSE into a country. Identification is
extremely important. We don’t have a national ID system in place. We're
moving in that direction, but how about if tomorrow we found a case of BSE
here and we couldn't trace it back? We couldn’t stand up in front of the
American public and say, "We don’t know where this animal came from." What
would be the reaction?
Disposal itself can cause emergency situations and we need an
infrastructure—not only for TSEs but for all animal diseases. The US and
Canada both need some kind of infrastructure for proper animal disposal, for
animal health and public health reasons.
Can it happen here in the US? Would we find it? To me, we could not have
been more fortunate. We got a warning shot right in our neighbor's back
yard. We now know that before May 20, we thought we had escaped the bullet
of the agent coming to North America. May 20 showed us we were wrong. We
know the agent entered North America in the indigenous population. We now
have time to re-evaluate the risk, to look the lessons not only in Canada,
but around the world. We can do it without having this crisis upon us.
We have import regulations, feed bans, etc. We've had import regulations
in place since 1989 on live ruminants and ruminant products. This shouldn't
be down-played. This is significant. Look at the way it spread through
Europe. That was from imports of high-risk products.
Surveillance: We've looked at all the groups you're supposed to look at
for BSE in all the high-risk categories of the downers, etc. We've looked
at about 20,000 for the last two years. We should go higher. We've had the
same feed ban date as Canada. We do have a few different exceptions.
We've done formal risk assessments—one by the USDA, one by the European
Union, and one by Harvard. Harvard's assessment was the most extensive.
They looked at the potential pathways. They found that ,although US is
resistant to BSE, there is still a potential for a low animal and human
exposure. We still allow the use of high-risk material in the US—brain,
spinal cord, and advanced meat recovery. Advanced meat recovery is the
process where the bones will go through a machine to remove more muscle
tissue. The allows some potential exposure if you have BSE infectivity in
the system because the spinal column (even if you remove the spinal cord)
still has dorsal ganglia that could be incorporated into AMR. That can
still introduce infectivity into the AMR product.
Harvard found the feed ban is key to protection and that the leaks will
allow amplification, at least in pockets. Why is the risk not zero?
Science, trading patterns, existing regulations and human error all
constitute a risk. The US, like Canada, does not have dedicated facilities
or transport.
The UK imports into the US. There were 496 total, and 173 of the UK
imports could have entered the US feed system. People don't like to hear
this, but it's possible that one of the UK imports in the US entered the
animal feed system and was exported to Canada. That's a possibility,
because they import 50% of their feed from the US.
From 1994, we imported 11 million head of cattle from Canada. Most of
these were feedlot animals for slaughter, but there were about 500,000
breeding animals. A number of Canada’s cull cows were slaughtered here and
could have introduced infectivity into our system. Even today we have
Canadian imports in the country, breeding animals that were brought in prior
to the ban and reside here.
We have feed ban exemptions: plate waste, poultry litter. We still allow
that if it comes off a human plate, or if it's trimmings, it can be
palletized and fed to ruminants. That might be a small amount, but it could
allow spinal cord in certain cuts to be fed back to ruminants. Poultry
litter or feather meal could be significant. Poultry is getting quite a bit
of ruminant material in the US because it cannot go back to ruminants.
Poultry and pigs are getting a substantial amount. Poultry litter is not
only what passes through the chicken, but think about how chickens eat.
They spill a lot on the floor. That stuff is still allowed to be fed back
to cattle. That's a direct break in the ban, except that it's legal.
Ruminants are getting ruminant material.
Unfiltered tallow: tallow is a lipid material. However, if it's not
filtered, there are protein residues. That's meat and bone meal. That's
allowed to be fed, so that's another legal exception where you can feed
ruminant meat and bone meal through unfiltered tallow. We don't have an SRM
ban and the 40 animals are the ones that if you have the agent, they
introduce the most infectivity back into the animal food chain when they're
rendered.
What's our on-farm compliance? We really don't know.

Dr. Thornsberry: What about blood meal?

Dr. Detwiler: Although there's not been infectivity found in cattle blood,
the one thing we have to be concerned about is the stunning method. You can
have micro-emboli of brain material in blood, and we do allow the legal
feeding of blood meal. You could have a contamination from these emboli.
Also the brain drips into the blood.
In the past a stun gun that injected air into the cranium had been used.
It was so high pressure that it scrambled the brain, pushed it through the
foramen magnum, rippled the spinal cord and went down into the azygous
venous system. Big parts of the brain and spinal cord were being found in
the heart, the lungs, the liver. The industry has gone away from using this
stun gun, but there's no regulation that prohibits it.

Firewalls
Identification and traceability aren’t going to stop the disease, but they
are essential for locating herd cohorts and identifying other animals that
may have been exposed. OIE requires birth cohorts and progeny of a BSE
positive cow to be removed.
Canada implemented what the international scientific committee told them to
do: they're taking all these tissues out of the food supply for people. You
can't remove the dorsal ganglia, the trigeminal ganglia out of the bony
tissues so you have both the skull and veretabral column be SRM. The distal
ileum is coming out of all ages.
AMR is one thing the US needs to look at. It's a wide dissemination of
nervous tissue in the meat product, especially if the process is not done
correctly. Even if you remove the spinal cord, you still have the dorsal
ganglia associated with the vertebral column and the dura that protects the
spinal cord. It would be prudent for the government to move now on
restricting ARM.
Harvard's first model did 1000 runs and found the mean ID-50s to humans was
35. Eleven of these were from brain and spinal cord. Twenty—more than
half—were from AMR. In the recent runs with Canada, they had some where the
ID-50s were up in the hundreds and, in the worst-case scenario- over 1000.
Half of those were from AMR. So even if you remove brain and spinal cord
from the human food chain, AMR will still deliver more potential doses of
infectivity to the public than brain and spinal cord combined.

Dr. Bartz: Exactly what is AMR? Why does it increase infectivity?

Dr. Detwiler: You put the vertebral column through a machine that beats
everything off of it.

Dr. Bartz: So you’re saying that if you pull out the spinal cord, when it
goes through the ARM, it's pulling off DRGs and the dura.

Dr. Detwiler: And then that gets incorporated into ground beef and it's
widely disseminated. In the United States, food has to be labeled. You
wouldn't go to a store and buy something that had brain and spinal cord in
it. It would have to be labeled so you would know you were eating it.

Dr. Thornsberry: I don't remember the percentage, but it was over half of
all advanced meat recovery material we were manufacturing was going into the
commodity-type systems.

Dr. Detwiler: That doesn't have to be labeled AMR, so you don’t know what's
in and what's not. That could be incorporated into any mixed meat product.
The public would feel betrayed and they wouldn't know what products to avoid
because of the labeling requirements.
McDonald's has never allowed it in their product, even before BSE risk.
They continue to not allow it. I don’t know about the other fast-food
chains.

Dr. Thornsberry: It looks like a paste. It doesn't have a consistency and
it's got a lot of water in it. It's a paste of junk. You combine that and
make a sausage or a salami. The average person wouldn't get a piece of AMR
meat, but might get some specialty meat that had AMR in it.

Dr. Detwiler: If it's in ground beef, it has to be done in a smaller
percentage. It technically could, in a lower percent, go into some ground
beef.
Some of these large dairies are looking for protein sources and they mix
their own on-farm. Two practitioners in California told me their dairies
are feeding pet food. What goes into pet food? A lot of the down cows. It
never crosses their mind that this could be ruminant material.
The 4D risk is significant. In the Harvard study the model showed that if
BSE was circulating in the cattle production system, 1500 ID-50s would come
from healthy cattle at slaughter, and 37,000 from the 4D. Think about it.
If a cow dies from BSE it is at the end stage disease where the level of
infectivity is highest. If there are leaks in the feed ban, this population
re-introduce the highest level of infectivity back to your cattle
population.

Dr. Spraker: What are the four Ds?

Dr. Detwiler: Dead, dying, down, diseased. They're a lump sum of the
animals that don’t make it for human consumption.

LESSONS LEARNED

We have to be careful that the absence of evidence is not evidence of
absence, and when we say there's absolutely no scientific evidence. We need
to be very careful when we talk to the public, and even producers, to make
sure there is scientific evidence, and if there's not, to tell that to the
public. A case in point is the UK with the link to human disease. In 1989,
some experts said that there was absolutely no scientific evidence that the
disease could go to humans. That was a correct statement then, however, it
was not the entire story. What should have followed was that there is no
evidence to definitively state that BSE could not cross species. The bottom
line was that we don’t have much evidence period. We have to be careful of
predictions from TSE’s in other species, as these long incubations dieeases
are like wolves in sheep’s clothing.
Prevention with adverse action or significant cost is very difficult to
sell. I spent my whole career with APHIS trying to sell prevention—first
with Scrapie and then with BSE. If BSE didn't go to humans, I don't think
we'd ever have sold the feed ban. Now the industry is glad we did.
Failure to prevent is an easy target, but how do you measure success?
Regulations: if people believe it will protect them, you don't need much
compliance. If people aren't cognizant or don't realize how it will protect
them, you need a lot of compliance. Driving is a good example. People
believe seat belts will at least reduce fatality. You don't need a cop in
your car to wear your seatbelt. But speeding is a different story. How
many people drive the speed limit? By statistics, the higher the speed
limit, the more you increase your chance of fatality. We need to know how
much your producers and clients believe in the feed ban.
Disposal: positive solutions for one problem may create more expensive
problems. It's easy to say "ban everything," but then what do you do with
it? That's something we need to educate the public on. It's not as easy as
it appears. You create environmental problems.

Dr. Keller: Do you know what they're doing with the rendered product? We
heard at one time they were using it in the UK to make asphalt.

Dr. Detwiler: There are a lot of different things now. Some places are
combining it with concrete and asphalt because it's a binder. Most of it
gets rendered first and then the rendered material is used. They're using
it as a fuel source—the meat and bone meal and the tallow. The tallow,
apparently, burns well. Some of the rendering plants in California
converted their systems when energy prices rose and ran off their own
tallow.
There's a cement company in the US that wants to burn meat and bone meal in
their kilns. Temperatures will go up into the thousands of degrees.
Regaining scientific credibility and public confidence may require extreme
measures. That's why I think it's so important for the countries not to
lose public confidence. Japan lost it, and they've had to take extreme
measures, like testing every cow for human consumption.

Dr. Thornsberry: Two years ago, I worked as a consultant in Missouri and I
had a couple of clients who owned large swine operations. They were
cleaning out their swine feed bins and feeding it to their cows. They were
feeding for a large corporate entity, and that entity would remove a group
of hogs and they might leave two or three tons of hog feed. The hog feed
contained meat and bone meal. It wasn't a real high level. Sometimes it
was porcine meal and sometimes it was bovine, but it was in there. I got in
contact with the state veterinarian and it went up to the FDA level
concerning the feeding of this and the feeding of poultry litter. I asked
if they'd made any rulings about the feeding of poultry litter because I
knew that 5% of our feed is lost without ever being consumed. The FDA
veterinarian’s comment to me was, "Dr. Thornsberry, until there are dead
bodies lying in the street, you will not see a change made on the feeding of
poultry litter."
I said, "You know there's a potential epidemiological link between the
spread of this disease and the feeding of poultry litter, and you're telling
me you have to have dead bodies before you make a ruling on it."
He said yes. What's going on with our government?
Dr. Thornsberry: We need to put the pressure on.

Dr. Detwiler: That's right, and there are different entities that are
saying this to the government. The Food Marketing Institute, the chain
restaurants, do have a position statement they've put together that say
they'd like to see the exemptions gone. They feel there are reasons to
close the loopholes.

Dr. Thornsberry: I got a memo this summer that Canada agreed not to use AMR
from that point on, but it is not prohibited in Canada.

Dr. Detwiler: Right, and that's crucial. Vertebral column is considered a
risk material and it cannot be used. The United States has not done that.
We're testing products to see if evidence of nervous tissue can be found,
but that's not every product. It's done on a random basis by FSIS. We are
still allowing AMR.


THE STATE REGULATORY VIEWPOINT

Dr. Keller: My focus is mainly on questions I have from a state
perspective. Although I had about 10 years of veterinary practice
experience, my education, with respect to prions, began when I started
working for the State of North Dakota in ~1997. Serving as ND’s Designated
Scrapie Epidemiologist (DSE) and working with their mandatory Chronic
Wasting Disease (CWD) surveillance program, has further increased my
appreciation for regulatory challenges associated with TSEs.
Recently, state animal health officials have had the task of drafting
comments for the proposed BSE rule, which attempts to address ‘minimal risk
countries.’

Background: In ND, the North Dakota Board of Animal Health is charged with
protecting the health of domestic animals and also non-traditional
livestock. In our state, that includes everything from captive birds to
farmed cervids, to lions and tigers and bears. When in a state regulatory
position, these responsibilities are not optional, they are required of
animal health officials by statute. We are also to prevent escape and
release of animals injurious or competitive with agriculture, horticulture,
forestry, and other natural resource interests; thereby addressing potential
environmental concerns.
We're also charged with taking any steps necessary to control, suppress,
eradicate, any and all contagious and infectious diseases. Since prion
diseases are indeed considered infectious, they fall under that statute.
The State Veterinarian also has the responsibility, if warranted, to
quarantine domestic animals and non-traditional livestock. He is to
regulate and prohibit arrival or departure from our state of any animals
that may be exposed or infected with the disease. Where this could get
interesting, is when a state doesn't think the federal regulations are
protecting the state’s animal health or is not responsive enough in
addressing urgent concerns.
Also, it is very difficult trying to regulate prion diseases when you have
limited ‘official live animal tests’ available, such as are available for TB
and Brucellosis where you can get more definitive answers quickly. In that
situation, prevention is always the best policy.
The industry groups and state agencies need to have a clearer understanding
of the prion agent, its pathogenicity, and how it's transmitted so that more
useful comments can be submitted on proposed rules.

We need to be concerned about what is an ‘acceptable risk’ of having
possible disease transmission to other animals. We also cannot over look
the economic damage done by diseases. Look at what one case of BSE did to
Canada from an economic standpoint! Another area of responsibility we are
charged with, is addressing zoonotic diseases. BSE is one prion disease
that is currently considered to be zoonotic.

We have zero tolerance in this country for fecal contamination of our food,
so what are we going to determine is an ‘acceptable risk’ for BSE
contamination? And if we do allow animals in from countries where there is
a risk, what are the mitigating factors or measures that need to be taken?
It is my opinion that we need to have extensive measures in place to prevent
the introduction of the disease and to maintain the identity of animals from
other countries, even if they are only considered to be of ‘minimal risk’.
When we talk about identifying a country as minimal risk because they've had
only one case, could this then be extrapolated to other countries? We may
end up comparing apples and oranges though. For example, if we have a
country in Europe that has one case of BSE, it's not really the same because
it's on a different continent with different neighbors. We may not even be
fully familiar with the extent of their country’s animal movements or the
quality of their surveillance.

Dr. Detwiler: That's a very good point. This rule will allow other
countries to apply for minimal-risk status, and I can tell you there are
countries other than Canada that have only one case—Greece, Austria,
Finland—that have much higher surveillance levels and have had measures in
place a lot longer. So this is a reality that this situation could happen.

Dr. Keller: Those countries are watching the situation closely and may be
waiting to make their application should the proposed BSE rule be approved
as is.
So what would happen if the US found one case of BSE? In our department,
there would be immediate attention to what we would say to the producers and
to the consuming public. We would need to have factual information
available to address food safety concerns. How would we do that? We would
need to be able to say we were doing a thorough epidemiologic investigation,
and quickly get information back to consumers and our international trading
partners as it became available. Two critical actions need to be done:
country of origin labeling through maintenance of import identification of
live animal and products and identification of individual animals. Another
question that needs to be posed is which is easier and more cost effective
to do first? There is identity on animals that come into this country, but
when we've looked into that, there's no requirement for anyone to maintain
that identity. In the past, this country has relied on the use of the
brucellosis tags, auction-market back tags and brand inspection. But this
does not allow for complete tracking of animals. Especially as the
Brucellosis program winds down. There is an obvious need for individual
animal identification.
The tattoos that have been suggested as a way to track animals long-term
are not a fail-safe method. Everyone has their own idea of what a tattoo
should be, what type of ink to use, etc. The reality is, you could have a
very good tattoo but it’s not user-friendly. We'd have trouble getting
compliance if we asked feedlots to rely on running every animal through a
chute to read a tattoo. Running multiple animals down an alley and using
RFID readers to collect their information as they run by, might be a
workable system. A second form of ID will be needed in case of ID failure.
What's the international standard when we talk about BSE? OIE does not
necessarily require seven years since the last case before you can move
animals, but OIE has certain requirements for regions to qualify as
minimal-risk regions in countries where BSE was identified fewer than seven
years ago. For a minimal-risk zone or in a country, these are the things
that have to be done: It has to have been based on fewer than one case per
million during each of the last four consecutive 12-month periods, within
the cattle population that’s over 24 months. These are very specific
requirements. Whenever we talk about being consistent with OIE, we really
need to consider that, since other countries usually recognize the status
assigned by the OIE.
What are the consequences of being more lenient than international
standards? It could be used in the marketplace against us and thus lead to
a loss of consumer confidence. Other countries would be quick to speak up
if they have more stringent standards in avoiding SRMs. When we have a
case, it may become a marketing tool for another country.
When our standards are lower, we tend to be the country that would be
exposed to the movement of animals that are at risk for bringing in BSE. We
could have a ripple effect where we would have loss of consumer confidence,
loss of markets, and devastation to the US cattle industry.
If we have another country that's not using SRM and we're looking at
identifying that country as a minimal risk area, but we bring in animals
that they're not using SRMs from, and the US is still allowing use of SRMS,
will that not possibly affect consumer confidence in this country or give
our trading partners a reason to question the rationale?

Dr. Detwiler: How about the animals we have now? As I mentioned, if
breeding animals were north of the border and they went to slaughter like
the cull dairy cows, they'd have their SRMs removed to protect the Canadian
public. Below the border, those same animals do not have their SRMs
removed.

Dr. Keller: I agree. The number of Canadian birth cohorts you refer to that
have entered the US is significant and has not yet been addressed by APHIS.
There again, another country could be quick to point out the lack of
attention by our epidemiologists to that group of animals, should we ever
have a case of BSE in a US born and raised bovine. The main point is that
we need to have a standard. The list of SRMs has expanded through research,
and I think we’re hearing today that it’s possible even more SRMs may be
added. Whatever the list of SRMs is, it needs to be consistent among
countries, and the OIE needs to define what the SRMs are.
Obviously there's going to be resistance to taking additional precautions
due to costs, but the costs will be even greater and farther reaching if we
don’t. We need to be proactive on this issue, not reactive.
The feeding of feather meal is also a major concern. Does feather meal
risk increase when you’re moving animals in that are from minimal-risk
countries? I believe it would be a potential route of amplification of the
prion in our ruminant feed supply. The Harvard Risk Assessment warned
against it.
Dr. Schuler, the ND State Veterinarian discovered that in the proposed BSE
rule, where they use the terminology "designated feedlot," that it means
only the feedlot name listed on a health certificate. It does not mean that
the animals going to a designated feedlot are actually going to go to
slaughter immediately under a verified protocol. When animals from another
country go to a designated feedlot, there's nothing to prevent their tags
from being removed and animals being deviated out of the slaughter channel
they were originally intended for.

Dr. Detwiler: How did they ever identify what animals ___________________

Dr. Keller: That's a very good question, and it goes back to "what is a
designated feedlot?" and "why doesn’t the ID have to be maintained on those
animals?" Actions must be taken to address the need for maintenance of ID.

Dr. Thornsberry: We're required in the state of Nebraska to write on the
health certificate that these animals are for feeding purposes only and must
go to slaughter. But that doesn’t mean that they do.

Dr. Detwiler: A lot of the feedlots will come in and remove the ear-tags
and put their own lot tags in order to make a consistent lot, so you lose
that ID. That was a problem when the US wanted to investigate what animals
were already in feedlots. There's no regulation for that animal to retail
that ID.

Dr. Thornsberry: I thought there was, for disease control.

Dr. Detwiler: There might be for Mexican steers, but not others.

Dr. Keller: Most people are not aware that all state animal health
officials know from the health certificates, is the first point of
destination. For example, feedlot heifers might be brought in from another
country and when the feedlot manager discovers some of them are bred, they
may be pulled out to be sold as breeding animals and no one may ever know.

Environmental Contamination
We're finding out on a smaller scale with CWD and Scrapie that
environmental contamination is becoming a big concern. It's something we’ve
never had to deal with. Even with anthrax, we never had as much interest
like we’re seeing with prions. If potential environmental contamination is
a problem, then how do you safely dispose of suspect animals? It takes a
lot of time and communication with the health department and the landfill
owners in trying to get everybody to a comfort level. Legal concerns do
need to be addressed.
It is a ripple effect. You may have feedlot contamination, and then you
have all the water that ran off. In many of these large feedlots, they're
using that water run-off to irrigate fields nearby.
Unfortunately, perception often wins over science in these situations,
whether it is a real risk or not!
Here's an example of a situation we dealt with in our state. A wild deer
was shot in a state that has areas endemic with CWD. This animal was
harvested just outside of that area. We received a call that this animal
had tested positive. The brain had been pulled in that state, but the
carcass was allowed to leave with the hunter before the results were
reported. By that time, the hunter had taken the meat to a ND processor for
final processing and packaging. Then we had to tell the processing plant
that they'd had a CWD-positive animal go through their facility and we gave
them advice on a recommended cleaning process.

Dr. Bartz: What is the recommended cleaning process?

Dr. Keller: There is no ‘officially approved’ disinfectant for surface
cleaning, but strong chlorine solutions, sodium hydroxide or Environ LPH
have been recommended.

Dr. Thornsberry: Steam cleaning is part of the process.

Dr. Keller: There has been discussion about the potential approval of
Environ LPH for surface cleaning.

Dr. Detwiler: Rick Race has a publication coming out on the different
versions of the LPh. The company's supposed to come back out with the old
formulation. It's not as corrosive.

Dr. Spraker: When you use it, hold your breath.

Dr. Keller: Another concern I have that's been talked about today already
is ‘what is the infectious dose’ and how does that affect the incubation
period? We have been focusing on animals, but I know that human health
concerns usually supercede animal health concerns in our state.
What's at stake? Supposedly an estimate came out of Canada saying they
were still losing $10 million per day. Extrapolating that to the number of
cattle in our country, you could be looking at up to $50 million a day. I
think that's a very conservative estimate.
Price of human life: should we be talking money when we're talking about a
disease with so many unknowns yet about its pathogenicity and infectivity?
It appears we can’t talk about zero tolerance anymore, because prions are
here in many forms (normal and abnormal). And we will not ever make all
prion diseases just ‘go away’, but we do need to put all the effective
mitigating factors in place that we can to decrease the introduction and
amplification of prion diseases. We need to look at what we can do to
‘prevent’ and then also be ready to deal with the disease if needed.

Dr. Detwiler: I went through one of the biggest cull cow plants up in
Canada. They have to do the full SRM—the vertebral column and everything.
I asked what it cost to implement that and it's about two cents per pound.
That's really not much when you consider the ramifications.

Dr. Keller: If there are going to be costs, they usually trickle down in
the cattle business to the producer. But there's probably not a producer
who wouldn't spend a few cents to save his industry and assure his customer
about the safety of the final product. From the estimates I’ve read, I don’
t think cost is as big an issue as some would like us to believe.

Dr. Thornsberry: If the USDA decides to open the border the first of March
with no restrictions, does North Dakota have the authority to turn away
Canadian cattle?

Dr. Keller: We cannot stop animals coming through our state, but we could
prevent them from unloading in North Dakota.

Dr. Thornsberry: So some state along the border could put a stop to cattle
coming into the state to stay. The interesting thing about Missouri, where
I live, is that Canada will not allow us to export any cattle. We are a
southern state with endemic Anaplasmosis and we have potential blue tongue
virus infection in our cattle herd. They have all these restrictions based
on those two disease control programs, but yet the government is going to
force us to take cattle from Canada. Our state veterinarian is under the
impression that, if they open the border, there's no way we can restrict
them coming in. In Missouri, it's a federal rule. My understanding was
that the state veterinarian could make a ruling that because of disease
control, nothing could come in to Missouri.

Dr. Keller: I'm really not the person to answer this from a legal
perspective, but we've always been counseled that state laws could be more
restrictive than federal laws.

Dr. Detwiler: If you recall some of the conference calls where the states
had more restrictive rules, the federal government said they would challenge
those in court.

Dr. Thornsberry: California has always been able to do that for years, and
everybody seems to abide by it.

Dr. Detwiler: Our attorney says that's wrong; they can't. I've heard it
both ways. I've heard it from the federal side that yes, you can be
restrictive on some diseases, and then they say no on others.

Dr. Thornsberry: I'd like Dr. Detwiler and Dr. Keller to comment on the
Harvard risk assessment. The USDA is kind of using that as a carte blanche
for opening up the border to Canada, and I think they've already made their
mind up. Yet the Harvard risk assessment, if you get into the depth of it a
little bit, says that yes, there is an inherent risk of bringing
prion-related diseases into this country if the border is opened. Nobody
can say that risk is not there. What we’re saying is if it does come in, we
have enough safe-guards in place to contain it. That one case in Japan
created the loss of 40% of their demand for beef products overnight, and it
lasted for several months until they instituted the program to test every
animal. I'm not sure we'd have that much of a ramification. However, just
saying we can identify the case is not sufficient. Our government should be
more oriented toward preventing a case from ever coming rather than letting
us figure out how to deal with it once it gets here.
I have a meat plant in Missouri and I have a zero tolerance for Listeria
spp., for E. coli O157:H7, and Salmonella, which are sometimes fatal to
children and older people. Everyone else survives. If they find that in my
plant, I'm shut down until I can prove to USDA that it's not there. And yet
we're going to open up our borders and allow a disease to come through
that's 100% fatal if a human being should contract the disease. There's an
inconsistency in the regulations concerning these animal diseases. One of
the comments we're probably going to make to the USDA as a veterinary group,
is that the Harvard risk assessment is fine, but once one case is here and
identified, we now have potentially seeded large areas with prions. And we
know they exist forever. How should the livestock industry view the Harvard
risk assessment?

Dr. Keller: I think we need to look at the big picture. There are bits and
pieces that people quote from the HRA, but even if you look at the executive
summary of the update, there are significant things it says. It's true that
the United States is robust and resistant. If the disease is introduced, it
will go away because of the system. Other countries have demonstrated that
by feed bans. But it also says we're not totally resistant to the
introduction, and that if you look at the possible scenarios, it could
potentially be a 20-year decline of the agent. It won’t be like it's here
today and gone tomorrow. That's one thing that's very significant—to say
that you have to expect a 20-year period to eliminate the agent.
The other thing is that there are weaknesses in the system, and it's so
important that we don't have high risk SRMs being introduced into our food
supply, because if the agent is in the United States, however small there's
the perception of a potential public health risk. And, the HRA indicated
that there is definitely a potential for further transmission to animals if
there are leaks in the feed ban.
The most significant item is that, in the worst-case scenario, if the agent
comes from Canada, it's below the level of detection of the USDA's current
surveillance system.

Dr. Thornsberry: That's very significant. From an epidemiological
standpoint, it blares out. It asks, “Why would you make this decision with
the current level of surveillance that we have in this country?”

Dr. Detwiler: To me, the concern is to look at what minimum risk is.
Unless it's further defined, how do you look at all the other countries?
Some have never been evaluated. Where do they fall in the scheme of things?
Those are concerns I have. We also really need to be aware that the risk
set a precedent for the US.

Dr. Keller: We know we're at risk for having our own case of BSE here in
the US, but I don't think two wrongs make a right. We need to be proactive
in getting something done to identify animals as they come in, if they do
come in. We need to be able to track animals that are considered minimal
risk. We need to hold all countries, including ourselves, to a standard to
determine prevalency of all diseases, including prion diseases. I don’t see
why we would accept animals from other countries until that is done! It's
easy to say you're free of something if you're not adequately testing for
it. We've seen that happen with other diseases.

Dr. Detwiler: I'll play devil's advocate. If the US found a case tomorrow,
what would we ask of the world? I think that's valid to ask. It's valid to
turn the mirror back on yourself.

Dr. Thornsberry: I guarantee you that we would immediately have individual
animal ID and it wouldn’t be two years later.

Dr. Keller: And it probably would not be ID that's initially acceptable to
the industry. Producers need to realize that if they don’t get involved, it
will become mandatory and then they will no longer have the opportunity for
producer input.

Dr. Thornsberry: It's almost in the process of being mandated now. I sat
on that task force. We're just months away. July 1, 2004, we're supposed
to issue premise IDs to every farm in the nation. By the first of January,
2005, we're supposed to have some sort of mandatory identification in place,
probably electronic. Right now they have no funds to do that, nor have they
been given permission, but they say that, because of disease control, they
have the right to mandate this program now.
The other issue that comes up is the concept of country-of-origin labeling.
At the time the Canadian BSE cow was identified, I did some investigation at
Sterling, Colorado, and a few other packing plants that were taking in
Canadian beef. There were over 500,000 pounds of Canadian-bred meat out of
two or three different plants in the Colorado area that were in the food
chain and could not be identified. Once the animal was brought into the
plant and slaughtered, that carcass just went right down the chain with all
the others. There was no way to identify where that meat went. If the
government at that time had wanted to put a retention on Canadian meat,
they'd have had to retain a whole bunch of our meat at the same time. That's
been one of the strong arguments for country-of-origin labeling, at least to
maintain some ability to track that carcass past the point that it's
slaughtered. Right now we don’t have that capability. Once it receives
that USDA stamp, it's over. That's an issue that will have to come into
play for disease control concerns.
We already have, as of the first of July, a country-of-origin labeling
program for Japan and South Korea to guarantee to them that no Canadian meat
will make it into their country through our system. But our consumers don’t
have the same choice. I think that's a travesty in our industry that we can
’t offer American consumers a choice. If we can offer it to Japan's
consumers and South Korea's consumers, it seems only logical we should be
able to offer it to US consumers, but we do not.

Dr. Spraker: Does the US import a lot of meat from Argentina and South
America?

Dr. Thornsberry: They don’t right now. They do import a lot of cooked
product. They have so much foot and mouth disease. I've been in a lot of
those countries and I don't see that they'll ever have that problem under
control.

Dr. Keller: We've had many foreign exchange students at out ranch, and
several from South America. They have insinuated that there is a great deal
of unrestricted movement between South American countries.

Dr. Thornsberry: I spent time in Venezuela on a ranch with 50,000 mother
cows and they couldn’t tell you where those cows were at any one time,
period. They have computers and they keep track of things, but the level of
technology is much more primitive.

Dr. Keller: Communication indicated that political pressure kept Argentina
animal health officials from reporting the FMD case. It was actually
producers who encouraged reporting their own FMD case. I am beginning to
wonder if in this country, too, it's going to take industry and producers to
encourage our officials to do the right thing for US animal health and US
agriculture.

Dr. Thornsberry: One group of people we haven't discussed today is the
consumers in this country. Over 90% of the meat in the United States is
purchased by women. I don't know if the USDA has allowed any input from
consumer groups on this subject.

Dr. Keller: We should not needlessly scare the public, but there is a
responsibility to take additional precautions in processing if needed. When
the education and research information does catch up with the buying and
consuming public, they're going to want to know what’s been done to address
concerns regarding SRMs and AMR meat.

Dr. Spraker: With CWD, so many men will turn a head in and say they can't
bring the meat in the house because the wife wants it tested first.

Dr. Detwiler: We get asked all the time why does the public react so
volatily to this? and the only thing that I can figure is that you can't
cook it away, you can't detect it with certainty in the live animal, you can
’t test the product for it. So the consumer relies on the industry and the
government. It's an issue where the consumer lacks control. With E. coli,
they know: if you cook it right and feed it to your kids, it's going to be
okay. They have some control.

Dr. Bartz: With many infectious diseases, in two weeks, you’re over it and
you get on with your life. But if you consume BSE-potentially-tainted beef,
you're going to be worried for the rest of your life.

Dr. Thornsberry: In 1994, I spent a month in Spain, Portugal and France,
and I know I ate beef when I was there..

Dr. Detwiler: You ate mutton.

Dr. Thornsberry: I ate mutton, I ate pork, I ate pig ears. I ate a lot of
traditional Spanish and Portuguese meat, and I ate beef. And Portugal and
France have had numerous cases of BSE. So have I been exposed to BSE? My
son went with me.

Dr. Spraker: We know how many deaths occur from alcohol and we know how
many people die from chewing tobacco. But we're not afraid of those. Why
not?

Dr. Keller: It's likely because the use is intentional with alcohol and
tobacco.

Dr. Detwiler: The risk communicators will tell you that if you make the
choice and take the risk, there's a different kind of perception.

Dr. Thornsberry: This has been an excellent round-table. I know I've
learned a lot…

[tape change]

…There does not seem to be very much risk of CWD being transferred to
other species except cervids. It hasn't been seen and the chances are not
great, although with any prion disease, the risk is possible.

Dr. Detwiler: If we let it go unchecked, we increase the biological load.
I think it's prudent for the government to keep the biological load down.

Dr. Spraker: Because we don’t know if, 10 years from now, there will be an
emergence of a new strain of CWD with a totally different host range.

Dr. Thornsberry: These prions are infective. That's the one thing that
we've come to a common knowledge of here today, that's not being released by
some of our cattle organizations. They are pushing the idea that a prion
disease is an end-stage disease: an animal gets it, he dies, and that's the
end of it. Being that these prions survive in the environment basically
forever and there's no way to disinfect them, no way to destroy them, no way
to get rid of them, the potential for BSE to be infective to any variety of
animals does exist. We know that certain risk factors are much more
inherent than others, but it is an infectious disease.
I have a small feed mill and there's no telling how many people have asked
me if they can feed dog food to their show steers to increase the fat in
their diet, not understanding that most dog food has some level of meat and
bone meal in it.

Dr. Spraker: And people eat dog food and cat food.

Dr. Thornsberry: The association of dog food manufacturers has fought
against a meat and bone meal ban and they don't even want to list it in
their contents. They believe if it's listed, people will quit buying it.
You have to fill out paperwork to buy fish food with meat and bone meal, but
I could buy a whole truck load of dog food with meat and bone meal in it and
feed it to my cattle. It is illegal.
There are some SRMs that are out there in this industry. Blood meal is one
of my favorite concerns. We have a lot of big corporations that are using
beef tallow in diets to increase the fat content in dairy rations. They're
doing it as much as a pound per cow per day.

Dr. Detwiler: The thing that worries me is that those SRM’s are also in
calf rations. The calves could have more susceptibility. I worry about the
increased amount of blood going into calf rations.

Dr. Thornsberry: The concept of the prions making it through the system is
very significant. Another issue we haven't discussed is the people taking
the cleanings out of poultry barns and scattering them on their pastures for
fertilizer. The prions don't ever go away, and the cattle eat the pasture
down to the ground. How many prions are they being exposed to? There are a
whole lot of issues there to deal with. Poultry manure can be composted and
used as fuel and other things. It has alternative uses.
The swine industry has made better strides in telling people not to feed
hog food to cattle, although I know it still goes on in my area.
Fortunately, most of the swine finishing diets don't have any bone meal.
There are some things we need to do in our country to do a better job of
surveillance. The veterinary industry needs to be proactive. I hope some
of those SRMs are identified and the livestock producers are made aware of
them and we get the information out: Don’t be feeding what's left over from
your hog feed bin to your cattle.

Dr. Keller: Is there an economic analysis being done on the current
situation in Canada, that could be used as an educational tool to encourage
immediate consideration of other ways to utilize high risk product?

Dr. Detwiler: They are doing something on economic alternatives and what
the costs would be.

Dr. Thornsberry: And there are a number of them. Any fat-soluble product
has a number of possibilities and we've identified several of them today.
So here's the plan. These proceedings will be made available to the
American Association of Bovine Practitioners, the Academy of Veterinary
Consultants, to veterinary schools and whoever would like to have them. I
will also try to translate and summarize the material into some sort of
publication we can give to the livestock industry. They obviously don't
want to know all about PRPC and all those things, but there is information
in these presentations that they need to know.
You may want to add something that you didn’t say that you think might be
pertinent. A lot of discussions like this have taken place, but nothing has
been made available to the livestock industry—nothing on a practical level
that a livestock producer can understand about the potential for CWD
spreading, etc. They do not understand what BSE is. I hope, from this, we
can give our producers a firm grasp of what the disease is and also let them
know how they can participate in the process. And if we do that, we will
have accomplished what I wanted to do with this program today.


Accomplished this day, Wednesday, December 11, 2003, Denver, Colorado


Terry S. Singeltary Sr.
P.O. Box 42
Bacliff, Texas USA 77518 (May 29, 2005)


----- Original Message -----
From: "Terry S. Singeltary Sr."
To:
Sent: Thursday, May 26, 2005 10:24 PM
Subject: BSE June 9: USDA roundtable discussion on beef safety open to
public


##################### Bovine Spongiform Encephalopathy
#####################


June 9: USDA roundtable discussion on beef safety open to public

5/25/2005, 10:42 AM CDT


Agriculture Secretary Mike Johanns recently announced USDA will
hold a roundtable discussion on June 9 regarding the safety of North
American beef and the changing infrastructure of the industry.

The event will bring together USDA experts, producers, packers,
other industry groups and academia to discuss the science of BSE and the
economic impacts on the US beef industry.

The roundtable discussion, "The Safety of North American Beef
and the Economic Effect of BSE on the US Beef Industry," will be open to the
public and held on Thursday, June 9, from 9:30-2:30 at the Andrew Boss
Laboratory, University of Minnesota, St. Paul campus, St. Paul, Minnesota.

Johanns noted that data illustrating the success of USDA's
enhanced BSE surveillance program will be part of the roundtable discussion.

"It is time to clearly present the science that underlies the
safety of North American beef and examine the changing infrastructure of the
industry," Johanns said.

USDA notes that the enhanced surveillance program targets the
population of animals in which BSE is most likely to be detected. That
includes non-ambulatory or downer animals, animals exhibiting signs of a
central nervous system disorder or any other signs that could be consistent
with BSE and animals that die from unknown causes.

More than 350,000 animals have been tested under the program and
all have been negative, USDA says.


http://www.agriculture.com/ag/story.jhtml;jsessionid=BXPT15IBME4IDQFIBQNSBHQ
?storyid=/templatedata/ag/story/data/agNews_050525crBSEMTG.xml&catref=ag1040


> The event will bring together USDA experts, producers, packers, other
industry groups and academia to discuss the science of BSE and the economic
impacts on the US beef industry. <


WELL, we know how that works ;

STRICTLY PRIVATE AND CONFIDENTIAL 25, AUGUST 1995

snip...

To minimise the risk of farmers' claims for compensation from feed
compounders.

To minimise the potential damage to compound feed markets through adverse
publicity.

To maximise freedom of action for feed compounders, notably by
maintaining the availability of meat and bone meal as a raw
material in animal feeds, and ensuring time is available to make any
changes which may be required.

snip...

THE FUTURE

4..........

MAFF remains under pressure in Brussels and is not skilled at
handling potentially explosive issues.

5. Tests _may_ show that ruminant feeds have been sold which
contain illegal traces of ruminant protein. More likely, a few positive
test results will turn up but proof that a particular feed mill knowingly
supplied it to a particular farm will be difficult if not impossible.

6. The threat remains real and it will be some years before feed
compounders are free of it. The longer we can avoid any direct
linkage between feed milling _practices_ and actual BSE cases,
the more likely it is that serious damage can be avoided. ...

SEE full text ;

http://www.bseinquiry.gov.uk/files/yb/1995/08/24002001.pdf


>"It is time to clearly present the science that underlies the safety of
North American beef and examine the changing infrastructure of the
industry," Johanns said. <

>USDA notes that the enhanced surveillance program targets the population of
animals in which BSE is most likely to be detected. That includes
non-ambulatory or downer animals, animals exhibiting signs of a central
nervous system disorder or any other signs that could be consistent with BSE
and animals that die from unknown causes. <


not true;

PLEASE note, the june 2004 BSE enhanced surveillance
was meaningless and ''NOT SCIENTIFIC'' without WB.

just ask the experts ;


-------- Original Message --------
Subject: Q&A Dr. Jean-Philippe Deslys USDA REFUSAL TO USE WB ON TEXAS COW
WITH BSE SYMPTOMS (FULL TEXT)
Date: Fri, 22 Apr 2005 11:53:47 -0500
From: "Terry S. Singeltary Sr."
Reply-To: Bovine Spongiform Encephalopathy
To: BSE-L@LISTS.UNI-KARLSRUHE.DE

##################### Bovine Spongiform Encephalopathy #####################

Q&A Dr. Jean-Philippe Deslys

1. What is the standard regime for testing of suspect animals in the EU?

The regime is an initial screening by a high-output test, the Bio-Rad test.
If a result raises suspicion, a confirmatory test is conducted with the
Western blot test.

2. How long has this been the case?

Its a fairly recent development. Only recently has the Western blot test
become sensitive enough, with the addition of phospohtungstic acid
precipitation step. The Bio-Rad test (which Deslys helped develop) is
extremely sensitive, and the standard Western blot is extremely reliable
with high-signal test results. However, it had to be made more sensitive for
low-signal (samples with low density of malformed prions) samples. It has
been made more sensitive.

Reproducibility is the problem with the IHC test. It is not standardized;
depending on the lab and its protocols, or even on the technician involved
in the test, one can get conflicting results.

3. Is there a way to measure the three tests in sensitivity, accuracy and
objectivity?

Historically, yes. The IHC was the gold standard at one point, but we have
shifted to the Western blot. It requires less work, it is more sensitive and
its results are reproducible. IHC relies on localization. If you have a weak
signal case, you may get lucky and test a spot with a high concentration of
prions. But the opposite it true too; you can miss an infection by testing a
sample with low concentrations. Western blot is much better for low signal
situations.

4. The USDA in 2003 used the Western blot to confirm the BSE case in
Washington state, and it sent samples to the U.K. for independent testing.
In the case this November, which it announced was negative, it instead used
the IHC test and did not send samples to the U.K. Is this good science?

Its not logical. If you have two consecutive questionable screenings, you
do another test. I can only advise, its managements duty at USDA to make
the decisions. But when you have a discrepancy between the rapid test and
the IHC, it is only logical to confirm it with another test.

5. We are hearing now about a new strain of BSE, atypical BSE or aBSE. Or
BaSE. We have heard that IHC, the so-called gold standard, cannot detect the
variant. Is this true?

Yes. There have been a few cases, one in Italy, one in Belgium, one here in
France. It seems to only affect very old animals. The distribution in the
brain is very different than we see with BSE, it looks very different. The
IHC test will come back negative.

This his a very recent phenomenon. I have no opinion on its virulence. We do
not know where it comes from. It could be a version of sporadic infection.
Western blot caught them, but we would not even know it existed if we
werent running systematic testing in the EU.

BSE was around for a long time before we caught it and by then, it was
everywhere. It had become highly infectious. It probably amplified due to
low-temperature rendering. The disease was recycled through the food chain,
and was given time to amplify. By the time it was identified, even good
cooking couldnt eliminate it.

I cant stress enough that systematic testing is necessary. Withdrawing all
positives from the food chain is the best way to break the cycle.

What can happen with testing of only cattle that are clearly at risk is that
several can remain undetected. Canada has tested about 30,000 head of cattle
and has three positives. That would indicate that there are probably
undiscovered cases. And what happens then is that the disease is allowed to
amplify. You have to maintain testing.

When people choose to protect their economic interests over public health,
it can have a boomerang effect. It happened all through Europe. They always
deny; its not OUR problem, it is our neighbors problem. And then a single
case is discovered and the public reacts. The economic results are
devastating. It would be better to just assume BSE is present and use
systematic testing as protection. That way, the public is reassured that it
is not entering the food supply.

By systematic testing, I mean doing as we do in the EU, which is to test
every animal over 30 months of age when it is slaughtered. In Europe, three
times as many cases of BSE have been caught by systematic testing as by
clinical testing (of clearly sick animals). In 2004, eight clinical cases
were discovered, 29 were discovered at rendering plants, and 17 at
slaughter. We should be using these tests as a weapon to protect the public
and to give them assurance that the food supply is being protected.

6. USDAs list of specified risk materials excludes some products, like
blood and bone meal, that are banned in the EU and UK. Is our feed supply
safe?

With SRMs, where do you stop? Tests have found prions in meat, nerves travel
through meat, and so on. The main infectivity is in the brain and the spinal
cord. A blood and bone meal ban in animal feed is not really necessary,
because except in cases of highly infective animals, it is unlikely that
they are dangerous in themselves. If you combine systematic testing and
targeted SRM removal, the brain and the spinal column in cattle over 30
months, you can have a compromise that is both safer and less costly than
expanded feed bans.

Certainly, you can stop the spread of BSE with a total ban on offal. But it
has to be a total ban. It cant be given to sheep or swine or poultry. It
would be very expensive and virtually impossible to accomplish. You can have
farmers using the wrong feed or transportation errors.

Systematic testing makes far more sense. I think of it as a thermometer. It
not only allows us to catch the disease, it also allows us to monitor its
progress. We can watch the levels of infectivity and if they start going up
instead of down, we can take measures.

To an extent, our environment is contaminated. About 10 percent of wild
animals test positive for TSEs. If you recycle these agents, they can evolve
and get more dangerous. This is probably what happened with BSE. It wasnt
very dangerous until it evolved to the disease we know today.

People complain that testing is very expensive. It is much more expensive to
kill and test whole herds.

7. In your opinion, is infected feed the sole method of transmission of BSE,
apart from the very rare maternal transmission?

Feed is the main problem. However, we are seeing some other possibilities,
including through fat and greases. Calves are fed milk extracts, with the
cream removed. To make it nutritious, they are using fat and grease from
cattle.

(FOLLOW QUESTION: Would that allow BSE to develop into an infective level in
cattle younger than 30 months, assuming they might be getting infected at a
younger age?)

8. You were involved in a study that tested two primates who were fed
infected brain tissue. One eventually died of TSE; the other survived. The
press reported that the main finding was that it would take something on the
order of 1.5 kilograms of infected matter to create an infection, but that
seems to be an oversimplification. Could you explain it further?

The findings suggest that as little as five grams is enough to infect. The
1.5 kilo figure is the amount of infected tissue that would have to be
ingested from an animal that would be below the threshold of infection, and
would test negative. In other words, even though a younger animal may be
developing the disease, it would take a considerable amount of tissue to
transmit the disease.

An animal could be just below the testing level, and not be particularly
dangerous. But that is why you have to keep testing. Once it reaches the
threshold, it can become highly infective.

9. BSE is a pretty horrifying disease, but overall, it has killed less than
200 humans, and only a handful in recent years. Listeria, by comparison,
kills thousands every year. Overall, how do you rate the threat from BSE?


The overall risk is not particularly high. Over two million infected animals
went into the food chain in Europe, 400,000 of them before the SRMs, the
brains and spinal column, were removed from the carcass. Less than 200 died,
and less than 4,000 are at risk of developing the disease. What we know now
is that one particle is not going to kill you. There has to be condensation
of the prions to be truly dangerous.

This is not a sterile world. But the danger is that now that the crisis
appears to be over, attention will turn elsewhere and that will allow the
disease to amplify again. Just as we stopped paying attention to AIDS when
medication seemed to control it, then were surprised when a new and more
infectious and aggressive strain appeared, we could be surprised by a more
serious strain of BSE. That is why I support systematic testing for the long
term. The object is to keep levels of BSE low, and to recognize the danger
if it suddenly pops back up. ...END

TSS

######### https://listserv.kaliv.uni-karlsruhe.de/warc/bse-l.html ##########

-------- Original Message --------
Subject: Re: Q&A Dr. Jean-Philippe Deslys USDA REFUSAL TO USE WB ON TEXAS
COW WITH BSE SYMPTOMS (FULL TEXT)
Date: Fri, 22 Apr 2005 12:14:14 -0500
From: "Terry S. Singeltary Sr."
Reply-To: Bovine Spongiform Encephalopathy
To: BSE-L@LISTS.UNI-KARLSRUHE.DE
References: <42692C1B.7090200@wt.net>

##################### Bovine Spongiform Encephalopathy #####################

IN FACT, i must bring this up again.
IN TEXAS, when they are really worried about a mad cow,
when the cow is clinical and stumbling and staggering, TEXAS
does not bother TESTING the cow at all. nope, they just send
it directly to be rendered head and all to get rid of all evidence.
the june 2004 enhanced bse cover-up was just that. the USA
could test every cow that goes to slaughter, and it would be meaningless
unless properly done with the most sensitive testing to date.
but not in TEXAS or any other state in the USA.............


FDA Statement

FOR IMMEDIATE RELEASE
Statement
May 4, 2004

Media Inquiries: 301-827-6242
Consumer Inquiries: 888-INFO-FDA


Statement on Texas Cow With Central Nervous System Symptoms

On Friday, April 30 th , the Food and Drug Administration learned that a cow
with central nervous system symptoms had been killed and shipped to a
processor for rendering into animal protein for use in animal feed.

FDA, which is responsible for the safety of animal feed, immediately began
an investigation. On Friday and throughout the weekend, FDA investigators
inspected the slaughterhouse, the rendering facility, the farm where the
animal came from, and the processor that initially received the cow from the
slaughterhouse.

FDA's investigation showed that the animal in question had already been
rendered into "meat and bone meal" (a type of protein animal feed). Over the
weekend FDA was able to track down all the implicated material. That
material is being held by the firm, which is cooperating fully with FDA.

Cattle with central nervous system symptoms are of particular interest
because cattle with bovine spongiform encephalopathy or BSE, also known as
"mad cow disease," can exhibit such symptoms. In this case, there is no way
now to test for BSE. But even if the cow had BSE, FDA's animal feed rule
would prohibit the feeding of its rendered protein to other ruminant animals
(e.g., cows, goats, sheep, bison).

FDA is sending a letter to the firm summarizing its findings and informing
the firm that FDA will not object to use of this material in swine feed
only. If it is not used in swine feed, this material will be destroyed. Pigs
have been shown not to be susceptible to BSE. If the firm agrees to use the
material for swine feed only, FDA will track the material all the way
through the supply chain from the processor to the farm to ensure that the
feed is properly monitored and used only as feed for pigs.

To protect the U.S. against BSE, FDA works to keep certain mammalian protein
out of animal feed for cattle and other ruminant animals. FDA established
its animal feed rule in 1997 after the BSE epidemic in the U.K. showed that
the disease spreads by feeding infected ruminant protein to cattle.

Under the current regulation, the material from this Texas cow is not
allowed in feed for cattle or other ruminant animals. FDA's action
specifying that the material go only into swine feed means also that it will
not be fed to poultry.

FDA is committed to protecting the U.S. from BSE and collaborates closely
with the U.S. Department of Agriculture on all BSE issues. The animal feed
rule provides crucial protection against the spread of BSE, but it is only
one of several such firewalls. FDA will soon be improving the animal feed
rule, to make this strong system even stronger.

####

http://www.fda.gov/bbs/topics/news/2004/NEW01061.html

TSS

------------------------------------------------------------------------
Date
------------------------------------------------------------------------

APHIS Statement: June 29 Inconclusive BSE Test is Negative
07/02/2004

APHIS Statement: First Inconclusive BSE Test is Negative
06/30/2004

APHIS Statement Regarding Second Inconclusive BSE Test
06/29/2004

APHIS Statement Regarding First Inconclusive BSE Test
06/25/2004

Week 25
(11/1511/21)
7,900
1
Negative
0
7,901

Week 5
(6/287/4)
3,500
1
Negative
0
3,501
Week 4
(6/216/27)
3,254
1
Negative
0
3,255

USDA orders silence on mad cow in Texas

By Steve Mitchell
United Press International
Published 5/11/2004 10:16 PM

WASHINGTON, May 11 (UPI) -- The U.S. Department of Agriculture has issued an
order instructing its inspectors in Texas, where federal mad cow disease
testing policies recently were violated, not to talk about the cattle
disorder with outside parties, United Press International has learned.

The order, sent May 6 by e-mail from the USDA's Dallas district office, was
issued in the wake of the April 27 case at Lone Star Beef in San Angelo, in
which a cow displaying signs of a brain disorder was not tested for mad cow
disease despite a federal policy to screen all such animals.

The deadly illness also is known as bovine spongiform encephalopathy.

Both the USDA and its Inspector General -- amid allegations that an offsite
supervisor overruled the opinion of the inspectors onsite and made the final
decision not to test the animal -- have opened up investigations to
determine why agency policy was violated.

The order, which was obtained by UPI, was issued by Ijaz Qazi, circuit
supervisor for the USDA's Food Safety and Inspection Service's Dallas
district, which covers the entire state. It reads: "All BSE inquiries MUST
be directed to Congressional Public Affairs Phone 202-720-9113 attention Rob
Larew OR Steve Khon. This is an urgent message. Any question contact me.
Ijaz Qazi."

Although the language might sound innocuous, experienced inspectors familiar
with USDA parlance have taken to referring to the notice as a "gag order."

The National Joint Council of Food Inspection Locals -- the national
inspectors union -- considers the order a violation of inspectors' free
speech rights and is considering legal action against the USDA for breaching
the labor agreement they have with the agency.

Inspectors alleged the order also suggests the agency is concerned about its
personnel leaking damaging information about either the Texas case or the
USDA's overall mad cow disease surveillance program, which has come under
fire since the discovery of an infected cow in Washington state last
December.

"Anytime the government suppresses an individual's freedom of speech, that's
unconstitutional," Gary Dahl, president of Local 925, the Colorado
inspectors union, told UPI.

Stanley Painter, chairman of the National Joint Council, said the USDA has
sent out notices in the past stating inspectors cannot talk to reporters.

"It's an intimidation thing," Painter told UPI. Inspectors have the right to
talk to anybody about any subject, as long as they clarify they are not
speaking on behalf of the USDA and they are not doing it on government time,
he said.

USDA spokesman Steven Cohen said he was not familiar with the notice from
the Dallas office. He said he would look into it, but did not respond by
UPI's publication time. In general, Cohen said, "There's an expectation any
statement on behalf of the agency would come from the office of
communications (in Washington.)"

Asked if employees could speak freely as long as they clarified that their
views did not reflect those of the agency, Cohen said, "We'd rather that
agency policy be communicated by those in a position to speak for the
agency."

Qazi told UPI the notice was not issued in conjunction with the Texas case
and it was routine agency practice that outside inquiries be referred to the
Washington office. He said inspectors are free to talk to outside parties,
including reporters, and he did not consider the e-mail a violation of the
labor agreement with the inspectors.

Painter said the USDA's efforts to keep its employees from talking about mad
cow would be better spent "with issues like protecting the consuming public
instead of trying to hide things." He added he would "just about bet his
last nickel" agency management was attempting to suppress information about
the Texas case.

"To keep federal employees from reporting government waste, misuse of
appropriations -- those types of things -- that's not a good thing either,"
Dahl said. "If there is something wrong, let's get it out in the open --
let's get it fixed. We're working for the public, the American consumers. I
think they have the right to know this," he said.

"And believe me there's so many indicators saying that the USDA's mad cow
testing program is broken," Dahl added.

At least one member of Congress, Sen. Tom Harkin, D-Iowa, agrees.

Harkin, a long-time critic of the USDA, sent a letter to Agriculture
Secretary Ann Veneman on Monday, saying the Texas incident "calls into
question the effectiveness and reliability of USDA's current and proposed
surveillance system."

The USDA has proposed testing more than 200,000 cows -- or 10 times its
current rate -- in an expanded program scheduled to begin June 1. Harkin
wrote in the five-page letter, however, that given the realities of the
cattle industry, it is "quite doubtful" the USDA will be able to test that
many cows, particularly because it had difficulty finding 20,000 last year.

"We simply cannot tolerate a BSE testing system that fails to give valid
answers to critical questions for U.S. consumers and foreign customers,"
Harkin said in the letter, which sharply criticizes the agency's failure to
address explicitly how its new surveillance program will be implemented.

"We look forward to receiving (Harkin's) letter and having the opportunity
to review it and respond to him," USDA spokesman Ed Loyd told UPI. "USDA has
acknowledged there was a failure in not testing that cow in Texas for BSE,
so we are all working to ensure that does not occur again."

Jim Rogers, a spokesman for USDA's Animal and Plant Health Inspection
Service, which oversees the agency's mad cow surveillance program, told UPI
the agency has tested about 15,500 animals since fiscal year 2004 began, on
Oct. 1, 2003. However, the agency has refused to identify the states and
facilities from which the cows originated. Rogers said UPI would have to
seek that information through the Freedom of Information Act.

The question is central to the USDA's implementation of its expanded
surveillance program. Downer cows -- those unable to stand or walk -- made
up the bulk of the animals the agency tested for mad cow in previous years,
but these were banned from being slaughtered for human consumption in
December. This means the agency inspectors no longer can obtain brain
samples from these cows at slaughterhouses as they could in the past.

Furthermore, the USDA has not provided any evidence it has worked out
agreements with rendering facilities or ranchers, where downers and dead
cows are now most likely to be found, to obtain the extra animals for
testing.

Loyd said the agency is "working very hard to get animals on the farm that
would never show up in a processing facility," and he was "not aware of any
issues" that would delay the launch of the new program.

However, he was unable to provide the names or locations of the rendering
facilities where the agency will be obtaining cow brains for BSE testing. He
said he would look into it but did not return two follow-up phone calls from
UPI before publication.


--

Steve Mitchell is UPI's Medical Correspondent. E-mail sciencemail@upi.com

Copyright © 2001-2004 United Press International

http://www.upi.com/view.cfm?StoryID=20040511-015527-4917r


USDA did not test possible mad cows

By Steve Mitchell
United Press International
Published 6/8/2004 9:30 PM

WASHINGTON, June 8 (UPI) -- The U.S. Department of Agriculture claims it
tested 500 cows with signs of a brain disorder for mad cow disease last
year, but agency documents obtained by United Press International show the
agency tested only half that number.

USDA officials said the difference is made up in animals tested at state
veterinary diagnostic laboratories, but these animals were not tested using
the "gold standard" test employed by the agency for confirming a case of the
deadly disease. Instead, the state labs used a less sensitive test that
experts say could miss mad cow cases.

In addition, the state lab figures were not included in a March 2004 USDA
document estimating the number of animals most likely to be infected among
U.S. herds, and apparently were not given to a congressional committee that
had requested agency data on the number of cows with brain disorder signs
that had been tested for the disease.

"This is just adding to the demise of USDA's credibility," said Felicia
Nestor, senior policy adviser to the Government Accountability Project, a
group in Washington, D.C., that works with federal whistleblowers.

"If the USDA is going to exclude from testing the animals most likely to
have the disease, that would seem to have a very negative impact on the
reliability of their conclusion," Nestor told UPI.

Nestor, who has monitored the USDA's mad cow surveillance program closely
for several years, asked, "Are they deliberately avoiding testing animals
that look like they have the disease?"

Concerns about the number of cows in U.S. herds with brain disorder symptoms
have been heightened due to the recent case in Texas, in which USDA
officials failed to test an animal with such symptoms, also known as central
nervous system or CNS signs. This was a violation of USDA policy, which
stipulates all CNS cows should be tested because they are considered the
most likely to be mad cow infected. To date, the Washington cow that tested
positive last December is the only confirmed case of mad cow disease -- also
known as bovine spongiform encephalopathy -- among U.S. herds.

The Texas incident has alarmed the public and members of Congress because
humans can contract a fatal brain disorder called variant Creutzfeldt-Jakob
disease from consuming meat infected with the mad cow pathogen. If the
USDA's surveillance program is allowing the riskiest cows to go untested, it
raises concerns about the ability of the monitoring system to detect the
disease reliably in U.S. herds, Rep. Henry Waxman, D-Calif., charged in a
May 13 letter to Agriculture Secretary Ann Veneman.

Dr. Peter Lurie, of the consumer group Public Citizen, said CNS cows should
be the one category that absolutely has to be tested to have a sound
surveillance system.

"CNS animals are far and away the most important animals to test," said
Lurie, who has done several analyses of the USDA's mad cow surveillance
program.

"If there's any category that needs 100 percent testing, that's it, because
they would be the most likely place to find mad cow in America," he told
UPI. "Any CNS cow that slips into the food supply represents a major case of
malpractice by USDA, and similarly, the failure to test the brain of that
animal to see if it was indeed infected is really a failure to protect the
public."

USDA officials said the agency has no estimate on how many CNS cows occur in
U.S. herds. But spokesman Ed Loyd has told UPI, and at least one other media
outlet, that 500 CNS cows were tested in fiscal year 2003. Yet agency
testing records for the first 10 months of FY 2003, obtained by UPI under
the Freedom of Information Act, show only 254 animals that fall under the
CNS category -- or about half the number Loyd cited.

After failing to respond to repeated requests from UPI for clarification of
the apparent discrepancy, Loyd finally offered the explanation that an
additional 45 CNS cows were tested by the USDA during the final two months
of FY 2003. The remainder, he said, was made up by CNS cases tested at
various state veterinary diagnostic laboratories.

"We also include data reported to us from state veterinary diagnostic
laboratories, and all of these are CNS cases that have been tested for BSE
using a histological examination," Loyd said.

"We were not using any other labs during this period, other than (the USDA
lab), to run the IHC tests for BSE, which is the gold standard," he said.
"This (state laboratory) information contributes important data to our
surveillance effort."

However, the state labs did not use the immunohistochemistry test, which the
USDA has called the "gold standard" for diagnosing mad cow disease. Instead,
the labs used a different test called histopathology, which the USDA itself
does not use to confirm a case, opting instead for the more sensitive IHC
test.

The histopathology test, unlike the IHC test, does not detect prions --
misfolded proteins that serve as a marker for infection and can be spotted
early on in the course of the illness. Rather, it screens for the
microscopic holes in the brain that are characteristic of advanced mad cow
disease.

According to the USDA's Web site, histopathology proves reliable only if the
brain sample is removed soon after the death of the animal. If there is too
much of a delay, the Web site states, it can be "very difficult to confirm a
diagnosis by histopathology" because the brain structures may have begun to
disintegrate.

That is one reason the agency began using the IHC test -- it can confirm a
diagnosis if the brain has begun disintegrating or been frozen for shipping.

The state labs used histopathology to screen 266 CNS cases in FY 2003, as
well as 257 cases in FY 2002, according to Loyd. He did not explain why this
information was not included in the testing records the agency provided to
UPI and has not responded to requests for the identity of the state labs.

Linda Detwiler, a former USDA veterinarian who oversaw the agency's mad cow
testing program, told UPI the histopathology test probably is adequate for
screening CNS cows. If they have mad cow disease, she said, it would likely
be an advanced stage that should be obvious.

Other mad cow disease experts, however, said having a back-up test such as
IHC would be advisable, because histopathology tests sometimes can miss
evidence of infection.

The Food and Agriculture Organization of the United Nations offers similar
recommendations in its protocol for conducing a histopathology test. The
protocol states that even if histopathology is negative, "further sampling
should be undertaken" in cases "where clinical signs have strongly suggested
BSE" -- a criteria that includes all of the cows tested at the state labs.

The USDA seems to agree on the need for a back-up test. Its expanded
surveillance program, which began June 1, calls for using IHC -- or another
test called Western blot -- to confirm any positives found on rapid tests.
The March 15 document that describes the new program does not mention using
histopathology to confirm cases of mad cow disease.

"Subtle changes can be missed on histopathology that would probably not be
as easy to miss using IHC," said Elizabeth Mumford, a veterinarian and BSE
expert at Safe Food Solutions in Bern, Switzerland, a company that provides
advice on reducing mad cow risk to industry and governments.

"Therefore I believe it is valuable to run (histopathology)," Mumford told
UPI.

She noted that in Europe, two tests -- neither one the histopathology
test -- are used to ensure no cases are missed. A rapid test is used
initially for screening, followed by IHC as a confirmatory test.

Markus Moser, a molecular biologist and chief executive officer of the Swiss
firm Prionics, which manufactures tests for detecting mad cow disease,
agrees about the possibility of a case being missed by histopathology.

"There were cases which were (histopathology) negative but still clearly
positive with the other (testing) methods," Moser said. "BSE testing based
on histology on sub-optimal tissue was probably one of the reasons why
Germany was allegedly BSE-free until our test discovered that they were not"
in 2000, Moser told UPI.

He agreed with Detwiler that histopathology should be suitable for most
cases of CNS cows, but added it still can fail to detect the disease in some
CNS cases -- particularly if the sample is not optimum.

"It is difficult, if not impossible, to distinguish the subtle changes in a
diseased brain from artifacts like ruptures in the tissue due to tissue
damage during the sampling, transport or preparation," he said.

Loyd asserted the additional CNS cases from the state labs actually yielded
a total of 565 such cows the USDA had tested -- 65 more than his original
figure of 500. Whether the USDA considers its total to be 500 or 565,
however, either figure would exceed the agency's own estimates for the total
number of such cows that it identifies annually.

According to data the USDA provided to the House Committee on Government
Reform, and numbers the agency included in the March document about its
expanded surveillance plan, only 201 to 249 CNS cows are identified at
slaughterhouses. Approximately 129 additional cases occur on farms annually.
At most, that yields a combined total of 378 CNS cows, or nearly 200 less
than the 565 Loyd claims the agency tested.

The USDA surveillance plan document makes no mention of the number of CNS
animals tested at state veterinary diagnostic labs. The figure also does not
appear to be included in the agency's estimates of the number of high-risk
animals that occur in the United States each year. The latter number was
used to help the USDA calculate the number of animals it will screen for mad
cow disease in its expanded surveillance plan.

USDA officials also did not include the state lab figures in response to a
question from the House Committee on Government Reform, a source close to
the issue told UPI. The committee, on which Waxman is the ranking Democrat,
had requested in a March 8 letter to Veneman that she provide "the number of
BSE tests that were conducted on cattle exhibiting central nervous system
symptoms" for each of the last five years.

Loyd did not respond to a request from UPI asking why agency officials did
not provide that information to the committee or include it in USDA's
explanation of its expanded surveillance plan.

The committee has taken note of the CNS issue and plans to delve into it
further in a hearing slated for sometime in the next few months.

"The committee will explore this and other issues surrounding USDA and BSE
testing at a hearing later this summer," Drew Crockett, spokesman for the
committee, told UPI.

--

Steve Mitchell is UPI's Medical Correspondent. E-mail sciencemail@upi.com

Copyright © 2001-2004 United Press International

http://www.upi.com/view.cfm?StoryID=20040608-014607-3865r

-------- Original Message -------- Subject: re-BSE prions propagate as

either variant CJD-like or sporadic CJD Date: Thu, 28 Nov 2002 10:23:43

-0000 From: "Asante, Emmanuel A" To:
"'flounder@wt.net'"

Dear Terry,

I have been asked by Professor Collinge to respond to your request. I am

a Senior Scientist in the MRC Prion Unit and the lead author on the

paper. I have attached a pdf copy of the paper for your attention. Thank

you for your interest in the paper.

In respect of your first question, the simple answer is, yes. As you

will find in the paper, we have managed to associate the alternate

phenotype to type 2 PrPSc, the commonest sporadic CJD.

It is too early to be able to claim any further sub-classification in

respect of Heidenhain variant CJD or Vicky Rimmer's version. It will

take further studies, which are on-going, to establish if there are

sub-types to our initial finding which we are now reporting. The main

point of the paper is that, as well as leading to the expected new

variant CJD phenotype, BSE transmission to the 129-methionine genotype

can lead to an alternate phenotype which is indistinguishable from type

2 PrPSc.

I hope reading the paper will enlighten you more on the subject. If I

can be of any further assistance please to not hesitate to ask. Best wishes.

Emmanuel Asante

<> ____________________________________

Dr. Emmanuel A Asante MRC Prion Unit & Neurogenetics Dept. Imperial

College School of Medicine (St. Mary's) Norfolk Place, LONDON W2 1PG

Tel: +44 (0)20 7594 3794 Fax: +44 (0)20 7706 3272 email:

e.asante@ic.ac.uk (until 9/12/02)

New e-mail: e.asante@prion.ucl.ac.uk (active from now)

____________________________________

snip...

full text ;

http://www.fda.gov/ohrms/dockets/ac/03/slides/3923s1_OPH.htm


AND the new findings of BASE in cattle in Italy of Identification of a
second bovine amyloidotic spongiform encephalopathy: Molecular
similarities with sporadic

Creutzfeldt-Jakob disease


http://www.pnas.org/cgi/content/abstract/0305777101v1


Adaptation of the bovine spongiform encephalopathy agent to primates
and comparison with Creutzfeldt- Jakob disease: Implications for
human health

THE findings from Corinne Ida Lasmézas*, [dagger] , Jean-Guy Fournier*,
Virginie Nouvel*,

Hermann Boe*, Domíníque Marcé*, François Lamoury*, Nicolas Kopp [Dagger

] , Jean-Jacques Hauw§, James Ironside¶, Moira Bruce [||] , Dominique

Dormont*, and Jean-Philippe Deslys* et al, that The agent responsible
for French iatrogenic growth hormone-linked CJD taken as a control is
very different from vCJD but is similar to that found in one case of
sporadic CJD and one sheep scrapie isolate;

http://www.pnas.org/cgi/content/full/041490898v1

Characterization of two distinct prion strains
derived from bovine spongiform encephalopathy
transmissions to inbred mice

http://vir.sgmjournals.org/cgi/content/abstract/85/8/2471


ALL animals for human/animal consumption must be tested for TSE.

ALL human TSEs must be made reportable Nationally and Internationally,OF ALL
AGES...TSS


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####################

#################### https://lists.aegee.org/bse-l.html ####################




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